WorldWideScience

Sample records for theseus assembly sequence

  1. Theseus Assembly Sequence #2

    Science.gov (United States)

    1996-01-01

    Crew members are seen here assembling the tail of the Theseus prototype research aircraft at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change

  2. Theseus Assembly Sequence #3

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft being assembled at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change measurements. Dryden's Project Manager was

  3. Theseus Assembly Sequence #1

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft being assembled at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change measurements. Dryden's Project Manager was

  4. Theseus in Flight

    Science.gov (United States)

    1996-01-01

    The twin pusher propeller-driven engines of the Theseus research aircraft can be clearly seen in this photo, taken during a 1996 research flight at NASA's Dryden Flight Research Center, Edwards, California. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite

  5. Theseus Engine Being Unloaded

    Science.gov (United States)

    1996-01-01

    Crew members are seen here unloading an engine of the Theseus prototype research aircraft at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change

  6. Theseus Tail Being Unloaded

    Science.gov (United States)

    1996-01-01

    The tail of the Theseus prototype research aircraft is seen here being unloaded at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change measurements

  7. Theseus Landing Following Maiden Flight

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft shows off its high aspect-ratio wing as it comes in for a landing on Rogers Dry Lake after its first test flight from NASA's Dryden Flight Research Center, Edwards, California, on May 24, 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able

  8. Theseus First Flight - May 24, 1996

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft shows off its high aspect-ratio wing as it lifts off from Rogers Dry Lake during its first test flight from NASA's Dryden Flight Research Center, Edwards, California, on May 24, 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to

  9. Theseus on Take-off for First Flight

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft takes off for its first test flight from NASA's Dryden Flight Research Center, Edwards, California, on May 24, 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental change measurements. Dryden

  10. Theseus Waits on Lakebed for First Flight

    Science.gov (United States)

    1996-01-01

    The Theseus prototype remotely-piloted aircraft (RPA) waits on the lakebed before its first test flight from NASA's Dryden Flight Research Center, Edwards, California, on May 24, 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental

  11. Theseus Take-off from Rogers Dry Lake

    Science.gov (United States)

    1996-01-01

    The Theseus prototype research aircraft shows off its high aspect-ratio wing in this rear view of the aircraft as it takes off on its first test flight from NASA's Dryden Flight Research Center, Edwards, California, on May 24, 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able

  12. Theseus Nose and Pod Cones Being Unloaded

    Science.gov (United States)

    1996-01-01

    Crew members are seen here unloading the nose and pod cones of the Theseus prototype research aircraft at NASA's Dryden Flight Research Center, Edwards, California, in May of 1996. The Theseus aircraft, built and operated by Aurora Flight Sciences Corporation, Manassas, Virginia, was a unique aircraft flown at NASA's Dryden Flight Research Center, Edwards, California, under a cooperative agreement between NASA and Aurora. Dryden hosted the Theseus program, providing hangar space and range safety for flight testing. Aurora Flight Sciences was responsible for the actual flight testing, vehicle flight safety, and operation of the aircraft. The Theseus remotely piloted aircraft flew its maiden flight on May 24, 1996, at Dryden. During its sixth flight on November 12, 1996, Theseus experienced an in-flight structural failure that resulted in the loss of the aircraft. As of the beginning of the year 2000, Aurora had not rebuilt the aircraft. Theseus was built for NASA under an innovative, $4.9 million fixed-price contract by Aurora Flight Sciences Corporation and its partners, West Virginia University, Morgantown, West Virginia, and Fairmont State College, Fairmont, West Virginia. The twin-engine, unpiloted vehicle had a 140-foot wingspan, and was constructed largely of composite materials. Powered by two 80-horsepower, turbocharged piston engines that drove twin 9-foot-diameter propellers, Theseus was designed to fly autonomously at high altitudes, with takeoff and landing under the active control of a ground-based pilot in a ground control station 'cockpit.' With the potential ability to carry 700 pounds of science instruments to altitudes above 60,000 feet for durations of greater than 24 hours, Theseus was intended to support research in areas such as stratospheric ozone depletion and the atmospheric effects of future high-speed civil transport aircraft engines. Instruments carried aboard Theseus also would be able to validate satellite-based global environmental

  13. The Theseus Autonomous Underwater Vehicle: A Canadian Success Story

    Science.gov (United States)

    1997-04-01

    P502414.PDF [Page: 1 of 9] P502414.PDF [Page: 2 of 9] P502414.PDF [Page: 3 of 9] The Theseus Autonomous Underwater Vehicle A Canadian Success Story...autonomous underwater vehicle, named Theseus , for laying optical fiber cables in ice- covered waters. In trials and missions conducted in 1996, this...stations. An acoustic telemetry system enables communication with Theseus from surface stations, and an optical telemetry system is used for system

  14. Sequence assembly

    DEFF Research Database (Denmark)

    Scheibye-Alsing, Karsten; Hoffmann, S.; Frankel, Annett Maria

    2009-01-01

    Despite the rapidly increasing number of sequenced and re-sequenced genomes, many issues regarding the computational assembly of large-scale sequencing data have remain unresolved. Computational assembly is crucial in large genome projects as well for the evolving high-throughput technologies and...... in genomic DNA, highly expressed genes and alternative transcripts in EST sequences. We summarize existing comparisons of different assemblers and provide a detailed descriptions and directions for download of assembly programs at: http://genome.ku.dk/resources/assembly/methods.html....

  15. Theseus is shaping up. 50 MW solar thermal power plant projected on Crete; Projekt Theseus nimmt Gestalt an. Solarthermisches 50-MW-Kraftwerk auf Kreta in Planung

    Energy Technology Data Exchange (ETDEWEB)

    Bachmann, O.; Brakmann, G. [Fichtner GmbH und Co. KG, Stuttgart (Germany)

    1998-10-01

    On the Greek island of Crete, with 8331 km{sup 2}, electric power is mainly generated by combustion of oil. This will change with Theseus (Thermal Solar European Power Station). Theseus is projected on a site with 2,200 sunshine hours per annum. The power station will cost around 125 Mio. ECU. It will have a capacity of 50 MWe. An auxiliary fossil-fuel boiler will provide steam for the turbines in the evening hours and in times of low insolation. Theseus is being constructed by a consortium under German leadership (orig.) [Deutsch] Auf der 8331 km{sup 2} grossen Insel Kreta wird die elektrische Energie bisher zum ueberwiegenden Teil durch Verbrennen von Oel erzeugt. Dies soll sich mit Theseus (Thermal Solar European Power Station) aendern. Theseus wird an einem Ort, der 2.200 Stunden pro Jahr Solarbetrieb gewaehrleistet, errichtet. Das rund 135 Mio. ECU teure Kraftwerk soll 50 MWe leisten und mit einem zusaetzlichen konventionellen, fossil gefeuerten Kessel ausgeruestet werden, der in den Abendstunden und bei geringer Sonnenstrahlung Dampf fuer die Turbinen erzeugt. Fuer die Realisierung von Theseus hat sich ein unter deutscher Leitung stehendes Konsortium gebildet. (orig./AKF)

  16. THESEUS: maximum likelihood superpositioning and analysis of macromolecular structures.

    Science.gov (United States)

    Theobald, Douglas L; Wuttke, Deborah S

    2006-09-01

    THESEUS is a command line program for performing maximum likelihood (ML) superpositions and analysis of macromolecular structures. While conventional superpositioning methods use ordinary least-squares (LS) as the optimization criterion, ML superpositions provide substantially improved accuracy by down-weighting variable structural regions and by correcting for correlations among atoms. ML superpositioning is robust and insensitive to the specific atoms included in the analysis, and thus it does not require subjective pruning of selected variable atomic coordinates. Output includes both likelihood-based and frequentist statistics for accurate evaluation of the adequacy of a superposition and for reliable analysis of structural similarities and differences. THESEUS performs principal components analysis for analyzing the complex correlations found among atoms within a structural ensemble. ANSI C source code and selected binaries for various computing platforms are available under the GNU open source license from http://monkshood.colorado.edu/theseus/ or http://www.theseus3d.org.

  17. THESEUS Deliverable ID2.5

    DEFF Research Database (Denmark)

    Nørgaard, Jørgen Harck; Andersen, Thomas Lykke; Burcharth, Hans F.

    2012-01-01

    As a consequence of climate changes, coastal defenses are worldwide at higher risk of failure. Climate changes can lead to problems in terms of function, stability, and safety of goods and persons. To overcome the problems, different scenarios can be adopted, such as; 1) no upgrade actions...... and repairing the damages 2) upgrade of existing defences by e.g. changing the structure dimensions 3) perform strategic retreat and decommissioning. Within the THESEUS project, the present work task, WT 2.5, focus on upgrading existing rubble mound breakwater defences. Solutions for upgrading parts of rubble...... mound structures are investigated by means of modifying the structure profile and/or adding structure elements. Results from this study will be incorporated in the THESEUS decision tool (DSS-tool) and in the guidelines in work package 5 (WP.5)....

  18. Genome Sequence Databases (Overview): Sequencing and Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Lapidus, Alla L.

    2009-01-01

    From the date its role in heredity was discovered, DNA has been generating interest among scientists from different fields of knowledge: physicists have studied the three dimensional structure of the DNA molecule, biologists tried to decode the secrets of life hidden within these long molecules, and technologists invent and improve methods of DNA analysis. The analysis of the nucleotide sequence of DNA occupies a special place among the methods developed. Thanks to the variety of sequencing technologies available, the process of decoding the sequence of genomic DNA (or whole genome sequencing) has become robust and inexpensive. Meanwhile the assembly of whole genome sequences remains a challenging task. In addition to the need to assemble millions of DNA fragments of different length (from 35 bp (Solexa) to 800 bp (Sanger)), great interest in analysis of microbial communities (metagenomes) of different complexities raises new problems and pushes some new requirements for sequence assembly tools to the forefront. The genome assembly process can be divided into two steps: draft assembly and assembly improvement (finishing). Despite the fact that automatically performed assembly (or draft assembly) is capable of covering up to 98% of the genome, in most cases, it still contains incorrectly assembled reads. The error rate of the consensus sequence produced at this stage is about 1/2000 bp. A finished genome represents the genome assembly of much higher accuracy (with no gaps or incorrectly assembled areas) and quality ({approx}1 error/10,000 bp), validated through a number of computer and laboratory experiments.

  19. Theseus, the 50 MW solar thermal power plant; Das solarthermische 50-MW-Kraftwerk Theseus

    Energy Technology Data Exchange (ETDEWEB)

    Brakmann, G. [Fichtner GmbH und Co. KG, Stuttgart (Germany). Solarenergieprojekte

    1998-04-01

    The Isle of Crete measures 8331 km{sup 2}, and this island renowned for its historical sites attracts millions of tourists every year. Like any other branch of industry, tourism, which is called a ``white`` industry, has an ever growing demand for electric power. Up to now, electricity generation on the island is based on fossil-fuelled thermal power plants. However, recent developments indicate that this technology might be overtaken soon by the novel Theseus power plant (Thermal Solar European Power Station) currently under construction. It is expected to usher in a new era of power generation on the Isle of Crete. (orig./CB) [Deutsch] Die 8 331 km{sup 2} grosse Insel Kreta wurde vor ueber 3 500 Jahren besiedelt. Der geschichtstraechtige Ort ist ein hochgeschaetztes Reiseziel von Millionen Griechenlandurlaubern. Wie jede Art von Industrie, so benoetigt auch die als `weisse Industrie` bezeichnete Touristikbranche immer mehr elektrische Energie. Diese wird derzeit auf Kreta ausschliesslich mit thermischen Kraftwerken, welche fossile Brennstoffe verbrennen, erzeugt. Aber die Vorherrschaft dieser Technologie kann schon bald mit dem neuen solarthermischen Kraftwerk Theseus (Thermal Solar European Power Station) gebrochen werden. Es soll in wenigen Jahren eine neue Aera der Energieerzeugung auf Kreta einlaeuten. (orig.)

  20. The THESEUS space mission concept: science case, design and expected performances

    DEFF Research Database (Denmark)

    Amati, L.; O’Brien, P.; Götz, D.

    2018-01-01

    THESEUS is a space mission concept aimed at exploiting Gamma-Ray Bursts for investigating the early Universe and at providing a substantial advancement of multi-messenger and time-domain astrophysics. These goals will be achieved through a unique combination of instruments allowing GRB and X...... with both imaging and spectroscopic capabilities. THESEUS will be perfectly suited for addressing the main open issues in cosmology such as, e.g., star formation rate and metallicity evolution of the inter-stellar and intra-galactic medium up to redshift 10, signatures of Pop III stars, sources and physics...... detected in the late ’20s/early ’30s by next generation facilities like aLIGO/ aVirgo, eLISA, KAGRA, and Einstein Telescope. THESEUS will also provide powerful synergies with the next generation of multi-wavelength observatories (e.g., LSST, ELT, SKA, CTA, ATHENA)....

  1. The Transient High Energy Sky and Early Universe Surveyor (THESEUS)

    Science.gov (United States)

    Amati, Lorenzo; O'Brien, Paul T.; Götz, Diego

    2016-07-01

    The Transient High Energy Sky and Early Universe Surveyor (THESEUS) is a mission concept under development by a large international collaboration aimed at exploiting gamma-ray bursts for investigating the early Universe. The main scientific objectives of THESEUS include: investigating the star formation rate and metallicity evolution of the ISM and IGM up to redshift 9-10, detecting the first generation (pop III) of stars, studying the sources and physics of re-ionization, detecting the faint end of galaxies luminosity function. These goals will be achieved through a unique combination of instruments allowing GRB detection and arcmin localization over a broad FOV (more than 1sr) and an energy band extending from several MeVs down to 0.3 keV with unprecedented sensitivity, as well as on-board prompt (few minutes) follow-up with a 0.6m class IR telescope with both imaging and spectroscopic capabilities. Such instrumentation will also allow THESEUS to unveil and study the population of soft and sub-energetic GRBs, and, more in general, to perform monitoring and survey of the X-ray sky with unprecedented sensitivity.

  2. Targeted assembly of short sequence reads.

    Directory of Open Access Journals (Sweden)

    René L Warren

    Full Text Available As next-generation sequence (NGS production continues to increase, analysis is becoming a significant bottleneck. However, in situations where information is required only for specific sequence variants, it is not necessary to assemble or align whole genome data sets in their entirety. Rather, NGS data sets can be mined for the presence of sequence variants of interest by localized assembly, which is a faster, easier, and more accurate approach. We present TASR, a streamlined assembler that interrogates very large NGS data sets for the presence of specific variants by only considering reads within the sequence space of input target sequences provided by the user. The NGS data set is searched for reads with an exact match to all possible short words within the target sequence, and these reads are then assembled stringently to generate a consensus of the target and flanking sequence. Typically, variants of a particular locus are provided as different target sequences, and the presence of the variant in the data set being interrogated is revealed by a successful assembly outcome. However, TASR can also be used to find unknown sequences that flank a given target. We demonstrate that TASR has utility in finding or confirming genomic mutations, polymorphisms, fusions and integration events. Targeted assembly is a powerful method for interrogating large data sets for the presence of sequence variants of interest. TASR is a fast, flexible and easy to use tool for targeted assembly.

  3. The Transient High-Energy Sky and Early Universe Surveyor (THESEUS)

    Science.gov (United States)

    Amati, L.; O'Brien, P.; Goetz, D.; Tenzer, C.; Bozzo, E.

    2017-10-01

    The Transient High Energy Sky and Early Universe Surveyor (THESEUS) is a mission concept developed by a large international collaboration aimed at exploiting Gamma-Ray Bursts for investigating the early Universe. The main scientic objectives of THESEUS, currently under evaluation by ESA within the selection process for next M5 mission, include: investigating the star formation rate and metallicity evolution of the ISM and IGM up to redshift 10, detecting the first generation (pop III) of stars, studying the sources and physics of re-ionization, detecting the faint end of galaxies luminosity function. These goals will be achieved through a unique combination of instruments allowing GRB detection and arcmin localization over a broad FOV (more than 1sr) and an energy band extending from several MeVs down to 0.3 keV with unprecedented sensitivity, as well as on-board prompt (few minutes) follow-up with a 0.7m class IR telescope with both imaging and spectroscopic capabilities. Such instrumentation will also allow THESEUS to perform a monitoring of the X-ray sky with unprecedented sensitivity, which will provide a perfect service and sinergy to next generation multi-wavalength (e.g., E-ELT, SKA, CTA, ATHENA) and multi-messenger (aLIGO, aVIRGO, eLISA, ET, neutrino detectors, ...) facilities.

  4. Solar thermal power stations for activities implemented jointly. The Theseus 50 MWe solar thermal power plant for the island of Crete, Greece

    Energy Technology Data Exchange (ETDEWEB)

    Brakmann, Georg [Fichtner, Stuttgart (Germany); Aringhoff, Rainer [Pilkington Solar International (United Kingdom); Cobi, Arend [PreussenElektra (Germany)

    1998-09-01

    THESEUS, the proposed commercial 50 MWe (net) Thermal Solar European Power Station for the Island of Crete is a solar hybrid plant with parabolic trough collectors and an advanced high efficiency Rankine reheat steam cycle. At the end of 1996 the DG XVII (Energy) of the European Commission has accepted the THERMIE application of the THESEUS consortium for the design phase. THESEUS reduces the required oil imports by 28 000 t/a, thereby saving the Greek economy every year 4 million ECU in foreign currency. During its 25 years technical lifetime 2.2 million tons of CO{sub 2} emissions will be avoided. Supply, construction, erection and operation of THESEUS creates 2 000 qualified employments (man-years). Because of the high manpower intensity of solar plants and their larger capital income from interest payments in contrast to the high fuel import intensity of fossil plants, THESEUS will generate larger tax revenues for Greece and for the supplier`s countries. The investment cost of THESEUS is some 135 million ECU. Even without any subsidies this would result in electricity generation cost of some 0.085 ECY/kWh, which is lower than the current average cost from the existing power plants of Crete. (author)

  5. THESEUS - achieving maximum possible road transport tanker safety by means of experimental accident simulation; THESEUS - Tankfahrzeuge mit hoechsterreichbarer Sicherheit durch experimentelle Unfallsimulation

    Energy Technology Data Exchange (ETDEWEB)

    Rompe, K.; Heuser, G.

    1996-03-01

    In spring 1990, the Federal German Minister for Education, Science, Research and Technology (BMBF) commissioned the team from the Federal Institute for Materials Research and Testing (BAM), DEKRA, Daimler-Benz, the Federal Road Research Institute (BASt) with the University of Cologne, Ellinghaus and TUeV Rheinland (project leader) to perform the research project `THESEUS` (the acronym THESEUS comes from the German `Tankfahrzeuge mit hoechst erreichbarer Sicherheit durch experimentelle Unfallsimulation`, which translates as `achieving maximum possible road transport tanker safety by means of experimental accident simulation`). Arting from an analysis of road transport tanker accidents, crash tests and overturn tests were performed and supplemented by investigations of road transport tanker components and accompanying complex calculations of the failure processes. Parallel to this, the static side-tilt stability of road transport tankers was determined on a tilting test platform. The causes of and constructive possibilities for avoiding overturning, which is the most common cause of accidents in which hazardous materials escape, wewre analysed in dynamic driving tests with supplementary computer simulation. Various measures for improving the safety of road transport tankers were determined and subjected to a cost-benefit analysis. (orig.) [Deutsch] Das Bundesministerium fuer Bildung, Wissenschaft, Forschung und Technologie hat seit 1990 das Forschungsproject `THESEUS` (Tankfahrzeuge mit hoechst erreichbarer Sicherheit durch experimentelle Unfallsimulation) der Arbeitsgemeinschaft aus TUeV Rheinland (Federfuehrung), Bundesanstalt fuer Materialforschung und -pruefung (BAM), DEKRA, Daimler Benz, Bundesanstalt fuer Strassenwesen (BASt), Universitaet Koeln und Ellinghaus gefoerdert. Ausgehend von der detaillierten Analyse von 231 Tankfahrzeugunfaellen wurden Tankfahrzeug-Crashversuche und Tankfahrzeug-Umsturzversuche durchgefuehrt. Diese Messungen an kompletten Fahrzeugen

  6. An investigation of Hebbian phase sequences as assembly graphs

    Directory of Open Access Journals (Sweden)

    Daniel Gomes Almeida Filho

    2014-04-01

    Full Text Available Hebb proposed that synapses between neurons that fire synchronously are strengthened, forming cell assemblies and phase sequences. The former, on a shorter scale, are ensembles of synchronized cells that function transiently as a closed processing system; the latter, on a larger scale, correspond to the sequential activation of cell assemblies able to represent percepts and behaviors. Nowadays, the recording of large neuronal populations allows for the detection of multiple cell assemblies. Within Hebb’s theory, the next logical step is the analysis of phase sequences. Here we detected phase sequences as consecutive assembly activation patterns, and then analyzed their graph attributes in relation to behavior. We investigated action potentials recorded from the adult rat hippocampus and neocortex before, during and after novel object exploration (experimental periods. Within assembly graphs, each assembly corresponded to a node, and each edge corresponded to the temporal sequence of consecutive node activations. The sum of all assembly activations was proportional to firing rates, but the activity of individual assemblies was not. Assembly repertoire was stable across experimental periods, suggesting that novel experience does not create new assemblies in the adult rat. Assembly graph attributes, on the other hand, varied significantly across behavioral states and experimental periods, and were separable enough to correctly classify experimental periods (Naïve Bayes classifier; maximum AUROCs ranging from 0.55 to 0.99 and behavioral states (waking, slow wave sleep, and rapid eye movement sleep; maximum AUROCs s ranging from 0.64 to 0.98. Our findings agree with Hebb’s view that assemblies correspond to primitive building blocks of representation, nearly unchanged in the adult, while phase sequences are labile across behavioral states and change after novel experience. The results are compatible with a role for phase sequences in behavior

  7. An assembly sequence planning method based on composite algorithm

    Directory of Open Access Journals (Sweden)

    Enfu LIU

    2016-02-01

    Full Text Available To solve the combination explosion problem and the blind searching problem in assembly sequence planning of complex products, an assembly sequence planning method based on composite algorithm is proposed. In the composite algorithm, a sufficient number of feasible assembly sequences are generated using formalization reasoning algorithm as the initial population of genetic algorithm. Then fuzzy knowledge of assembly is integrated into the planning process of genetic algorithm and ant algorithm to get the accurate solution. At last, an example is conducted to verify the feasibility of composite algorithm.

  8. Theseus: tethered distributed robotics (TDR)

    Science.gov (United States)

    Digney, Bruce L.; Penzes, Steven G.

    2003-09-01

    The Defence Research and Development Canada's (DRDC) Autonomous Intelligent System's program conducts research to increase the independence and effectiveness of military vehicles and systems. DRDC-Suffield's Autonomous Land Systems (ALS) is creating new concept vehicles and autonomous control systems for use in outdoor areas, urban streets, urban interiors and urban subspaces. This paper will first give an overview of the ALS program and then give a specific description of the work being done for mobility in urban subspaces. Discussed will be the Theseus: Thethered Distributed Robotics (TDR) system, which will not only manage an unavoidable tether but exploit it for mobility and navigation. Also discussed will be the prototype robot called the Hedgehog, which uses conformal 3D mobility in ducts, sewer pipes, collapsed rubble voids and chimneys.

  9. Towards human exploration of space: the THESEUS review series on neurophysiology research priorities.

    Science.gov (United States)

    White, Olivier; Clément, Gilles; Fortrat, Jacques-Olivier; Pavy-LeTraon, Anne; Thonnard, Jean-Louis; Blanc, Stéphane; Wuyts, Floris L; Paloski, William H

    2016-01-01

    The THESEUS project (Towards Human Exploration of Space: a European Strategy), initiated within the seventh Framework Programme by the European Commission, aimed at providing a cross-cutting, life-science-based roadmap for Europe's strategy towards human exploration of long space missions, and its relevance to applications on Earth. This topic was investigated by experts in the field, in the framework of the THESEUS project whose aim was to develop an integrated life sciences research roadmap regarding human space exploration. In particular, decades of research have shown that altered gravity impairs neurological responses at large, such as perception, sleep, motor control, and cognitive factors. International experts established a list of key issues that should be addressed in that context and provided several recommendations such as a maximal exploitation of currently available resources on Earth and in space.

  10. Single molecule sequencing-guided scaffolding and correction of draft assemblies.

    Science.gov (United States)

    Zhu, Shenglong; Chen, Danny Z; Emrich, Scott J

    2017-12-06

    Although single molecule sequencing is still improving, the lengths of the generated sequences are inevitably an advantage in genome assembly. Prior work that utilizes long reads to conduct genome assembly has mostly focused on correcting sequencing errors and improving contiguity of de novo assemblies. We propose a disassembling-reassembling approach for both correcting structural errors in the draft assembly and scaffolding a target assembly based on error-corrected single molecule sequences. To achieve this goal, we formulate a maximum alternating path cover problem. We prove that this problem is NP-hard, and solve it by a 2-approximation algorithm. Our experimental results show that our approach can improve the structural correctness of target assemblies in the cost of some contiguity, even with smaller amounts of long reads. In addition, our reassembling process can also serve as a competitive scaffolder relative to well-established assembly benchmarks.

  11. Oxford Nanopore MinION Sequencing and Genome Assembly

    Directory of Open Access Journals (Sweden)

    Hengyun Lu

    2016-10-01

    Full Text Available The revolution of genome sequencing is continuing after the successful second-generation sequencing (SGS technology. The third-generation sequencing (TGS technology, led by Pacific Biosciences (PacBio, is progressing rapidly, moving from a technology once only capable of providing data for small genome analysis, or for performing targeted screening, to one that promises high quality de novo assembly and structural variation detection for human-sized genomes. In 2014, the MinION, the first commercial sequencer using nanopore technology, was released by Oxford Nanopore Technologies (ONT. MinION identifies DNA bases by measuring the changes in electrical conductivity generated as DNA strands pass through a biological pore. Its portability, affordability, and speed in data production makes it suitable for real-time applications, the release of the long read sequencer MinION has thus generated much excitement and interest in the genomics community. While de novo genome assemblies can be cheaply produced from SGS data, assembly continuity is often relatively poor, due to the limited ability of short reads to handle long repeats. Assembly quality can be greatly improved by using TGS long reads, since repetitive regions can be easily expanded into using longer sequencing lengths, despite having higher error rates at the base level. The potential of nanopore sequencing has been demonstrated by various studies in genome surveillance at locations where rapid and reliable sequencing is needed, but where resources are limited.

  12. Optimum Assembly Sequence Planning System Using Discrete Artificial Bee Colony Algorithm

    Directory of Open Access Journals (Sweden)

    Özkan Özmen

    2018-01-01

    Full Text Available Assembly refers both to the process of combining parts to create a structure and to the product resulting therefrom. The complexity of this process increases with the number of pieces in the assembly. This paper presents the assembly planning system design (APSD program, a computer program developed based on a matrix-based approach and the discrete artificial bee colony (DABC algorithm, which determines the optimum assembly sequence among numerous feasible assembly sequences (FAS. Specifically, the assembly sequences of three-dimensional (3D parts prepared in the computer-aided design (CAD software AutoCAD are first coded using the matrix-based methodology and the resulting FAS are assessed and the optimum assembly sequence is selected according to the assembly time optimisation criterion using DABC. The results of comparison of the performance of the proposed method with other methods proposed in the literature verify its superiority in finding the sequence with the lowest overall time. Further, examination of the results of application of APSD to assemblies consisting of parts in different numbers and shapes shows that it can select the optimum sequence from among hundreds of FAS.

  13. Towards human exploration of space: The THESEUS review series on immunology research priorities

    DEFF Research Database (Denmark)

    Jean-Pol, Frippiat; Crucian, Brian E; de Quervain, Dominique

    2016-01-01

    to maintain immune homeostasis under such challenges. In the framework of the THESEUS project whose aim was to develop an integrated life sciences research roadmap regarding human space exploration, experts working in the field of space immunology, and related disciplines, established a questionnaire sent...

  14. Identification of optimum sequencing depth especially for de novo genome assembly of small genomes using next generation sequencing data.

    Science.gov (United States)

    Desai, Aarti; Marwah, Veer Singh; Yadav, Akshay; Jha, Vineet; Dhaygude, Kishor; Bangar, Ujwala; Kulkarni, Vivek; Jere, Abhay

    2013-01-01

    Next Generation Sequencing (NGS) is a disruptive technology that has found widespread acceptance in the life sciences research community. The high throughput and low cost of sequencing has encouraged researchers to undertake ambitious genomic projects, especially in de novo genome sequencing. Currently, NGS systems generate sequence data as short reads and de novo genome assembly using these short reads is computationally very intensive. Due to lower cost of sequencing and higher throughput, NGS systems now provide the ability to sequence genomes at high depth. However, currently no report is available highlighting the impact of high sequence depth on genome assembly using real data sets and multiple assembly algorithms. Recently, some studies have evaluated the impact of sequence coverage, error rate and average read length on genome assembly using multiple assembly algorithms, however, these evaluations were performed using simulated datasets. One limitation of using simulated datasets is that variables such as error rates, read length and coverage which are known to impact genome assembly are carefully controlled. Hence, this study was undertaken to identify the minimum depth of sequencing required for de novo assembly for different sized genomes using graph based assembly algorithms and real datasets. Illumina reads for E.coli (4.6 MB) S.kudriavzevii (11.18 MB) and C.elegans (100 MB) were assembled using SOAPdenovo, Velvet, ABySS, Meraculous and IDBA-UD. Our analysis shows that 50X is the optimum read depth for assembling these genomes using all assemblers except Meraculous which requires 100X read depth. Moreover, our analysis shows that de novo assembly from 50X read data requires only 6-40 GB RAM depending on the genome size and assembly algorithm used. We believe that this information can be extremely valuable for researchers in designing experiments and multiplexing which will enable optimum utilization of sequencing as well as analysis resources.

  15. Plantagora: modeling whole genome sequencing and assembly of plant genomes.

    Directory of Open Access Journals (Sweden)

    Roger Barthelson

    Full Text Available BACKGROUND: Genomics studies are being revolutionized by the next generation sequencing technologies, which have made whole genome sequencing much more accessible to the average researcher. Whole genome sequencing with the new technologies is a developing art that, despite the large volumes of data that can be produced, may still fail to provide a clear and thorough map of a genome. The Plantagora project was conceived to address specifically the gap between having the technical tools for genome sequencing and knowing precisely the best way to use them. METHODOLOGY/PRINCIPAL FINDINGS: For Plantagora, a platform was created for generating simulated reads from several different plant genomes of different sizes. The resulting read files mimicked either 454 or Illumina reads, with varying paired end spacing. Thousands of datasets of reads were created, most derived from our primary model genome, rice chromosome one. All reads were assembled with different software assemblers, including Newbler, Abyss, and SOAPdenovo, and the resulting assemblies were evaluated by an extensive battery of metrics chosen for these studies. The metrics included both statistics of the assembly sequences and fidelity-related measures derived by alignment of the assemblies to the original genome source for the reads. The results were presented in a website, which includes a data graphing tool, all created to help the user compare rapidly the feasibility and effectiveness of different sequencing and assembly strategies prior to testing an approach in the lab. Some of our own conclusions regarding the different strategies were also recorded on the website. CONCLUSIONS/SIGNIFICANCE: Plantagora provides a substantial body of information for comparing different approaches to sequencing a plant genome, and some conclusions regarding some of the specific approaches. Plantagora also provides a platform of metrics and tools for studying the process of sequencing and assembly

  16. SeqLib: a C ++ API for rapid BAM manipulation, sequence alignment and sequence assembly.

    Science.gov (United States)

    Wala, Jeremiah; Beroukhim, Rameen

    2017-03-01

    We present SeqLib, a C ++ API and command line tool that provides a rapid and user-friendly interface to BAM/SAM/CRAM files, global sequence alignment operations and sequence assembly. Four C libraries perform core operations in SeqLib: HTSlib for BAM access, BWA-MEM and BLAT for sequence alignment and Fermi for error correction and sequence assembly. Benchmarking indicates that SeqLib has lower CPU and memory requirements than leading C ++ sequence analysis APIs. We demonstrate an example of how minimal SeqLib code can extract, error-correct and assemble reads from a CRAM file and then align with BWA-MEM. SeqLib also provides additional capabilities, including chromosome-aware interval queries and read plotting. Command line tools are available for performing integrated error correction, micro-assemblies and alignment. SeqLib is available on Linux and OSX for the C ++98 standard and later at github.com/walaj/SeqLib. SeqLib is released under the Apache2 license. Additional capabilities for BLAT alignment are available under the BLAT license. jwala@broadinstitue.org ; rameen@broadinstitute.org. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

  17. THESEUS 1, FERONIA and relatives: a family of cell wall-sensing receptor kinases?

    Science.gov (United States)

    Cheung, Alice Y; Wu, Hen-Ming

    2011-12-01

    The plant cell wall provides form and integrity to the cell as well as a dynamic interface between a cell and its environment. Therefore mechanisms capable of policing changes in the cell wall, signaling cellular responses including those that would feedback regulate cell wall properties are expected to play important roles in facilitating growth and ensuring survival. Discoveries in the last few years that the Arabidopsis THESEUS 1 receptor-like kinase (RLK) may function as a sensor for cell wall defects to regulate growth and that its relatives FERONIA and ANXURs regulate pollen tube integrity imply strongly that they play key roles in cell wall-related processes. Furthermore, FERONIA acts as a cell surface regulator for RAC/ROP GTPases and activates production of reactive oxygen species which are, respectively, important molecular switches and mediators for diverse processes. These findings position the THESEUS 1/FERONIA family RLKs as surface regulators and potential cell wall sensors capable of broadly and profoundly impacting cellular pathways in response to diverse signals. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. GABenchToB: a genome assembly benchmark tuned on bacteria and benchtop sequencers.

    Directory of Open Access Journals (Sweden)

    Sebastian Jünemann

    Full Text Available De novo genome assembly is the process of reconstructing a complete genomic sequence from countless small sequencing reads. Due to the complexity of this task, numerous genome assemblers have been developed to cope with different requirements and the different kinds of data provided by sequencers within the fast evolving field of next-generation sequencing technologies. In particular, the recently introduced generation of benchtop sequencers, like Illumina's MiSeq and Ion Torrent's Personal Genome Machine (PGM, popularized the easy, fast, and cheap sequencing of bacterial organisms to a broad range of academic and clinical institutions. With a strong pragmatic focus, here, we give a novel insight into the line of assembly evaluation surveys as we benchmark popular de novo genome assemblers based on bacterial data generated by benchtop sequencers. Therefore, single-library assemblies were generated, assembled, and compared to each other by metrics describing assembly contiguity and accuracy, and also by practice-oriented criteria as for instance computing time. In addition, we extensively analyzed the effect of the depth of coverage on the genome assemblies within reasonable ranges and the k-mer optimization problem of de Bruijn Graph assemblers. Our results show that, although both MiSeq and PGM allow for good genome assemblies, they require different approaches. They not only pair with different assembler types, but also affect assemblies differently regarding the depth of coverage where oversampling can become problematic. Assemblies vary greatly with respect to contiguity and accuracy but also by the requirement on the computing power. Consequently, no assembler can be rated best for all preconditions. Instead, the given kind of data, the demands on assembly quality, and the available computing infrastructure determines which assembler suits best. The data sets, scripts and all additional information needed to replicate our results are freely

  19. Solving Assembly Sequence Planning using Angle Modulated Simulated Kalman Filter

    Science.gov (United States)

    Mustapa, Ainizar; Yusof, Zulkifli Md.; Adam, Asrul; Muhammad, Badaruddin; Ibrahim, Zuwairie

    2018-03-01

    This paper presents an implementation of Simulated Kalman Filter (SKF) algorithm for optimizing an Assembly Sequence Planning (ASP) problem. The SKF search strategy contains three simple steps; predict-measure-estimate. The main objective of the ASP is to determine the sequence of component installation to shorten assembly time or save assembly costs. Initially, permutation sequence is generated to represent each agent. Each agent is then subjected to a precedence matrix constraint to produce feasible assembly sequence. Next, the Angle Modulated SKF (AMSKF) is proposed for solving ASP problem. The main idea of the angle modulated approach in solving combinatorial optimization problem is to use a function, g(x), to create a continuous signal. The performance of the proposed AMSKF is compared against previous works in solving ASP by applying BGSA, BPSO, and MSPSO. Using a case study of ASP, the results show that AMSKF outperformed all the algorithms in obtaining the best solution.

  20. Nonhybrid, finished microbial genome assemblies from long-read SMRT sequencing data.

    Science.gov (United States)

    Chin, Chen-Shan; Alexander, David H; Marks, Patrick; Klammer, Aaron A; Drake, James; Heiner, Cheryl; Clum, Alicia; Copeland, Alex; Huddleston, John; Eichler, Evan E; Turner, Stephen W; Korlach, Jonas

    2013-06-01

    We present a hierarchical genome-assembly process (HGAP) for high-quality de novo microbial genome assemblies using only a single, long-insert shotgun DNA library in conjunction with Single Molecule, Real-Time (SMRT) DNA sequencing. Our method uses the longest reads as seeds to recruit all other reads for construction of highly accurate preassembled reads through a directed acyclic graph-based consensus procedure, which we follow with assembly using off-the-shelf long-read assemblers. In contrast to hybrid approaches, HGAP does not require highly accurate raw reads for error correction. We demonstrate efficient genome assembly for several microorganisms using as few as three SMRT Cell zero-mode waveguide arrays of sequencing and for BACs using just one SMRT Cell. Long repeat regions can be successfully resolved with this workflow. We also describe a consensus algorithm that incorporates SMRT sequencing primary quality values to produce de novo genome sequence exceeding 99.999% accuracy.

  1. Tablet—next generation sequence assembly visualization

    Science.gov (United States)

    Milne, Iain; Bayer, Micha; Cardle, Linda; Shaw, Paul; Stephen, Gordon; Wright, Frank; Marshall, David

    2010-01-01

    Summary: Tablet is a lightweight, high-performance graphical viewer for next-generation sequence assemblies and alignments. Supporting a range of input assembly formats, Tablet provides high-quality visualizations showing data in packed or stacked views, allowing instant access and navigation to any region of interest, and whole contig overviews and data summaries. Tablet is both multi-core aware and memory efficient, allowing it to handle assemblies containing millions of reads, even on a 32-bit desktop machine. Availability: Tablet is freely available for Microsoft Windows, Apple Mac OS X, Linux and Solaris. Fully bundled installers can be downloaded from http://bioinf.scri.ac.uk/tablet in 32- and 64-bit versions. Contact: tablet@scri.ac.uk PMID:19965881

  2. A Case Study into Microbial Genome Assembly Gap Sequences and Finishing Strategies.

    Science.gov (United States)

    Utturkar, Sagar M; Klingeman, Dawn M; Hurt, Richard A; Brown, Steven D

    2017-01-01

    This study characterized regions of DNA which remained unassembled by either PacBio and Illumina sequencing technologies for seven bacterial genomes. Two genomes were manually finished using bioinformatics and PCR/Sanger sequencing approaches and regions not assembled by automated software were analyzed. Gaps present within Illumina assemblies mostly correspond to repetitive DNA regions such as multiple rRNA operon sequences. PacBio gap sequences were evaluated for several properties such as GC content, read coverage, gap length, ability to form strong secondary structures, and corresponding annotations. Our hypothesis that strong secondary DNA structures blocked DNA polymerases and contributed to gap sequences was not accepted. PacBio assemblies had few limitations overall and gaps were explained as cumulative effect of lower than average sequence coverage and repetitive sequences at contig termini. An important aspect of the present study is the compilation of biological features that interfered with assembly and included active transposons, multiple plasmid sequences, phage DNA integration, and large sequence duplication. Our targeted genome finishing approach and systematic evaluation of the unassembled DNA will be useful for others looking to close, finish, and polish microbial genome sequences.

  3. Automatic Enhancement of the Reference Set for Multi-Criteria Sorting in The Frame of Theseus Method

    Directory of Open Access Journals (Sweden)

    Fernandez Eduardo

    2014-05-01

    Full Text Available Some recent works have established the importance of handling abundant reference information in multi-criteria sorting problems. More valid information allows a better characterization of the agent’s assignment policy, which can lead to an improved decision support. However, sometimes information for enhancing the reference set may be not available, or may be too expensive. This paper explores an automatic mode of enhancing the reference set in the framework of the THESEUS multi-criteria sorting method. Some performance measures are defined in order to test results of the enhancement. Several theoretical arguments and practical experiments are provided here, supporting a basic advantage of the automatic enhancement: a reduction of the vagueness measure that improves the THESEUS accuracy, without additional efforts from the decision agent. The experiments suggest that the errors coming from inadequate automatic assignments can be kept at a manageable level.

  4. INTEGRATION OF SHIP HULL ASSEMBLY SEQUENCE PLANNING, SCHEDULING AND BUDGETING

    Directory of Open Access Journals (Sweden)

    Remigiusz Romuald Iwańkowicz

    2015-02-01

    Full Text Available The specificity of the yard work requires the particularly careful treatment of the issues of scheduling and budgeting in the production planning processes. The article presents the method of analysis of the assembly sequence taking into account the duration of individual activities and the demand for resources. A method of the critical path and resource budgeting were used. Modelling of the assembly was performed using the acyclic graphs. It has been shown that the assembly sequences can have very different feasible budget regions. The proposed model is applied to the assembly processes of large-scale welded structures, including the hulls of ships. The presented computational examples have a simulation character. They show the usefulness of the model and the possibility to use it in a variety of analyses.

  5. De novo assembly of human genomes with massively parallel short read sequencing

    DEFF Research Database (Denmark)

    Li, Ruiqiang; Zhu, Hongmei; Ruan, Jue

    2010-01-01

    genomes from short read sequences. We successfully assembled both the Asian and African human genome sequences, achieving an N50 contig size of 7.4 and 5.9 kilobases (kb) and scaffold of 446.3 and 61.9 kb, respectively. The development of this de novo short read assembly method creates new opportunities...... for building reference sequences and carrying out accurate analyses of unexplored genomes in a cost-effective way....

  6. Continuity and Discontinuity, Change and Duration: Hobbes' Riddle of the Theseus and the Diversity of Historical Logics.

    Science.gov (United States)

    Blum, Mark E.

    1996-01-01

    Reiterates the need for an understanding of the concepts of continuity and change, not simply in the representation of historical events, but in the writing and study of history. Uses Thomas Hobbes's riddle of Theseus to illustrate the need for multiple readings and critical analysis in history instruction. (MJP)

  7. Fast and simple protein-alignment-guided assembly of orthologous gene families from microbiome sequencing reads.

    Science.gov (United States)

    Huson, Daniel H; Tappu, Rewati; Bazinet, Adam L; Xie, Chao; Cummings, Michael P; Nieselt, Kay; Williams, Rohan

    2017-01-25

    Microbiome sequencing projects typically collect tens of millions of short reads per sample. Depending on the goals of the project, the short reads can either be subjected to direct sequence analysis or be assembled into longer contigs. The assembly of whole genomes from metagenomic sequencing reads is a very difficult problem. However, for some questions, only specific genes of interest need to be assembled. This is then a gene-centric assembly where the goal is to assemble reads into contigs for a family of orthologous genes. We present a new method for performing gene-centric assembly, called protein-alignment-guided assembly, and provide an implementation in our metagenome analysis tool MEGAN. Genes are assembled on the fly, based on the alignment of all reads against a protein reference database such as NCBI-nr. Specifically, the user selects a gene family based on a classification such as KEGG and all reads binned to that gene family are assembled. Using published synthetic community metagenome sequencing reads and a set of 41 gene families, we show that the performance of this approach compares favorably with that of full-featured assemblers and that of a recently published HMM-based gene-centric assembler, both in terms of the number of reference genes detected and of the percentage of reference sequence covered. Protein-alignment-guided assembly of orthologous gene families complements whole-metagenome assembly in a new and very useful way.

  8. Reducing assembly complexity of microbial genomes with single-molecule sequencing

    Science.gov (United States)

    Genome assembly algorithms cannot fully reconstruct microbial chromosomes from the DNA reads output by first or second-generation sequencing instruments. Therefore, most genomes are left unfinished due to the significant resources required to manually close gaps left in the draft assemblies. Single-...

  9. Graph mining for next generation sequencing: leveraging the assembly graph for biological insights.

    Science.gov (United States)

    Warnke-Sommer, Julia; Ali, Hesham

    2016-05-06

    The assembly of Next Generation Sequencing (NGS) reads remains a challenging task. This is especially true for the assembly of metagenomics data that originate from environmental samples potentially containing hundreds to thousands of unique species. The principle objective of current assembly tools is to assemble NGS reads into contiguous stretches of sequence called contigs while maximizing for both accuracy and contig length. The end goal of this process is to produce longer contigs with the major focus being on assembly only. Sequence read assembly is an aggregative process, during which read overlap relationship information is lost as reads are merged into longer sequences or contigs. The assembly graph is information rich and capable of capturing the genomic architecture of an input read data set. We have developed a novel hybrid graph in which nodes represent sequence regions at different levels of granularity. This model, utilized in the assembly and analysis pipeline Focus, presents a concise yet feature rich view of a given input data set, allowing for the extraction of biologically relevant graph structures for graph mining purposes. Focus was used to create hybrid graphs to model metagenomics data sets obtained from the gut microbiomes of five individuals with Crohn's disease and eight healthy individuals. Repetitive and mobile genetic elements are found to be associated with hybrid graph structure. Using graph mining techniques, a comparative study of the Crohn's disease and healthy data sets was conducted with focus on antibiotics resistance genes associated with transposase genes. Results demonstrated significant differences in the phylogenetic distribution of categories of antibiotics resistance genes in the healthy and diseased patients. Focus was also evaluated as a pure assembly tool and produced excellent results when compared against the Meta-velvet, Omega, and UD-IDBA assemblers. Mining the hybrid graph can reveal biological phenomena captured

  10. The fast changing landscape of sequencing technologies and their impact on microbial genome assemblies and annotation.

    Science.gov (United States)

    Mavromatis, Konstantinos; Land, Miriam L; Brettin, Thomas S; Quest, Daniel J; Copeland, Alex; Clum, Alicia; Goodwin, Lynne; Woyke, Tanja; Lapidus, Alla; Klenk, Hans Peter; Cottingham, Robert W; Kyrpides, Nikos C

    2012-01-01

    The emergence of next generation sequencing (NGS) has provided the means for rapid and high throughput sequencing and data generation at low cost, while concomitantly creating a new set of challenges. The number of available assembled microbial genomes continues to grow rapidly and their quality reflects the quality of the sequencing technology used, but also of the analysis software employed for assembly and annotation. In this work, we have explored the quality of the microbial draft genomes across various sequencing technologies. We have compared the draft and finished assemblies of 133 microbial genomes sequenced at the Department of Energy-Joint Genome Institute and finished at the Los Alamos National Laboratory using a variety of combinations of sequencing technologies, reflecting the transition of the institute from Sanger-based sequencing platforms to NGS platforms. The quality of the public assemblies and of the associated gene annotations was evaluated using various metrics. Results obtained with the different sequencing technologies, as well as their effects on downstream processes, were analyzed. Our results demonstrate that the Illumina HiSeq 2000 sequencing system, the primary sequencing technology currently used for de novo genome sequencing and assembly at JGI, has various advantages in terms of total sequence throughput and cost, but it also introduces challenges for the downstream analyses. In all cases assembly results although on average are of high quality, need to be viewed critically and consider sources of errors in them prior to analysis. These data follow the evolution of microbial sequencing and downstream processing at the JGI from draft genome sequences with large gaps corresponding to missing genes of significant biological role to assemblies with multiple small gaps (Illumina) and finally to assemblies that generate almost complete genomes (Illumina+PacBio).

  11. Optimizing and benchmarking de novo transcriptome sequencing: from library preparation to assembly evaluation.

    Science.gov (United States)

    Hara, Yuichiro; Tatsumi, Kaori; Yoshida, Michio; Kajikawa, Eriko; Kiyonari, Hiroshi; Kuraku, Shigehiro

    2015-11-18

    RNA-seq enables gene expression profiling in selected spatiotemporal windows and yields massive sequence information with relatively low cost and time investment, even for non-model species. However, there remains a large room for optimizing its workflow, in order to take full advantage of continuously developing sequencing capacity. Transcriptome sequencing for three embryonic stages of Madagascar ground gecko (Paroedura picta) was performed with the Illumina platform. The output reads were assembled de novo for reconstructing transcript sequences. In order to evaluate the completeness of transcriptome assemblies, we prepared a reference gene set consisting of vertebrate one-to-one orthologs. To take advantage of increased read length of >150 nt, we demonstrated shortened RNA fragmentation time, which resulted in a dramatic shift of insert size distribution. To evaluate products of multiple de novo assembly runs incorporating reads with different RNA sources, read lengths, and insert sizes, we introduce a new reference gene set, core vertebrate genes (CVG), consisting of 233 genes that are shared as one-to-one orthologs by all vertebrate genomes examined (29 species)., The completeness assessment performed by the computational pipelines CEGMA and BUSCO referring to CVG, demonstrated higher accuracy and resolution than with the gene set previously established for this purpose. As a result of the assessment with CVG, we have derived the most comprehensive transcript sequence set of the Madagascar ground gecko by means of assembling individual libraries followed by clustering the assembled sequences based on their overall similarities. Our results provide several insights into optimizing de novo RNA-seq workflow, including the coordination between library insert size and read length, which manifested in improved connectivity of assemblies. The approach and assembly assessment with CVG demonstrated here would be applicable to transcriptome analysis of other species as

  12. Sequencing and De Novo Transcriptome Assembly of Brachypodium sylvaticum (Poaceae

    Directory of Open Access Journals (Sweden)

    Samuel E. Fox

    2013-03-01

    Full Text Available Premise of the study: We report the de novo assembly and characterization of the transcriptomes of Brachypodium sylvaticum (slender false-brome accessions from native populations of Spain and Greece, and an invasive population west of Corvallis, Oregon, USA. Methods and Results: More than 350 million sequence reads from the mRNA libraries prepared from three B. sylvaticum genotypes were assembled into 120,091 (Corvallis, 104,950 (Spain, and 177,682 (Greece transcript contigs. In comparison with the B. distachyon Bd21 reference genome and GenBank protein sequences, we estimate >90% exome coverage for B. sylvaticum. The transcripts were assigned Gene Ontology and InterPro annotations. Brachypodium sylvaticum sequence reads aligned against the Bd21 genome revealed 394,654 single-nucleotide polymorphisms (SNPs and >20,000 simple sequence repeat (SSR DNA sites. Conclusions: To our knowledge, this is the first report of transcriptome sequencing of invasive plant species with a closely related sequenced reference genome. The sequences and identified SNP variant and SSR sites will provide tools for developing novel genetic markers for use in genotyping and characterization of invasive behavior of B. sylvaticum.

  13. New tool to assemble repetitive regions using next-generation sequencing data

    Science.gov (United States)

    Kuśmirek, Wiktor; Nowak, Robert M.; Neumann, Łukasz

    2017-08-01

    The next generation sequencing techniques produce a large amount of sequencing data. Some part of the genome are composed of repetitive DNA sequences, which are very problematic for the existing genome assemblers. We propose a modification of the algorithm for a DNA assembly, which uses the relative frequency of reads to properly reconstruct repetitive sequences. The new approach was implemented and tested, as a demonstration of the capability of our software we present some results for model organisms. The new implementation, using a three-layer software architecture was selected, where the presentation layer, data processing layer, and data storage layer were kept separate. Source code as well as demo application with web interface and the additional data are available at project web-page: http://dnaasm.sourceforge.net.

  14. The sequence and de novo assembly of the giant panda genome

    Science.gov (United States)

    Li, Ruiqiang; Fan, Wei; Tian, Geng; Zhu, Hongmei; He, Lin; Cai, Jing; Huang, Quanfei; Cai, Qingle; Li, Bo; Bai, Yinqi; Zhang, Zhihe; Zhang, Yaping; Wang, Wen; Li, Jun; Wei, Fuwen; Li, Heng; Jian, Min; Li, Jianwen; Zhang, Zhaolei; Nielsen, Rasmus; Li, Dawei; Gu, Wanjun; Yang, Zhentao; Xuan, Zhaoling; Ryder, Oliver A.; Leung, Frederick Chi-Ching; Zhou, Yan; Cao, Jianjun; Sun, Xiao; Fu, Yonggui; Fang, Xiaodong; Guo, Xiaosen; Wang, Bo; Hou, Rong; Shen, Fujun; Mu, Bo; Ni, Peixiang; Lin, Runmao; Qian, Wubin; Wang, Guodong; Yu, Chang; Nie, Wenhui; Wang, Jinhuan; Wu, Zhigang; Liang, Huiqing; Min, Jiumeng; Wu, Qi; Cheng, Shifeng; Ruan, Jue; Wang, Mingwei; Shi, Zhongbin; Wen, Ming; Liu, Binghang; Ren, Xiaoli; Zheng, Huisong; Dong, Dong; Cook, Kathleen; Shan, Gao; Zhang, Hao; Kosiol, Carolin; Xie, Xueying; Lu, Zuhong; Zheng, Hancheng; Li, Yingrui; Steiner, Cynthia C.; Lam, Tommy Tsan-Yuk; Lin, Siyuan; Zhang, Qinghui; Li, Guoqing; Tian, Jing; Gong, Timing; Liu, Hongde; Zhang, Dejin; Fang, Lin; Ye, Chen; Zhang, Juanbin; Hu, Wenbo; Xu, Anlong; Ren, Yuanyuan; Zhang, Guojie; Bruford, Michael W.; Li, Qibin; Ma, Lijia; Guo, Yiran; An, Na; Hu, Yujie; Zheng, Yang; Shi, Yongyong; Li, Zhiqiang; Liu, Qing; Chen, Yanling; Zhao, Jing; Qu, Ning; Zhao, Shancen; Tian, Feng; Wang, Xiaoling; Wang, Haiyin; Xu, Lizhi; Liu, Xiao; Vinar, Tomas; Wang, Yajun; Lam, Tak-Wah; Yiu, Siu-Ming; Liu, Shiping; Zhang, Hemin; Li, Desheng; Huang, Yan; Wang, Xia; Yang, Guohua; Jiang, Zhi; Wang, Junyi; Qin, Nan; Li, Li; Li, Jingxiang; Bolund, Lars; Kristiansen, Karsten; Wong, Gane Ka-Shu; Olson, Maynard; Zhang, Xiuqing; Li, Songgang; Yang, Huanming; Wang, Jian; Wang, Jun

    2013-01-01

    Using next-generation sequencing technology alone, we have successfully generated and assembled a draft sequence of the giant panda genome. The assembled contigs (2.25 gigabases (Gb)) cover approximately 94% of the whole genome, and the remaining gaps (0.05 Gb) seem to contain carnivore-specific repeats and tandem repeats. Comparisons with the dog and human showed that the panda genome has a lower divergence rate. The assessment of panda genes potentially underlying some of its unique traits indicated that its bamboo diet might be more dependent on its gut microbiome than its own genetic composition. We also identified more than 2.7 million heterozygous single nucleotide polymorphisms in the diploid genome. Our data and analyses provide a foundation for promoting mammalian genetic research, and demonstrate the feasibility for using next-generation sequencing technologies for accurate, cost-effective and rapid de novo assembly of large eukaryotic genomes. PMID:20010809

  15. Syntactic sequencing in Hebbian cell assemblies.

    Science.gov (United States)

    Wennekers, Thomas; Palm, Günther

    2009-12-01

    Hebbian cell assemblies provide a theoretical framework for the modeling of cognitive processes that grounds them in the underlying physiological neural circuits. Recently we have presented an extension of cell assemblies by operational components which allows to model aspects of language, rules, and complex behaviour. In the present work we study the generation of syntactic sequences using operational cell assemblies timed by unspecific trigger signals. Syntactic patterns are implemented in terms of hetero-associative transition graphs in attractor networks which cause a directed flow of activity through the neural state space. We provide regimes for parameters that enable an unspecific excitatory control signal to switch reliably between attractors in accordance with the implemented syntactic rules. If several target attractors are possible in a given state, noise in the system in conjunction with a winner-takes-all mechanism can randomly choose a target. Disambiguation can also be guided by context signals or specific additional external signals. Given a permanently elevated level of external excitation the model can enter an autonomous mode, where it generates temporal grammatical patterns continuously.

  16. Long-read sequencing and de novo assembly of a Chinese genome

    Science.gov (United States)

    Short-read sequencing has enabled the de novo assembly of several individual human genomes, but with inherent limitations in characterizing repeat elements. Here we sequence a Chinese individual HX1 by single-molecule real-time (SMRT) long-read sequencing, construct a physical map by NanoChannel arr...

  17. Norgal: extraction and de novo assembly of mitochondrial DNA from whole-genome sequencing data.

    Science.gov (United States)

    Al-Nakeeb, Kosai; Petersen, Thomas Nordahl; Sicheritz-Pontén, Thomas

    2017-11-21

    Whole-genome sequencing (WGS) projects provide short read nucleotide sequences from nuclear and possibly organelle DNA depending on the source of origin. Mitochondrial DNA is present in animals and fungi, while plants contain DNA from both mitochondria and chloroplasts. Current techniques for separating organelle reads from nuclear reads in WGS data require full reference or partial seed sequences for assembling. Norgal (de Novo ORGAneLle extractor) avoids this requirement by identifying a high frequency subset of k-mers that are predominantly of mitochondrial origin and performing a de novo assembly on a subset of reads that contains these k-mers. The method was applied to WGS data from a panda, brown algae seaweed, butterfly and filamentous fungus. We were able to extract full circular mitochondrial genomes and obtained sequence identities to the reference sequences in the range from 98.5 to 99.5%. We also assembled the chloroplasts of grape vines and cucumbers using Norgal together with seed-based de novo assemblers. Norgal is a pipeline that can extract and assemble full or partial mitochondrial and chloroplast genomes from WGS short reads without prior knowledge. The program is available at: https://bitbucket.org/kosaidtu/norgal .

  18. A practical comparison of de novo genome assembly software tools for next-generation sequencing technologies.

    Directory of Open Access Journals (Sweden)

    Wenyu Zhang

    Full Text Available The advent of next-generation sequencing technologies is accompanied with the development of many whole-genome sequence assembly methods and software, especially for de novo fragment assembly. Due to the poor knowledge about the applicability and performance of these software tools, choosing a befitting assembler becomes a tough task. Here, we provide the information of adaptivity for each program, then above all, compare the performance of eight distinct tools against eight groups of simulated datasets from Solexa sequencing platform. Considering the computational time, maximum random access memory (RAM occupancy, assembly accuracy and integrity, our study indicate that string-based assemblers, overlap-layout-consensus (OLC assemblers are well-suited for very short reads and longer reads of small genomes respectively. For large datasets of more than hundred millions of short reads, De Bruijn graph-based assemblers would be more appropriate. In terms of software implementation, string-based assemblers are superior to graph-based ones, of which SOAPdenovo is complex for the creation of configuration file. Our comparison study will assist researchers in selecting a well-suited assembler and offer essential information for the improvement of existing assemblers or the developing of novel assemblers.

  19. NxRepair: error correction in de novo sequence assembly using Nextera mate pairs

    Directory of Open Access Journals (Sweden)

    Rebecca R. Murphy

    2015-06-01

    Full Text Available Scaffolding errors and incorrect repeat disambiguation during de novo assembly can result in large scale misassemblies in draft genomes. Nextera mate pair sequencing data provide additional information to resolve assembly ambiguities during scaffolding. Here, we introduce NxRepair, an open source toolkit for error correction in de novo assemblies that uses Nextera mate pair libraries to identify and correct large-scale errors. We show that NxRepair can identify and correct large scaffolding errors, without use of a reference sequence, resulting in quantitative improvements in the assembly quality. NxRepair can be downloaded from GitHub or PyPI, the Python Package Index; a tutorial and user documentation are also available.

  20. Virtual Genome Walking across the 32 Gb Ambystoma mexicanum genome; assembling gene models and intronic sequence.

    Science.gov (United States)

    Evans, Teri; Johnson, Andrew D; Loose, Matthew

    2018-01-12

    Large repeat rich genomes present challenges for assembly using short read technologies. The 32 Gb axolotl genome is estimated to contain ~19 Gb of repetitive DNA making an assembly from short reads alone effectively impossible. Indeed, this model species has been sequenced to 20× coverage but the reads could not be conventionally assembled. Using an alternative strategy, we have assembled subsets of these reads into scaffolds describing over 19,000 gene models. We call this method Virtual Genome Walking as it locally assembles whole genome reads based on a reference transcriptome, identifying exons and iteratively extending them into surrounding genomic sequence. These assemblies are then linked and refined to generate gene models including upstream and downstream genomic, and intronic, sequence. Our assemblies are validated by comparison with previously published axolotl bacterial artificial chromosome (BAC) sequences. Our analyses of axolotl intron length, intron-exon structure, repeat content and synteny provide novel insights into the genic structure of this model species. This resource will enable new experimental approaches in axolotl, such as ChIP-Seq and CRISPR and aid in future whole genome sequencing efforts. The assembled sequences and annotations presented here are freely available for download from https://tinyurl.com/y8gydc6n . The software pipeline is available from https://github.com/LooseLab/iterassemble .

  1. Harvesting Statistical Metadata from an Online Repository for Data Analysis and Visualization : Concept Application on Theseus

    OpenAIRE

    Gebresilassie, Sem

    2014-01-01

    Theses and publications from Finnish universities of applied sciences are accessible from an open online repository called Theseus. This repository has an application programming interface (API) that provides tools for harvesting its contents. By properly utilizing this API, it is possible to gather and reuse metadata of thesis documents for any other objective. This thesis mainly intends to explain how to gather the author name, title, submission year, keywords, subjects, department, u...

  2. Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

    DEFF Research Database (Denmark)

    Maretty, Lasse; Jensen, Jacob Malte; Petersen, Bent

    2017-01-01

    Hundreds of thousands of human genomes are now being sequenced to characterize genetic variation and use this information to augment association mapping studies of complex disorders and other phenotypic traits. Genetic variation is identified mainly by mapping short reads to the reference genome......-coverage sequencing with mate-pair libraries extending up to 20 kilobases. We report de novo assemblies of 150 individuals (50 trios) from the GenomeDenmark project. The quality of these assemblies is similar to those obtained using the more expensive long-read technology. We use the assemblies to identify a rich set...... or by performing local assembly. However, these approaches are biased against discovery of structural variants and variation in the more complex parts of the genome. Hence, large-scale de novo assembly is needed. Here we show that it is possible to construct excellent de novo assemblies from high...

  3. Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

    DEFF Research Database (Denmark)

    Maretty, Lasse; Jensen, Jacob Malte; Petersen, Bent

    2017-01-01

    Hundreds of thousands of human genomes are now being sequenced to characterize genetic variation and use this information to augment association mapping studies of complex disorders and other phenotypic traits. Genetic variation is identified mainly by mapping short reads to the reference genome...... or by performing local assembly. However, these approaches are biased against discovery of structural variants and variation in the more complex parts of the genome. Hence, large-scale de novo assembly is needed. Here we show that it is possible to construct excellent de novo assemblies from high......-coverage sequencing with mate-pair libraries extending up to 20 kilobases. We report de novo assemblies of 150 individuals (50 trios) from the GenomeDenmark project. The quality of these assemblies is similar to those obtained using the more expensive long-read technology. We use the assemblies to identify a rich set...

  4. The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads.

    Science.gov (United States)

    Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo; Zhu, Shilin; Shi, Daihu; McDill, Joshua; Yang, Linfeng; Hawkins, Simon; Neutelings, Godfrey; Datla, Raju; Lambert, Georgina; Galbraith, David W; Grassa, Christopher J; Geraldes, Armando; Cronk, Quentin C; Cullis, Christopher; Dash, Prasanta K; Kumar, Polumetla A; Cloutier, Sylvie; Sharpe, Andrew G; Wong, Gane K-S; Wang, Jun; Deyholos, Michael K

    2012-11-01

    Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp to 10 kb were sequenced using an Illumina genome analyzer. A de novo assembly, comprised exclusively of deep-coverage (approximately 94× raw, approximately 69× filtered) short-sequence reads (44-100 bp), produced a set of scaffolds with N(50) =694 kb, including contigs with N(50)=20.1 kb. The contig assembly contained 302 Mb of non-redundant sequence representing an estimated 81% genome coverage. Up to 96% of published flax ESTs aligned to the whole-genome shotgun scaffolds. However, comparisons with independently sequenced BACs and fosmids showed some mis-assembly of regions at the genome scale. A total of 43384 protein-coding genes were predicted in the whole-genome shotgun assembly, and up to 93% of published flax ESTs, and 86% of A. thaliana genes aligned to these predicted genes, indicating excellent coverage and accuracy at the gene level. Analysis of the synonymous substitution rates (K(s) ) observed within duplicate gene pairs was consistent with a recent (5-9 MYA) whole-genome duplication in flax. Within the predicted proteome, we observed enrichment of many conserved domains (Pfam-A) that may contribute to the unique properties of this crop, including agglutinin proteins. Together these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

  5. IDBA-UD: a de novo assembler for single-cell and metagenomic sequencing data with highly uneven depth.

    Science.gov (United States)

    Peng, Yu; Leung, Henry C M; Yiu, S M; Chin, Francis Y L

    2012-06-01

    Next-generation sequencing allows us to sequence reads from a microbial environment using single-cell sequencing or metagenomic sequencing technologies. However, both technologies suffer from the problem that sequencing depth of different regions of a genome or genomes from different species are highly uneven. Most existing genome assemblers usually have an assumption that sequencing depths are even. These assemblers fail to construct correct long contigs. We introduce the IDBA-UD algorithm that is based on the de Bruijn graph approach for assembling reads from single-cell sequencing or metagenomic sequencing technologies with uneven sequencing depths. Several non-trivial techniques have been employed to tackle the problems. Instead of using a simple threshold, we use multiple depthrelative thresholds to remove erroneous k-mers in both low-depth and high-depth regions. The technique of local assembly with paired-end information is used to solve the branch problem of low-depth short repeat regions. To speed up the process, an error correction step is conducted to correct reads of high-depth regions that can be aligned to highconfident contigs. Comparison of the performances of IDBA-UD and existing assemblers (Velvet, Velvet-SC, SOAPdenovo and Meta-IDBA) for different datasets, shows that IDBA-UD can reconstruct longer contigs with higher accuracy. The IDBA-UD toolkit is available at our website http://www.cs.hku.hk/~alse/idba_ud

  6. Programming molecular self-assembly of intrinsically disordered proteins containing sequences of low complexity

    Science.gov (United States)

    Simon, Joseph R.; Carroll, Nick J.; Rubinstein, Michael; Chilkoti, Ashutosh; López, Gabriel P.

    2017-06-01

    Dynamic protein-rich intracellular structures that contain phase-separated intrinsically disordered proteins (IDPs) composed of sequences of low complexity (SLC) have been shown to serve a variety of important cellular functions, which include signalling, compartmentalization and stabilization. However, our understanding of these structures and our ability to synthesize models of them have been limited. We present design rules for IDPs possessing SLCs that phase separate into diverse assemblies within droplet microenvironments. Using theoretical analyses, we interpret the phase behaviour of archetypal IDP sequences and demonstrate the rational design of a vast library of multicomponent protein-rich structures that ranges from uniform nano-, meso- and microscale puncta (distinct protein droplets) to multilayered orthogonally phase-separated granular structures. The ability to predict and program IDP-rich assemblies in this fashion offers new insights into (1) genetic-to-molecular-to-macroscale relationships that encode hierarchical IDP assemblies, (2) design rules of such assemblies in cell biology and (3) molecular-level engineering of self-assembled recombinant IDP-rich materials.

  7. PNA Directed Sequence Addressed Self-Assembly of DNA Nanostructures

    DEFF Research Database (Denmark)

    Nielsen, Peter E.

    2008-01-01

    sequence specifically recognize another PNA oligomer. We describe how such three domain PNAs have utility for assembling dsDNA grid and clover leaf structures, and in combination with SNAP-tag technol. of protein dsDNA structures. (c) 2008 American Institute of Physics. [on SciFinder (R)] Udgivelsesdato...

  8. Comprehensive evaluation of non-hybrid genome assembly tools for third-generation PacBio long-read sequence data.

    Science.gov (United States)

    Jayakumar, Vasanthan; Sakakibara, Yasubumi

    2017-11-03

    Long reads obtained from third-generation sequencing platforms can help overcome the long-standing challenge of the de novo assembly of sequences for the genomic analysis of non-model eukaryotic organisms. Numerous long-read-aided de novo assemblies have been published recently, which exhibited superior quality of the assembled genomes in comparison with those achieved using earlier second-generation sequencing technologies. Evaluating assemblies is important in guiding the appropriate choice for specific research needs. In this study, we evaluated 10 long-read assemblers using a variety of metrics on Pacific Biosciences (PacBio) data sets from different taxonomic categories with considerable differences in genome size. The results allowed us to narrow down the list to a few assemblers that can be effectively applied to eukaryotic assembly projects. Moreover, we highlight how best to use limited genomic resources for effectively evaluating the genome assemblies of non-model organisms. © The Author 2017. Published by Oxford University Press.

  9. A holistic evaluation of risks in coastal regions under changing climatic, environmental and socioeconomic conditions: the Theseus Decision Support System.

    Science.gov (United States)

    Losada, I. J.; Garcia Alonso, E.; Mendez, F. J.; Zanuttigh, B.; Nicholls, R. J.; Thompson, R.; Vanderlinden, J. P.; Fernandez, F.; Ondiviela, B.; Diaz-Simal, P.; Bagli, S.

    2012-04-01

    There is a general acceptance that global changes associated with natural hazards and socioeconomic processes are occurring at a faster pace than ever, with deep implications in terms of risk exposure and environmental impact. The capacity of coastal areas to adapt and react to these changes will be a key factor in the future preservation of life standards and represents a great challenge for politicians, scientists and professionals at any level. Within the large scope of Theseus Project (EU 7th Framework Program), one of the main objectives is to design a tool to help decision makers in defining optimal strategies to minimize risks within a certain city or coastal area in a three-fold sense: economic losses, human damages and environmental impacts. The resulting software, the Theseus-DSS, links the most relevant physical processes (waves, sea-levels, hard and soft structures, coastal erosion and inland flooding) with the potential impact zones (marine and inland), considering their functions (ecosystems) and uses (economic units), and the dependence of this functions and uses upon the prevailing physical conditions. The new software tries to fill a gap among the existing tools, based on the following pillars: • Seamless integration of disciplines: physics, engineering, ecology, social sciences and economy. • Intermediate spatial scales (1- 10 km) and medium-to- long time spans (1-10 years). • Decision-making based on a balance between deterministic models and expert, discussion-based assumptions. The user of the Theseus-DSS will be able either to check the consequences of predefined scenarios at a particular study site, or to create user-defined scenarios, run them and compare the results with other scenarios. The results are expressed, locally and at an aggregate level, in the three aforementioned dimensions: economic losses (€/year), mean annual expected live losses (persons/year) and impact on habitats (null, low, medium and high).

  10. Colloidal polymers with controlled sequence and branching constructed from magnetic field assembled nanoparticles.

    Science.gov (United States)

    Bannwarth, Markus B; Utech, Stefanie; Ebert, Sandro; Weitz, David A; Crespy, Daniel; Landfester, Katharina

    2015-03-24

    The assembly of nanoparticles into polymer-like architectures is challenging and usually requires highly defined colloidal building blocks. Here, we show that the broad size-distribution of a simple dispersion of magnetic nanocolloids can be exploited to obtain various polymer-like architectures. The particles are assembled under an external magnetic field and permanently linked by thermal sintering. The remarkable variety of polymer-analogue architectures that arises from this simple process ranges from statistical and block copolymer-like sequencing to branched chains and networks. This library of architectures can be realized by controlling the sequencing of the particles and the junction points via a size-dependent self-assembly of the single building blocks.

  11. Genome-wide SNP identification by high-throughput sequencing and selective mapping allows sequence assembly positioning using a framework genetic linkage map

    Directory of Open Access Journals (Sweden)

    Xu Xiangming

    2010-12-01

    Full Text Available Abstract Background Determining the position and order of contigs and scaffolds from a genome assembly within an organism's genome remains a technical challenge in a majority of sequencing projects. In order to exploit contemporary technologies for DNA sequencing, we developed a strategy for whole genome single nucleotide polymorphism sequencing allowing the positioning of sequence contigs onto a linkage map using the bin mapping method. Results The strategy was tested on a draft genome of the fungal pathogen Venturia inaequalis, the causal agent of apple scab, and further validated using sequence contigs derived from the diploid plant genome Fragaria vesca. Using our novel method we were able to anchor 70% and 92% of sequences assemblies for V. inaequalis and F. vesca, respectively, to genetic linkage maps. Conclusions We demonstrated the utility of this approach by accurately determining the bin map positions of the majority of the large sequence contigs from each genome sequence and validated our method by mapping single sequence repeat markers derived from sequence contigs on a full mapping population.

  12. Two-Stage orders sequencing system for mixed-model assembly

    Science.gov (United States)

    Zemczak, M.; Skolud, B.; Krenczyk, D.

    2015-11-01

    In the paper, the authors focus on the NP-hard problem of orders sequencing, formulated similarly to Car Sequencing Problem (CSP). The object of the research is the assembly line in an automotive industry company, on which few different models of products, each in a certain number of versions, are assembled on the shared resources, set in a line. Such production type is usually determined as a mixed-model production, and arose from the necessity of manufacturing customized products on the basis of very specific orders from single clients. The producers are nowadays obliged to provide each client the possibility to determine a huge amount of the features of the product they are willing to buy, as the competition in the automotive market is large. Due to the previously mentioned nature of the problem (NP-hard), in the given time period only satisfactory solutions are sought, as the optimal solution method has not yet been found. Most of the researchers that implemented inaccurate methods (e.g. evolutionary algorithms) to solving sequencing problems dropped the research after testing phase, as they were not able to obtain reproducible results, and met problems while determining the quality of the received solutions. Therefore a new approach to solving the problem, presented in this paper as a sequencing system is being developed. The sequencing system consists of a set of determined rules, implemented into computer environment. The system itself works in two stages. First of them is connected with the determination of a place in the storage buffer to which certain production orders should be sent. In the second stage of functioning, precise sets of sequences are determined and evaluated for certain parts of the storage buffer under certain criteria.

  13. Handling the Multiplicity of Solutions in a Moea Based PDA-THESEUS Framework for Multi-Criteria Sorting

    Directory of Open Access Journals (Sweden)

    Covantes Edgar

    2016-11-01

    Full Text Available This paper proposes the combination of the THESEUS multi-criteria sorting method with an evolutionary optimization-based preference-disaggregation analysis. The main features of the combined method are studied by performing an extensive computer experiment that explores many models of preferences and sizes of problems as well as different degrees of decision-maker involvement. As a result of the experiment, the effectiveness of the combined framework and the importance of the decision-maker’s involvement are characterized.

  14. Moleculo Long-Read Sequencing Facilitates Assembly and Genomic Binning from Complex Soil Metagenomes

    Energy Technology Data Exchange (ETDEWEB)

    White, Richard Allen; Bottos, Eric M.; Roy Chowdhury, Taniya; Zucker, Jeremy D.; Brislawn, Colin J.; Nicora, Carrie D.; Fansler, Sarah J.; Glaesemann, Kurt R.; Glass, Kevin; Jansson, Janet K.; Langille, Morgan

    2016-06-28

    ABSTRACT

    Soil metagenomics has been touted as the “grand challenge” for metagenomics, as the high microbial diversity and spatial heterogeneity of soils make them unamenable to current assembly platforms. Here, we aimed to improve soil metagenomic sequence assembly by applying the Moleculo synthetic long-read sequencing technology. In total, we obtained 267 Gbp of raw sequence data from a native prairie soil; these data included 109.7 Gbp of short-read data (~100 bp) from the Joint Genome Institute (JGI), an additional 87.7 Gbp of rapid-mode read data (~250 bp), plus 69.6 Gbp (>1.5 kbp) from Moleculo sequencing. The Moleculo data alone yielded over 5,600 reads of >10 kbp in length, and over 95% of the unassembled reads mapped to contigs of >1.5 kbp. Hybrid assembly of all data resulted in more than 10,000 contigs over 10 kbp in length. We mapped three replicate metatranscriptomes derived from the same parent soil to the Moleculo subassembly and found that 95% of the predicted genes, based on their assignments to Enzyme Commission (EC) numbers, were expressed. The Moleculo subassembly also enabled binning of >100 microbial genome bins. We obtained via direct binning the first complete genome, that of “CandidatusPseudomonas sp. strain JKJ-1” from a native soil metagenome. By mapping metatranscriptome sequence reads back to the bins, we found that several bins corresponding to low-relative-abundanceAcidobacteriawere highly transcriptionally active, whereas bins corresponding to high-relative-abundanceVerrucomicrobiawere not. These results demonstrate that Moleculo sequencing provides a significant advance for resolving complex soil microbial communities.

    IMPORTANCESoil microorganisms carry out key processes for life on our planet, including cycling of carbon and other nutrients and supporting growth of plants. However, there is poor molecular-level understanding of their

  15. BioNano genome mapping of individual chromosomes supports physical mapping and sequence assembly in complex plant genomes.

    Science.gov (United States)

    Staňková, Helena; Hastie, Alex R; Chan, Saki; Vrána, Jan; Tulpová, Zuzana; Kubaláková, Marie; Visendi, Paul; Hayashi, Satomi; Luo, Mingcheng; Batley, Jacqueline; Edwards, David; Doležel, Jaroslav; Šimková, Hana

    2016-07-01

    The assembly of a reference genome sequence of bread wheat is challenging due to its specific features such as the genome size of 17 Gbp, polyploid nature and prevalence of repetitive sequences. BAC-by-BAC sequencing based on chromosomal physical maps, adopted by the International Wheat Genome Sequencing Consortium as the key strategy, reduces problems caused by the genome complexity and polyploidy, but the repeat content still hampers the sequence assembly. Availability of a high-resolution genomic map to guide sequence scaffolding and validate physical map and sequence assemblies would be highly beneficial to obtaining an accurate and complete genome sequence. Here, we chose the short arm of chromosome 7D (7DS) as a model to demonstrate for the first time that it is possible to couple chromosome flow sorting with genome mapping in nanochannel arrays and create a de novo genome map of a wheat chromosome. We constructed a high-resolution chromosome map composed of 371 contigs with an N50 of 1.3 Mb. Long DNA molecules achieved by our approach facilitated chromosome-scale analysis of repetitive sequences and revealed a ~800-kb array of tandem repeats intractable to current DNA sequencing technologies. Anchoring 7DS sequence assemblies obtained by clone-by-clone sequencing to the 7DS genome map provided a valuable tool to improve the BAC-contig physical map and validate sequence assembly on a chromosome-arm scale. Our results indicate that creating genome maps for the whole wheat genome in a chromosome-by-chromosome manner is feasible and that they will be an affordable tool to support the production of improved pseudomolecules. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  16. The THESEUS project -- 50 MWe solar thermal power for Crete

    Energy Technology Data Exchange (ETDEWEB)

    Schillig, F.; Geyer, M.; Kistner, R.; Aringhoff, R.; Nava, P.; Brakmann, G.

    1998-07-01

    A consortium of European industry, utilities and research institutions from Greece, Germany, Spain and Italy attempts to implement a 52 MWe solar thermal power plant with parabolic trough technology on the Greek island of Crete sponsored by the EU' s THERMIE program. The increased demand for electricity on the island, a consequence of the growing allurement of the island as a tourist resort, makes it necessary to expand the installed capacity on Crete during the next years. According to the capacity expansion plans of Greek' s utility PPC a 160 MWe heavy fuel-fired power plant complex--two 30 MWe diesel units and two 50 MWe steam turbine units--is foreseen to be built by the year 2002. In this paper a description of the technical, economical and environmental aspects of the THESEUS project is provided. Moreover a market entry strategy for solar thermal power generation is discussed.

  17. Draft Sequencing of the Heterozygous Diploid Genome of Satsuma (Citrus unshiu Marc. Using a Hybrid Assembly Approach

    Directory of Open Access Journals (Sweden)

    Tokurou Shimizu

    2017-12-01

    Full Text Available Satsuma (Citrus unshiu Marc. is one of the most abundantly produced mandarin varieties of citrus, known for its seedless fruit production and as a breeding parent of citrus. De novo assembly of the heterozygous diploid genome of Satsuma (“Miyagawa Wase” was conducted by a hybrid assembly approach using short-read sequences, three mate-pair libraries, and a long-read sequence of PacBio by the PLATANUS assembler. The assembled sequence, with a total size of 359.7 Mb at the N50 length of 386,404 bp, consisted of 20,876 scaffolds. Pseudomolecules of Satsuma constructed by aligning the scaffolds to three genetic maps showed genome-wide synteny to the genomes of Clementine, pummelo, and sweet orange. Gene prediction by modeling with MAKER-P proposed 29,024 genes and 37,970 mRNA; additionally, gene prediction analysis found candidates for novel genes in several biosynthesis pathways for gibberellin and violaxanthin catabolism. BUSCO scores for the assembled scaffold and predicted transcripts, and another analysis by BAC end sequence mapping indicated the assembled genome consistency was close to those of the haploid Clementine, pummel, and sweet orange genomes. The number of repeat elements and long terminal repeat retrotransposon were comparable to those of the seven citrus genomes; this suggested no significant failure in the assembly at the repeat region. A resequencing application using the assembled sequence confirmed that both kunenbo-A and Satsuma are offsprings of Kishu, and Satsuma is a back-crossed offspring of Kishu. These results illustrated the performance of the hybrid assembly approach and its ability to construct an accurate heterozygous diploid genome.

  18. Multi-objective Analysis for a Sequencing Planning of Mixed-model Assembly Line

    Science.gov (United States)

    Shimizu, Yoshiaki; Waki, Toshiya; Yoo, Jae Kyu

    Diversified customer demands are raising importance of just-in-time and agile manufacturing much more than before. Accordingly, introduction of mixed-model assembly lines becomes popular to realize the small-lot-multi-kinds production. Since it produces various kinds on the same assembly line, a rational management is of special importance. With this point of view, this study focuses on a sequencing problem of mixed-model assembly line including a paint line as its preceding process. By taking into account the paint line together, reducing work-in-process (WIP) inventory between these heterogeneous lines becomes a major concern of the sequencing problem besides improving production efficiency. Finally, we have formulated the sequencing problem as a bi-objective optimization problem to prevent various line stoppages, and to reduce the volume of WIP inventory simultaneously. Then we have proposed a practical method for the multi-objective analysis. For this purpose, we applied the weighting method to derive the Pareto front. Actually, the resulting problem is solved by a meta-heuristic method like SA (Simulated Annealing). Through numerical experiments, we verified the validity of the proposed approach, and discussed the significance of trade-off analysis between the conflicting objectives.

  19. Cost-effective sequencing of full-length cDNA clones powered by a de novo-reference hybrid assembly.

    Science.gov (United States)

    Kuroshu, Reginaldo M; Watanabe, Junichi; Sugano, Sumio; Morishita, Shinichi; Suzuki, Yutaka; Kasahara, Masahiro

    2010-05-07

    Sequencing full-length cDNA clones is important to determine gene structures including alternative splice forms, and provides valuable resources for experimental analyses to reveal the biological functions of coded proteins. However, previous approaches for sequencing cDNA clones were expensive or time-consuming, and therefore, a fast and efficient sequencing approach was demanded. We developed a program, MuSICA 2, that assembles millions of short (36-nucleotide) reads collected from a single flow cell lane of Illumina Genome Analyzer to shotgun-sequence approximately 800 human full-length cDNA clones. MuSICA 2 performs a hybrid assembly in which an external de novo assembler is run first and the result is then improved by reference alignment of shotgun reads. We compared the MuSICA 2 assembly with 200 pooled full-length cDNA clones finished independently by the conventional primer-walking using Sanger sequencers. The exon-intron structure of the coding sequence was correct for more than 95% of the clones with coding sequence annotation when we excluded cDNA clones insufficiently represented in the shotgun library due to PCR failure (42 out of 200 clones excluded), and the nucleotide-level accuracy of coding sequences of those correct clones was over 99.99%. We also applied MuSICA 2 to full-length cDNA clones from Toxoplasma gondii, to confirm that its ability was competent even for non-human species. The entire sequencing and shotgun assembly takes less than 1 week and the consumables cost only approximately US$3 per clone, demonstrating a significant advantage over previous approaches.

  20. THESEUS: the first-scale 50 MEw THErmal Solar EUropean power station for the Island of Crete, Greece

    Energy Technology Data Exchange (ETDEWEB)

    Aringhof, Rainer [Pilkington Solar International (United States)

    1996-03-01

    The first solar power plant to receive funding from the European Union is to be constructed on the Island of Crete, Greece. This utility-scale 50 MWe plant is based on technology known as THERMIE (THErmal Solar European power station) and it represents a major breakthrough for Europe`s small solar thermal power community. The proposed THESEUS project uses an advanced parabolic trough collector field as the primary heat source and will be constructed between 1977 and 1999. (UK)

  1. Improving transcriptome assembly through error correction of high-throughput sequence reads

    Directory of Open Access Journals (Sweden)

    Matthew D. MacManes

    2013-07-01

    Full Text Available The study of functional genomics, particularly in non-model organisms, has been dramatically improved over the last few years by the use of transcriptomes and RNAseq. While these studies are potentially extremely powerful, a computationally intensive procedure, the de novo construction of a reference transcriptome must be completed as a prerequisite to further analyses. The accurate reference is critically important as all downstream steps, including estimating transcript abundance are critically dependent on the construction of an accurate reference. Though a substantial amount of research has been done on assembly, only recently have the pre-assembly procedures been studied in detail. Specifically, several stand-alone error correction modules have been reported on and, while they have shown to be effective in reducing errors at the level of sequencing reads, how error correction impacts assembly accuracy is largely unknown. Here, we show via use of a simulated and empiric dataset, that applying error correction to sequencing reads has significant positive effects on assembly accuracy, and should be applied to all datasets. A complete collection of commands which will allow for the production of Reptile corrected reads is available at https://github.com/macmanes/error_correction/tree/master/scripts and as File S1.

  2. 2D nanomaterials assembled from sequence-defined molecules

    International Nuclear Information System (INIS)

    Mu, Peng; State University of New York; Zhou, Guangwen; Chen, Chun-Long

    2017-01-01

    Two dimensional (2D) nanomaterials have attracted broad interest owing to their unique physical and chemical properties with potential applications in electronics, chemistry, biology, medicine and pharmaceutics. Due to the current limitations of traditional 2D nanomaterials (e.g., graphene and graphene oxide) in tuning surface chemistry and compositions, 2D nanomaterials assembled from sequence-defined molecules (e.g., DNAs, proteins, peptides and peptoids) have recently been developed. They represent an emerging class of 2D nanomaterials with attractive physical and chemical properties. Here, we summarize the recent progress in the synthesis and applications of this type of sequence-defined 2D nanomaterials. We also discuss the challenges and opportunities in this new field.

  3. High-coverage sequencing and annotated assembly of the genome of the Australian dragon lizard Pogona vitticeps.

    Science.gov (United States)

    Georges, Arthur; Li, Qiye; Lian, Jinmin; O'Meally, Denis; Deakin, Janine; Wang, Zongji; Zhang, Pei; Fujita, Matthew; Patel, Hardip R; Holleley, Clare E; Zhou, Yang; Zhang, Xiuwen; Matsubara, Kazumi; Waters, Paul; Graves, Jennifer A Marshall; Sarre, Stephen D; Zhang, Guojie

    2015-01-01

    The lizards of the family Agamidae are one of the most prominent elements of the Australian reptile fauna. Here, we present a genomic resource built on the basis of a wild-caught male ZZ central bearded dragon Pogona vitticeps. The genomic sequence for P. vitticeps, generated on the Illumina HiSeq 2000 platform, comprised 317 Gbp (179X raw read depth) from 13 insert libraries ranging from 250 bp to 40 kbp. After filtering for low-quality and duplicated reads, 146 Gbp of data (83X) was available for assembly. Exceptionally high levels of heterozygosity (0.85 % of single nucleotide polymorphisms plus sequence insertions or deletions) complicated assembly; nevertheless, 96.4 % of reads mapped back to the assembled scaffolds, indicating that the assembly included most of the sequenced genome. Length of the assembly was 1.8 Gbp in 545,310 scaffolds (69,852 longer than 300 bp), the longest being 14.68 Mbp. N50 was 2.29 Mbp. Genes were annotated on the basis of de novo prediction, similarity to the green anole Anolis carolinensis, Gallus gallus and Homo sapiens proteins, and P. vitticeps transcriptome sequence assemblies, to yield 19,406 protein-coding genes in the assembly, 63 % of which had intact open reading frames. Our assembly captured 99 % (246 of 248) of core CEGMA genes, with 93 % (231) being complete. The quality of the P. vitticeps assembly is comparable or superior to that of other published squamate genomes, and the annotated P. vitticeps genome can be accessed through a genome browser available at https://genomics.canberra.edu.au.

  4. Atkinesin-13A Modulates Cell-Wall Synthesis and Cell Expansion in Arabidopsis thaliana via the THESEUS1 Pathway

    Science.gov (United States)

    Fujikura, Ushio; Elsaesser, Lore; Breuninger, Holger; Sánchez-Rodríguez, Clara; Ivakov, Alexander; Laux, Thomas; Findlay, Kim; Persson, Staffan; Lenhard, Michael

    2014-01-01

    Growth of plant organs relies on cell proliferation and expansion. While an increasingly detailed picture about the control of cell proliferation is emerging, our knowledge about the control of cell expansion remains more limited. We demonstrate here that the internal-motor kinesin AtKINESIN-13A (AtKIN13A) limits cell expansion and cell size in Arabidopsis thaliana, with loss-of-function atkin13a mutants forming larger petals with larger cells. The homolog, AtKINESIN-13B, also affects cell expansion and double mutants display growth, gametophytic and early embryonic defects, indicating a redundant role of the two genes. AtKIN13A is known to depolymerize microtubules and influence Golgi motility and distribution. Consistent with this function, AtKIN13A interacts genetically with ANGUSTIFOLIA, encoding a regulator of Golgi dynamics. Reduced AtKIN13A activity alters cell wall structure as assessed by Fourier-transformed infrared-spectroscopy and triggers signalling via the THESEUS1-dependent cell-wall integrity pathway, which in turn promotes the excess cell expansion in the atkin13a mutant. Thus, our results indicate that the intracellular activity of AtKIN13A regulates cell expansion and wall architecture via THESEUS1, providing a compelling case of interplay between cell wall integrity sensing and expansion. PMID:25232944

  5. Assessment of metagenomic assembly using simulated next generation sequencing data

    DEFF Research Database (Denmark)

    Mende, Daniel R; Waller, Alison S; Sunagawa, Shinichi

    2012-01-01

    with platform-specific (Sanger, pyrosequencing, Illumina) base-error models, and simulated metagenomes of differing community complexities. We first evaluated the effect of rigorous quality control on Illumina data. Although quality filtering removed a large proportion of the data, it greatly improved...... the accuracy and contig lengths of resulting assemblies. We then compared the quality-trimmed Illumina assemblies to those from Sanger and pyrosequencing. For the simple community (10 genomes) all sequencing technologies assembled a similar amount and accurately represented the expected functional composition...... the Sanger reads still represented the overall functional composition reasonably well. We further examined the effect of scaffolding of contigs using paired-end Illumina reads. It dramatically increased contig lengths of the simple community and yielded minor improvements to the more complex communities...

  6. Assembly of the Complete Sitka Spruce Chloroplast Genome Using 10X Genomics' GemCode Sequencing Data.

    Directory of Open Access Journals (Sweden)

    Lauren Coombe

    Full Text Available The linked read sequencing library preparation platform by 10X Genomics produces barcoded sequencing libraries, which are subsequently sequenced using the Illumina short read sequencing technology. In this new approach, long fragments of DNA are partitioned into separate micro-reactions, where the same index sequence is incorporated into each of the sequencing fragment inserts derived from a given long fragment. In this study, we exploited this property by using reads from index sequences associated with a large number of reads, to assemble the chloroplast genome of the Sitka spruce tree (Picea sitchensis. Here we report on the first Sitka spruce chloroplast genome assembled exclusively from P. sitchensis genomic libraries prepared using the 10X Genomics protocol. We show that the resulting 124,049 base pair long genome shares high sequence similarity with the related white spruce and Norway spruce chloroplast genomes, but diverges substantially from a previously published P. sitchensis- P. thunbergii chimeric genome. The use of reads from high-frequency indices enabled separation of the nuclear genome reads from that of the chloroplast, which resulted in the simplification of the de Bruijn graphs used at the various stages of assembly.

  7. Transcriptome sequencing in an ecologically important tree species: assembly, annotation, and marker discovery

    Directory of Open Access Journals (Sweden)

    Benkman Craig W

    2010-03-01

    Full Text Available Abstract Background Massively parallel sequencing of cDNA is now an efficient route for generating enormous sequence collections that represent expressed genes. This approach provides a valuable starting point for characterizing functional genetic variation in non-model organisms, especially where whole genome sequencing efforts are currently cost and time prohibitive. The large and complex genomes of pines (Pinus spp. have hindered the development of genomic resources, despite the ecological and economical importance of the group. While most genomic studies have focused on a single species (P. taeda, genomic level resources for other pines are insufficiently developed to facilitate ecological genomic research. Lodgepole pine (P. contorta is an ecologically important foundation species of montane forest ecosystems and exhibits substantial adaptive variation across its range in western North America. Here we describe a sequencing study of expressed genes from P. contorta, including their assembly and annotation, and their potential for molecular marker development to support population and association genetic studies. Results We obtained 586,732 sequencing reads from a 454 GS XLR70 Titanium pyrosequencer (mean length: 306 base pairs. A combination of reference-based and de novo assemblies yielded 63,657 contigs, with 239,793 reads remaining as singletons. Based on sequence similarity with known proteins, these sequences represent approximately 17,000 unique genes, many of which are well covered by contig sequences. This sequence collection also included a surprisingly large number of retrotransposon sequences, suggesting that they are highly transcriptionally active in the tissues we sampled. We located and characterized thousands of simple sequence repeats and single nucleotide polymorphisms as potential molecular markers in our assembled and annotated sequences. High quality PCR primers were designed for a substantial number of the SSR loci

  8. Sequencing and de novo assembly of 150 genomes from Denmark as a population reference

    DEFF Research Database (Denmark)

    Maretty, Lasse; Jensen, Jacob Malte; Petersen, Bent

    2017-01-01

    or by performing local assembly. However, these approaches are biased against discovery of structural variants and variation in the more complex parts of the genome. Hence, large-scale de novo assembly is needed. Here we show that it is possible to construct excellent de novo assemblies from high......-coverage sequencing with mate-pair libraries extending up to 20 kilobases. We report de novo assemblies of 150 individuals (50 trios) from the GenomeDenmark project. The quality of these assemblies is similar to those obtained using the more expensive long-read technology. We use the assemblies to identify a rich set...

  9. INTEGRATED APPROACH TO GENERATION OF PRECEDENCE RELATIONS AND PRECEDENCE GRAPHS FOR ASSEMBLY SEQUENCE PLANNING

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    An integrated approach to generation of precedence relations and precedence graphs for assembly sequence planning is presented, which contains more assembly flexibility. The approach involves two stages. Based on the assembly model, the components in the assembly can be divided into partially constrained components and completely constrained components in the first stage, and then geometric precedence relation for every component is generated automatically. According to the result of the first stage, the second stage determines and constructs all precedence graphs. The algorithms of these two stages proposed are verified by two assembly examples.

  10. Choice of reference sequence and assembler for alignment of Listeria monocytogenes short-read sequence data greatly influences rates of error in SNP analyses.

    Directory of Open Access Journals (Sweden)

    Arthur W Pightling

    Full Text Available The wide availability of whole-genome sequencing (WGS and an abundance of open-source software have made detection of single-nucleotide polymorphisms (SNPs in bacterial genomes an increasingly accessible and effective tool for comparative analyses. Thus, ensuring that real nucleotide differences between genomes (i.e., true SNPs are detected at high rates and that the influences of errors (such as false positive SNPs, ambiguously called sites, and gaps are mitigated is of utmost importance. The choices researchers make regarding the generation and analysis of WGS data can greatly influence the accuracy of short-read sequence alignments and, therefore, the efficacy of such experiments. We studied the effects of some of these choices, including: i depth of sequencing coverage, ii choice of reference-guided short-read sequence assembler, iii choice of reference genome, and iv whether to perform read-quality filtering and trimming, on our ability to detect true SNPs and on the frequencies of errors. We performed benchmarking experiments, during which we assembled simulated and real Listeria monocytogenes strain 08-5578 short-read sequence datasets of varying quality with four commonly used assemblers (BWA, MOSAIK, Novoalign, and SMALT, using reference genomes of varying genetic distances, and with or without read pre-processing (i.e., quality filtering and trimming. We found that assemblies of at least 50-fold coverage provided the most accurate results. In addition, MOSAIK yielded the fewest errors when reads were aligned to a nearly identical reference genome, while using SMALT to align reads against a reference sequence that is ∼0.82% distant from 08-5578 at the nucleotide level resulted in the detection of the greatest numbers of true SNPs and the fewest errors. Finally, we show that whether read pre-processing improves SNP detection depends upon the choice of reference sequence and assembler. In total, this study demonstrates that researchers

  11. Assembly and melting of DNA nanotubes from single-sequence tiles

    International Nuclear Information System (INIS)

    Sobey, T L; Renner, S; Simmel, F C

    2009-01-01

    DNA melting and renaturation studies are an extremely valuable tool to study the kinetics and thermodynamics of duplex dissociation and reassociation reactions. These are important not only in a biological or biotechnological context, but also for DNA nanotechnology which aims at the construction of molecular materials by DNA self-assembly. We here study experimentally the formation and melting of a DNA nanotube structure, which is composed of many copies of an oligonucleotide containing several palindromic sequences. This is done using temperature-controlled UV absorption measurements correlated with atomic force microscopy, fluorescence microscopy and transmission electron microscopy techniques. In the melting studies, important factors such as DNA strand concentration, hierarchy of assembly and annealing protocol are investigated. Assembly and melting of the nanotubes are shown to proceed via different pathways. Whereas assembly occurs in several hierarchical steps related to the formation of tiles, lattices and tubes, melting of DNA nanotubes appears to occur in a single step. This is proposed to relate to fundamental differences between closed, three-dimensional tube-like structures and open, two-dimensional lattices. DNA melting studies can lead to a better understanding of the many factors that affect the assembly process which will be essential for the assembly of increasingly complex DNA nanostructures.

  12. Harnessing NGS and Big Data Optimally: Comparison of miRNA Prediction from Assembled versus Non-assembled Sequencing Data--The Case of the Grass Aegilops tauschii Complex Genome.

    Science.gov (United States)

    Budak, Hikmet; Kantar, Melda

    2015-07-01

    MicroRNAs (miRNAs) are small, endogenous, non-coding RNA molecules that regulate gene expression at the post-transcriptional level. As high-throughput next generation sequencing (NGS) and Big Data rapidly accumulate for various species, efforts for in silico identification of miRNAs intensify. Surprisingly, the effect of the input genomics sequence on the robustness of miRNA prediction was not evaluated in detail to date. In the present study, we performed a homology-based miRNA and isomiRNA prediction of the 5D chromosome of bread wheat progenitor, Aegilops tauschii, using two distinct sequence data sets as input: (1) raw sequence reads obtained from 454-GS FLX Titanium sequencing platform and (2) an assembly constructed from these reads. We also compared this method with a number of available plant sequence datasets. We report here the identification of 62 and 22 miRNAs from raw reads and the assembly, respectively, of which 16 were predicted with high confidence from both datasets. While raw reads promoted sensitivity with the high number of miRNAs predicted, 55% (12 out of 22) of the assembly-based predictions were supported by previous observations, bringing specificity forward compared to the read-based predictions, of which only 37% were supported. Importantly, raw reads could identify several repeat-related miRNAs that could not be detected with the assembly. However, raw reads could not capture 6 miRNAs, for which the stem-loops could only be covered by the relatively longer sequences from the assembly. In summary, the comparison of miRNA datasets obtained by these two strategies revealed that utilization of raw reads, as well as assemblies for in silico prediction, have distinct advantages and disadvantages. Consideration of these important nuances can benefit future miRNA identification efforts in the current age of NGS and Big Data driven life sciences innovation.

  13. Nanopore sequencing technology and tools for genome assembly: computational analysis of the current state, bottlenecks and future directions.

    Science.gov (United States)

    Senol Cali, Damla; Kim, Jeremie S; Ghose, Saugata; Alkan, Can; Mutlu, Onur

    2018-04-02

    Nanopore sequencing technology has the potential to render other sequencing technologies obsolete with its ability to generate long reads and provide portability. However, high error rates of the technology pose a challenge while generating accurate genome assemblies. The tools used for nanopore sequence analysis are of critical importance, as they should overcome the high error rates of the technology. Our goal in this work is to comprehensively analyze current publicly available tools for nanopore sequence analysis to understand their advantages, disadvantages and performance bottlenecks. It is important to understand where the current tools do not perform well to develop better tools. To this end, we (1) analyze the multiple steps and the associated tools in the genome assembly pipeline using nanopore sequence data, and (2) provide guidelines for determining the appropriate tools for each step. Based on our analyses, we make four key observations: (1) the choice of the tool for basecalling plays a critical role in overcoming the high error rates of nanopore sequencing technology. (2) Read-to-read overlap finding tools, GraphMap and Minimap, perform similarly in terms of accuracy. However, Minimap has a lower memory usage, and it is faster than GraphMap. (3) There is a trade-off between accuracy and performance when deciding on the appropriate tool for the assembly step. The fast but less accurate assembler Miniasm can be used for quick initial assembly, and further polishing can be applied on top of it to increase the accuracy, which leads to faster overall assembly. (4) The state-of-the-art polishing tool, Racon, generates high-quality consensus sequences while providing a significant speedup over another polishing tool, Nanopolish. We analyze various combinations of different tools and expose the trade-offs between accuracy, performance, memory usage and scalability. We conclude that our observations can guide researchers and practitioners in making conscious

  14. THESEUS - a research project to improve the safety standard of tank vehicles for dangerous goods

    International Nuclear Information System (INIS)

    Guenther, B.

    1992-01-01

    A research project reffered to as THESEUS was initiated by the Federal Ministry of Research and Technology of Germany. The intent of the investigation is to generate measures designed to enhance the safety standard of commercial transports of dangerous goods in tank vehicles. Hereby, the analysis of real accidents by teams within the project will provide the relevant parameters for the experimental and theoretical investigation of vehicles, tank components and safety devices. The project started in summer 1990. This paper will focus main features and the work done so far. Special consideration will be made to the failure behaviour of tank components as the authors field of activity. (orig.)

  15. Multi-platform next-generation sequencing of the domestic turkey (Meleagris gallopavo: genome assembly and analysis.

    Directory of Open Access Journals (Sweden)

    Rami A Dalloul

    2010-09-01

    Full Text Available A synergistic combination of two next-generation sequencing platforms with a detailed comparative BAC physical contig map provided a cost-effective assembly of the genome sequence of the domestic turkey (Meleagris gallopavo. Heterozygosity of the sequenced source genome allowed discovery of more than 600,000 high quality single nucleotide variants. Despite this heterozygosity, the current genome assembly (∼1.1 Gb includes 917 Mb of sequence assigned to specific turkey chromosomes. Annotation identified nearly 16,000 genes, with 15,093 recognized as protein coding and 611 as non-coding RNA genes. Comparative analysis of the turkey, chicken, and zebra finch genomes, and comparing avian to mammalian species, supports the characteristic stability of avian genomes and identifies genes unique to the avian lineage. Clear differences are seen in number and variety of genes of the avian immune system where expansions and novel genes are less frequent than examples of gene loss. The turkey genome sequence provides resources to further understand the evolution of vertebrate genomes and genetic variation underlying economically important quantitative traits in poultry. This integrated approach may be a model for providing both gene and chromosome level assemblies of other species with agricultural, ecological, and evolutionary interest.

  16. When less is more: 'slicing' sequencing data improves read decoding accuracy and de novo assembly quality.

    Science.gov (United States)

    Lonardi, Stefano; Mirebrahim, Hamid; Wanamaker, Steve; Alpert, Matthew; Ciardo, Gianfranco; Duma, Denisa; Close, Timothy J

    2015-09-15

    As the invention of DNA sequencing in the 70s, computational biologists have had to deal with the problem of de novo genome assembly with limited (or insufficient) depth of sequencing. In this work, we investigate the opposite problem, that is, the challenge of dealing with excessive depth of sequencing. We explore the effect of ultra-deep sequencing data in two domains: (i) the problem of decoding reads to bacterial artificial chromosome (BAC) clones (in the context of the combinatorial pooling design we have recently proposed), and (ii) the problem of de novo assembly of BAC clones. Using real ultra-deep sequencing data, we show that when the depth of sequencing increases over a certain threshold, sequencing errors make these two problems harder and harder (instead of easier, as one would expect with error-free data), and as a consequence the quality of the solution degrades with more and more data. For the first problem, we propose an effective solution based on 'divide and conquer': we 'slice' a large dataset into smaller samples of optimal size, decode each slice independently, and then merge the results. Experimental results on over 15 000 barley BACs and over 4000 cowpea BACs demonstrate a significant improvement in the quality of the decoding and the final assembly. For the second problem, we show for the first time that modern de novo assemblers cannot take advantage of ultra-deep sequencing data. Python scripts to process slices and resolve decoding conflicts are available from http://goo.gl/YXgdHT; software Hashfilter can be downloaded from http://goo.gl/MIyZHs stelo@cs.ucr.edu or timothy.close@ucr.edu Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  17. THESEUS - the first utility-scale 50 MWe. THErmal Solar EUropean power Station for the island of Crete, Greece

    Energy Technology Data Exchange (ETDEWEB)

    Cobi, A. [PreussenElektra AG, Hannover (Germany); Tzatzanis, A.

    1997-12-31

    When the European Commission`s Directorate for Energy (DG XVII) opened the second application window for THERMIE proposals under the Fourth Framework Programme on September 15, 1995, Europe`s solar thermal power community discovered a remarkable modification to the previous call for proposals. For the first time since solar thermal electricity was introduced for THERMIE demonstration projects, it now became eligible for EU financial support. Right now, the European Commission committed funding for the design and engineering phase of the THESEUS project implementation. (orig.)

  18. Tips and tricks for the assembly of a Corynebacterium pseudotuberculosis genome using a semiconductor sequencer

    DEFF Research Database (Denmark)

    Ramos, Rommel Thiago Jucá; Carneiro, Adriana Ribeiro; Soares, Siomar de Castro

    2013-01-01

    that enable reference-based assembly, such as the one used in the present study, Corynebacterium pseudotuberculosis biovar equi, which causes high economic losses in the US equine industry. The quality treatment strategy incorporated into the assembly pipeline enabled a 16-fold greater use of the sequencing...... was validated by comparative genomics with other species of the genus Corynebacterium. The present study presents a modus operandi that enables a greater and better use of data obtained from semiconductor sequencing for obtaining the complete genome from a prokaryotic microorganism, C. pseudotuberculosis, which...

  19. Genome sequencing of bacteria: sequencing, de novo assembly and rapid analysis using open source tools.

    Science.gov (United States)

    Kisand, Veljo; Lettieri, Teresa

    2013-04-01

    De novo genome sequencing of previously uncharacterized microorganisms has the potential to open up new frontiers in microbial genomics by providing insight into both functional capabilities and biodiversity. Until recently, Roche 454 pyrosequencing was the NGS method of choice for de novo assembly because it generates hundreds of thousands of long reads (tools for processing NGS data are increasingly free and open source and are often adopted for both their high quality and role in promoting academic freedom. The error rate of pyrosequencing the Alcanivorax borkumensis genome was such that thousands of insertions and deletions were artificially introduced into the finished genome. Despite a high coverage (~30 fold), it did not allow the reference genome to be fully mapped. Reads from regions with errors had low quality, low coverage, or were missing. The main defect of the reference mapping was the introduction of artificial indels into contigs through lower than 100% consensus and distracting gene calling due to artificial stop codons. No assembler was able to perform de novo assembly comparable to reference mapping. Automated annotation tools performed similarly on reference mapped and de novo draft genomes, and annotated most CDSs in the de novo assembled draft genomes. Free and open source software (FOSS) tools for assembly and annotation of NGS data are being developed rapidly to provide accurate results with less computational effort. Usability is not high priority and these tools currently do not allow the data to be processed without manual intervention. Despite this, genome assemblers now readily assemble medium short reads into long contigs (>97-98% genome coverage). A notable gap in pyrosequencing technology is the quality of base pair calling and conflicting base pairs between single reads at the same nucleotide position. Regardless, using draft whole genomes that are not finished and remain fragmented into tens of contigs allows one to characterize

  20. Norgal: Extraction and de novo assembly of mitochondrial DNA from whole-genome sequencing data

    DEFF Research Database (Denmark)

    Al-Nakeeb, Kosai Ali Ahmed; Petersen, Thomas Nordahl; Sicheritz-Pontén, Thomas

    2017-01-01

    and performing a de novo assembly on a subset of reads that contains these k-mers. The method was applied to WGS data from a panda, brown algae seaweed, butterfly and filamentous fungus. We were able to extract full circular mitochondrial genomes and obtained sequence identities to the reference sequences...

  1. Bioethics, biotechnology products and humans: Europe between the skilled Theseus and the Labyrinth-Minotaur's syndrome.

    Science.gov (United States)

    Frati, P

    1999-01-01

    Following the approval on May 1998 of the European Union Common position no. 19/98 regarding the legal protection of biotechnological inventions, the debate on bioethical aspects of biotechnologies is increased. The European Union document clearly protects the patentability of inventions (that concerns more than a particular plant or animal variety or a single procedure if they are of industrial interest), but not the finding or discovery of that is in the nature, e.g. a gene. Some safeguards (the dignity and integrity of the person and of the human embryo, the plant diversity, etc.) and exclusions from patentability (plant and animal varieties, processes for the production of plants or animals, the human body at any stage of growth, cloning of human beings, modifications of germ line, use of human embryos for industrial or commercial purposes as well as the inventions whose publication or exploitation would offend against public policy or morality, according to the Article 53a of the European Patenting Convention) are also indicated. Ethical issues discussed include the nature of human life and its protection, the safeguard of plant-animal biological diversity, the safeguard of human dignity and nature, whereas on several aspects (e. g. limits of the use of genetic material, xenotransplantation, etc.) the Parliament Assembly of the Council of Europe has requested a discussion or a moratorium (April, 1999). In this case an evaluation on the basis of the ethical beneficial principles should be performed and society should decide whether to master technologies and emulate the positive action of the hero Theseus against the Labyrinth-Minotaur syndrome or to renounce or "misuse" technologies like Daedalus and Icarus, who met a tragic end.

  2. De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

    Science.gov (United States)

    Nowrousian, Minou; Stajich, Jason E; Chu, Meiling; Engh, Ines; Espagne, Eric; Halliday, Karen; Kamerewerd, Jens; Kempken, Frank; Knab, Birgit; Kuo, Hsiao-Che; Osiewacz, Heinz D; Pöggeler, Stefanie; Read, Nick D; Seiler, Stephan; Smith, Kristina M; Zickler, Denise; Kück, Ulrich; Freitag, Michael

    2010-04-08

    Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb) with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data can be used for

  3. De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

    Directory of Open Access Journals (Sweden)

    Minou Nowrousian

    2010-04-01

    Full Text Available Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data

  4. RePS: a sequence assembler that masks exact repeats identified from the shotgun data

    DEFF Research Database (Denmark)

    Wang, Jun; Wong, Gane Ka-Shu; Ni, Peixiang

    2002-01-01

    We describe a sequence assembler, RePS (repeat-masked Phrap with scaffolding), that explicitly identifies exact 20mer repeats from the shotgun data and removes them prior to the assembly. The established software is used to compute meaningful error probabilities for each base. Clone......-end-pairing information is used to construct scaffolds that order and orient the contigs. We show with real data for human and rice that reasonable assemblies are possible even at coverages of only 4x to 6x, despite having up to 42.2% in exact repeats. Udgivelsesdato: 2002-May...

  5. Growth of rat dorsal root ganglion neurons on a novel self-assembling scaffold containing IKVAV sequence

    Energy Technology Data Exchange (ETDEWEB)

    Zou Zhenwei; Zheng Qixin [Department of Orthopaedics, Union Hospital, Tongji Medical college of Huazhong University of science and technology, Wuhan, 430022 (China); Wu Yongchao, E-mail: wuyongchao@hotmail.com [Department of Orthopaedics, Union Hospital, Tongji Medical college of Huazhong University of science and technology, Wuhan, 430022 (China); Song Yulin; Wu Bin [Department of Orthopaedics, Union Hospital, Tongji Medical college of Huazhong University of science and technology, Wuhan, 430022 (China)

    2009-08-31

    The potential benefits of self-assembly in synthesizing materials for the treatment of both peripheral and central nervous system disorders are tremendous. In this study, we synthesized peptide-amphiphile (PA) molecules containing IKVAV sequence and induced self-assembly of the PA solutions in vitro to form nanofiber gels. Then, we tested the characterization of gels by transmission electron microscopy and demonstrated the biocompatibility of this gel towards rat dorsal root ganglion neurons. The nanofiber gel was formed by self-assembly of IKVAV PA molecules, which was triggered by metal ions. The fibers were 7-8 nm in diameter and with lengths of hundreds of nanometers. Gels were shown to be non-toxic to neurons and able to promote neurons adhesion and neurite sprouting. The results indicated that the self-assembling scaffold containing IKVAV sequence had excellent biocompatibility with adult sensory neurons and could be useful in nerve tissue engineering.

  6. mPUMA: a computational approach to microbiota analysis by de novo assembly of operational taxonomic units based on protein-coding barcode sequences.

    Science.gov (United States)

    Links, Matthew G; Chaban, Bonnie; Hemmingsen, Sean M; Muirhead, Kevin; Hill, Janet E

    2013-08-15

    Formation of operational taxonomic units (OTU) is a common approach to data aggregation in microbial ecology studies based on amplification and sequencing of individual gene targets. The de novo assembly of OTU sequences has been recently demonstrated as an alternative to widely used clustering methods, providing robust information from experimental data alone, without any reliance on an external reference database. Here we introduce mPUMA (microbial Profiling Using Metagenomic Assembly, http://mpuma.sourceforge.net), a software package for identification and analysis of protein-coding barcode sequence data. It was developed originally for Cpn60 universal target sequences (also known as GroEL or Hsp60). Using an unattended process that is independent of external reference sequences, mPUMA forms OTUs by DNA sequence assembly and is capable of tracking OTU abundance. mPUMA processes microbial profiles both in terms of the direct DNA sequence as well as in the translated amino acid sequence for protein coding barcodes. By forming OTUs and calculating abundance through an assembly approach, mPUMA is capable of generating inputs for several popular microbiota analysis tools. Using SFF data from sequencing of a synthetic community of Cpn60 sequences derived from the human vaginal microbiome, we demonstrate that mPUMA can faithfully reconstruct all expected OTU sequences and produce compositional profiles consistent with actual community structure. mPUMA enables analysis of microbial communities while empowering the discovery of novel organisms through OTU assembly.

  7. MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads (Metagenomics Informatics Challenges Workshop: 10K Genomes at a Time)

    Energy Technology Data Exchange (ETDEWEB)

    Sakakibara, Yasumbumi

    2011-10-13

    Keio University's Yasumbumi Sakakibara on "MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads" at the Metagenomics Informatics Challenges Workshop held at the DOE JGI on October 12-13, 2011.

  8. Multifunctional hybrid networks based on self assembling peptide sequences

    Science.gov (United States)

    Sathaye, Sameer

    The overall aim of this dissertation is to achieve a comprehensive correlation between the molecular level changes in primary amino acid sequences of amphiphilic beta-hairpin peptides and their consequent solution-assembly properties and bulk network hydrogel behavior. This has been accomplished using two broad approaches. In the first approach, amino acid substitutions were made to peptide sequence MAX1 such that the hydrophobic surfaces of the folded beta-hairpins from the peptides demonstrate shape specificity in hydrophobic interactions with other beta-hairpins during the assembly process, thereby causing changes to the peptide nanostructure and bulk rheological properties of hydrogels formed from the peptides. Steric lock and key complementary hydrophobic interactions were designed to occur between two beta-hairpin molecules of a single molecule, LNK1 during beta-sheet fibrillar assembly of LNK1. Experimental results from circular dichroism, transmission electron microscopy and oscillatory rheology collectively indicate that the molecular design of the LNK1 peptide can be assigned the cause of the drastically different behavior of the networks relative to MAX1. The results indicate elimination or significant reduction of fibrillar branching due to steric complementarity in LNK1 that does not exist in MAX1, thus supporting the original hypothesis. As an extension of the designed steric lock and key complementarity between two beta-hairpin molecules of the same peptide molecule. LNK1, three new pairs of peptide molecules LP1-KP1, LP2-KP2 and LP3-KP3 that resemble complementary 'wedge' and 'trough' shapes when folded into beta-hairpins were designed and studied. All six peptides individually and when blended with their corresponding shape complement formed fibrillar nanostructures with non-uniform thickness values. Loose packing in the assembled structures was observed in all the new peptides as compared to the uniform tight packing in MAX1 by SANS analysis. This

  9. SEQUENCING AND DE NOVO DRAFT ASSEMBLIES OF A FATHEAD MINNOW (Pimpehales promelas) reference genome

    Data.gov (United States)

    U.S. Environmental Protection Agency — The dataset provides the URLs for accessing the genome sequence data and two draft assemblies as well as fathead minnow genotyping data associated with estimating...

  10. De novo assembly, characterization and functional annotation of pineapple fruit transcriptome through massively parallel sequencing.

    Science.gov (United States)

    Ong, Wen Dee; Voo, Lok-Yung Christopher; Kumar, Vijay Subbiah

    2012-01-01

    Pineapple (Ananas comosus var. comosus), is an important tropical non-climacteric fruit with high commercial potential. Understanding the mechanism and processes underlying fruit ripening would enable scientists to enhance the improvement of quality traits such as, flavor, texture, appearance and fruit sweetness. Although, the pineapple is an important fruit, there is insufficient transcriptomic or genomic information that is available in public databases. Application of high throughput transcriptome sequencing to profile the pineapple fruit transcripts is therefore needed. To facilitate this, we have performed transcriptome sequencing of ripe yellow pineapple fruit flesh using Illumina technology. About 4.7 millions Illumina paired-end reads were generated and assembled using the Velvet de novo assembler. The assembly produced 28,728 unique transcripts with a mean length of approximately 200 bp. Sequence similarity search against non-redundant NCBI database identified a total of 16,932 unique transcripts (58.93%) with significant hits. Out of these, 15,507 unique transcripts were assigned to gene ontology terms. Functional annotation against Kyoto Encyclopedia of Genes and Genomes pathway database identified 13,598 unique transcripts (47.33%) which were mapped to 126 pathways. The assembly revealed many transcripts that were previously unknown. The unique transcripts derived from this work have rapidly increased of the number of the pineapple fruit mRNA transcripts as it is now available in public databases. This information can be further utilized in gene expression, genomics and other functional genomics studies in pineapple.

  11. Towards human exploration of space: The THESEUS review series on immunology research priorities.

    Science.gov (United States)

    Frippiat, Jean-Pol; Crucian, Brian E; de Quervain, Dominique J-F; Grimm, Daniela; Montano, Nicola; Praun, Siegfried; Roozendaal, Benno; Schelling, Gustav; Thiel, Manfred; Ullrich, Oliver; Choukèr, Alexander

    2016-01-01

    Dysregulation of the immune system occurs during spaceflight and may represent a crew health risk during exploration missions because astronauts are challenged by many stressors. Therefore, it is crucial to understand the biology of immune modulation under spaceflight conditions in order to be able to maintain immune homeostasis under such challenges. In the framework of the THESEUS project whose aim was to develop an integrated life sciences research roadmap regarding human space exploration, experts working in the field of space immunology, and related disciplines, established a questionnaire sent to scientists around the world. From the review of collected answers, they deduced a list of key issues and provided several recommendations such as a maximal exploitation of currently available resources on Earth and in space, and to increase increments duration for some ISS crew members to 12 months or longer. These recommendations should contribute to improve our knowledge about spaceflight effects on the immune system and the development of countermeasures that, beyond astronauts, could have a societal impact.

  12. The de novo assembly of mitochondrial genomes of the extinct passenger pigeon (Ectopistes migratorius with next generation sequencing.

    Directory of Open Access Journals (Sweden)

    Chih-Ming Hung

    Full Text Available The information from ancient DNA (aDNA provides an unparalleled opportunity to infer phylogenetic relationships and population history of extinct species and to investigate genetic evolution directly. However, the degraded and fragmented nature of aDNA has posed technical challenges for studies based on conventional PCR amplification. In this study, we present an approach based on next generation sequencing to efficiently sequence the complete mitochondrial genome (mitogenome of two extinct passenger pigeons (Ectopistes migratorius using de novo assembly of massive short (90 bp, paired-end or single-end reads. Although varying levels of human contamination and low levels of postmortem nucleotide lesion were observed, they did not impact sequencing accuracy. Our results demonstrated that the de novo assembly of shotgun sequence reads could be a potent approach to sequence mitogenomes, and offered an efficient way to infer evolutionary history of extinct species.

  13. The De Novo Assembly of Mitochondrial Genomes of the Extinct Passenger Pigeon (Ectopistes migratorius) with Next Generation Sequencing

    Science.gov (United States)

    Hung, Chih-Ming; Lin, Rong-Chien; Chu, Jui-Hua; Yeh, Chia-Fen; Yao, Chiou-Ju; Li, Shou-Hsien

    2013-01-01

    The information from ancient DNA (aDNA) provides an unparalleled opportunity to infer phylogenetic relationships and population history of extinct species and to investigate genetic evolution directly. However, the degraded and fragmented nature of aDNA has posed technical challenges for studies based on conventional PCR amplification. In this study, we present an approach based on next generation sequencing to efficiently sequence the complete mitochondrial genome (mitogenome) of two extinct passenger pigeons (Ectopistes migratorius) using de novo assembly of massive short (90 bp), paired-end or single-end reads. Although varying levels of human contamination and low levels of postmortem nucleotide lesion were observed, they did not impact sequencing accuracy. Our results demonstrated that the de novo assembly of shotgun sequence reads could be a potent approach to sequence mitogenomes, and offered an efficient way to infer evolutionary history of extinct species. PMID:23437111

  14. Characterization of Liaoning cashmere goat transcriptome: sequencing, de novo assembly, functional annotation and comparative analysis.

    Directory of Open Access Journals (Sweden)

    Hongliang Liu

    Full Text Available Liaoning cashmere goat is a famous goat breed for cashmere wool. In order to increase the transcriptome data and accelerate genetic improvement for this breed, we performed de novo transcriptome sequencing to generate the first expressed sequence tag dataset for the Liaoning cashmere goat, using next-generation sequencing technology.Transcriptome sequencing of Liaoning cashmere goat on a Roche 454 platform yielded 804,601 high-quality reads. Clustering and assembly of these reads produced a non-redundant set of 117,854 unigenes, comprising 13,194 isotigs and 104,660 singletons. Based on similarity searches with known proteins, 17,356 unigenes were assigned to 6,700 GO categories, and the terms were summarized into three main GO categories and 59 sub-categories. 3,548 and 46,778 unigenes had significant similarity to existing sequences in the KEGG and COG databases, respectively. Comparative analysis revealed that 42,254 unigenes were aligned to 17,532 different sequences in NCBI non-redundant nucleotide databases. 97,236 (82.51% unigenes were mapped to the 30 goat chromosomes. 35,551 (30.17% unigenes were matched to 11,438 reported goat protein-coding genes. The remaining non-matched unigenes were further compared with cattle and human reference genes, 67 putative new goat genes were discovered. Additionally, 2,781 potential simple sequence repeats were initially identified from all unigenes.The transcriptome of Liaoning cashmere goat was deep sequenced, de novo assembled, and annotated, providing abundant data to better understand the Liaoning cashmere goat transcriptome. The potential simple sequence repeats provide a material basis for future genetic linkage and quantitative trait loci analyses.

  15. Sequencing and De novo Draft Assemblies of the Fathead Minnow (Pimphales promelas)Reference Genome

    Science.gov (United States)

    This study was undertaken to develop genome-scale resources for the fathead minnow (Pimphales promelas) an important model organism widely used in both aquatic ecotoxicology research and in regulatory toxicity testing. We report on the first sequencing and two draft assemblies fo...

  16. Coevolutionary constraints in the sequence-space of macromolecular complexes reflect their self-assembly pathways.

    Science.gov (United States)

    Mallik, Saurav; Kundu, Sudip

    2017-07-01

    Is the order in which biomolecular subunits self-assemble into functional macromolecular complexes imprinted in their sequence-space? Here, we demonstrate that the temporal order of macromolecular complex self-assembly can be efficiently captured using the landscape of residue-level coevolutionary constraints. This predictive power of coevolutionary constraints is irrespective of the structural, functional, and phylogenetic classification of the complex and of the stoichiometry and quaternary arrangement of the constituent monomers. Combining this result with a number of structural attributes estimated from the crystal structure data, we find indications that stronger coevolutionary constraints at interfaces formed early in the assembly hierarchy probably promotes coordinated fixation of mutations that leads to high-affinity binding with higher surface area, increased surface complementarity and elevated number of molecular contacts, compared to those that form late in the assembly. Proteins 2017; 85:1183-1189. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  17. The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads

    DEFF Research Database (Denmark)

    Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo

    2012-01-01

    Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp...... these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species....

  18. AFEAP cloning: a precise and efficient method for large DNA sequence assembly.

    Science.gov (United States)

    Zeng, Fanli; Zang, Jinping; Zhang, Suhua; Hao, Zhimin; Dong, Jingao; Lin, Yibin

    2017-11-14

    Recent development of DNA assembly technologies has spurred myriad advances in synthetic biology, but new tools are always required for complicated scenarios. Here, we have developed an alternative DNA assembly method named AFEAP cloning (Assembly of Fragment Ends After PCR), which allows scarless, modular, and reliable construction of biological pathways and circuits from basic genetic parts. The AFEAP method requires two-round of PCRs followed by ligation of the sticky ends of DNA fragments. The first PCR yields linear DNA fragments and is followed by a second asymmetric (one primer) PCR and subsequent annealing that inserts overlapping overhangs at both sides of each DNA fragment. The overlapping overhangs of the neighboring DNA fragments annealed and the nick was sealed by T4 DNA ligase, followed by bacterial transformation to yield the desired plasmids. We characterized the capability and limitations of new developed AFEAP cloning and demonstrated its application to assemble DNA with varying scenarios. Under the optimized conditions, AFEAP cloning allows assembly of an 8 kb plasmid from 1-13 fragments with high accuracy (between 80 and 100%), and 8.0, 11.6, 19.6, 28, and 35.6 kb plasmids from five fragments at 91.67, 91.67, 88.33, 86.33, and 81.67% fidelity, respectively. AFEAP cloning also is capable to construct bacterial artificial chromosome (BAC, 200 kb) with a fidelity of 46.7%. AFEAP cloning provides a powerful, efficient, seamless, and sequence-independent DNA assembly tool for multiple fragments up to 13 and large DNA up to 200 kb that expands synthetic biologist's toolbox.

  19. Mathematical model and metaheuristics for simultaneous balancing and sequencing of a robotic mixed-model assembly line

    Science.gov (United States)

    Li, Zixiang; Janardhanan, Mukund Nilakantan; Tang, Qiuhua; Nielsen, Peter

    2018-05-01

    This article presents the first method to simultaneously balance and sequence robotic mixed-model assembly lines (RMALB/S), which involves three sub-problems: task assignment, model sequencing and robot allocation. A new mixed-integer programming model is developed to minimize makespan and, using CPLEX solver, small-size problems are solved for optimality. Two metaheuristics, the restarted simulated annealing algorithm and co-evolutionary algorithm, are developed and improved to address this NP-hard problem. The restarted simulated annealing method replaces the current temperature with a new temperature to restart the search process. The co-evolutionary method uses a restart mechanism to generate a new population by modifying several vectors simultaneously. The proposed algorithms are tested on a set of benchmark problems and compared with five other high-performing metaheuristics. The proposed algorithms outperform their original editions and the benchmarked methods. The proposed algorithms are able to solve the balancing and sequencing problem of a robotic mixed-model assembly line effectively and efficiently.

  20. Discovery, genotyping and characterization of structural variation and novel sequence at single nucleotide resolution from de novo genome assemblies on a population scale

    DEFF Research Database (Denmark)

    Liu, Siyang; Huang, Shujia; Rao, Junhua

    2015-01-01

    present a novel approach implemented in a single software package, AsmVar, to discover, genotype and characterize different forms of structural variation and novel sequence from population-scale de novo genome assemblies up to nucleotide resolution. Application of AsmVar to several human de novo genome......) as well as large deletions. However, these approaches consistently display a substantial bias against the recovery of complex structural variants and novel sequence in individual genomes and do not provide interpretation information such as the annotation of ancestral state and formation mechanism. We...... assemblies captures a wide spectrum of structural variants and novel sequences present in the human population in high sensitivity and specificity. Our method provides a direct solution for investigating structural variants and novel sequences from de novo genome assemblies, facilitating the construction...

  1. Ammattikorkeakoulujen Julkaisuarkisto Theseus : käyttöönotto- ja perustamisvaiheet sekä nykyiset käytänteet ammattikorkeakouluissa

    OpenAIRE

    Ranta, Anna-Kaisa; Lahdenperä, Silja

    2012-01-01

    Tässä opinnäytetyössä perehdytään Julkaisuarkisto Theseuksen perustamisvaiheisiin ja käyttöönottoon Suomen ammattikorkeakouluissa. Theseuksen historia käydään läpi Open Access -hankkeen alkuvaiheista nykypäivään asti ja lopuksi luodaan katsaus myös tulevaisuuteen. Lisäksi työssä tutkitaan ammattikorkeakoulujen erilaisia käytänteitä Theseuksen käytön suhteen. Aloitteen työn tekemiseksi on tehnyt Theseus-ohjausryhmä ja työn toimeksiantaja on AMKIT-konsortion johtoryhmä. Tietolähteinä käytet...

  2. Draft Genome Sequence of a “Candidatus Liberibacter europaeus” Strain Assembled from Broom Psyllids (Arytainilla spartiophila) from New Zealand

    Science.gov (United States)

    Thompson, Sarah M.; Kalamorz, Falk; David, Charles; Addison, Shea M.; Smith, Grant R.

    2018-01-01

    ABSTRACT Here, we report the draft genome sequence of “Candidatus Liberibacter europaeus” ASNZ1, assembled from broom psyllids (Arytainilla spartiophila) from New Zealand. The assembly comprises 15 contigs, with a total length of 1.33 Mb and a G+C content of 33.5%. PMID:29773636

  3. De novo transcriptome sequencing and assembly from apomictic and sexual Eragrostis curvula genotypes.

    Directory of Open Access Journals (Sweden)

    Ingrid Garbus

    Full Text Available A long-standing goal in plant breeding has been the ability to confer apomixis to agriculturally relevant species, which would require a deeper comprehension of the molecular basis of apomictic regulatory mechanisms. Eragrostis curvula (Schrad. Nees is a perennial grass that includes both sexual and apomictic cytotypes. The availability of a reference transcriptome for this species would constitute a very important tool toward the identification of genes controlling key steps of the apomictic pathway. Here, we used Roche/454 sequencing technologies to generate reads from inflorescences of E. curvula apomictic and sexual genotypes that were de novo assembled into a reference transcriptome. Near 90% of the 49568 assembled isotigs showed sequence similarity to sequences deposited in the public databases. A gene ontology analysis categorized 27448 isotigs into at least one of the three main GO categories. We identified 11475 SSRs, and several of them were assayed in E curvula germoplasm using SSR-based primers, providing a valuable set of molecular markers that could allow direct allele selection. The differential contribution to each library of the spliced forms of several transcripts revealed the existence of several isotigs produced via alternative splicing of single genes. The reference transcriptome presented and validated in this work will be useful for the identification of a wide range of gene(s related to agronomic traits of E. curvula, including those controlling key steps of the apomictic pathway in this species, allowing the extrapolation of the findings to other plant species.

  4. Knowledge-based decision support for Space Station assembly sequence planning

    Science.gov (United States)

    1991-04-01

    A complete Personal Analysis Assistant (PAA) for Space Station Freedom (SSF) assembly sequence planning consists of three software components: the system infrastructure, intra-flight value added, and inter-flight value added. The system infrastructure is the substrate on which software elements providing inter-flight and intra-flight value-added functionality are built. It provides the capability for building representations of assembly sequence plans and specification of constraints and analysis options. Intra-flight value-added provides functionality that will, given the manifest for each flight, define cargo elements, place them in the National Space Transportation System (NSTS) cargo bay, compute performance measure values, and identify violated constraints. Inter-flight value-added provides functionality that will, given major milestone dates and capability requirements, determine the number and dates of required flights and develop a manifest for each flight. The current project is Phase 1 of a projected two phase program and delivers the system infrastructure. Intra- and inter-flight value-added were to be developed in Phase 2, which has not been funded. Based on experience derived from hundreds of projects conducted over the past seven years, ISX developed an Intelligent Systems Engineering (ISE) methodology that combines the methods of systems engineering and knowledge engineering to meet the special systems development requirements posed by intelligent systems, systems that blend artificial intelligence and other advanced technologies with more conventional computing technologies. The ISE methodology defines a phased program process that begins with an application assessment designed to provide a preliminary determination of the relative technical risks and payoffs associated with a potential application, and then moves through requirements analysis, system design, and development.

  5. Gene Discovery in the Apicomplexa as Revealed by EST Sequencing and Assembly of a Comparative Gene Database

    Science.gov (United States)

    Li, Li; Brunk, Brian P.; Kissinger, Jessica C.; Pape, Deana; Tang, Keliang; Cole, Robert H.; Martin, John; Wylie, Todd; Dante, Mike; Fogarty, Steven J.; Howe, Daniel K.; Liberator, Paul; Diaz, Carmen; Anderson, Jennifer; White, Michael; Jerome, Maria E.; Johnson, Emily A.; Radke, Jay A.; Stoeckert, Christian J.; Waterston, Robert H.; Clifton, Sandra W.; Roos, David S.; Sibley, L. David

    2003-01-01

    Large-scale EST sequencing projects for several important parasites within the phylum Apicomplexa were undertaken for the purpose of gene discovery. Included were several parasites of medical importance (Plasmodium falciparum, Toxoplasma gondii) and others of veterinary importance (Eimeria tenella, Sarcocystis neurona, and Neospora caninum). A total of 55,192 ESTs, deposited into dbEST/GenBank, were included in the analyses. The resulting sequences have been clustered into nonredundant gene assemblies and deposited into a relational database that supports a variety of sequence and text searches. This database has been used to compare the gene assemblies using BLAST similarity comparisons to the public protein databases to identify putative genes. Of these new entries, ∼15%–20% represent putative homologs with a conservative cutoff of p neurona: , , , , , , , , , , , , , –, –, –, –, –. Eimeria tenella: –, –, –, –, –, –, –, –, – , –, –, –, –, –, –, –, –, –, –, –. Neospora caninum: –, –, , – , –, –.] PMID:12618375

  6. Sequencing of BAC pools by different next generation sequencing platforms and strategies

    Directory of Open Access Journals (Sweden)

    Scholz Uwe

    2011-10-01

    Full Text Available Abstract Background Next generation sequencing of BACs is a viable option for deciphering the sequence of even large and highly repetitive genomes. In order to optimize this strategy, we examined the influence of read length on the quality of Roche/454 sequence assemblies, to what extent Illumina/Solexa mate pairs (MPs improve the assemblies by scaffolding and whether barcoding of BACs is dispensable. Results Sequencing four BACs with both FLX and Titanium technologies revealed similar sequencing accuracy, but showed that the longer Titanium reads produce considerably less misassemblies and gaps. The 454 assemblies of 96 barcoded BACs were improved by scaffolding 79% of the total contig length with MPs from a non-barcoded library. Assembly of the unmasked 454 sequences without separation by barcodes revealed chimeric contig formation to be a major problem, encompassing 47% of the total contig length. Masking the sequences reduced this fraction to 24%. Conclusion Optimal BAC pool sequencing should be based on the longest available reads, with barcoding essential for a comprehensive assessment of both repetitive and non-repetitive sequence information. When interest is restricted to non-repetitive regions and repeats are masked prior to assembly, barcoding is non-essential. In any case, the assemblies can be improved considerably by scaffolding with non-barcoded BAC pool MPs.

  7. Distilled single-cell genome sequencing and de novo assembly for sparse microbial communities.

    Science.gov (United States)

    Taghavi, Zeinab; Movahedi, Narjes S; Draghici, Sorin; Chitsaz, Hamidreza

    2013-10-01

    Identification of every single genome present in a microbial sample is an important and challenging task with crucial applications. It is challenging because there are typically millions of cells in a microbial sample, the vast majority of which elude cultivation. The most accurate method to date is exhaustive single-cell sequencing using multiple displacement amplification, which is simply intractable for a large number of cells. However, there is hope for breaking this barrier, as the number of different cell types with distinct genome sequences is usually much smaller than the number of cells. Here, we present a novel divide and conquer method to sequence and de novo assemble all distinct genomes present in a microbial sample with a sequencing cost and computational complexity proportional to the number of genome types, rather than the number of cells. The method is implemented in a tool called Squeezambler. We evaluated Squeezambler on simulated data. The proposed divide and conquer method successfully reduces the cost of sequencing in comparison with the naïve exhaustive approach. Squeezambler and datasets are available at http://compbio.cs.wayne.edu/software/squeezambler/.

  8. De novo sequencing, assembly and characterization of antennal transcriptome of Anomala corpulenta Motschulsky (Coleoptera: Rutelidae.

    Directory of Open Access Journals (Sweden)

    Haoliang Chen

    Full Text Available Anomala corpulenta is an important insect pest and can cause enormous economic losses in agriculture, horticulture and forestry. It is widely distributed in China, and both larvae and adults can cause serious damage. It is difficult to control this pest because the larvae live underground. Any new control strategy should exploit alternatives to heavily and frequently used chemical insecticides. However, little genetic research has been carried out on A. corpulenta due to the lack of genomic resources. Genomic resources could be produced by next generation sequencing technologies with low cost and in a short time. In this study, we performed de novo sequencing, assembly and characterization of the antennal transcriptome of A. corpulenta.Illumina sequencing technology was used to sequence the antennal transcriptome of A. corpulenta. Approximately 76.7 million total raw reads and about 68.9 million total clean reads were obtained, and then 35,656 unigenes were assembled. Of these unigenes, 21,463 of them could be annotated in the NCBI nr database, and, among the annotated unigenes, 11,154 and 6,625 unigenes could be assigned to GO and COG, respectively. Additionally, 16,350 unigenes could be annotated in the Swiss-Prot database, and 14,499 unigenes could map onto 258 pathways in the KEGG Pathway database. We also found 24 unigenes related to OBPs, 6 to CSPs, and in total 167 unigenes related to chemodetection. We analyzed 4 OBPs and 3CSPs sequences and their RT-qPCR results agreed well with their FPKM values.We produced the first large-scale antennal transcriptome of A. corpulenta, which is a species that has little genomic information in public databases. The identified chemodetection unigenes can promote the molecular mechanistic study of behavior in A. corpulenta. These findings provide a general sequence resource for molecular genetics research on A. corpulenta.

  9. Sequence Identification, Recombinant Production, and Analysis of the Self-Assembly of Egg Stalk Silk Proteins from Lacewing Chrysoperla carnea.

    Science.gov (United States)

    Neuenfeldt, Martin; Scheibel, Thomas

    2017-06-13

    Egg stalk silks of the common green lacewing Chrysoperla carnea likely comprise at least three different silk proteins. Based on the natural spinning process, it was hypothesized that these proteins self-assemble without shear stress, as adult lacewings do not use a spinneret. To examine this, the first sequence identification and determination of the gene expression profile of several silk proteins and various transcript variants thereof was conducted, and then the three major proteins were recombinantly produced in Escherichia coli encoded by their native complementary DNA (cDNA) sequences. Circular dichroism measurements indicated that the silk proteins in aqueous solutions had a mainly intrinsically disordered structure. The largest silk protein, which we named ChryC1, exhibited a lower critical solution temperature (LCST) behavior and self-assembled into fibers or film morphologies, depending on the conditions used. The second silk protein, ChryC2, self-assembled into nanofibrils and subsequently formed hydrogels. Circular dichroism and Fourier transform infrared spectroscopy confirmed conformational changes of both proteins into beta sheet rich structures upon assembly. ChryC3 did not self-assemble into any morphology under the tested conditions. Thereby, through this work, it could be shown that recombinant lacewing silk proteins can be produced and further used for studying the fiber formation of lacewing egg stalks.

  10. Path planning algorithms for assembly sequence planning. [in robot kinematics

    Science.gov (United States)

    Krishnan, S. S.; Sanderson, Arthur C.

    1991-01-01

    Planning for manipulation in complex environments often requires reasoning about the geometric and mechanical constraints which are posed by the task. In planning assembly operations, the automatic generation of operations sequences depends on the geometric feasibility of paths which permit parts to be joined into subassemblies. Feasible locations and collision-free paths must be present for part motions, robot and grasping motions, and fixtures. This paper describes an approach to reasoning about the feasibility of straight-line paths among three-dimensional polyhedral parts using an algebra of polyhedral cones. A second method recasts the feasibility conditions as constraints in a nonlinear optimization framework. Both algorithms have been implemented and results are presented.

  11. Computational complexity of algorithms for sequence comparison, short-read assembly and genome alignment.

    Science.gov (United States)

    Baichoo, Shakuntala; Ouzounis, Christos A

    A multitude of algorithms for sequence comparison, short-read assembly and whole-genome alignment have been developed in the general context of molecular biology, to support technology development for high-throughput sequencing, numerous applications in genome biology and fundamental research on comparative genomics. The computational complexity of these algorithms has been previously reported in original research papers, yet this often neglected property has not been reviewed previously in a systematic manner and for a wider audience. We provide a review of space and time complexity of key sequence analysis algorithms and highlight their properties in a comprehensive manner, in order to identify potential opportunities for further research in algorithm or data structure optimization. The complexity aspect is poised to become pivotal as we will be facing challenges related to the continuous increase of genomic data on unprecedented scales and complexity in the foreseeable future, when robust biological simulation at the cell level and above becomes a reality. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Scar-less multi-part DNA assembly design automation

    Science.gov (United States)

    Hillson, Nathan J.

    2016-06-07

    The present invention provides a method of a method of designing an implementation of a DNA assembly. In an exemplary embodiment, the method includes (1) receiving a list of DNA sequence fragments to be assembled together and an order in which to assemble the DNA sequence fragments, (2) designing DNA oligonucleotides (oligos) for each of the DNA sequence fragments, and (3) creating a plan for adding flanking homology sequences to each of the DNA oligos. In an exemplary embodiment, the method includes (1) receiving a list of DNA sequence fragments to be assembled together and an order in which to assemble the DNA sequence fragments, (2) designing DNA oligonucleotides (oligos) for each of the DNA sequence fragments, and (3) creating a plan for adding optimized overhang sequences to each of the DNA oligos.

  13. SWAP-Assembler: scalable and efficient genome assembly towards thousands of cores.

    Science.gov (United States)

    Meng, Jintao; Wang, Bingqiang; Wei, Yanjie; Feng, Shengzhong; Balaji, Pavan

    2014-01-01

    There is a widening gap between the throughput of massive parallel sequencing machines and the ability to analyze these sequencing data. Traditional assembly methods requiring long execution time and large amount of memory on a single workstation limit their use on these massive data. This paper presents a highly scalable assembler named as SWAP-Assembler for processing massive sequencing data using thousands of cores, where SWAP is an acronym for Small World Asynchronous Parallel model. In the paper, a mathematical description of multi-step bi-directed graph (MSG) is provided to resolve the computational interdependence on merging edges, and a highly scalable computational framework for SWAP is developed to automatically preform the parallel computation of all operations. Graph cleaning and contig extension are also included for generating contigs with high quality. Experimental results show that SWAP-Assembler scales up to 2048 cores on Yanhuang dataset using only 26 minutes, which is better than several other parallel assemblers, such as ABySS, Ray, and PASHA. Results also show that SWAP-Assembler can generate high quality contigs with good N50 size and low error rate, especially it generated the longest N50 contig sizes for Fish and Yanhuang datasets. In this paper, we presented a highly scalable and efficient genome assembly software, SWAP-Assembler. Compared with several other assemblers, it showed very good performance in terms of scalability and contig quality. This software is available at: https://sourceforge.net/projects/swapassembler.

  14. Next-generation transcriptome assembly

    Energy Technology Data Exchange (ETDEWEB)

    Martin, Jeffrey A.; Wang, Zhong

    2011-09-01

    Transcriptomics studies often rely on partial reference transcriptomes that fail to capture the full catalog of transcripts and their variations. Recent advances in sequencing technologies and assembly algorithms have facilitated the reconstruction of the entire transcriptome by deep RNA sequencing (RNA-seq), even without a reference genome. However, transcriptome assembly from billions of RNA-seq reads, which are often very short, poses a significant informatics challenge. This Review summarizes the recent developments in transcriptome assembly approaches - reference-based, de novo and combined strategies-along with some perspectives on transcriptome assembly in the near future.

  15. Draft Genome Sequences of 12 Dry-Heat-Resistant Bacillus Strains Isolated from the Cleanrooms Where the Viking Spacecraft Were Assembled.

    Science.gov (United States)

    Seuylemezian, Arman; Cooper, Kerry; Schubert, Wayne; Vaishampayan, Parag

    2018-03-22

    Spore-forming microorganisms are of concern for forward contamination because they can survive harsh interplanetary travel. Here, we report the draft genome sequences of 12 spore-forming strains isolated from the Manned Spacecraft Operations Building (MSOB) and the Vehicle Assembly Building (VAB) in Cape Canaveral, FL, where the Viking spacecraft were assembled. Copyright © 2018 Seuylemezian et al.

  16. Sequencing and de novo assembly of the Asian clam (Corbicula fluminea transcriptome using the Illumina GAIIx method.

    Directory of Open Access Journals (Sweden)

    Huihui Chen

    Full Text Available BACKGROUND: The Asian clam (Corbicula fluminea is currently one of the most economically important aquatic species in China and has been used as a test organism in many environmental studies. However, the lack of genomic resources, such as sequenced genome, expressed sequence tags (ESTs and transcriptome sequences has hindered the research on C. fluminea. Recent advances in large-scale RNA-Seq enable generation of genomic resources in a short time, and provide large expression datasets for functional genomic analysis. METHODOLOGY/PRINCIPAL FINDINGS: We used a next-generation high-throughput DNA sequencing technique with an Illumina GAIIx method to analyze the transcriptome from the whole bodies of C. fluminea. More than 62,250,336 high-quality reads were generated based on the raw data, and 134,684 unigenes with a mean length of 791 bp were assembled using the Velvet and Oases software. All of the assembly unigenes were annotated by running BLASTx and BLASTn similarity searches on the Nt, Nr, Swiss-Prot, COG and KEGG databases. In addition, the Clusters of Orthologous Groups (COGs, Gene Ontology (GO terms and Kyoto Encyclopedia of Gene and Genome (KEGG annotations were also assigned to each unigene transcript. To provide a preliminary verification of the assembly and annotation results, and search for potential environmental pollution biomarkers, 15 functional genes (five antioxidase genes, two cytochrome P450 genes, three GABA receptor-related genes and five heat shock protein genes were cloned and identified. Expressions of the 15 selected genes following fluoxetine exposure confirmed that the genes are indeed linked to environmental stress. CONCLUSIONS/SIGNIFICANCE: The C. fluminea transcriptome advances the underlying molecular understanding of this freshwater clam, provides a basis for further exploration of C. fluminea as an environmental test organism and promotes further studies on other bivalve organisms.

  17. De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from Total DNA Sequences

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    Shairul Izan

    2017-08-01

    Full Text Available Whole Genome Shotgun (WGS sequences of plant species often contain an abundance of reads that are derived from the chloroplast genome. Up to now these reads have generally been identified and assembled into chloroplast genomes based on homology to chloroplasts from related species. This re-sequencing approach may select against structural differences between the genomes especially in non-model species for which no close relatives have been sequenced before. The alternative approach is to de novo assemble the chloroplast genome from total genomic DNA sequences. In this study, we used k-mer frequency tables to identify and extract the chloroplast reads from the WGS reads and assemble these using a highly integrated and automated custom pipeline. Our strategy includes steps aimed at optimizing assemblies and filling gaps which are left due to coverage variation in the WGS dataset. We have successfully de novo assembled three complete chloroplast genomes from plant species with a range of nuclear genome sizes to demonstrate the universality of our approach: Solanum lycopersicum (0.9 Gb, Aegilops tauschii (4 Gb and Paphiopedilum henryanum (25 Gb. We also highlight the need to optimize the choice of k and the amount of data used. This new and cost-effective method for de novo short read assembly will facilitate the study of complete chloroplast genomes with more accurate analyses and inferences, especially in non-model plant genomes.

  18. Sequencing and De Novo Assembly of the Toxicodendron radicans (Poison Ivy) Transcriptome.

    Science.gov (United States)

    Weisberg, Alexandra J; Kim, Gunjune; Westwood, James H; Jelesko, John G

    2017-11-10

    Contact with poison ivy plants is widely dreaded because they produce a natural product called urushiol that is responsible for allergenic contact delayed-dermatitis symptoms lasting for weeks. For this reason, the catchphrase most associated with poison ivy is "leaves of three, let it be", which serves the purpose of both identification and an appeal for avoidance. Ironically, despite this notoriety, there is a dearth of specific knowledge about nearly all other aspects of poison ivy physiology and ecology. As a means of gaining a more molecular-oriented understanding of poison ivy physiology and ecology, Next Generation DNA sequencing technology was used to develop poison ivy root and leaf RNA-seq transcriptome resources. De novo assembled transcriptomes were analyzed to generate a core set of high quality expressed transcripts present in poison ivy tissue. The predicted protein sequences were evaluated for similarity to SwissProt homologs and InterProScan domains, as well as assigned both GO terms and KEGG annotations. Over 23,000 simple sequence repeats were identified in the transcriptome, and corresponding oligo nucleotide primer pairs were designed. A pan-transcriptome analysis of existing Anacardiaceae transcriptomes revealed conserved and unique transcripts among these species.

  19. Transcriptome sequencing and de novo assembly in arecanut, Areca catechu L elucidates the secondary metabolite pathway genes

    Directory of Open Access Journals (Sweden)

    Ramaswamy Manimekalai

    2018-03-01

    Full Text Available Areca catechu L. belongs to the Arecaceae family which comprises many economically important palms. The palm is a source of alkaloids and carotenoids. The lack of ample genetic information in public databases has been a constraint for the genetic improvement of arecanut. To gain molecular insight into the palm, high throughput RNA sequencing and de novo assembly of arecanut leaf transcriptome was undertaken in the present study. A total 56,321,907 paired end reads of 101 bp length consisting of 11.343 Gb nucleotides were generated. De novo assembly resulted in 48,783 good quality transcripts, of which 67% of transcripts could be annotated against NCBI non – redundant database. The Gene Ontology (GO analysis with UniProt database identified 9222 biological process, 11268 molecular function and 7574 cellular components GO terms. Large scale expression profiling through Fragments per Kilobase per Million mapped reads (FPKM showed major genes involved in different metabolic pathways of the plant. Metabolic pathway analysis of the assembled transcripts identified 124 plant related pathways. The transcripts related to carotenoid and alkaloid biosynthetic pathways had more number of reads and FPKM values suggesting higher expression of these genes. The arecanut transcript sequences generated in the study showed high similarity with coconut, oil palm and date palm sequences retrieved from public domains. We also identified 6853 genic SSR regions in the arecanut. The possible primers were designed for SSR detection and this would simplify the future efforts in genetic characterization of arecanut.

  20. Bacteriophage Assembly

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    Anastasia A. Aksyuk

    2011-02-01

    Full Text Available Bacteriophages have been a model system to study assembly processes for over half a century. Formation of infectious phage particles involves specific protein-protein and protein-nucleic acid interactions, as well as large conformational changes of assembly precursors. The sequence and molecular mechanisms of phage assembly have been elucidated by a variety of methods. Differences and similarities of assembly processes in several different groups of bacteriophages are discussed in this review. The general principles of phage assembly are applicable to many macromolecular complexes.

  1. Genome Sequencing

    DEFF Research Database (Denmark)

    Sato, Shusei; Andersen, Stig Uggerhøj

    2014-01-01

    The current Lotus japonicus reference genome sequence is based on a hybrid assembly of Sanger TAC/BAC, Sanger shotgun and Illumina shotgun sequencing data generated from the Miyakojima-MG20 accession. It covers nearly all expressed L. japonicus genes and has been annotated mainly based on transcr......The current Lotus japonicus reference genome sequence is based on a hybrid assembly of Sanger TAC/BAC, Sanger shotgun and Illumina shotgun sequencing data generated from the Miyakojima-MG20 accession. It covers nearly all expressed L. japonicus genes and has been annotated mainly based...

  2. The carbohydrate-binding module (CBM)-like sequence is crucial for rice CWA1/BC1 function in proper assembly of secondary cell wall materials.

    Science.gov (United States)

    Sato, Kanna; Ito, Sachiko; Fujii, Takeo; Suzuki, Ryu; Takenouchi, Sachi; Nakaba, Satoshi; Funada, Ryo; Sano, Yuzou; Kajita, Shinya; Kitano, Hidemi; Katayama, Yoshihiro

    2010-11-01

    We recently reported that the cwa1 mutation disturbed the deposition and assembly of secondary cell wall materials in the cortical fiber of rice internodes. Genetic analysis revealed that cwa1 is allelic to bc1, which encodes glycosylphosphatidylinositol (GPI)-anchored COBRA-like protein with the highest homology to Arabidopsis COBRA-like 4 (COBL4) and maize Brittle Stalk 2 (Bk2). Our results suggested that CWA1/BC1 plays a role in assembling secondary cell wall materials at appropriate sites, enabling synthesis of highly ordered secondary cell wall structure with solid and flexible internodes in rice. The N-terminal amino acid sequence of CWA1/BC1, as well as its orthologs (COBL4, Bk2) and other BC1-like proteins in rice, shows weak similarity to a family II carbohydrate-binding module (CBM2) of several bacterial cellulases. To investigate the importance of the CBM-like sequence of CWA1/BC1 in the assembly of secondary cell wall materials, Trp residues in the CBM-like sequence, which is important for carbohydrate binding, were substituted for Val residues and introduced into the cwa1 mutant. CWA1/BC1 with the mutated sequence did not complement the abnormal secondary cell walls seen in the cwa1 mutant, indicating that the CBM-like sequence is essential for the proper function of CWA1/BC1, including assembly of secondary cell wall materials.

  3. Transcriptome Sequencing, De Novo Assembly and Differential Gene Expression Analysis of the Early Development of Acipenser baeri.

    Directory of Open Access Journals (Sweden)

    Wei Song

    Full Text Available The molecular mechanisms that drive the development of the endangered fossil fish species Acipenser baeri are difficult to study due to the lack of genomic data. Recent advances in sequencing technologies and the reducing cost of sequencing offer exclusive opportunities for exploring important molecular mechanisms underlying specific biological processes. This manuscript describes the large scale sequencing and analyses of mRNA from Acipenser baeri collected at five development time points using the Illumina Hiseq2000 platform. The sequencing reads were de novo assembled and clustered into 278167 unigenes, of which 57346 (20.62% had 45837 known homologues proteins in Uniprot protein databases while 11509 proteins matched with at least one sequence of assembled unigenes. The remaining 79.38% of unigenes could stand for non-coding unigenes or unigenes specific to A. baeri. A number of 43062 unigenes were annotated into functional categories via Gene Ontology (GO annotation whereas 29526 unigenes were associated with 329 pathways by mapping to KEGG database. Subsequently, 3479 differentially expressed genes were scanned within developmental stages and clustered into 50 gene expression profiles. Genes preferentially expressed at each stage were also identified. Through GO and KEGG pathway enrichment analysis, relevant physiological variations during the early development of A. baeri could be better cognized. Accordingly, the present study gives insights into the transcriptome profile of the early development of A. baeri, and the information contained in this large scale transcriptome will provide substantial references for A. baeri developmental biology and promote its aquaculture research.

  4. Assembling large, complex environmental metagenomes

    Energy Technology Data Exchange (ETDEWEB)

    Howe, A. C. [Michigan State Univ., East Lansing, MI (United States). Microbiology and Molecular Genetics, Plant Soil and Microbial Sciences; Jansson, J. [USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Earth Sciences Division; Malfatti, S. A. [USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States); Tringe, S. G. [USDOE Joint Genome Institute (JGI), Walnut Creek, CA (United States); Tiedje, J. M. [Michigan State Univ., East Lansing, MI (United States). Microbiology and Molecular Genetics, Plant Soil and Microbial Sciences; Brown, C. T. [Michigan State Univ., East Lansing, MI (United States). Microbiology and Molecular Genetics, Computer Science and Engineering

    2012-12-28

    The large volumes of sequencing data required to sample complex environments deeply pose new challenges to sequence analysis approaches. De novo metagenomic assembly effectively reduces the total amount of data to be analyzed but requires significant computational resources. We apply two pre-assembly filtering approaches, digital normalization and partitioning, to make large metagenome assemblies more computationaly tractable. Using a human gut mock community dataset, we demonstrate that these methods result in assemblies nearly identical to assemblies from unprocessed data. We then assemble two large soil metagenomes from matched Iowa corn and native prairie soils. The predicted functional content and phylogenetic origin of the assembled contigs indicate significant taxonomic differences despite similar function. The assembly strategies presented are generic and can be extended to any metagenome; full source code is freely available under a BSD license.

  5. An efficient approach to BAC based assembly of complex genomes.

    Science.gov (United States)

    Visendi, Paul; Berkman, Paul J; Hayashi, Satomi; Golicz, Agnieszka A; Bayer, Philipp E; Ruperao, Pradeep; Hurgobin, Bhavna; Montenegro, Juan; Chan, Chon-Kit Kenneth; Staňková, Helena; Batley, Jacqueline; Šimková, Hana; Doležel, Jaroslav; Edwards, David

    2016-01-01

    There has been an exponential growth in the number of genome sequencing projects since the introduction of next generation DNA sequencing technologies. Genome projects have increasingly involved assembly of whole genome data which produces inferior assemblies compared to traditional Sanger sequencing of genomic fragments cloned into bacterial artificial chromosomes (BACs). While whole genome shotgun sequencing using next generation sequencing (NGS) is relatively fast and inexpensive, this method is extremely challenging for highly complex genomes, where polyploidy or high repeat content confounds accurate assembly, or where a highly accurate 'gold' reference is required. Several attempts have been made to improve genome sequencing approaches by incorporating NGS methods, to variable success. We present the application of a novel BAC sequencing approach which combines indexed pools of BACs, Illumina paired read sequencing, a sequence assembler specifically designed for complex BAC assembly, and a custom bioinformatics pipeline. We demonstrate this method by sequencing and assembling BAC cloned fragments from bread wheat and sugarcane genomes. We demonstrate that our assembly approach is accurate, robust, cost effective and scalable, with applications for complete genome sequencing in large and complex genomes.

  6. Comprehensive transcriptome assembly of Chickpea (Cicer arietinum L. using sanger and next generation sequencing platforms: development and applications.

    Directory of Open Access Journals (Sweden)

    Himabindu Kudapa

    Full Text Available A comprehensive transcriptome assembly of chickpea has been developed using 134.95 million Illumina single-end reads, 7.12 million single-end FLX/454 reads and 139,214 Sanger expressed sequence tags (ESTs from >17 genotypes. This hybrid transcriptome assembly, referred to as Cicer arietinumTranscriptome Assembly version 2 (CaTA v2, available at http://data.comparative-legumes.org/transcriptomes/cicar/lista_cicar-201201, comprising 46,369 transcript assembly contigs (TACs has an N50 length of 1,726 bp and a maximum contig size of 15,644 bp. Putative functions were determined for 32,869 (70.8% of the TACs and gene ontology assignments were determined for 21,471 (46.3%. The new transcriptome assembly was compared with the previously available chickpea transcriptome assemblies as well as to the chickpea genome. Comparative analysis of CaTA v2 against transcriptomes of three legumes - Medicago, soybean and common bean, resulted in 27,771 TACs common to all three legumes indicating strong conservation of genes across legumes. CaTA v2 was also used for identification of simple sequence repeats (SSRs and intron spanning regions (ISRs for developing molecular markers. ISRs were identified by aligning TACs to the Medicago genome, and their putative mapping positions at chromosomal level were identified using transcript map of chickpea. Primer pairs were designed for 4,990 ISRs, each representing a single contig for which predicted positions are inferred and distributed across eight linkage groups. A subset of randomly selected ISRs representing all eight chickpea linkage groups were validated on five chickpea genotypes and showed 20% polymorphism with average polymorphic information content (PIC of 0.27. In summary, the hybrid transcriptome assembly developed and novel markers identified can be used for a variety of applications such as gene discovery, marker-trait association, diversity analysis etc., to advance genetics research and breeding

  7. Human Contamination in Public Genome Assemblies.

    Science.gov (United States)

    Kryukov, Kirill; Imanishi, Tadashi

    2016-01-01

    Contamination in genome assembly can lead to wrong or confusing results when using such genome as reference in sequence comparison. Although bacterial contamination is well known, the problem of human-originated contamination received little attention. In this study we surveyed 45,735 available genome assemblies for evidence of human contamination. We used lineage specificity to distinguish between contamination and conservation. We found that 154 genome assemblies contain fragments that with high confidence originate as contamination from human DNA. Majority of contaminating human sequences were present in the reference human genome assembly for over a decade. We recommend that existing contaminated genomes should be revised to remove contaminated sequence, and that new assemblies should be thoroughly checked for presence of human DNA before submitting them to public databases.

  8. Orthology Guided Assembly in highly heterozygous crops

    DEFF Research Database (Denmark)

    Ruttink, Tom; Sterck, Lieven; Rohde, Antje

    2013-01-01

    to outbreeding crop species hamper De Bruijn Graph-based de novo assembly algorithms, causing transcript fragmentation and the redundant assembly of allelic contigs. If multiple genotypes are sequenced to study genetic diversity, primary de novo assembly is best performed per genotype to limit the level......Despite current advances in next-generation sequencing data analysis procedures, de novo assembly of a reference sequence required for SNP discovery and expression analysis is still a major challenge in genetically uncharacterized, highly heterozygous species. High levels of polymorphism inherent...... of polymorphism and avoid transcript fragmentation. Here, we propose an Orthology Guided Assembly procedure that first uses sequence similarity (tBLASTn) to proteins of a model species to select allelic and fragmented contigs from all genotypes and then performs CAP3 clustering on a gene-by-gene basis. Thus, we...

  9. Optimizing Transcriptome Assemblies for Eleusine indica Leaf and Seedling by Combining Multiple Assemblies from Three De Novo Assemblers

    Directory of Open Access Journals (Sweden)

    Shu Chen

    2015-03-01

    Full Text Available Due to rapid advances in sequencing technology, increasing amounts of genomic and transcriptomic data are available for plant species, presenting enormous challenges for biocomputing analysis. A crucial first step for a successful transcriptomics-based study is the building of a high-quality assembly. Here, we utilized three different de novo assemblers (Trinity, Velvet, and CLC and the EvidentialGene pipeline tr2aacds to assemble two optimized transcript sets for the notorious weed species, . Two RNA sequencing (RNA-seq datasets from leaf and aboveground seedlings were processed using three assemblers, which resulted in 20 assemblies for each dataset. The contig numbers and N50 values of each assembly were compared to study the effect of read number, k-mer size, and in silico normalization on assembly output. The 20 assemblies were then processed through the tr2aacds pipeline to remove redundant transcripts and to select the transcript set with the best coding potential. Each assembly contributed a considerable proportion to the final transcript combination with the exception of the CLC-k14. Thus each assembler and parameter set did assemble better contigs for certain transcripts. The redundancy, total contig number, N50, fully assembled contig number, and transcripts related to target-site herbicide resistance were evaluated for the EvidentialGene and Trinity assemblies. Comparing the EvidentialGene set with the Trinity assembly revealed improved quality and reduced redundancy in both leaf and seedling EvidentialGene sets. The optimized transcriptome references will be useful for studying herbicide resistance in and the evolutionary process in the three allotetraploid offspring.

  10. Sequencing and de novo assembly of the transcriptome of the glassy-winged sharpshooter (Homalodisca vitripennis.

    Directory of Open Access Journals (Sweden)

    Raja Sekhar Nandety

    Full Text Available BACKGROUND: The glassy-winged sharpshooter Homalodisca vitripennis (Hemiptera: Cicadellidae, is a xylem-feeding leafhopper and important vector of the bacterium Xylella fastidiosa; the causal agent of Pierce's disease of grapevines. The functional complexity of the transcriptome of H. vitripennis has not been elucidated thus far. It is a necessary blueprint for an understanding of the development of H. vitripennis and for designing efficient biorational control strategies including those based on RNA interference. RESULTS: Here we elucidate and explore the transcriptome of adult H. vitripennis using high-throughput paired end deep sequencing and de novo assembly. A total of 32,803,656 paired-end reads were obtained with an average transcript length of 624 nucleotides. We assembled 32.9 Mb of the transcriptome of H. vitripennis that spanned across 47,265 loci and 52,708 transcripts. Comparison of our non-redundant database showed that 45% of the deduced proteins of H. vitripennis exhibit identity (e-value ≤1(-5 with known proteins. We assigned Gene Ontology (GO terms, Kyoto Encyclopedia of Genes and Genomes (KEGG annotations, and potential Pfam domains to each transcript isoform. In order to gain insight into the molecular basis of key regulatory genes of H. vitripennis, we characterized predicted proteins involved in the metabolism of juvenile hormone, and biogenesis of small RNAs (Dicer and Piwi sequences from the transcriptomic sequences. Analysis of transposable element sequences of H. vitripennis indicated that the genome is less expanded in comparison to many other insects with approximately 1% of the transcriptome carrying transposable elements. CONCLUSIONS: Our data significantly enhance the molecular resources available for future study and control of this economically important hemipteran. This transcriptional information not only provides a more nuanced understanding of the underlying biological and physiological mechanisms that

  11. Construction of high resolution genetic linkage maps to improve the soybean genome sequence assembly Glyma1.01

    Science.gov (United States)

    A landmark in soybean research, Glyma1.01, the first whole genome sequence of variety Williams 82 (Glycine max L. Merr.) was completed in 2010 and is widely used. However, because the assembly was primarily built based on the linkage maps constructed with a limited number of markers and recombinant...

  12. A sweetpotato gene index established by de novo assembly of pyrosequencing and Sanger sequences and mining for gene-based microsatellite markers

    Directory of Open Access Journals (Sweden)

    Solis Julio

    2010-10-01

    Full Text Available Abstract Background Sweetpotato (Ipomoea batatas (L. Lam., a hexaploid outcrossing crop, is an important staple and food security crop in developing countries in Africa and Asia. The availability of genomic resources for sweetpotato is in striking contrast to its importance for human nutrition. Previously existing sequence data were restricted to around 22,000 expressed sequence tag (EST sequences and ~ 1,500 GenBank sequences. We have used 454 pyrosequencing to augment the available gene sequence information to enhance functional genomics and marker design for this plant species. Results Two quarter 454 pyrosequencing runs used two normalized cDNA collections from stems and leaves from drought-stressed sweetpotato clone Tanzania and yielded 524,209 reads, which were assembled together with 22,094 publically available expressed sequence tags into 31,685 sets of overlapping DNA segments and 34,733 unassembled sequences. Blastx comparisons with the UniRef100 database allowed annotation of 23,957 contigs and 15,342 singletons resulting in 24,657 putatively unique genes. Further, 27,119 sequences had no match to protein sequences of UniRef100database. On the basis of this gene index, we have identified 1,661 gene-based microsatellite sequences, of which 223 were selected for testing and 195 were successfully amplified in a test panel of 6 hexaploid (I. batatas and 2 diploid (I. trifida accessions. Conclusions The sweetpotato gene index is a useful source for functionally annotated sweetpotato gene sequences that contains three times more gene sequence information for sweetpotato than previous EST assemblies. A searchable version of the gene index, including a blastn function, is available at http://www.cipotato.org/sweetpotato_gene_index.

  13. De novo assembly, gene annotation and marker development using Illumina paired-end transcriptome sequences in celery (Apium graveolens L..

    Directory of Open Access Journals (Sweden)

    Nan Fu

    Full Text Available BACKGROUND: Celery is an increasing popular vegetable species, but limited transcriptome and genomic data hinder the research to it. In addition, a lack of celery molecular markers limits the process of molecular genetic breeding. High-throughput transcriptome sequencing is an efficient method to generate a large transcriptome sequence dataset for gene discovery, molecular marker development and marker-assisted selection breeding. PRINCIPAL FINDINGS: Celery transcriptomes from four tissues were sequenced using Illumina paired-end sequencing technology. De novo assembling was performed to generate a collection of 42,280 unigenes (average length of 502.6 bp that represent the first transcriptome of the species. 78.43% and 48.93% of the unigenes had significant similarity with proteins in the National Center for Biotechnology Information (NCBI non-redundant protein database (Nr and Swiss-Prot database respectively, and 10,473 (24.77% unigenes were assigned to Clusters of Orthologous Groups (COG. 21,126 (49.97% unigenes harboring Interpro domains were annotated, in which 15,409 (36.45% were assigned to Gene Ontology(GO categories. Additionally, 7,478 unigenes were mapped onto 228 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG. Large numbers of simple sequence repeats (SSRs were indentified, and then the rate of successful amplication and polymorphism were investigated among 31 celery accessions. CONCLUSIONS: This study demonstrates the feasibility of generating a large scale of sequence information by Illumina paired-end sequencing and efficient assembling. Our results provide a valuable resource for celery research. The developed molecular markers are the foundation of further genetic linkage analysis and gene localization, and they will be essential to accelerate the process of breeding.

  14. Assembly of Repeat Content Using Next Generation Sequencing Data

    Energy Technology Data Exchange (ETDEWEB)

    labutti, Kurt; Kuo, Alan; Grigoriev, Igor; Copeland, Alex

    2014-03-17

    Repetitive organisms pose a challenge for short read assembly, and typically only unique regions and repeat regions shorter than the read length, can be accurately assembled. Recently, we have been investigating the use of Pacific Biosciences reads for de novo fungal assembly. We will present an assessment of the quality and degree of repeat reconstruction possible in a fungal genome using long read technology. We will also compare differences in assembly of repeat content using short read and long read technology.

  15. Spike protein assembly into the coronavirion: exploring the limits of its sequence requirements

    International Nuclear Information System (INIS)

    Bosch, Berend Jan; Haan, Cornelis A.M. de; Smits, Saskia L.; Rottier, Peter J.M.

    2005-01-01

    The coronavirus spike (S) protein, required for receptor binding and membrane fusion, is incorporated into the assembling virion by interactions with the viral membrane (M) protein. Earlier we showed that the ectodomain of the S protein is not involved in this process. Here we further defined the requirements of the S protein for virion incorporation. We show that the cytoplasmic domain, not the transmembrane domain, determines the association with the M protein and suffices to effect the incorporation into viral particles of chimeric spikes as well as of foreign viral glycoproteins. The essential sequence was mapped to the membrane-proximal region of the cytoplasmic domain, which is also known to be of critical importance for the fusion function of the S protein. Consistently, only short C-terminal truncations of the S protein were tolerated when introduced into the virus by targeted recombination. The important role of the about 38-residues cytoplasmic domain in the assembly of and membrane fusion by this approximately 1300 amino acids long protein is discussed

  16. Sequencing and assembly of low copy and genic regions of isolated Triticum aestivum chromosome arm 7DS

    Czech Academy of Sciences Publication Activity Database

    Berkman, O. J.; Skarshewski, A.; Lorenc, M. T.; Kubaláková, Marie; Šimková, Hana; Batley, J.; Doležel, Jaroslav; Edwards, D.

    2011-01-01

    Roč. 9, č. 7 (2011), s. 768-775 ISSN 1467-7644 R&D Projects: GA ČR GA521/07/1573; GA MŠk ED0007/01/01 Institutional research plan: CEZ:AV0Z50380511 Keywords : wheat genome sequence * chromosome 7 * genome assembly Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.442, year: 2011

  17. Technology for assembling and welding of top and bottom nozzles in fuel assembly

    International Nuclear Information System (INIS)

    Xia Chenglie; Wan Longfu

    1989-10-01

    The construction character, technology and sequence of assembling and welding, assembling jig used for preventing from deformation, and acceptance test of welding technology for top and bottom nozzles are presented

  18. Comparing de novo assemblers for 454 transcriptome data.

    Science.gov (United States)

    Kumar, Sujai; Blaxter, Mark L

    2010-10-16

    Roche 454 pyrosequencing has become a method of choice for generating transcriptome data from non-model organisms. Once the tens to hundreds of thousands of short (250-450 base) reads have been produced, it is important to correctly assemble these to estimate the sequence of all the transcripts. Most transcriptome assembly projects use only one program for assembling 454 pyrosequencing reads, but there is no evidence that the programs used to date are optimal. We have carried out a systematic comparison of five assemblers (CAP3, MIRA, Newbler, SeqMan and CLC) to establish best practices for transcriptome assemblies, using a new dataset from the parasitic nematode Litomosoides sigmodontis. Although no single assembler performed best on all our criteria, Newbler 2.5 gave longer contigs, better alignments to some reference sequences, and was fast and easy to use. SeqMan assemblies performed best on the criterion of recapitulating known transcripts, and had more novel sequence than the other assemblers, but generated an excess of small, redundant contigs. The remaining assemblers all performed almost as well, with the exception of Newbler 2.3 (the version currently used by most assembly projects), which generated assemblies that had significantly lower total length. As different assemblers use different underlying algorithms to generate contigs, we also explored merging of assemblies and found that the merged datasets not only aligned better to reference sequences than individual assemblies, but were also more consistent in the number and size of contigs. Transcriptome assemblies are smaller than genome assemblies and thus should be more computationally tractable, but are often harder because individual contigs can have highly variable read coverage. Comparing single assemblers, Newbler 2.5 performed best on our trial data set, but other assemblers were closely comparable. Combining differently optimal assemblies from different programs however gave a more credible

  19. Toward allotetraploid cotton genome assembly: integration of a high-density molecular genetic linkage map with DNA sequence information

    Science.gov (United States)

    2012-01-01

    Background Cotton is the world’s most important natural textile fiber and a significant oilseed crop. Decoding cotton genomes will provide the ultimate reference and resource for research and utilization of the species. Integration of high-density genetic maps with genomic sequence information will largely accelerate the process of whole-genome assembly in cotton. Results In this paper, we update a high-density interspecific genetic linkage map of allotetraploid cultivated cotton. An additional 1,167 marker loci have been added to our previously published map of 2,247 loci. Three new marker types, InDel (insertion-deletion) and SNP (single nucleotide polymorphism) developed from gene information, and REMAP (retrotransposon-microsatellite amplified polymorphism), were used to increase map density. The updated map consists of 3,414 loci in 26 linkage groups covering 3,667.62 cM with an average inter-locus distance of 1.08 cM. Furthermore, genome-wide sequence analysis was finished using 3,324 informative sequence-based markers and publicly-available Gossypium DNA sequence information. A total of 413,113 EST and 195 BAC sequences were physically anchored and clustered by 3,324 sequence-based markers. Of these, 14,243 ESTs and 188 BACs from different species of Gossypium were clustered and specifically anchored to the high-density genetic map. A total of 2,748 candidate unigenes from 2,111 ESTs clusters and 63 BACs were mined for functional annotation and classification. The 337 ESTs/genes related to fiber quality traits were integrated with 132 previously reported cotton fiber quality quantitative trait loci, which demonstrated the important roles in fiber quality of these genes. Higher-level sequence conservation between different cotton species and between the A- and D-subgenomes in tetraploid cotton was found, indicating a common evolutionary origin for orthologous and paralogous loci in Gossypium. Conclusion This study will serve as a valuable genomic resource

  20. Assembly of the PLT device

    International Nuclear Information System (INIS)

    Marino, R.

    1975-11-01

    The assembly of the PLT device began in June 1974 with a preassembly of the mechanical structure at a remote site. The preassembly sequence incorporated final fabrication procedures with an initial staging operation. This successful staging/fabrication procedure proved to be an invaluable asset when the final assembly was started in August 1974. The assembly continued with the initial reassembly of the previously tested structural components at the final machine site. Construction was interrupted at several points to allow for toroidal field coil, vacuum vessel, and poloidal coil installation. Two phases of toroidal field coil power tests were included in the assembly sequence prior to, and just after the vacuum vessel insertion

  1. Exact algorithms for haplotype assembly from whole-genome sequence data.

    Science.gov (United States)

    Chen, Zhi-Zhong; Deng, Fei; Wang, Lusheng

    2013-08-15

    Haplotypes play a crucial role in genetic analysis and have many applications such as gene disease diagnoses, association studies, ancestry inference and so forth. The development of DNA sequencing technologies makes it possible to obtain haplotypes from a set of aligned reads originated from both copies of a chromosome of a single individual. This approach is often known as haplotype assembly. Exact algorithms that can give optimal solutions to the haplotype assembly problem are highly demanded. Unfortunately, previous algorithms for this problem either fail to output optimal solutions or take too long time even executed on a PC cluster. We develop an approach to finding optimal solutions for the haplotype assembly problem under the minimum-error-correction (MEC) model. Most of the previous approaches assume that the columns in the input matrix correspond to (putative) heterozygous sites. This all-heterozygous assumption is correct for most columns, but it may be incorrect for a small number of columns. In this article, we consider the MEC model with or without the all-heterozygous assumption. In our approach, we first use new methods to decompose the input read matrix into small independent blocks and then model the problem for each block as an integer linear programming problem, which is then solved by an integer linear programming solver. We have tested our program on a single PC [a Linux (x64) desktop PC with i7-3960X CPU], using the filtered HuRef and the NA 12878 datasets (after applying some variant calling methods). With the all-heterozygous assumption, our approach can optimally solve the whole HuRef data set within a total time of 31 h (26 h for the most difficult block of the 15th chromosome and only 5 h for the other blocks). To our knowledge, this is the first time that MEC optimal solutions are completely obtained for the filtered HuRef dataset. Moreover, in the general case (without the all-heterozygous assumption), for the HuRef dataset our

  2. V-GAP: Viral genome assembly pipeline

    KAUST Repository

    Nakamura, Yoji

    2015-10-22

    Next-generation sequencing technologies have allowed the rapid determination of the complete genomes of many organisms. Although shotgun sequences from large genome organisms are still difficult to reconstruct perfect contigs each of which represents a full chromosome, those from small genomes have been assembled successfully into a very small number of contigs. In this study, we show that shotgun reads from phage genomes can be reconstructed into a single contig by controlling the number of read sequences used in de novo assembly. We have developed a pipeline to assemble small viral genomes with good reliability using a resampling method from shotgun data. This pipeline, named V-GAP (Viral Genome Assembly Pipeline), will contribute to the rapid genome typing of viruses, which are highly divergent, and thus will meet the increasing need for viral genome comparisons in metagenomic studies.

  3. V-GAP: Viral genome assembly pipeline

    KAUST Repository

    Nakamura, Yoji; Yasuike, Motoshige; Nishiki, Issei; Iwasaki, Yuki; Fujiwara, Atushi; Kawato, Yasuhiko; Nakai, Toshihiro; Nagai, Satoshi; Kobayashi, Takanori; Gojobori, Takashi; Ototake, Mitsuru

    2015-01-01

    Next-generation sequencing technologies have allowed the rapid determination of the complete genomes of many organisms. Although shotgun sequences from large genome organisms are still difficult to reconstruct perfect contigs each of which represents a full chromosome, those from small genomes have been assembled successfully into a very small number of contigs. In this study, we show that shotgun reads from phage genomes can be reconstructed into a single contig by controlling the number of read sequences used in de novo assembly. We have developed a pipeline to assemble small viral genomes with good reliability using a resampling method from shotgun data. This pipeline, named V-GAP (Viral Genome Assembly Pipeline), will contribute to the rapid genome typing of viruses, which are highly divergent, and thus will meet the increasing need for viral genome comparisons in metagenomic studies.

  4. Human Assisted Assembly Processes

    Energy Technology Data Exchange (ETDEWEB)

    CALTON,TERRI L.; PETERS,RALPH R.

    2000-01-01

    Automatic assembly sequencing and visualization tools are valuable in determining the best assembly sequences, but without Human Factors and Figure Models (HFFMs) it is difficult to evaluate or visualize human interaction. In industry, accelerating technological advances and shorter market windows have forced companies to turn to an agile manufacturing paradigm. This trend has promoted computerized automation of product design and manufacturing processes, such as automated assembly planning. However, all automated assembly planning software tools assume that the individual components fly into their assembled configuration and generate what appear to be a perfectly valid operations, but in reality the operations cannot physically be carried out by a human. Similarly, human figure modeling algorithms may indicate that assembly operations are not feasible and consequently force design modifications; however, if they had the capability to quickly generate alternative assembly sequences, they might have identified a feasible solution. To solve this problem HFFMs must be integrated with automated assembly planning to allow engineers to verify that assembly operations are possible and to see ways to make the designs even better. Factories will very likely put humans and robots together in cooperative environments to meet the demands for customized products, for purposes including robotic and automated assembly. For robots to work harmoniously within an integrated environment with humans the robots must have cooperative operational skills. For example, in a human only environment, humans may tolerate collisions with one another if they did not cause much pain. This level of tolerance may or may not apply to robot-human environments. Humans expect that robots will be able to operate and navigate in their environments without collisions or interference. The ability to accomplish this is linked to the sensing capabilities available. Current work in the field of cooperative

  5. Next-Generation Sequencing of the Chrysanthemum nankingense (Asteraceae) Transcriptome Permits Large-Scale Unigene Assembly and SSR Marker Discovery

    Science.gov (United States)

    Wang, Haibin; Jiang, Jiafu; Chen, Sumei; Qi, Xiangyu; Peng, Hui; Li, Pirui; Song, Aiping; Guan, Zhiyong; Fang, Weimin; Liao, Yuan; Chen, Fadi

    2013-01-01

    Background Simple sequence repeats (SSRs) are ubiquitous in eukaryotic genomes. Chrysanthemum is one of the largest genera in the Asteraceae family. Only few Chrysanthemum expressed sequence tag (EST) sequences have been acquired to date, so the number of available EST-SSR markers is very low. Methodology/Principal Findings Illumina paired-end sequencing technology produced over 53 million sequencing reads from C. nankingense mRNA. The subsequent de novo assembly yielded 70,895 unigenes, of which 45,789 (64.59%) unigenes showed similarity to the sequences in NCBI database. Out of 45,789 sequences, 107 have hits to the Chrysanthemum Nr protein database; 679 and 277 sequences have hits to the database of Helianthus and Lactuca species, respectively. MISA software identified a large number of putative EST-SSRs, allowing 1,788 primer pairs to be designed from the de novo transcriptome sequence and a further 363 from archival EST sequence. Among 100 primer pairs randomly chosen, 81 markers have amplicons and 20 are polymorphic for genotypes analysis in Chrysanthemum. The results showed that most (but not all) of the assays were transferable across species and that they exposed a significant amount of allelic diversity. Conclusions/Significance SSR markers acquired by transcriptome sequencing are potentially useful for marker-assisted breeding and genetic analysis in the genus Chrysanthemum and its related genera. PMID:23626799

  6. Spaced Seed Data Structures for De Novo Assembly

    Directory of Open Access Journals (Sweden)

    Inanç Birol

    2015-01-01

    Full Text Available De novo assembly of the genome of a species is essential in the absence of a reference genome sequence. Many scalable assembly algorithms use the de Bruijn graph (DBG paradigm to reconstruct genomes, where a table of subsequences of a certain length is derived from the reads, and their overlaps are analyzed to assemble sequences. Despite longer subsequences unlocking longer genomic features for assembly, associated increase in compute resources limits the practicability of DBG over other assembly archetypes already designed for longer reads. Here, we revisit the DBG paradigm to adapt it to the changing sequencing technology landscape and introduce three data structure designs for spaced seeds in the form of paired subsequences. These data structures address memory and run time constraints imposed by longer reads. We observe that when a fixed distance separates seed pairs, it provides increased sequence specificity with increased gap length. Further, we note that Bloom filters would be suitable to implicitly store spaced seeds and be tolerant to sequencing errors. Building on this concept, we describe a data structure for tracking the frequencies of observed spaced seeds. These data structure designs will have applications in genome, transcriptome and metagenome assemblies, and read error correction.

  7. Comparing de novo assemblers for 454 transcriptome data

    Directory of Open Access Journals (Sweden)

    Blaxter Mark L

    2010-10-01

    Full Text Available Abstract Background Roche 454 pyrosequencing has become a method of choice for generating transcriptome data from non-model organisms. Once the tens to hundreds of thousands of short (250-450 base reads have been produced, it is important to correctly assemble these to estimate the sequence of all the transcripts. Most transcriptome assembly projects use only one program for assembling 454 pyrosequencing reads, but there is no evidence that the programs used to date are optimal. We have carried out a systematic comparison of five assemblers (CAP3, MIRA, Newbler, SeqMan and CLC to establish best practices for transcriptome assemblies, using a new dataset from the parasitic nematode Litomosoides sigmodontis. Results Although no single assembler performed best on all our criteria, Newbler 2.5 gave longer contigs, better alignments to some reference sequences, and was fast and easy to use. SeqMan assemblies performed best on the criterion of recapitulating known transcripts, and had more novel sequence than the other assemblers, but generated an excess of small, redundant contigs. The remaining assemblers all performed almost as well, with the exception of Newbler 2.3 (the version currently used by most assembly projects, which generated assemblies that had significantly lower total length. As different assemblers use different underlying algorithms to generate contigs, we also explored merging of assemblies and found that the merged datasets not only aligned better to reference sequences than individual assemblies, but were also more consistent in the number and size of contigs. Conclusions Transcriptome assemblies are smaller than genome assemblies and thus should be more computationally tractable, but are often harder because individual contigs can have highly variable read coverage. Comparing single assemblers, Newbler 2.5 performed best on our trial data set, but other assemblers were closely comparable. Combining differently optimal assemblies

  8. Targeted isolation, sequence assembly and characterization of two white spruce (Picea glauca BAC clones for terpenoid synthase and cytochrome P450 genes involved in conifer defence reveal insights into a conifer genome

    Directory of Open Access Journals (Sweden)

    Ritland Carol

    2009-08-01

    Full Text Available Abstract Background Conifers are a large group of gymnosperm trees which are separated from the angiosperms by more than 300 million years of independent evolution. Conifer genomes are extremely large and contain considerable amounts of repetitive DNA. Currently, conifer sequence resources exist predominantly as expressed sequence tags (ESTs and full-length (FLcDNAs. There is no genome sequence available for a conifer or any other gymnosperm. Conifer defence-related genes often group into large families with closely related members. The goals of this study are to assess the feasibility of targeted isolation and sequence assembly of conifer BAC clones containing specific genes from two large gene families, and to characterize large segments of genomic DNA sequence for the first time from a conifer. Results We used a PCR-based approach to identify BAC clones for two target genes, a terpene synthase (3-carene synthase; 3CAR and a cytochrome P450 (CYP720B4 from a non-arrayed genomic BAC library of white spruce (Picea glauca. Shotgun genomic fragments isolated from the BAC clones were sequenced to a depth of 15.6- and 16.0-fold coverage, respectively. Assembly and manual curation yielded sequence scaffolds of 172 kbp (3CAR and 94 kbp (CYP720B4 long. Inspection of the genomic sequences revealed the intron-exon structures, the putative promoter regions and putative cis-regulatory elements of these genes. Sequences related to transposable elements (TEs, high complexity repeats and simple repeats were prevalent and comprised approximately 40% of the sequenced genomic DNA. An in silico simulation of the effect of sequencing depth on the quality of the sequence assembly provides direction for future efforts of conifer genome sequencing. Conclusion We report the first targeted cloning, sequencing, assembly, and annotation of large segments of genomic DNA from a conifer. We demonstrate that genomic BAC clones for individual members of multi-member gene

  9. Targeted isolation, sequence assembly and characterization of two white spruce (Picea glauca) BAC clones for terpenoid synthase and cytochrome P450 genes involved in conifer defence reveal insights into a conifer genome.

    Science.gov (United States)

    Hamberger, Björn; Hall, Dawn; Yuen, Mack; Oddy, Claire; Hamberger, Britta; Keeling, Christopher I; Ritland, Carol; Ritland, Kermit; Bohlmann, Jörg

    2009-08-06

    Conifers are a large group of gymnosperm trees which are separated from the angiosperms by more than 300 million years of independent evolution. Conifer genomes are extremely large and contain considerable amounts of repetitive DNA. Currently, conifer sequence resources exist predominantly as expressed sequence tags (ESTs) and full-length (FL)cDNAs. There is no genome sequence available for a conifer or any other gymnosperm. Conifer defence-related genes often group into large families with closely related members. The goals of this study are to assess the feasibility of targeted isolation and sequence assembly of conifer BAC clones containing specific genes from two large gene families, and to characterize large segments of genomic DNA sequence for the first time from a conifer. We used a PCR-based approach to identify BAC clones for two target genes, a terpene synthase (3-carene synthase; 3CAR) and a cytochrome P450 (CYP720B4) from a non-arrayed genomic BAC library of white spruce (Picea glauca). Shotgun genomic fragments isolated from the BAC clones were sequenced to a depth of 15.6- and 16.0-fold coverage, respectively. Assembly and manual curation yielded sequence scaffolds of 172 kbp (3CAR) and 94 kbp (CYP720B4) long. Inspection of the genomic sequences revealed the intron-exon structures, the putative promoter regions and putative cis-regulatory elements of these genes. Sequences related to transposable elements (TEs), high complexity repeats and simple repeats were prevalent and comprised approximately 40% of the sequenced genomic DNA. An in silico simulation of the effect of sequencing depth on the quality of the sequence assembly provides direction for future efforts of conifer genome sequencing. We report the first targeted cloning, sequencing, assembly, and annotation of large segments of genomic DNA from a conifer. We demonstrate that genomic BAC clones for individual members of multi-member gene families can be isolated in a gene-specific fashion. The

  10. De novo assembly and characterization of the garlic (Allium sativum) bud transcriptome by Illumina sequencing.

    Science.gov (United States)

    Sun, Xiudong; Zhou, Shumei; Meng, Fanlu; Liu, Shiqi

    2012-10-01

    Garlic is widely used as a spice throughout the world for the culinary value of its flavor and aroma, which are created by the chemical transformation of a series of organic sulfur compounds. To analyze the transcriptome of Allium sativum and discover the genes involved in sulfur metabolism, cDNAs derived from the total RNA of Allium sativum buds were analyzed by Illumina sequencing. Approximately 26.67 million 90 bp paired-end clean reads were achieved in two libraries. A total of 127,933 unigenes were generated by de novo assembly and were compared with the sequences in public databases. Of these, 45,286 unigenes had significant hits to the sequences in the Nr database, 29,514 showed significant similarity to known proteins in the Swiss-Prot database and, 20,706 and 21,952 unigenes had significant similarity to existing sequences in the KEGG and COG databases, respectively. Moreover, genes involved in organic sulfur biosynthesis were identified. These unigenes data will provide the foundation for research on gene expression, genomics and functional genomics in Allium sativum. Key message The obtained unigenes will provide the foundation for research on functional genomics in Allium sativum and its closely related species, and fill the gap of the existing plant EST database.

  11. De novo assembly of highly diverse viral populations

    Directory of Open Access Journals (Sweden)

    Yang Xiao

    2012-09-01

    Full Text Available Abstract Background Extensive genetic diversity in viral populations within infected hosts and the divergence of variants from existing reference genomes impede the analysis of deep viral sequencing data. A de novo population consensus assembly is valuable both as a single linear representation of the population and as a backbone on which intra-host variants can be accurately mapped. The availability of consensus assemblies and robustly mapped variants are crucial to the genetic study of viral disease progression, transmission dynamics, and viral evolution. Existing de novo assembly techniques fail to robustly assemble ultra-deep sequence data from genetically heterogeneous populations such as viruses into full-length genomes due to the presence of extensive genetic variability, contaminants, and variable sequence coverage. Results We present VICUNA, a de novo assembly algorithm suitable for generating consensus assemblies from genetically heterogeneous populations. We demonstrate its effectiveness on Dengue, Human Immunodeficiency and West Nile viral populations, representing a range of intra-host diversity. Compared to state-of-the-art assemblers designed for haploid or diploid systems, VICUNA recovers full-length consensus and captures insertion/deletion polymorphisms in diverse samples. Final assemblies maintain a high base calling accuracy. VICUNA program is publicly available at: http://www.broadinstitute.org/scientific-community/science/projects/viral-genomics/ viral-genomics-analysis-software. Conclusions We developed VICUNA, a publicly available software tool, that enables consensus assembly of ultra-deep sequence derived from diverse viral populations. While VICUNA was developed for the analysis of viral populations, its application to other heterogeneous sequence data sets such as metagenomic or tumor cell population samples may prove beneficial in these fields of research.

  12. Cyprinus carpio Genome sequencing and assembly

    NARCIS (Netherlands)

    Kolder, I.C.R.M.; Plas-Duivesteijn, van der Suzanne J.; Tan, G.; Wiegertjes, G.; Forlenza, M.; Guler, A.T.; Travin, D.Y.; Nakao, M.; Moritomo, T.; Irnazarow, I.; Jansen, H.J.

    2013-01-01

    Sequencing of the common carp (Cyprinus carpio carpio Linnaeus, 1758) genome, with the objective of establishing carp as a model organism to supplement the closely related zebrafish (Danio rerio). The sequenced individual is a homozygous female (by gynogenesis) of R3 x R8 carp, the heterozygous

  13. Sequencing, de novo assembling, and annotating the genome of the endangered Chinese crocodile lizard Shinisaurus crocodilurus.

    Science.gov (United States)

    Gao, Jian; Li, Qiye; Wang, Zongji; Zhou, Yang; Martelli, Paolo; Li, Fang; Xiong, Zijun; Wang, Jian; Yang, Huanming; Zhang, Guojie

    2017-07-01

    The Chinese crocodile lizard, Shinisaurus crocodilurus, is the only living representative of the monotypic family Shinisauridae under the order Squamata. It is an obligate semi-aquatic, viviparous, diurnal species restricted to specific portions of mountainous locations in southwestern China and northeastern Vietnam. However, in the past several decades, this species has undergone a rapid decrease in population size due to illegal poaching and habitat disruption, making this unique reptile species endangered and listed in the Convention on International Trade in Endangered Species of Wild Fauna and Flora Appendix II since 1990. A proposal to uplist it to Appendix I was passed at the Convention on International Trade in Endangered Species of Wild Fauna and Flora Seventeenth meeting of the Conference of the Parties in 2016. To promote the conservation of this species, we sequenced the genome of a male Chinese crocodile lizard using a whole-genome shotgun strategy on the Illumina HiSeq 2000 platform. In total, we generated ∼291 Gb of raw sequencing data (×149 depth) from 13 libraries with insert sizes ranging from 250 bp to 40 kb. After filtering for polymerase chain reaction-duplicated and low-quality reads, ∼137 Gb of clean data (×70 depth) were obtained for genome assembly. We yielded a draft genome assembly with a total length of 2.24 Gb and an N50 scaffold size of 1.47 Mb. The assembled genome was predicted to contain 20 150 protein-coding genes and up to 1114 Mb (49.6%) of repetitive elements. The genomic resource of the Chinese crocodile lizard will contribute to deciphering the biology of this organism and provides an essential tool for conservation efforts. It also provides a valuable resource for future study of squamate evolution. © The Authors 2017. Published by Oxford University Press.

  14. De Novo Assembly of Human Herpes Virus Type 1 (HHV-1) Genome, Mining of Non-Canonical Structures and Detection of Novel Drug-Resistance Mutations Using Short- and Long-Read Next Generation Sequencing Technologies.

    Science.gov (United States)

    Karamitros, Timokratis; Harrison, Ian; Piorkowska, Renata; Katzourakis, Aris; Magiorkinis, Gkikas; Mbisa, Jean Lutamyo

    2016-01-01

    Human herpesvirus type 1 (HHV-1) has a large double-stranded DNA genome of approximately 152 kbp that is structurally complex and GC-rich. This makes the assembly of HHV-1 whole genomes from short-read sequencing data technically challenging. To improve the assembly of HHV-1 genomes we have employed a hybrid genome assembly protocol using data from two sequencing technologies: the short-read Roche 454 and the long-read Oxford Nanopore MinION sequencers. We sequenced 18 HHV-1 cell culture-isolated clinical specimens collected from immunocompromised patients undergoing antiviral therapy. The susceptibility of the samples to several antivirals was determined by plaque reduction assay. Hybrid genome assembly resulted in a decrease in the number of contigs in 6 out of 7 samples and an increase in N(G)50 and N(G)75 of all 7 samples sequenced by both technologies. The approach also enhanced the detection of non-canonical contigs including a rearrangement between the unique (UL) and repeat (T/IRL) sequence regions of one sample that was not detectable by assembly of 454 reads alone. We detected several known and novel resistance-associated mutations in UL23 and UL30 genes. Genome-wide genetic variability ranged from genomes will be useful in determining genetic determinants of drug resistance, virulence, pathogenesis and viral evolution. The numerous, complex repeat regions of the HHV-1 genome currently remain a barrier towards this goal.

  15. The A, C, G, and T of Genome Assembly.

    Science.gov (United States)

    Wajid, Bilal; Sohail, Muhammad U; Ekti, Ali R; Serpedin, Erchin

    2016-01-01

    Genome assembly in its two decades of history has produced significant research, in terms of both biotechnology and computational biology. This contribution delineates sequencing platforms and their characteristics, examines key steps involved in filtering and processing raw data, explains assembly frameworks, and discusses quality statistics for the assessment of the assembled sequence. Furthermore, the paper explores recent Ubuntu-based software environments oriented towards genome assembly as well as some avenues for future research.

  16. Effect of amino acid sequence and pH on nanofiber formation of self-assembling peptides EAK16-II and EAK16-IV.

    Science.gov (United States)

    Hong, Yooseong; Legge, Raymond L; Zhang, S; Chen, P

    2003-01-01

    Atomic force microscopy (AFM) and axisymmetric drop shape analysis-profile (ASDA-P) were used to investigate the mechanism of self-assembly of peptides. The peptides chosen consisted of 16 alternating hydrophobic and hydrophilic amino acids, where the hydrophilic residues possess alternating negative and positive charges. Two types of peptides, AEAEAKAKAEAEAKAK (EAK16-II) and AEAEAEAEAKAKAKAK (EAK16-IV), were investigated in terms of nanostructure formation through self-assembly. The experimental results, which focused on the effects of the amino acid sequence and pH, show that the nanostructures formed by the peptides are dependent on the amino acid sequence and the pH of the solution. For pH conditions around neutrality, one of the peptides used in this study, EAK16-IV, forms globular assemblies and has lower surface tension at air-water interfaces than another peptide, EAK16-II, which forms fibrillar assemblies at the same pH. When the pH is lowered below 6.5 or raised above 7.5, there is a transition from globular to fibrillar structures for EAK16-IV, but EAK16-II does not show any structural transition. Surface tension measurements using ADSA-P showed different surface activities of peptides at air-water interfaces. EAK16-II does not show a significant difference in surface tension for the pH range between 4 and 9. However, EAK16-IV shows a noticeable decrease in surface tension at pH around neutrality, indicating that the formation of globular assemblies is related to the molecular hydrophobicity.

  17. Evaluation of GRCh38 and de novo haploid genome assemblies demonstrates the enduring quality of the reference assembly.

    Science.gov (United States)

    Schneider, Valerie A; Graves-Lindsay, Tina; Howe, Kerstin; Bouk, Nathan; Chen, Hsiu-Chuan; Kitts, Paul A; Murphy, Terence D; Pruitt, Kim D; Thibaud-Nissen, Françoise; Albracht, Derek; Fulton, Robert S; Kremitzki, Milinn; Magrini, Vincent; Markovic, Chris; McGrath, Sean; Steinberg, Karyn Meltz; Auger, Kate; Chow, William; Collins, Joanna; Harden, Glenn; Hubbard, Timothy; Pelan, Sarah; Simpson, Jared T; Threadgold, Glen; Torrance, James; Wood, Jonathan M; Clarke, Laura; Koren, Sergey; Boitano, Matthew; Peluso, Paul; Li, Heng; Chin, Chen-Shan; Phillippy, Adam M; Durbin, Richard; Wilson, Richard K; Flicek, Paul; Eichler, Evan E; Church, Deanna M

    2017-05-01

    The human reference genome assembly plays a central role in nearly all aspects of today's basic and clinical research. GRCh38 is the first coordinate-changing assembly update since 2009; it reflects the resolution of roughly 1000 issues and encompasses modifications ranging from thousands of single base changes to megabase-scale path reorganizations, gap closures, and localization of previously orphaned sequences. We developed a new approach to sequence generation for targeted base updates and used data from new genome mapping technologies and single haplotype resources to identify and resolve larger assembly issues. For the first time, the reference assembly contains sequence-based representations for the centromeres. We also expanded the number of alternate loci to create a reference that provides a more robust representation of human population variation. We demonstrate that the updates render the reference an improved annotation substrate, alter read alignments in unchanged regions, and impact variant interpretation at clinically relevant loci. We additionally evaluated a collection of new de novo long-read haploid assemblies and conclude that although the new assemblies compare favorably to the reference with respect to continuity, error rate, and gene completeness, the reference still provides the best representation for complex genomic regions and coding sequences. We assert that the collected updates in GRCh38 make the newer assembly a more robust substrate for comprehensive analyses that will promote our understanding of human biology and advance our efforts to improve health. © 2017 Schneider et al.; Published by Cold Spring Harbor Laboratory Press.

  18. Sequencing and de novo transcriptome assembly of the Chinese giant salamander (Andrias davidianus

    Directory of Open Access Journals (Sweden)

    Yong Huang

    2017-06-01

    Full Text Available Next-generation technologies for determination of genomics and transcriptomics composition have a wide range of applications. Andrias davidianus, has become an endangered amphibian species of salamander endemic in China. However, there is a lack of the molecular information. In this study, we obtained the RNA-Seq data from a pool of A. davidianus tissue including spleen, liver, muscle, kidney, skin, testis, gut and heart using Illumina HiSeq 2500 platform. A total of 15,398,997,600 bp were obtained, corresponding to 102,659,984 raw reads. A total of 102,659,984 reads were filtered after removing low-quality reads and trimming the adapter sequences. The Trinity program was used to de novo assemble 132,912 unigenes with an average length of 690 bp and N50 of 1263 bp. Unigenes were annotated through number of databases. These transcriptomic data of A. davidianus should open the door to molecular evolution studies based on the entire transcriptome or targeted genes of interest to sequence. The raw data in this study can be available in NCBI SRA database with accession number of SRP099564.

  19. De novo assembly and next-generation sequencing to analyse full-length gene variants from codon-barcoded libraries.

    Science.gov (United States)

    Cho, Namjin; Hwang, Byungjin; Yoon, Jung-ki; Park, Sangun; Lee, Joongoo; Seo, Han Na; Lee, Jeewon; Huh, Sunghoon; Chung, Jinsoo; Bang, Duhee

    2015-09-21

    Interpreting epistatic interactions is crucial for understanding evolutionary dynamics of complex genetic systems and unveiling structure and function of genetic pathways. Although high resolution mapping of en masse variant libraries renders molecular biologists to address genotype-phenotype relationships, long-read sequencing technology remains indispensable to assess functional relationship between mutations that lie far apart. Here, we introduce JigsawSeq for multiplexed sequence identification of pooled gene variant libraries by combining a codon-based molecular barcoding strategy and de novo assembly of short-read data. We first validate JigsawSeq on small sub-pools and observed high precision and recall at various experimental settings. With extensive simulations, we then apply JigsawSeq to large-scale gene variant libraries to show that our method can be reliably scaled using next-generation sequencing. JigsawSeq may serve as a rapid screening tool for functional genomics and offer the opportunity to explore evolutionary trajectories of protein variants.

  20. The A, C, G, and T of Genome Assembly

    Directory of Open Access Journals (Sweden)

    Bilal Wajid

    2016-01-01

    Full Text Available Genome assembly in its two decades of history has produced significant research, in terms of both biotechnology and computational biology. This contribution delineates sequencing platforms and their characteristics, examines key steps involved in filtering and processing raw data, explains assembly frameworks, and discusses quality statistics for the assessment of the assembled sequence. Furthermore, the paper explores recent Ubuntu-based software environments oriented towards genome assembly as well as some avenues for future research.

  1. De novo assembly and phasing of a Korean human genome.

    Science.gov (United States)

    Seo, Jeong-Sun; Rhie, Arang; Kim, Junsoo; Lee, Sangjin; Sohn, Min-Hwan; Kim, Chang-Uk; Hastie, Alex; Cao, Han; Yun, Ji-Young; Kim, Jihye; Kuk, Junho; Park, Gun Hwa; Kim, Juhyeok; Ryu, Hanna; Kim, Jongbum; Roh, Mira; Baek, Jeonghun; Hunkapiller, Michael W; Korlach, Jonas; Shin, Jong-Yeon; Kim, Changhoon

    2016-10-13

    Advances in genome assembly and phasing provide an opportunity to investigate the diploid architecture of the human genome and reveal the full range of structural variation across population groups. Here we report the de novo assembly and haplotype phasing of the Korean individual AK1 (ref. 1) using single-molecule real-time sequencing, next-generation mapping, microfluidics-based linked reads, and bacterial artificial chromosome (BAC) sequencing approaches. Single-molecule sequencing coupled with next-generation mapping generated a highly contiguous assembly, with a contig N50 size of 17.9 Mb and a scaffold N50 size of 44.8 Mb, resolving 8 chromosomal arms into single scaffolds. The de novo assembly, along with local assemblies and spanning long reads, closes 105 and extends into 72 out of 190 euchromatic gaps in the reference genome, adding 1.03 Mb of previously intractable sequence. High concordance between the assembly and paired-end sequences from 62,758 BAC clones provides strong support for the robustness of the assembly. We identify 18,210 structural variants by direct comparison of the assembly with the human reference, identifying thousands of breakpoints that, to our knowledge, have not been reported before. Many of the insertions are reflected in the transcriptome and are shared across the Asian population. We performed haplotype phasing of the assembly with short reads, long reads and linked reads from whole-genome sequencing and with short reads from 31,719 BAC clones, thereby achieving phased blocks with an N50 size of 11.6 Mb. Haplotigs assembled from single-molecule real-time reads assigned to haplotypes on phased blocks covered 89% of genes. The haplotigs accurately characterized the hypervariable major histocompatability complex region as well as demonstrating allele configuration in clinically relevant genes such as CYP2D6. This work presents the most contiguous diploid human genome assembly so far, with extensive investigation of

  2. Research on Key Technologies of Unit-Based CNC Machine Tool Assembly Design

    Directory of Open Access Journals (Sweden)

    Zhongqi Sheng

    2014-01-01

    Full Text Available Assembly is the part that produces the maximum workload and consumed time during product design and manufacturing process. CNC machine tool is the key basic equipment in manufacturing industry and research on assembly design technologies of CNC machine tool has theoretical significance and practical value. This study established a simplified ASRG for CNC machine tool. The connection between parts, semantic information of transmission, and geometric constraint information were quantified to assembly connection strength to depict the assembling difficulty level. The transmissibility based on trust relationship was applied on the assembly connection strength. Assembly unit partition based on assembly connection strength was conducted, and interferential assembly units were identified and revised. The assembly sequence planning and optimization of parts in each assembly unit and between assembly units was conducted using genetic algorithm. With certain type of high speed CNC turning center, as an example, this paper explored into the assembly modeling, assembly unit partition, and assembly sequence planning and optimization and realized the optimized assembly sequence of headstock of CNC machine tool.

  3. Sequencing of chloroplast genome using whole cellular DNA and Solexa sequencing technology

    Directory of Open Access Journals (Sweden)

    Jian eWu

    2012-11-01

    Full Text Available Sequencing of the chloroplast genome using traditional sequencing methods has been difficult because of its size (>120 kb and the complicated procedures required to prepare templates. To explore the feasibility of sequencing the chloroplast genome using DNA extracted from whole cells and Solexa sequencing technology, we sequenced whole cellular DNA isolated from leaves of three Brassica rapa accessions with one lane per accession. In total, 246 Mb, 362Mb, 361 Mb sequence data were generated for the three accessions Chiifu-401-42, Z16 and FT, respectively. Microreads were assembled by reference-guided assembly using the cpDNA sequences of B. rapa, Arabidopsis thaliana, and Nicotiana tabacum. We achieved coverage of more than 99.96% of the cp genome in the three tested accessions using the B. rapa sequence as the reference. When A. thaliana or N. tabacum sequences were used as references, 99.7–99.8% or 95.5–99.7% of the B. rapa chloroplast genome was covered, respectively. These results demonstrated that sequencing of whole cellular DNA isolated from young leaves using the Illumina Genome Analyzer is an efficient method for high-throughput sequencing of chloroplast genome.

  4. Detection of M-Sequences from Spike Sequence in Neuronal Networks

    Directory of Open Access Journals (Sweden)

    Yoshi Nishitani

    2012-01-01

    Full Text Available In circuit theory, it is well known that a linear feedback shift register (LFSR circuit generates pseudorandom bit sequences (PRBS, including an M-sequence with the maximum period of length. In this study, we tried to detect M-sequences known as a pseudorandom sequence generated by the LFSR circuit from time series patterns of stimulated action potentials. Stimulated action potentials were recorded from dissociated cultures of hippocampal neurons grown on a multielectrode array. We could find several M-sequences from a 3-stage LFSR circuit (M3. These results show the possibility of assembling LFSR circuits or its equivalent ones in a neuronal network. However, since the M3 pattern was composed of only four spike intervals, the possibility of an accidental detection was not zero. Then, we detected M-sequences from random spike sequences which were not generated from an LFSR circuit and compare the result with the number of M-sequences from the originally observed raster data. As a result, a significant difference was confirmed: a greater number of “0–1” reversed the 3-stage M-sequences occurred than would have accidentally be detected. This result suggests that some LFSR equivalent circuits are assembled in neuronal networks.

  5. An Improved Genome Assembly of Azadirachta indica A. Juss.

    Directory of Open Access Journals (Sweden)

    Neeraja M. Krishnan

    2016-07-01

    Full Text Available Neem (Azadirachta indica A. Juss., an evergreen tree of the Meliaceae family, is known for its medicinal, cosmetic, pesticidal and insecticidal properties. We had previously sequenced and published the draft genome of a neem plant, using mainly short read sequencing data. In this report, we present an improved genome assembly generated using additional short reads from Illumina and long reads from Pacific Biosciences SMRT sequencer. We assembled short reads and error-corrected long reads using Platanus, an assembler designed to perform well for heterozygous genomes. The updated genome assembly (v2.0 yielded 3- and 3.5-fold increase in N50 and N75, respectively; 2.6-fold decrease in the total number of scaffolds; 1.25-fold increase in the number of valid transcriptome alignments; 13.4-fold less misassembly and 1.85-fold increase in the percentage repeat, over the earlier assembly (v1.0. The current assembly also maps better to the genes known to be involved in the terpenoid biosynthesis pathway. Together, the data represent an improved assembly of the A. indica genome.

  6. Hapsembler: An Assembler for Highly Polymorphic Genomes

    Science.gov (United States)

    Donmez, Nilgun; Brudno, Michael

    As whole genome sequencing has become a routine biological experiment, algorithms for assembly of whole genome shotgun data has become a topic of extensive research, with a plethora of off-the-shelf methods that can reconstruct the genomes of many organisms. Simultaneously, several recently sequenced genomes exhibit very high polymorphism rates. For these organisms genome assembly remains a challenge as most assemblers are unable to handle highly divergent haplotypes in a single individual. In this paper we describe Hapsembler, an assembler for highly polymorphic genomes, which makes use of paired reads. Our experiments show that Hapsembler produces accurate and contiguous assemblies of highly polymorphic genomes, while performing on par with the leading tools on haploid genomes. Hapsembler is available for download at http://compbio.cs.toronto.edu/hapsembler.

  7. Evaluation of nine popular de novo assemblers in microbial genome assembly.

    Science.gov (United States)

    Forouzan, Esmaeil; Maleki, Masoumeh Sadat Mousavi; Karkhane, Ali Asghar; Yakhchali, Bagher

    2017-12-01

    Next generation sequencing (NGS) technologies are revolutionizing biology, with Illumina being the most popular NGS platform. Short read assembly is a critical part of most genome studies using NGS. Hence, in this study, the performance of nine well-known assemblers was evaluated in the assembly of seven different microbial genomes. Effect of different read coverage and k-mer parameters on the quality of the assembly were also evaluated on both simulated and actual read datasets. Our results show that the performance of assemblers on real and simulated datasets could be significantly different, mainly because of coverage bias. According to outputs on actual read datasets, for all studied read coverages (of 7×, 25× and 100×), SPAdes and IDBA-UD clearly outperformed other assemblers based on NGA50 and accuracy metrics. Velvet is the most conservative assembler with the lowest NGA50 and error rate. Copyright © 2017. Published by Elsevier B.V.

  8. De Novo Assembly of the Pea (Pisum sativum L. Nodule Transcriptome

    Directory of Open Access Journals (Sweden)

    Vladimir A. Zhukov

    2015-01-01

    Full Text Available The large size and complexity of the garden pea (Pisum sativum L. genome hamper its sequencing and the discovery of pea gene resources. Although transcriptome sequencing provides extensive information about expressed genes, some tissue-specific transcripts can only be identified from particular organs under appropriate conditions. In this study, we performed RNA sequencing of polyadenylated transcripts from young pea nodules and root tips on an Illumina GAIIx system, followed by de novo transcriptome assembly using the Trinity program. We obtained more than 58,000 and 37,000 contigs from “Nodules” and “Root Tips” assemblies, respectively. The quality of the assemblies was assessed by comparison with pea expressed sequence tags and transcriptome sequencing project data available from NCBI website. The “Nodules” assembly was compared with the “Root Tips” assembly and with pea transcriptome sequencing data from projects indicating tissue specificity. As a result, approximately 13,000 nodule-specific contigs were found and annotated by alignment to known plant protein-coding sequences and by Gene Ontology searching. Of these, 581 sequences were found to possess full CDSs and could thus be considered as novel nodule-specific transcripts of pea. The information about pea nodule-specific gene sequences can be applied for gene-based markers creation, polymorphism studies, and real-time PCR.

  9. Sequence finishing and mapping of Drosophila melanogasterheterochromatin

    Energy Technology Data Exchange (ETDEWEB)

    Hoskins, Roger A.; Carlson, Joseph W.; Kennedy, Cameron; Acevedo,David; Evans-Holm, Martha; Frise, Erwin; Wan, Kenneth H.; Park, Soo; Mendez-Lago, Maria; Rossi, Fabrizio; Villasante, Alfredo; Dimitri,Patrizio; Karpen, Gary H.; Celniker, Susan E.

    2007-06-15

    Genome sequences for most metazoans are incomplete due tothe presence of repeated DNA in the pericentromeric heterochromatin. Theheterochromatic regions of D. melanogaster contain 20 Mb of sequenceamenable to mapping, sequence assembly and finishing. Here we describethe generation of 15 Mb of finished or improved heterochromatic sequenceusing available clone resources and assembly and mapping methods. We alsoconstructed a BAC-based physical map that spans approximately 13 Mb ofthe pericentromeric heterochromatin, and a cytogenetic map that positionsapproximately 11 Mb of BAC contigs and sequence scaffolds in specificchromosomal locations. The integrated sequence assembly and maps greatlyimprove our understanding of the structure and composition of this poorlyunderstood fraction of a metazoan genome and provide a framework forfunctional analyses.

  10. AutoAssemblyD: a graphical user interface system for several genome assemblers.

    Science.gov (United States)

    Veras, Adonney Allan de Oliveira; de Sá, Pablo Henrique Caracciolo Gomes; Azevedo, Vasco; Silva, Artur; Ramos, Rommel Thiago Jucá

    2013-01-01

    Next-generation sequencing technologies have increased the amount of biological data generated. Thus, bioinformatics has become important because new methods and algorithms are necessary to manipulate and process such data. However, certain challenges have emerged, such as genome assembly using short reads and high-throughput platforms. In this context, several algorithms have been developed, such as Velvet, Abyss, Euler-SR, Mira, Edna, Maq, SHRiMP, Newbler, ALLPATHS, Bowtie and BWA. However, most such assemblers do not have a graphical interface, which makes their use difficult for users without computing experience given the complexity of the assembler syntax. Thus, to make the operation of such assemblers accessible to users without a computing background, we developed AutoAssemblyD, which is a graphical tool for genome assembly submission and remote management by multiple assemblers through XML templates. AssemblyD is freely available at https://sourceforge.net/projects/autoassemblyd. It requires Sun jdk 6 or higher.

  11. Genome Sequence, Assembly and Characterization of Two Metschnikowia fructicola Strains Used as Biocontrol Agents of Postharvest Diseases

    Directory of Open Access Journals (Sweden)

    Edoardo Piombo

    2018-04-01

    Full Text Available The yeast Metschnikowia fructicola was reported as an efficient biological control agent of postharvest diseases of fruits and vegetables, and it is the bases of the commercial formulated product “Shemer.” Several mechanisms of action by which M. fructicola inhibits postharvest pathogens were suggested including iron-binding compounds, induction of defense signaling genes, production of fungal cell wall degrading enzymes and relatively high amounts of superoxide anions. We assembled the whole genome sequence of two strains of M. fructicola using PacBio and Illumina shotgun sequencing technologies. Using the PacBio, a high-quality draft genome consisting of 93 contigs, with an estimated genome size of approximately 26 Mb, was obtained. Comparative analysis of M. fructicola proteins with the other three available closely related genomes revealed a shared core of homologous proteins coded by 5,776 genes. Comparing the genomes of the two M. fructicola strains using a SNP calling approach resulted in the identification of 564,302 homologous SNPs with 2,004 predicted high impact mutations. The size of the genome is exceptionally high when compared with those of available closely related organisms, and the high rate of homology among M. fructicola genes points toward a recent whole-genome duplication event as the cause of this large genome. Based on the assembled genome, sequences were annotated with a gene description and gene ontology (GO term and clustered in functional groups. Analysis of CAZymes family genes revealed 1,145 putative genes, and transcriptomic analysis of CAZyme expression levels in M. fructicola during its interaction with either grapefruit peel tissue or Penicillium digitatum revealed a high level of CAZyme gene expression when the yeast was placed in wounded fruit tissue.

  12. Quality Assessment of Domesticated Animal Genome Assemblies

    DEFF Research Database (Denmark)

    Seemann, Stefan E; Anthon, Christian; Palasca, Oana

    2015-01-01

    affected by the lack of genomic sequence. Herein, we quantify the quality of the genome assemblies of 20 domesticated animals and related species by assessing a range of measurable parameters, and we show that there is a positive correlation between the fraction of mappable reads from RNAseq data...... domesticated animal genomes still need to be sequenced deeper in order to produce high-quality assemblies. In the meanwhile, ironically, the extent to which RNAseq and other next-generation data is produced frequently far exceeds that of the genomic sequence. Furthermore, basic comparative analysis is often...

  13. Assembly of 500,000 inter-specific catfish expressed sequence tags and large scale gene-associated marker development for whole genome association studies

    Energy Technology Data Exchange (ETDEWEB)

    Catfish Genome Consortium; Wang, Shaolin; Peatman, Eric; Abernathy, Jason; Waldbieser, Geoff; Lindquist, Erika; Richardson, Paul; Lucas, Susan; Wang, Mei; Li, Ping; Thimmapuram, Jyothi; Liu, Lei; Vullaganti, Deepika; Kucuktas, Huseyin; Murdock, Christopher; Small, Brian C; Wilson, Melanie; Liu, Hong; Jiang, Yanliang; Lee, Yoona; Chen, Fei; Lu, Jianguo; Wang, Wenqi; Xu, Peng; Somridhivej, Benjaporn; Baoprasertkul, Puttharat; Quilang, Jonas; Sha, Zhenxia; Bao, Baolong; Wang, Yaping; Wang, Qun; Takano, Tomokazu; Nandi, Samiran; Liu, Shikai; Wong, Lilian; Kaltenboeck, Ludmilla; Quiniou, Sylvie; Bengten, Eva; Miller, Norman; Trant, John; Rokhsar, Daniel; Liu, Zhanjiang

    2010-03-23

    Background-Through the Community Sequencing Program, a catfish EST sequencing project was carried out through a collaboration between the catfish research community and the Department of Energy's Joint Genome Institute. Prior to this project, only a limited EST resource from catfish was available for the purpose of SNP identification. Results-A total of 438,321 quality ESTs were generated from 8 channel catfish (Ictalurus punctatus) and 4 blue catfish (Ictalurus furcatus) libraries, bringing the number of catfish ESTs to nearly 500,000. Assembly of all catfish ESTs resulted in 45,306 contigs and 66,272 singletons. Over 35percent of the unique sequences had significant similarities to known genes, allowing the identification of 14,776 unique genes in catfish. Over 300,000 putative SNPs have been identified, of which approximately 48,000 are high-quality SNPs identified from contigs with at least four sequences and the minor allele presence of at least two sequences in the contig. The EST resource should be valuable for identification of microsatellites, genome annotation, large-scale expression analysis, and comparative genome analysis. Conclusions-This project generated a large EST resource for catfish that captured the majority of the catfish transcriptome. The parallel analysis of ESTs from two closely related Ictalurid catfishes should also provide powerful means for the evaluation of ancient and recent gene duplications, and for the development of high-density microarrays in catfish. The inter- and intra-specific SNPs identified from all catfish EST dataset assembly will greatly benefit the catfish introgression breeding program and whole genome association studies.

  14. De Novo Sequencing and Assembly Analysis of Transcriptome in Pinus bungeana Zucc. ex Endl.

    Directory of Open Access Journals (Sweden)

    Qifei Cai

    2018-03-01

    Full Text Available To enrich the molecular data of Pinus bungeana Zucc. ex Endl. and study the regulating factors of different morphology controled by apical dominance. In this study, de novo assembly of transcriptome annotation was performed for two varieties of Pinus bungeana Zucc. ex Endl. that are obviously different in morphology. More than 147 million reads were produced, which were assembled into 88,092 unigenes. Based on a similarity search, 11,692 unigenes showed significant similarity to proteins from Picea sitchensis (Bong. Carr. From this collection of unigenes, a large number of molecular markers were identified, including 2829 simple sequence repeats (SSRs. A total of 158 unigenes expressed differently between two varieties, including 98 up-regulated and 60 down-regulated unigenes. Furthermore, among the differently expressed genes (DEGs, five genes which may impact the plant morphology were further validated by reverse transcription quantitative polymerase chain reaction (RT-qPCR. The five genes related to cytokinin oxidase/dehydrogenase (CKX, two-component response regulator ARR-A family (ARR-A, plant hormone signal transduction (AHP, and MADS-box transcription factors have a close relationship with apical dominance. This new dataset will be a useful resource for future genetic and genomic studies in Pinus bungeana Zucc. ex Endl.

  15. Biomedical Applications of Self-Assembling Peptides

    NARCIS (Netherlands)

    Radmalekshahi, Mazda; Lempsink, Ludwijn; Amidi, Maryam; Hennink, Wim E.; Mastrobattista, Enrico

    2016-01-01

    Self-assembling peptides have gained increasing attention as versatile molecules to generate diverse supramolecular structures with tunable functionality. Because of the possibility to integrate a wide range of functional domains into self-assembling peptides including cell attachment sequences,

  16. Single-molecule sequencing and Hi-C-based proximity-guided assembly of amaranth (Amaranthus hypochondriacus) chromosomes provide insights into genome evolution

    KAUST Repository

    Lightfoot, D. J.; Jarvis, David Erwin; Ramaraj, T.; Lee, R.; Jellen, E. N.; Maughan, P. J.

    2017-01-01

    Background: Amaranth (Amaranthus hypochondriacus) was a food staple among the ancient civilizations of Central and South America that has recently received increased attention due to the high nutritional value of the seeds, with the potential to help alleviate malnutrition and food security concerns, particularly in arid and semiarid regions of the developing world. Here, we present a reference-quality assembly of the amaranth genome which will assist the agronomic development of the species.Results: Utilizing single-molecule, real-time sequencing (Pacific Biosciences) and chromatin interaction mapping (Hi-C) to close assembly gaps and scaffold contigs, respectively, we improved our previously reported Illumina-based assembly to produce a chromosome-scale assembly with a scaffold N50 of 24.4 Mb. The 16 largest scaffolds contain 98% of the assembly and likely represent the haploid chromosomes (n = 16). To demonstrate the accuracy and utility of this approach, we produced physical and genetic maps and identified candidate genes for the betalain pigmentation pathway. The chromosome-scale assembly facilitated a genome-wide syntenic comparison of amaranth with other Amaranthaceae species, revealing chromosome loss and fusion events in amaranth that explain the reduction from the ancestral haploid chromosome number (n = 18) for a tetraploid member of the Amaranthaceae. as major evolutionary events in the 2n = 32 amaranths and clearly establish the homoeologous relationship among most of the subgenome chromosomes, which will facilitate future investigations of intragenomic changes that occurred post polyploidization.

  17. Challenges, Solutions, and Quality Metrics of Personal Genome Assembly in Advancing Precision Medicine

    Directory of Open Access Journals (Sweden)

    Wenming Xiao

    2016-04-01

    Full Text Available Even though each of us shares more than 99% of the DNA sequences in our genome, there are millions of sequence codes or structure in small regions that differ between individuals, giving us different characteristics of appearance or responsiveness to medical treatments. Currently, genetic variants in diseased tissues, such as tumors, are uncovered by exploring the differences between the reference genome and the sequences detected in the diseased tissue. However, the public reference genome was derived with the DNA from multiple individuals. As a result of this, the reference genome is incomplete and may misrepresent the sequence variants of the general population. The more reliable solution is to compare sequences of diseased tissue with its own genome sequence derived from tissue in a normal state. As the price to sequence the human genome has dropped dramatically to around $1000, it shows a promising future of documenting the personal genome for every individual. However, de novo assembly of individual genomes at an affordable cost is still challenging. Thus, till now, only a few human genomes have been fully assembled. In this review, we introduce the history of human genome sequencing and the evolution of sequencing platforms, from Sanger sequencing to emerging “third generation sequencing” technologies. We present the currently available de novo assembly and post-assembly software packages for human genome assembly and their requirements for computational infrastructures. We recommend that a combined hybrid assembly with long and short reads would be a promising way to generate good quality human genome assemblies and specify parameters for the quality assessment of assembly outcomes. We provide a perspective view of the benefit of using personal genomes as references and suggestions for obtaining a quality personal genome. Finally, we discuss the usage of the personal genome in aiding vaccine design and development, monitoring host

  18. Optimization of de novo transcriptome assembly from high-throughput short read sequencing data improves functional annotation for non-model organisms

    Directory of Open Access Journals (Sweden)

    Haznedaroglu Berat Z

    2012-07-01

    Full Text Available Abstract Background The k-mer hash length is a key factor affecting the output of de novo transcriptome assembly packages using de Bruijn graph algorithms. Assemblies constructed with varying single k-mer choices might result in the loss of unique contiguous sequences (contigs and relevant biological information. A common solution to this problem is the clustering of single k-mer assemblies. Even though annotation is one of the primary goals of a transcriptome assembly, the success of assembly strategies does not consider the impact of k-mer selection on the annotation output. This study provides an in-depth k-mer selection analysis that is focused on the degree of functional annotation achieved for a non-model organism where no reference genome information is available. Individual k-mers and clustered assemblies (CA were considered using three representative software packages. Pair-wise comparison analyses (between individual k-mers and CAs were produced to reveal missing Kyoto Encyclopedia of Genes and Genomes (KEGG ortholog identifiers (KOIs, and to determine a strategy that maximizes the recovery of biological information in a de novo transcriptome assembly. Results Analyses of single k-mer assemblies resulted in the generation of various quantities of contigs and functional annotations within the selection window of k-mers (k-19 to k-63. For each k-mer in this window, generated assemblies contained certain unique contigs and KOIs that were not present in the other k-mer assemblies. Producing a non-redundant CA of k-mers 19 to 63 resulted in a more complete functional annotation than any single k-mer assembly. However, a fraction of unique annotations remained (~0.19 to 0.27% of total KOIs in the assemblies of individual k-mers (k-19 to k-63 that were not present in the non-redundant CA. A workflow to recover these unique annotations is presented. Conclusions This study demonstrated that different k-mer choices result in various quantities

  19. Meta-IDBA: a de Novo assembler for metagenomic data.

    Science.gov (United States)

    Peng, Yu; Leung, Henry C M; Yiu, S M; Chin, Francis Y L

    2011-07-01

    Next-generation sequencing techniques allow us to generate reads from a microbial environment in order to analyze the microbial community. However, assembling of a set of mixed reads from different species to form contigs is a bottleneck of metagenomic research. Although there are many assemblers for assembling reads from a single genome, there are no assemblers for assembling reads in metagenomic data without reference genome sequences. Moreover, the performances of these assemblers on metagenomic data are far from satisfactory, because of the existence of common regions in the genomes of subspecies and species, which make the assembly problem much more complicated. We introduce the Meta-IDBA algorithm for assembling reads in metagenomic data, which contain multiple genomes from different species. There are two core steps in Meta-IDBA. It first tries to partition the de Bruijn graph into isolated components of different species based on an important observation. Then, for each component, it captures the slight variants of the genomes of subspecies from the same species by multiple alignments and represents the genome of one species, using a consensus sequence. Comparison of the performances of Meta-IDBA and existing assemblers, such as Velvet and Abyss for different metagenomic datasets shows that Meta-IDBA can reconstruct longer contigs with similar accuracy. Meta-IDBA toolkit is available at our website http://www.cs.hku.hk/~alse/metaidba. chin@cs.hku.hk.

  20. The chaperonin-60 universal target is a barcode for bacteria that enables de novo assembly of metagenomic sequence data.

    Science.gov (United States)

    Links, Matthew G; Dumonceaux, Tim J; Hemmingsen, Sean M; Hill, Janet E

    2012-01-01

    Barcoding with molecular sequences is widely used to catalogue eukaryotic biodiversity. Studies investigating the community dynamics of microbes have relied heavily on gene-centric metagenomic profiling using two genes (16S rRNA and cpn60) to identify and track Bacteria. While there have been criteria formalized for barcoding of eukaryotes, these criteria have not been used to evaluate gene targets for other domains of life. Using the framework of the International Barcode of Life we evaluated DNA barcodes for Bacteria. Candidates from the 16S rRNA gene and the protein coding cpn60 gene were evaluated. Within complete bacterial genomes in the public domain representing 983 species from 21 phyla, the largest difference between median pairwise inter- and intra-specific distances ("barcode gap") was found from cpn60. Distribution of sequence diversity along the ∼555 bp cpn60 target region was remarkably uniform. The barcode gap of the cpn60 universal target facilitated the faithful de novo assembly of full-length operational taxonomic units from pyrosequencing data from a synthetic microbial community. Analysis supported the recognition of both 16S rRNA and cpn60 as DNA barcodes for Bacteria. The cpn60 universal target was found to have a much larger barcode gap than 16S rRNA suggesting cpn60 as a preferred barcode for Bacteria. A large barcode gap for cpn60 provided a robust target for species-level characterization of data. The assembly of consensus sequences for barcodes was shown to be a reliable method for the identification and tracking of novel microbes in metagenomic studies.

  1. Combining de novo and reference-guided assembly with scaffold_builder

    NARCIS (Netherlands)

    Silva, G.G.; Dutilh, B.E.; Matthews, T.D.; Elkins, K.; Schmieder, R.; Dinsdale, E.A.; Edwards, R.A.

    2013-01-01

    Genome sequencing has become routine, however genome assembly still remains a challenge despite the computational advances in the last decade. In particular, the abundance of repeat elements in genomes makes it difficult to assemble them into a single complete sequence. Identical repeats shorter

  2. Improving Phrap-based assembly of the rat using "reliable" overlaps.

    Directory of Open Access Journals (Sweden)

    Michael Roberts

    2008-03-01

    Full Text Available The assembly methods used for whole-genome shotgun (WGS data have a major impact on the quality of resulting draft genomes. We present a novel algorithm to generate a set of "reliable" overlaps based on identifying repeat k-mers. To demonstrate the benefits of using reliable overlaps, we have created a version of the Phrap assembly program that uses only overlaps from a specific list. We call this version PhrapUMD. Integrating PhrapUMD and our "reliable-overlap" algorithm with the Baylor College of Medicine assembler, Atlas, we assemble the BACs from the Rattus norvegicus genome project. Starting with the same data as the Nov. 2002 Atlas assembly, we compare our results and the Atlas assembly to the 4.3 Mb of rat sequence in the 21 BACs that have been finished. Our version of the draft assembly of the 21 BACs increases the coverage of finished sequence from 93.4% to 96.3%, while simultaneously reducing the base error rate from 4.5 to 1.1 errors per 10,000 bases. There are a number of ways of assessing the relative merits of assemblies when the finished sequence is available. If one views the overall quality of an assembly as proportional to the inverse of the product of the error rate and sequence missed, then the assembly presented here is seven times better. The UMD Overlapper with options for reliable overlaps is available from the authors at http://www.genome.umd.edu. We also provide the changes to the Phrap source code enabling it to use only the reliable overlaps.

  3. Detection of a Usp-like gene in Calotropis procera plant from the de novo assembled genome contigs of the high-throughput sequencing dataset

    KAUST Repository

    Shokry, Ahmed M.

    2014-02-01

    The wild plant species Calotropis procera (C. procera) has many potential applications and beneficial uses in medicine, industry and ornamental field. It also represents an excellent source of genes for drought and salt tolerance. Genes encoding proteins that contain the conserved universal stress protein (USP) domain are known to provide organisms like bacteria, archaea, fungi, protozoa and plants with the ability to respond to a plethora of environmental stresses. However, information on the possible occurrence of Usp in C. procera is not available. In this study, we uncovered and characterized a one-class A Usp-like (UspA-like, NCBI accession No. KC954274) gene in this medicinal plant from the de novo assembled genome contigs of the high-throughput sequencing dataset. A number of GenBank accessions for Usp sequences were blasted with the recovered de novo assembled contigs. Homology modelling of the deduced amino acids (NCBI accession No. AGT02387) was further carried out using Swiss-Model, accessible via the EXPASY. Superimposition of C. procera USPA-like full sequence model on Thermus thermophilus USP UniProt protein (PDB accession No. Q5SJV7) was constructed using RasMol and Deep-View programs. The functional domains of the novel USPA-like amino acids sequence were identified from the NCBI conserved domain database (CDD) that provide insights into sequence structure/function relationships, as well as domain models imported from a number of external source databases (Pfam, SMART, COG, PRK, TIGRFAM). © 2014 Académie des sciences.

  4. Optimized core loading sequence for Ukraine WWER-1000 reactors

    International Nuclear Information System (INIS)

    Dye, M.; Shah, H.

    2015-01-01

    Fuel Assemblies (WFAs) experienced mechanical damage of the grids during loading at both South Ukraine 2 (SU2) and South Ukraine 3 (SU3). The grids were damaged due to high lateral loads exceeding their strength limit. The high lateral loads were caused by a combination of distortion and stiffness of the mixed core fuel assemblies and significant fuel assembly-to-fuel assembly interaction combined with the core loading sequence being used. To prevent damage of the WFA grids during core loading, Westinghouse has developed a loading sequence technique and loading aides (smooth sided dummies and top nozzle loading guides) designed to minimize fuel assembly-to-fuel assembly interaction while maximizing the potential for successful loading (i.e., no fuel assembly damage and minimized loading time). The loading sequence technique accounts for cycle-specific core loading patterns and is based on previous Westinghouse WWER core loading experience and fundamental principles. The loading aids are developed to “open-up” the target core location or to provide guidance into a target core location. The Westinghouse optimized core loading sequence and smooth sided dummies were utilized during the successful loading of SU3 Cycle 25 mixed core in March 2015, with no instances of fuel assembly damage and yet still provided considerable time savings relative to the 2012 and 2013 SU3 reload campaigns. (authors)

  5. Zseq: An Approach for Preprocessing Next-Generation Sequencing Data.

    Science.gov (United States)

    Alkhateeb, Abedalrhman; Rueda, Luis

    2017-08-01

    Next-generation sequencing technology generates a huge number of reads (short sequences), which contain a vast amount of genomic data. The sequencing process, however, comes with artifacts. Preprocessing of sequences is mandatory for further downstream analysis. We present Zseq, a linear method that identifies the most informative genomic sequences and reduces the number of biased sequences, sequence duplications, and ambiguous nucleotides. Zseq finds the complexity of the sequences by counting the number of unique k-mers in each sequence as its corresponding score and also takes into the account other factors such as ambiguous nucleotides or high GC-content percentage in k-mers. Based on a z-score threshold, Zseq sweeps through the sequences again and filters those with a z-score less than the user-defined threshold. Zseq algorithm is able to provide a better mapping rate; it reduces the number of ambiguous bases significantly in comparison with other methods. Evaluation of the filtered reads has been conducted by aligning the reads and assembling the transcripts using the reference genome as well as de novo assembly. The assembled transcripts show a better discriminative ability to separate cancer and normal samples in comparison with another state-of-the-art method. Moreover, de novo assembled transcripts from the reads filtered by Zseq have longer genomic sequences than other tested methods. Estimating the threshold of the cutoff point is introduced using labeling rules with optimistic results.

  6. Assembly of viral genomes from metagenomes

    Directory of Open Access Journals (Sweden)

    Saskia L Smits

    2014-12-01

    Full Text Available Viral infections remain a serious global health issue. Metagenomic approaches are increasingly used in the detection of novel viral pathogens but also to generate complete genomes of uncultivated viruses. In silico identification of complete viral genomes from sequence data would allow rapid phylogenetic characterization of these new viruses. Often, however, complete viral genomes are not recovered, but rather several distinct contigs derived from a single entity, some of which have no sequence homology to any known proteins. De novo assembly of single viruses from a metagenome is challenging, not only because of the lack of a reference genome, but also because of intrapopulation variation and uneven or insufficient coverage. Here we explored different assembly algorithms, remote homology searches, genome-specific sequence motifs, k-mer frequency ranking, and coverage profile binning to detect and obtain viral target genomes from metagenomes. All methods were tested on 454-generated sequencing datasets containing three recently described RNA viruses with a relatively large genome which were divergent to previously known viruses from the viral families Rhabdoviridae and Coronaviridae. Depending on specific characteristics of the target virus and the metagenomic community, different assembly and in silico gap closure strategies were successful in obtaining near complete viral genomes.

  7. Brain transcriptome sequencing and assembly of three songbird model systems for the study of social behavior

    Directory of Open Access Journals (Sweden)

    Christopher N. Balakrishnan

    2014-05-01

    Full Text Available Emberizid sparrows (emberizidae have played a prominent role in the study of avian vocal communication and social behavior. We present here brain transcriptomes for three emberizid model systems, song sparrow Melospiza melodia, white-throated sparrow Zonotrichia albicollis, and Gambel’s white-crowned sparrow Zonotrichia leucophrys gambelii. Each of the assemblies covered fully or in part, over 89% of the previously annotated protein coding genes in the zebra finch Taeniopygia guttata, with 16,846, 15,805, and 16,646 unique BLAST hits in song, white-throated and white-crowned sparrows, respectively. As in previous studies, we find tissue of origin (auditory forebrain versus hypothalamus and whole brain as an important determinant of overall expression profile. We also demonstrate the successful isolation of RNA and RNA-sequencing from post-mortem samples from building strikes and suggest that such an approach could be useful when traditional sampling opportunities are limited. These transcriptomes will be an important resource for the study of social behavior in birds and for data driven annotation of forthcoming whole genome sequences for these and other bird species.

  8. Assembling draft genomes using contiBAIT

    OpenAIRE

    O'Neill, Kieran; Hills, Mark; Gottlieb, Mike; Borkowski, Matthew; Karsan, Aly; Lansdorp, Peter M.

    2017-01-01

    A Summary: Massively parallel sequencing is now widely used, but data interpretation is only as good as the reference assembly to which it is aligned. While the number of reference assemblies has rapidly expanded, most of these remain at intermediate stages of completion, either as scaffold builds, or as chromosome builds (consisting of correctly ordered, but not necessarily correctly oriented scaffolds separated by gaps). Completion of de novo assemblies remains difficult, as regions that ar...

  9. Reliable Detection of Herpes Simplex Virus Sequence Variation by High-Throughput Resequencing.

    Science.gov (United States)

    Morse, Alison M; Calabro, Kaitlyn R; Fear, Justin M; Bloom, David C; McIntyre, Lauren M

    2017-08-16

    High-throughput sequencing (HTS) has resulted in data for a number of herpes simplex virus (HSV) laboratory strains and clinical isolates. The knowledge of these sequences has been critical for investigating viral pathogenicity. However, the assembly of complete herpesviral genomes, including HSV, is complicated due to the existence of large repeat regions and arrays of smaller reiterated sequences that are commonly found in these genomes. In addition, the inherent genetic variation in populations of isolates for viruses and other microorganisms presents an additional challenge to many existing HTS sequence assembly pipelines. Here, we evaluate two approaches for the identification of genetic variants in HSV1 strains using Illumina short read sequencing data. The first, a reference-based approach, identifies variants from reads aligned to a reference sequence and the second, a de novo assembly approach, identifies variants from reads aligned to de novo assembled consensus sequences. Of critical importance for both approaches is the reduction in the number of low complexity regions through the construction of a non-redundant reference genome. We compared variants identified in the two methods. Our results indicate that approximately 85% of variants are identified regardless of the approach. The reference-based approach to variant discovery captures an additional 15% representing variants divergent from the HSV1 reference possibly due to viral passage. Reference-based approaches are significantly less labor-intensive and identify variants across the genome where de novo assembly-based approaches are limited to regions where contigs have been successfully assembled. In addition, regions of poor quality assembly can lead to false variant identification in de novo consensus sequences. For viruses with a well-assembled reference genome, a reference-based approach is recommended.

  10. De novo assembly of a haplotype-resolved human genome.

    Science.gov (United States)

    Cao, Hongzhi; Wu, Honglong; Luo, Ruibang; Huang, Shujia; Sun, Yuhui; Tong, Xin; Xie, Yinlong; Liu, Binghang; Yang, Hailong; Zheng, Hancheng; Li, Jian; Li, Bo; Wang, Yu; Yang, Fang; Sun, Peng; Liu, Siyang; Gao, Peng; Huang, Haodong; Sun, Jing; Chen, Dan; He, Guangzhu; Huang, Weihua; Huang, Zheng; Li, Yue; Tellier, Laurent C A M; Liu, Xiao; Feng, Qiang; Xu, Xun; Zhang, Xiuqing; Bolund, Lars; Krogh, Anders; Kristiansen, Karsten; Drmanac, Radoje; Drmanac, Snezana; Nielsen, Rasmus; Li, Songgang; Wang, Jian; Yang, Huanming; Li, Yingrui; Wong, Gane Ka-Shu; Wang, Jun

    2015-06-01

    The human genome is diploid, and knowledge of the variants on each chromosome is important for the interpretation of genomic information. Here we report the assembly of a haplotype-resolved diploid genome without using a reference genome. Our pipeline relies on fosmid pooling together with whole-genome shotgun strategies, based solely on next-generation sequencing and hierarchical assembly methods. We applied our sequencing method to the genome of an Asian individual and generated a 5.15-Gb assembled genome with a haplotype N50 of 484 kb. Our analysis identified previously undetected indels and 7.49 Mb of novel coding sequences that could not be aligned to the human reference genome, which include at least six predicted genes. This haplotype-resolved genome represents the most complete de novo human genome assembly to date. Application of our approach to identify individual haplotype differences should aid in translating genotypes to phenotypes for the development of personalized medicine.

  11. The sequence d(CGGCGGCCGC) self-assembles into a two dimensional rhombic DNA lattice

    International Nuclear Information System (INIS)

    Venkadesh, S.; Mandal, P.K.; Gautham, N.

    2011-01-01

    Highlights: → This is the first crystal structure of a four-way junction with sticky ends. → Four junction structures bind to each other and form a rhombic cavity. → Each rhombus binds to others to form 'infinite' 2D tiles. → This is an example of bottom-up fabrication of a DNA nano-lattice. -- Abstract: We report here the crystal structure of the partially self-complementary decameric sequence d(CGGCGGCCGC), which self assembles to form a four-way junction with sticky ends. Each junction binds to four others through Watson-Crick base pairing at the sticky ends to form a rhombic structure. The rhombuses bind to each other and form two dimensional tiles. The tiles stack to form the crystal. The crystal diffracted in the space group P1 to a resolution of 2.5 A. The junction has the anti-parallel stacked-X conformation like other junction structures, though the formation of the rhombic net noticeably alters the details of the junction geometry.

  12. Harnessing Whole Genome Sequencing in Medical Mycology.

    Science.gov (United States)

    Cuomo, Christina A

    2017-01-01

    Comparative genome sequencing studies of human fungal pathogens enable identification of genes and variants associated with virulence and drug resistance. This review describes current approaches, resources, and advances in applying whole genome sequencing to study clinically important fungal pathogens. Genomes for some important fungal pathogens were only recently assembled, revealing gene family expansions in many species and extreme gene loss in one obligate species. The scale and scope of species sequenced is rapidly expanding, leveraging technological advances to assemble and annotate genomes with higher precision. By using iteratively improved reference assemblies or those generated de novo for new species, recent studies have compared the sequence of isolates representing populations or clinical cohorts. Whole genome approaches provide the resolution necessary for comparison of closely related isolates, for example, in the analysis of outbreaks or sampled across time within a single host. Genomic analysis of fungal pathogens has enabled both basic research and diagnostic studies. The increased scale of sequencing can be applied across populations, and new metagenomic methods allow direct analysis of complex samples.

  13. Automated ensemble assembly and validation of microbial genomes

    Science.gov (United States)

    2014-01-01

    Background The continued democratization of DNA sequencing has sparked a new wave of development of genome assembly and assembly validation methods. As individual research labs, rather than centralized centers, begin to sequence the majority of new genomes, it is important to establish best practices for genome assembly. However, recent evaluations such as GAGE and the Assemblathon have concluded that there is no single best approach to genome assembly. Instead, it is preferable to generate multiple assemblies and validate them to determine which is most useful for the desired analysis; this is a labor-intensive process that is often impossible or unfeasible. Results To encourage best practices supported by the community, we present iMetAMOS, an automated ensemble assembly pipeline; iMetAMOS encapsulates the process of running, validating, and selecting a single assembly from multiple assemblies. iMetAMOS packages several leading open-source tools into a single binary that automates parameter selection and execution of multiple assemblers, scores the resulting assemblies based on multiple validation metrics, and annotates the assemblies for genes and contaminants. We demonstrate the utility of the ensemble process on 225 previously unassembled Mycobacterium tuberculosis genomes as well as a Rhodobacter sphaeroides benchmark dataset. On these real data, iMetAMOS reliably produces validated assemblies and identifies potential contamination without user intervention. In addition, intelligent parameter selection produces assemblies of R. sphaeroides comparable to or exceeding the quality of those from the GAGE-B evaluation, affecting the relative ranking of some assemblers. Conclusions Ensemble assembly with iMetAMOS provides users with multiple, validated assemblies for each genome. Although computationally limited to small or mid-sized genomes, this approach is the most effective and reproducible means for generating high-quality assemblies and enables users to

  14. PAVE: Program for assembling and viewing ESTs

    Directory of Open Access Journals (Sweden)

    Bomhoff Matthew

    2009-08-01

    Full Text Available Abstract Background New sequencing technologies are rapidly emerging. Many laboratories are simultaneously working with the traditional Sanger ESTs and experimenting with ESTs generated by the 454 Life Science sequencers. Though Sanger ESTs have been used to generate contigs for many years, no program takes full advantage of the 5' and 3' mate-pair information, hence, many tentative transcripts are assembled into two separate contigs. The new 454 technology has the benefit of high-throughput expression profiling, but introduces time and space problems for assembling large contigs. Results The PAVE (Program for Assembling and Viewing ESTs assembler takes advantage of the 5' and 3' mate-pair information by requiring that the mate-pairs be assembled into the same contig and joined by n's if the two sub-contigs do not overlap. It handles the depth of 454 data sets by "burying" similar ESTs during assembly, which retains the expression level information while circumventing time and space problems. PAVE uses MegaBLAST for the clustering step and CAP3 for assembly, however it assembles incrementally to enforce the mate-pair constraint, bury ESTs, and reduce incorrect joins and splits. The PAVE data management system uses a MySQL database to store multiple libraries of ESTs along with their metadata; the management system allows multiple assemblies with variations on libraries and parameters. Analysis routines provide standard annotation for the contigs including a measure of differentially expressed genes across the libraries. A Java viewer program is provided for display and analysis of the results. Our results clearly show the benefit of using the PAVE assembler to explicitly use mate-pair information and bury ESTs for large contigs. Conclusion The PAVE assembler provides a software package for assembling Sanger and/or 454 ESTs. The assembly software, data management software, Java viewer and user's guide are freely available.

  15. PAVE: program for assembling and viewing ESTs.

    Science.gov (United States)

    Soderlund, Carol; Johnson, Eric; Bomhoff, Matthew; Descour, Anne

    2009-08-26

    New sequencing technologies are rapidly emerging. Many laboratories are simultaneously working with the traditional Sanger ESTs and experimenting with ESTs generated by the 454 Life Science sequencers. Though Sanger ESTs have been used to generate contigs for many years, no program takes full advantage of the 5' and 3' mate-pair information, hence, many tentative transcripts are assembled into two separate contigs. The new 454 technology has the benefit of high-throughput expression profiling, but introduces time and space problems for assembling large contigs. The PAVE (Program for Assembling and Viewing ESTs) assembler takes advantage of the 5' and 3' mate-pair information by requiring that the mate-pairs be assembled into the same contig and joined by n's if the two sub-contigs do not overlap. It handles the depth of 454 data sets by "burying" similar ESTs during assembly, which retains the expression level information while circumventing time and space problems. PAVE uses MegaBLAST for the clustering step and CAP3 for assembly, however it assembles incrementally to enforce the mate-pair constraint, bury ESTs, and reduce incorrect joins and splits. The PAVE data management system uses a MySQL database to store multiple libraries of ESTs along with their metadata; the management system allows multiple assemblies with variations on libraries and parameters. Analysis routines provide standard annotation for the contigs including a measure of differentially expressed genes across the libraries. A Java viewer program is provided for display and analysis of the results. Our results clearly show the benefit of using the PAVE assembler to explicitly use mate-pair information and bury ESTs for large contigs. The PAVE assembler provides a software package for assembling Sanger and/or 454 ESTs. The assembly software, data management software, Java viewer and user's guide are freely available.

  16. CAPRRESI: Chimera Assembly by Plasmid Recovery and Restriction Enzyme Site Insertion.

    Science.gov (United States)

    Santillán, Orlando; Ramírez-Romero, Miguel A; Dávila, Guillermo

    2017-06-25

    Here, we present chimera assembly by plasmid recovery and restriction enzyme site insertion (CAPRRESI). CAPRRESI benefits from many strengths of the original plasmid recovery method and introduces restriction enzyme digestion to ease DNA ligation reactions (required for chimera assembly). For this protocol, users clone wildtype genes into the same plasmid (pUC18 or pUC19). After the in silico selection of amino acid sequence regions where chimeras should be assembled, users obtain all the synonym DNA sequences that encode them. Ad hoc Perl scripts enable users to determine all synonym DNA sequences. After this step, another Perl script searches for restriction enzyme sites on all synonym DNA sequences. This in silico analysis is also performed using the ampicillin resistance gene (ampR) found on pUC18/19 plasmids. Users design oligonucleotides inside synonym regions to disrupt wildtype and ampR genes by PCR. After obtaining and purifying complementary DNA fragments, restriction enzyme digestion is accomplished. Chimera assembly is achieved by ligating appropriate complementary DNA fragments. pUC18/19 vectors are selected for CAPRRESI because they offer technical advantages, such as small size (2,686 base pairs), high copy number, advantageous sequencing reaction features, and commercial availability. The usage of restriction enzymes for chimera assembly eliminates the need for DNA polymerases yielding blunt-ended products. CAPRRESI is a fast and low-cost method for fusing protein-coding genes.

  17. Transcriptome sequencing of different narrow-leafed lupin tissue types provides a comprehensive uni-gene assembly and extensive gene-based molecular markers

    Science.gov (United States)

    Kamphuis, Lars G; Hane, James K; Nelson, Matthew N; Gao, Lingling; Atkins, Craig A; Singh, Karam B

    2015-01-01

    Narrow-leafed lupin (NLL; Lupinus angustifolius L.) is an important grain legume crop that is valuable for sustainable farming and is becoming recognized as a human health food. NLL breeding is directed at improving grain production, disease resistance, drought tolerance and health benefits. However, genetic and genomic studies have been hindered by a lack of extensive genomic resources for the species. Here, the generation, de novo assembly and annotation of transcriptome datasets derived from five different NLL tissue types of the reference accession cv. Tanjil are described. The Tanjil transcriptome was compared to transcriptomes of an early domesticated cv. Unicrop, a wild accession P27255, as well as accession 83A:476, together being the founding parents of two recombinant inbred line (RIL) populations. In silico predictions for transcriptome-derived gene-based length and SNP polymorphic markers were conducted and corroborated using a survey assembly sequence for NLL cv. Tanjil. This yielded extensive indel and SNP polymorphic markers for the two RIL populations. A total of 335 transcriptome-derived markers and 66 BAC-end sequence-derived markers were evaluated, and 275 polymorphic markers were selected to genotype the reference NLL 83A:476 × P27255 RIL population. This significantly improved the completeness, marker density and quality of the reference NLL genetic map. PMID:25060816

  18. Sequence-Dependent Self-Assembly and Structural Diversity of Islet Amyloid Polypeptide-Derived β-Sheet Fibrils

    International Nuclear Information System (INIS)

    Wang, Shih-Ting; Lin, Yiyang; Spencer, Ryan K.; Thomas, Michael R.; Nguyen, Andy I.

    2017-01-01

    Determining the structural origins of amyloid fibrillation is essential for understanding both the pathology of amyloidosis and the rational design of inhibitors to prevent or reverse amyloid formation. In this work, the decisive roles of peptide structures on amyloid self-assembly and morphological diversity were investigated by the design of eight amyloidogenic peptides derived from islet amyloid polypeptide. Among the segments, two distinct morphologies were highlighted in the form of twisted and planar (untwisted) ribbons with varied diameters, thicknesses, and lengths. In particular, transformation of amyloid fibrils from twisted ribbons into untwisted structures was triggered by substitution of the C-terminal serine with threonine, where the side chain methyl group was responsible for the distinct morphological change. This effect was confirmed following serine substitution with alanine and valine and was ascribed to the restriction of intersheet torsional strain through the increased hydrophobic interactions and hydrogen bonding. We also studied the variation of fibril morphology (i.e., association and helicity) and peptide aggregation propensity by increasing the hydrophobicity of the peptide side group, capping the N-terminus, and extending sequence length. Lastly, we anticipate that our insights into sequence-dependent fibrillation and morphological diversity will shed light on the structural interpretation of amyloidogenesis and development of structure-specific imaging agents and aggregation inhibitors.

  19. Assembly and comparative analysis of complete mitochondrial genome sequence of an economic plant Salix suchowensis

    Directory of Open Access Journals (Sweden)

    Ning Ye

    2017-03-01

    Full Text Available Willow is a widely used dioecious woody plant of Salicaceae family in China. Due to their high biomass yields, willows are promising sources for bioenergy crops. In this study, we assembled the complete mitochondrial (mt genome sequence of S. suchowensis with the length of 644,437 bp using Roche-454 GS FLX Titanium sequencing technologies. Base composition of the S. suchowensis mt genome is A (27.43%, T (27.59%, C (22.34%, and G (22.64%, which shows a prevalent GC content with that of other angiosperms. This long circular mt genome encodes 58 unique genes (32 protein-coding genes, 23 tRNA genes and 3 rRNA genes, and 9 of the 32 protein-coding genes contain 17 introns. Through the phylogenetic analysis of 35 species based on 23 protein-coding genes, it is supported that Salix as a sister to Populus. With the detailed phylogenetic information and the identification of phylogenetic position, some ribosomal protein genes and succinate dehydrogenase genes are found usually lost during evolution. As a native shrub willow species, this worthwhile research of S. suchowensis mt genome will provide more desirable information for better understanding the genomic breeding and missing pieces of sex determination evolution in the future.

  20. Mind the gap; seven reasons to close fragmented genome assemblies.

    Science.gov (United States)

    Thomma, Bart P H J; Seidl, Michael F; Shi-Kunne, Xiaoqian; Cook, David E; Bolton, Melvin D; van Kan, Jan A L; Faino, Luigi

    2016-05-01

    Like other domains of life, research into the biology of filamentous microbes has greatly benefited from the advent of whole-genome sequencing. Next-generation sequencing (NGS) technologies have revolutionized sequencing, making genomic sciences accessible to many academic laboratories including those that study non-model organisms. Thus, hundreds of fungal genomes have been sequenced and are publically available today, although these initiatives have typically yielded considerably fragmented genome assemblies that often lack large contiguous genomic regions. Many important genomic features are contained in intergenic DNA that is often missing in current genome assemblies, and recent studies underscore the significance of non-coding regions and repetitive elements for the life style, adaptability and evolution of many organisms. The study of particular types of genetic elements, such as telomeres, centromeres, repetitive elements, effectors, and clusters of co-regulated genes, but also of phenomena such as structural rearrangements, genome compartmentalization and epigenetics, greatly benefits from having a contiguous and high-quality, preferably even complete and gapless, genome assembly. Here we discuss a number of important reasons to produce gapless, finished, genome assemblies to help answer important biological questions. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Illustrating how mechanical assemblies work

    KAUST Repository

    Mitra, Niloy J.; Yang, Yongliang; Yan, Dongming; Li, Wilmot; Agrawala, Maneesh

    2010-01-01

    How things work visualizations use a variety of visual techniques to depict the operation of complex mechanical assemblies. We present an automated approach for generating such visualizations. Starting with a 3D CAD model of an assembly, we first infer the motions of individual parts and the interactions between parts based on their geometry and a few user specified constraints. We then use this information to generate visualizations that incorporate motion arrows, frame sequences and animation to convey the causal chain of motions and mechanical interactions between parts. We present results for a wide variety of assemblies. © 2010 ACM.

  2. Illustrating how mechanical assemblies work

    KAUST Repository

    Mitra, Niloy J.

    2010-07-26

    How things work visualizations use a variety of visual techniques to depict the operation of complex mechanical assemblies. We present an automated approach for generating such visualizations. Starting with a 3D CAD model of an assembly, we first infer the motions of individual parts and the interactions between parts based on their geometry and a few user specified constraints. We then use this information to generate visualizations that incorporate motion arrows, frame sequences and animation to convey the causal chain of motions and mechanical interactions between parts. We present results for a wide variety of assemblies. © 2010 ACM.

  3. Reference genome sequence of the model plant Setaria.

    Science.gov (United States)

    Bennetzen, Jeffrey L; Schmutz, Jeremy; Wang, Hao; Percifield, Ryan; Hawkins, Jennifer; Pontaroli, Ana C; Estep, Matt; Feng, Liang; Vaughn, Justin N; Grimwood, Jane; Jenkins, Jerry; Barry, Kerrie; Lindquist, Erika; Hellsten, Uffe; Deshpande, Shweta; Wang, Xuewen; Wu, Xiaomei; Mitros, Therese; Triplett, Jimmy; Yang, Xiaohan; Ye, Chu-Yu; Mauro-Herrera, Margarita; Wang, Lin; Li, Pinghua; Sharma, Manoj; Sharma, Rita; Ronald, Pamela C; Panaud, Olivier; Kellogg, Elizabeth A; Brutnell, Thomas P; Doust, Andrew N; Tuskan, Gerald A; Rokhsar, Daniel; Devos, Katrien M

    2012-05-13

    We generated a high-quality reference genome sequence for foxtail millet (Setaria italica). The ∼400-Mb assembly covers ∼80% of the genome and >95% of the gene space. The assembly was anchored to a 992-locus genetic map and was annotated by comparison with >1.3 million expressed sequence tag reads. We produced more than 580 million RNA-Seq reads to facilitate expression analyses. We also sequenced Setaria viridis, the ancestral wild relative of S. italica, and identified regions of differential single-nucleotide polymorphism density, distribution of transposable elements, small RNA content, chromosomal rearrangement and segregation distortion. The genus Setaria includes natural and cultivated species that demonstrate a wide capacity for adaptation. The genetic basis of this adaptation was investigated by comparing five sequenced grass genomes. We also used the diploid Setaria genome to evaluate the ongoing genome assembly of a related polyploid, switchgrass (Panicum virgatum).

  4. Reference genome sequence of the model plant Setaria

    Energy Technology Data Exchange (ETDEWEB)

    Bennetzen, Jeffrey L [ORNL; Schmutz, Jeremy [Hudson Alpha Institute of Biotechnology; Wang, Hao [University of Georgia, Athens, GA; Percifield, Ryan [University of Georgia, Athens, GA; Hawkins, Jennifer [University of Georgia, Athens, GA; Pontaroli, Ana C. [University of Georgia, Athens, GA; Estep, Matt [University of Georgia, Athens, GA; Feng, Liang [University of Georgia, Athens, GA; Vaughn, Justin N [ORNL; Grimwood, Jane [Hudson Alpha Institute of Biotechnology; Jenkins, Jerry [Hudson Alpha Institute of Biotechnology; Barry, Kerrie [U.S. Department of Energy, Joint Genome Institute; Lindquist, Erika [U.S. Department of Energy, Joint Genome Institute; Hellsten, Uffe [U.S. Department of Energy, Joint Genome Institute; Deshpande, Shweta [U.S. Department of Energy, Joint Genome Institute; Wang, Xuewen [University of Georgia, Athens, GA; Wu, Xiaomei [University of Georgia, Athens, GA; Mitros, Therese [University of California, Berkeley; Triplett, Jimmy [University of Missouri, St. Louis; Yang, Xiaohan [ORNL; Ye, Chuyu [ORNL; Mauro-Herrera, Margarita [Oklahoma State University; Wang, Lin [Cornell University; Li, Pinghua [Cornell University; Sharma, Manoj [University of California, Davis; Sharma, Rita [University of California, Davis; Ronald, Pamela [University of California, Davis; Panaud, Olivier [Universite de Perpignan, Perpignan, France; Kellogg, Elizabeth A. [University of Missouri, St. Louis; Brutnell, Thomas P. [Cornell University; Doust, Andrew N. [Oklahoma State University; Tuskan, Gerald A [ORNL; Rokhsar, Daniel [U.S. Department of Energy, Joint Genome Institute; Devos, Katrien M [ORNL

    2012-01-01

    We generated a high-quality reference genome sequence for foxtail millet (Setaria italica). The ~400-Mb assembly covers ~80% of the genome and >95% of the gene space. The assembly was anchored to a 992-locus genetic map and was annotated by comparison with >1.3 million expressed sequence tag reads. We produced more than 580 million RNA-Seq reads to facilitate expression analyses. We also sequenced Setaria viridis, the ancestral wild relative of S. italica, and identified regions of differential single-nucleotide polymorphism density, distribution of transposable elements, small RNA content, chromosomal rearrangement and segregation distortion. The genus Setaria includes natural and cultivated species that demonstrate a wide capacity for adaptation. The genetic basis of this adaptation was investigated by comparing five sequenced grass genomes. We also used the diploid Setaria genome to evaluate the ongoing genome assembly of a related polyploid, switchgrass (Panicum virgatum).

  5. Reference genome sequence of the model plant Setaria

    Energy Technology Data Exchange (ETDEWEB)

    Bennetzen, Jeffrey L [ORNL; Yang, Xiaohan [ORNL; Ye, Chuyu [ORNL; Tuskan, Gerald A [ORNL

    2012-01-01

    We generated a high-quality reference genome sequence for foxtail millet (Setaria italica). The {approx}400-Mb assembly covers {approx}80% of the genome and >95% of the gene space. The assembly was anchored to a 992-locus genetic map and was annotated by comparison with >1.3 million expressed sequence tag reads. We produced more than 580 million RNA-Seq reads to facilitate expression analyses. We also sequenced Setaria viridis, the ancestral wild relative of S. italica, and identified regions of differential single-nucleotide polymorphism density, distribution of transposable elements, small RNA content, chromosomal rearrangement and segregation distortion. The genus Setaria includes natural and cultivated species that demonstrate a wide capacity for adaptation. The genetic basis of this adaptation was investigated by comparing five sequenced grass genomes. We also used the diploid Setaria genome to evaluate the ongoing genome assembly of a related polyploid, switchgrass (Panicum virgatum).

  6. Elevation or Suppression? The Resolved Star Formation Main Sequence of Galaxies with Two Different Assembly Modes

    Science.gov (United States)

    Liu, Qing; Wang, Enci; Lin, Zesen; Gao, Yulong; Liu, Haiyang; Berhane Teklu, Berzaf; Kong, Xu

    2018-04-01

    We investigate the spatially resolved star formation main sequence in star-forming galaxies using Integral Field Spectroscopic observations from the Mapping Nearby Galaxies at the Apache Point Observatory survey. We demonstrate that the correlation between the stellar mass surface density (Σ*) and star formation rate surface density (ΣSFR) holds down to the sub-galactic scale, leading to the sub-galactic main sequence (SGMS). By dividing galaxies into two populations based on their recent mass assembly modes, we find the resolved main sequence in galaxies with the “outside-in” mode is steeper than that in galaxies with the “inside-out” mode. This is also confirmed on a galaxy-by-galaxy level, where we find the distributions of SGMS slopes for individual galaxies are clearly separated for the two populations. When normalizing and stacking the SGMS of individual galaxies on one panel for the two populations, we find that the inner regions of galaxies with the “inside-out” mode statistically exhibit a suppression in star formation, with a less significant trend in the outer regions of galaxies with the “outside-in” mode. In contrast, the inner regions of galaxies with “outside-in” mode and the outer regions of galaxies with “inside-out” mode follow a slightly sublinear scaling relation with a slope ∼0.9, which is in good agreement with previous findings, suggesting that they are experiencing a universal regulation without influences of additional physical processes.

  7. Comparing memory-efficient genome assemblers on stand-alone and cloud infrastructures.

    Science.gov (United States)

    Kleftogiannis, Dimitrios; Kalnis, Panos; Bajic, Vladimir B

    2013-01-01

    A fundamental problem in bioinformatics is genome assembly. Next-generation sequencing (NGS) technologies produce large volumes of fragmented genome reads, which require large amounts of memory to assemble the complete genome efficiently. With recent improvements in DNA sequencing technologies, it is expected that the memory footprint required for the assembly process will increase dramatically and will emerge as a limiting factor in processing widely available NGS-generated reads. In this report, we compare current memory-efficient techniques for genome assembly with respect to quality, memory consumption and execution time. Our experiments prove that it is possible to generate draft assemblies of reasonable quality on conventional multi-purpose computers with very limited available memory by choosing suitable assembly methods. Our study reveals the minimum memory requirements for different assembly programs even when data volume exceeds memory capacity by orders of magnitude. By combining existing methodologies, we propose two general assembly strategies that can improve short-read assembly approaches and result in reduction of the memory footprint. Finally, we discuss the possibility of utilizing cloud infrastructures for genome assembly and we comment on some findings regarding suitable computational resources for assembly.

  8. Comparing Memory-Efficient Genome Assemblers on Stand-Alone and Cloud Infrastructures

    KAUST Repository

    Kleftogiannis, Dimitrios A.

    2013-09-27

    A fundamental problem in bioinformatics is genome assembly. Next-generation sequencing (NGS) technologies produce large volumes of fragmented genome reads, which require large amounts of memory to assemble the complete genome efficiently. With recent improvements in DNA sequencing technologies, it is expected that the memory footprint required for the assembly process will increase dramatically and will emerge as a limiting factor in processing widely available NGS-generated reads. In this report, we compare current memory-efficient techniques for genome assembly with respect to quality, memory consumption and execution time. Our experiments prove that it is possible to generate draft assemblies of reasonable quality on conventional multi-purpose computers with very limited available memory by choosing suitable assembly methods. Our study reveals the minimum memory requirements for different assembly programs even when data volume exceeds memory capacity by orders of magnitude. By combining existing methodologies, we propose two general assembly strategies that can improve short-read assembly approaches and result in reduction of the memory footprint. Finally, we discuss the possibility of utilizing cloud infrastructures for genome assembly and we comment on some findings regarding suitable computational resources for assembly.

  9. Prediction of phenotypes of missense mutations in human proteins from biological assemblies.

    Science.gov (United States)

    Wei, Qiong; Xu, Qifang; Dunbrack, Roland L

    2013-02-01

    Single nucleotide polymorphisms (SNPs) are the most frequent variation in the human genome. Nonsynonymous SNPs that lead to missense mutations can be neutral or deleterious, and several computational methods have been presented that predict the phenotype of human missense mutations. These methods use sequence-based and structure-based features in various combinations, relying on different statistical distributions of these features for deleterious and neutral mutations. One structure-based feature that has not been studied significantly is the accessible surface area within biologically relevant oligomeric assemblies. These assemblies are different from the crystallographic asymmetric unit for more than half of X-ray crystal structures. We find that mutations in the core of proteins or in the interfaces in biological assemblies are significantly more likely to be disease-associated than those on the surface of the biological assemblies. For structures with more than one protein in the biological assembly (whether the same sequence or different), we find the accessible surface area from biological assemblies provides a statistically significant improvement in prediction over the accessible surface area of monomers from protein crystal structures (P = 6e-5). When adding this information to sequence-based features such as the difference between wildtype and mutant position-specific profile scores, the improvement from biological assemblies is statistically significant but much smaller (P = 0.018). Combining this information with sequence-based features in a support vector machine leads to 82% accuracy on a balanced dataset of 50% disease-associated mutations from SwissVar and 50% neutral mutations from human/primate sequence differences in orthologous proteins. Copyright © 2012 Wiley Periodicals, Inc.

  10. De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from total DNA Sequences.

    NARCIS (Netherlands)

    Izan, Shairul; Esselink, G.; Visser, R.G.F.; Smulders, M.J.M.; Borm, T.J.A.

    2017-01-01

    Whole Genome Shotgun (WGS) sequences of plant species often contain an abundance of reads that are derived from the chloroplast genome. Up to now these reads have generally been identified and assembled into chloroplast genomes based on homology to chloroplasts from related species. This

  11. Technical operations procedure for assembly and emplacement of the soil temperature test--test assembly

    International Nuclear Information System (INIS)

    Weber, A.P.

    1978-01-01

    A description is given of the plan for assembly, instrumentation, emplacement, and operational checkout of the soil temperature test assembly and dry well liner. The activities described cover all operations necessary to accomplish the receiving inspection, instrumentation and pre-construction handling of the dry well liner, plus all operations performed with the test article. Actual details of construction work are not covered by this procedure. Each part and/or section of this procedure is a separate function to be accomplished as required by the nature of the operation. The organization of the procedure is not intended to imply a special operational sequence or schedular requirement. Specific procedure operational sections include: receiving inspection; liner assembly operations; construction operations (by others); prepare shield plug; test article assembly and installation; and operational checkout

  12. Assembly-history dynamics of a pitcher-plant protozoan community in experimental microcosms.

    Directory of Open Access Journals (Sweden)

    Kohmei Kadowaki

    Full Text Available History drives community assembly through differences both in density (density effects and in the sequence in which species arrive (sequence effects. Density effects arise from predictable population dynamics, which are free of history, but sequence effects are due to a density-free mechanism, arising solely from the order and timing of immigration events. Few studies have determined how components of immigration history (timing, number of individuals, frequency alter local dynamics to determine community assembly, beyond addressing when immigration history produces historically contingent assembly.We varied density and sequence effects independently in a two-way factorial design to follow community assembly in a three-species aquatic protozoan community. A superior competitor, Colpoda steinii, mediated alternative community states; early arrival or high introduction density allowed this species to outcompete or suppress the other competitors (Poterioochromonas malhamensis and Eimeriidae gen. sp.. Multivariate analysis showed that density effects caused greater variation in community states, whereas sequence effects altered the mean community composition.A significant interaction between density and sequence effects suggests that we should refine our understanding of priority effects. These results highlight a practical need to understand not only the "ingredients" (species in ecological communities but their "recipes" as well.

  13. Two low coverage bird genomes and a comparison of reference-guided versus de novo genome assemblies.

    Science.gov (United States)

    Card, Daren C; Schield, Drew R; Reyes-Velasco, Jacobo; Fujita, Matthew K; Andrew, Audra L; Oyler-McCance, Sara J; Fike, Jennifer A; Tomback, Diana F; Ruggiero, Robert P; Castoe, Todd A

    2014-01-01

    As a greater number and diversity of high-quality vertebrate reference genomes become available, it is increasingly feasible to use these references to guide new draft assemblies for related species. Reference-guided assembly approaches may substantially increase the contiguity and completeness of a new genome using only low levels of genome coverage that might otherwise be insufficient for de novo genome assembly. We used low-coverage (∼3.5-5.5x) Illumina paired-end sequencing to assemble draft genomes of two bird species (the Gunnison Sage-Grouse, Centrocercus minimus, and the Clark's Nutcracker, Nucifraga columbiana). We used these data to estimate de novo genome assemblies and reference-guided assemblies, and compared the information content and completeness of these assemblies by comparing CEGMA gene set representation, repeat element content, simple sequence repeat content, and GC isochore structure among assemblies. Our results demonstrate that even lower-coverage genome sequencing projects are capable of producing informative and useful genomic resources, particularly through the use of reference-guided assemblies.

  14. Two low coverage bird genomes and a comparison of reference-guided versus de novo genome assemblies

    Science.gov (United States)

    Card, Daren C.; Schield, Drew R.; Reyes-Velasco, Jacobo; Fujita, Matthre K.; Andrew, Audra L.; Oyler-McCance, Sara J.; Fike, Jennifer A.; Tomback, Diana F.; Ruggiero, Robert P.; Castoe, Todd A.

    2014-01-01

    As a greater number and diversity of high-quality vertebrate reference genomes become available, it is increasingly feasible to use these references to guide new draft assemblies for related species. Reference-guided assembly approaches may substantially increase the contiguity and completeness of a new genome using only low levels of genome coverage that might otherwise be insufficient for de novo genome assembly. We used low-coverage (~3.5–5.5x) Illumina paired-end sequencing to assemble draft genomes of two bird species (the Gunnison Sage-Grouse, Centrocercus minimus, and the Clark's Nutcracker, Nucifraga columbiana). We used these data to estimate de novo genome assemblies and reference-guided assemblies, and compared the information content and completeness of these assemblies by comparing CEGMA gene set representation, repeat element content, simple sequence repeat content, and GC isochore structure among assemblies. Our results demonstrate that even lower-coverage genome sequencing projects are capable of producing informative and useful genomic resources, particularly through the use of reference-guided assemblies.

  15. Transcriptome sequencing of lentil based on second-generation technology permits large-scale unigene assembly and SSR marker discovery

    Directory of Open Access Journals (Sweden)

    Materne Michael

    2011-05-01

    Full Text Available Abstract Background Lentil (Lens culinaris Medik. is a cool-season grain legume which provides a rich source of protein for human consumption. In terms of genomic resources, lentil is relatively underdeveloped, in comparison to other Fabaceae species, with limited available data. There is hence a significant need to enhance such resources in order to identify novel genes and alleles for molecular breeding to increase crop productivity and quality. Results Tissue-specific cDNA samples from six distinct lentil genotypes were sequenced using Roche 454 GS-FLX Titanium technology, generating c. 1.38 × 106 expressed sequence tags (ESTs. De novo assembly generated a total of 15,354 contigs and 68,715 singletons. The complete unigene set was sequence-analysed against genome drafts of the model legume species Medicago truncatula and Arabidopsis thaliana to identify 12,639, and 7,476 unique matches, respectively. When compared to the genome of Glycine max, a total of 20,419 unique hits were observed corresponding to c. 31% of the known gene space. A total of 25,592 lentil unigenes were subsequently annoated from GenBank. Simple sequence repeat (SSR-containing ESTs were identified from consensus sequences and a total of 2,393 primer pairs were designed. A subset of 192 EST-SSR markers was screened for validation across a panel 12 cultivated lentil genotypes and one wild relative species. A total of 166 primer pairs obtained successful amplification, of which 47.5% detected genetic polymorphism. Conclusions A substantial collection of ESTs has been developed from sequence analysis of lentil genotypes using second-generation technology, permitting unigene definition across a broad range of functional categories. As well as providing resources for functional genomics studies, the unigene set has permitted significant enhancement of the number of publicly-available molecular genetic markers as tools for improvement of this species.

  16. Optimizing de novo common wheat transcriptome assembly using short-read RNA-Seq data

    Directory of Open Access Journals (Sweden)

    Duan Jialei

    2012-08-01

    Full Text Available Abstract Background Rapid advances in next-generation sequencing methods have provided new opportunities for transcriptome sequencing (RNA-Seq. The unprecedented sequencing depth provided by RNA-Seq makes it a powerful and cost-efficient method for transcriptome study, and it has been widely used in model organisms and non-model organisms to identify and quantify RNA. For non-model organisms lacking well-defined genomes, de novo assembly is typically required for downstream RNA-Seq analyses, including SNP discovery and identification of genes differentially expressed by phenotypes. Although RNA-Seq has been successfully used to sequence many non-model organisms, the results of de novo assembly from short reads can still be improved by using recent bioinformatic developments. Results In this study, we used 212.6 million pair-end reads, which accounted for 16.2 Gb, to assemble the hexaploid wheat transcriptome. Two state-of-the-art assemblers, Trinity and Trans-ABySS, which use the single and multiple k-mer methods, respectively, were used, and the whole de novo assembly process was divided into the following four steps: pre-assembly, merging different samples, removal of redundancy and scaffolding. We documented every detail of these steps and how these steps influenced assembly performance to gain insight into transcriptome assembly from short reads. After optimization, the assembled transcripts were comparable to Sanger-derived ESTs in terms of both continuity and accuracy. We also provided considerable new wheat transcript data to the community. Conclusions It is feasible to assemble the hexaploid wheat transcriptome from short reads. Special attention should be paid to dealing with multiple samples to balance the spectrum of expression levels and redundancy. To obtain an accurate overview of RNA profiling, removal of redundancy may be crucial in de novo assembly.

  17. Self-assembled Nanomaterials for Chemotherapeutic Applications

    Science.gov (United States)

    Shieh, Aileen

    The self-assembly of short designed peptides into functional nanostructures is becoming a growing interest in a wide range of fields from optoelectronic devices to nanobiotechnology. In the medical field, self-assembled peptides have especially attracted attention with several of its attractive features for applications in drug delivery, tissue regeneration, biological engineering as well as cosmetic industry and also the antibiotics field. We here describe the self-assembly of peptide conjugated with organic chromophore to successfully deliver sequence independent micro RNAs into human non-small cell lung cancer cell lines. The nanofiber used as the delivery vehicle is completely non-toxic and biodegradable, and exhibit enhanced permeability effect for targeting malignant tumors. The transfection efficiency with nanofiber as the delivery vehicle is comparable to that of the commercially available RNAiMAX lipofectamine while the toxicity is significantly lower. We also conjugated the peptide sequence with camptothecin (CPT) and observed the self-assembly of nanotubes for chemotherapeutic applications. The peptide scaffold is non-toxic and biodegradable, and drug loading of CPT is high, which minimizes the issue of systemic toxicity caused by extensive burden from the elimination of drug carriers. In addition, the peptide assembly drastically increases the solubility and stability of CPT under physiological conditions in vitro, while active CPT is gradually released from the peptide chain under the slight acidic tumor cell environment. Cytotoxicity results on human colorectal cancer cells and non-small cell lung cancer cell lines display promising anti-cancer properties compared to the parental CPT drug, which cannot be used clinically due to its poor solubility and lack of stability in physiological conditions. Moreover, the peptide sequence conjugated with 5-fluorouracil formed a hydrogel with promising topical chemotherapeutic applications that also display

  18. Illustrating how mechanical assemblies work

    KAUST Repository

    Mitra, Niloy J.; Yang, Yongliang; Yan, Dongming; Li, Wilmot; Agrawala, Maneesh

    2013-01-01

    How-things-work visualizations use a variety of visual techniques to depict the operation of complex mechanical assemblies. We present an automated approach for generating such visualizations. Starting with a 3D CAD model of an assembly, we first infer the motions of the individual parts and the interactions across the parts based on their geometry and a few user-specified constraints. We then use this information to generate visualizations that incorporate motion arrows, frame sequences, and animation to convey the causal chain of motions and mechanical interactions across parts. We demonstrate our system on a wide variety of assemblies. © 2013 ACM 0001-0782/13/01.

  19. Extreme-Scale De Novo Genome Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Georganas, Evangelos [Intel Corporation, Santa Clara, CA (United States); Hofmeyr, Steven [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Egan, Rob [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Computational Research Division; Buluc, Aydin [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Oliker, Leonid [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.; Rokhsar, Daniel [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Computational Research Division; Yelick, Katherine [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Joint Genome Inst.

    2017-09-26

    De novo whole genome assembly reconstructs genomic sequence from short, overlapping, and potentially erroneous DNA segments and is one of the most important computations in modern genomics. This work presents HipMER, a high-quality end-to-end de novo assembler designed for extreme scale analysis, via efficient parallelization of the Meraculous code. Genome assembly software has many components, each of which stresses different components of a computer system. This chapter explains the computational challenges involved in each step of the HipMer pipeline, the key distributed data structures, and communication costs in detail. We present performance results of assembling the human genome and the large hexaploid wheat genome on large supercomputers up to tens of thousands of cores.

  20. Population structure of pigs determined by single nucleotide polymorphisms observed in assembled expressed sequence tags.

    Science.gov (United States)

    Matsumoto, Toshimi; Okumura, Naohiko; Uenishi, Hirohide; Hayashi, Takeshi; Hamasima, Noriyuki; Awata, Takashi

    2012-01-01

    We have collected more than 190000 porcine expressed sequence tags (ESTs) from full-length complementary DNA (cDNA) libraries and identified more than 2800 single nucleotide polymorphisms (SNPs). In this study, we tentatively chose 222 SNPs observed in assembled ESTs to study pigs of different breeds; 104 were selected by comparing the cDNA sequences of a Meishan pig and samples of three-way cross pigs (Landrace, Large White, and Duroc: LWD), and 118 were selected from LWD samples. To evaluate the genetic variation between the chosen SNPs from pig breeds, we determined the genotypes for 192 pig samples (11 pig groups) from our DNA reference panel with matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Of the 222 reference SNPs, 186 were successfully genotyped. A neighbor-joining tree showed that the pig groups were classified into two large clusters, namely, Euro-American and East Asian pig populations. F-statistics and the analysis of molecular variance of Euro-American pig groups revealed that approximately 25% of the genetic variations occurred because of intergroup differences. As the F(IS) values were less than the F(ST) values(,) the clustering, based on the Bayesian inference, implied that there was strong genetic differentiation among pig groups and less divergence within the groups in our samples. © 2011 The Authors. Animal Science Journal © 2011 Japanese Society of Animal Science.

  1. The diploid genome sequence of an Asian individual

    DEFF Research Database (Denmark)

    Wang, Jun; Wang, Wei; Li, Ruiqiang

    2008-01-01

    Here we present the first diploid genome sequence of an Asian individual. The genome was sequenced to 36-fold average coverage using massively parallel sequencing technology. We aligned the short reads onto the NCBI human reference genome to 99.97% coverage, and guided by the reference genome, we...... used uniquely mapped reads to assemble a high-quality consensus sequence for 92% of the Asian individual's genome. We identified approximately 3 million single-nucleotide polymorphisms (SNPs) inside this region, of which 13.6% were not in the dbSNP database. Genotyping analysis showed that SNP...... identification had high accuracy and consistency, indicating the high sequence quality of this assembly. We also carried out heterozygote phasing and haplotype prediction against HapMap CHB and JPT haplotypes (Chinese and Japanese, respectively), sequence comparison with the two available individual genomes (J...

  2. THESEUS: A wavelength division multiplexed/microwave subcarrier multiplexed optical network, its ATM switch applications and device requirements

    Science.gov (United States)

    Xin, Wei

    1997-10-01

    A Terabit Hybrid Electro-optical /underline[Se]lf- routing Ultrafast Switch (THESEUS) has been proposed. It is a self-routing wavelength division multiplexed (WDM) / microwave subcarrier multiplexed (SCM) asynchronous transfer mode (ATM) switch for the multirate ATM networks. It has potential to be extended to a large ATM switch as 1000 x 1000 without internal blocking. Among the advantages of the hybrid implementation are flexibility in service upgrade, relaxed tolerances on optical filtering, protocol simplification and less processing overhead. For a small ATM switch, the subcarrier can be used as output buffers to solve output contention. A mathematical analysis was conducted to evaluate different buffer configurations. A testbed has been successfully constructed. Multirate binary data streams have been switched through the testbed and error free reception ([<]10-9 bit error rate) has been achieved. A simple, intuitive theoretical model has been developed to describe the heterodyne optical beat interference. A new concept of interference time and interference length has been introduced. An experimental confirmation has been conducted. The experimental results match the model very well. It shows that a large portion of optical bandwidth is wasted due to the beat interference. Based on the model, several improvement approaches have been proposed. The photo-generated carrier lifetime of silicon germanium has been measured using time-resolved reflectivity measurement. Via oxygen ion implantation, the carrier lifetime has been reduced to as short as 1 ps, corresponding to 1 THz of photodetector bandwidth. It has also been shown that copper dopants act as recombination centers in the silicon germanium.

  3. BAUM: Improving genome assembly by adaptive unique mapping and local overlap-layout-consensus approach.

    Science.gov (United States)

    Wang, Anqi; Wang, Zhanyu; Li, Zheng; Li, Lei M

    2018-01-15

    It is highly desirable to assemble genomes of high continuity and consistency at low cost. The current bottleneck of draft genome continuity using the Second Generation Sequencing (SGS) reads is primarily caused by uncertainty among repetitive sequences. Even though the Single-Molecule Real-Time sequencing technology is very promising to overcome the uncertainty issue, its relatively high cost and error rate add burden on budget or computation. Many long-read assemblers take the overlap-layout-consensus (OLC) paradigm, which is less sensitive to sequencing errors, heterozygosity and variability of coverage. However, current assemblers of SGS data do not sufficiently take advantage of the OLC approach. Aiming at minimizing uncertainty, the proposed method BAUM, breaks the whole genome into regions by adaptive unique mapping; then the local OLC is used to assemble each region in parallel. BAUM can: (1) perform reference-assisted assembly based on the genome of a close species; (2) or improve the results of existing assemblies that are obtained based on short or long sequencing reads. The tests on two eukaryote genomes, a wild rice Oryza longistaminata and a parrot Melopsittacus undulatus, show that BAUM achieved substantial improvement on genome size and continuity. Besides, BAUM reconstructed a considerable amount of repetitive regions that failed to be assembled by existing short read assemblers. We also propose statistical approaches to control the uncertainty in different steps of BAUM. http://www.zhanyuwang.xin/wordpress/index.php/2017/07/21/baum. lilei@amss.ac.cn. Supplementary data are available at Bioinformatics online. © The Author (2018). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  4. A Framework for Geometric Reasoning About Human Figures and Factors in Assembly Processes

    Energy Technology Data Exchange (ETDEWEB)

    Calton, Terri L.

    1999-07-20

    Automatic assembly sequencing and visualization tools are valuable in determining the best assembly sequences, but without Human Factors and Figure Models (HFFMs) it is difficult to evaluate or visualize human interaction. In industry, accelerating technological advances and shorter market windows have forced companies to turn to an agile manufacturing paradigm. This trend has promoted computerized automation of product design and manufacturing processes, such as automated assembly planning. However, all automated assembly planning software tools assume that the individual components fly into their assembled configuration and generate what appear to be perfectly valid operations, but in reality some operations cannot physically be carried out by a human. For example, the use of a ratchet may be reasoned feasible for an assembly operation; however, when a hand is placed on the tool the operation is no longer feasible, perhaps because of inaccessibility, insufficient strength or human interference with assembly components. Similarly, human figure modeling algorithms may indicate that assembly operations are not feasible and consequently force design modifications, however, if they had the capability to quickly generate alternative assembly sequences, they might have identified a feasible solution. To solve this problem, HFFMs must be integrated with automated assembly planning which allows engineers to quickly verify that assembly operations are possible and to see ways to make the designs even better. This paper presents a framework for integrating geometry-based assembly planning algorithms with commercially available human figure modeling software packages. Experimental results to selected applications along with lessons learned are presented.

  5. Genome survey sequencing and genetic background characterization of Gracilariopsis lemaneiformis (Rhodophyta) based on next-generation sequencing.

    Science.gov (United States)

    Zhou, Wei; Hu, Yiyi; Sui, Zhenghong; Fu, Feng; Wang, Jinguo; Chang, Lianpeng; Guo, Weihua; Li, Binbin

    2013-01-01

    Gracilariopsis lemaneiformis has a high economic value and is one of the most important aquaculture species in China. Despite it is economic importance, it has remained largely unstudied at the genomic level. In this study, we conducted a genome survey of Gp. lemaneiformis using next-generation sequencing (NGS) technologies. In total, 18.70 Gb of high-quality sequence data with an estimated genome size of 97 Mb were obtained by HiSeq 2000 sequencing for Gp. lemaneiformis. These reads were assembled into 160,390 contigs with a N50 length of 3.64 kb, which were further assembled into 125,685 scaffolds with a total length of 81.17 Mb. Genome analysis predicted 3490 genes and a GC% content of 48%. The identified genes have an average transcript length of 1,429 bp, an average coding sequence size of 1,369 bp, 1.36 exons per gene, exon length of 1,008 bp, and intron length of 191 bp. From the initial assembled scaffold, transposable elements constituted 54.64% (44.35 Mb) of the genome, and 7737 simple sequence repeats (SSRs) were identified. Among these SSRs, the trinucleotide repeat type was the most abundant (up to 73.20% of total SSRs), followed by the di- (17.41%), tetra- (5.49%), hexa- (2.90%), and penta- (1.00%) nucleotide repeat type. These characteristics suggest that Gp. lemaneiformis is a model organism for genetic study. This is the first report of genome-wide characterization within this taxon.

  6. Genome Survey Sequencing and Genetic Background Characterization of Gracilariopsis lemaneiformis (Rhodophyta) Based on Next-Generation Sequencing

    Science.gov (United States)

    Sui, Zhenghong; Fu, Feng; Wang, Jinguo; Chang, Lianpeng; Guo, Weihua; Li, Binbin

    2013-01-01

    Gracilariopsis lemaneiformis has a high economic value and is one of the most important aquaculture species in China. Despite it is economic importance, it has remained largely unstudied at the genomic level. In this study, we conducted a genome survey of Gp. lemaneiformis using next-generation sequencing (NGS) technologies. In total, 18.70 Gb of high-quality sequence data with an estimated genome size of 97 Mb were obtained by HiSeq 2000 sequencing for Gp. lemaneiformis. These reads were assembled into 160,390 contigs with a N50 length of 3.64 kb, which were further assembled into 125,685 scaffolds with a total length of 81.17 Mb. Genome analysis predicted 3490 genes and a GC% content of 48%. The identified genes have an average transcript length of 1,429 bp, an average coding sequence size of 1,369 bp, 1.36 exons per gene, exon length of 1,008 bp, and intron length of 191 bp. From the initial assembled scaffold, transposable elements constituted 54.64% (44.35 Mb) of the genome, and 7737 simple sequence repeats (SSRs) were identified. Among these SSRs, the trinucleotide repeat type was the most abundant (up to 73.20% of total SSRs), followed by the di- (17.41%), tetra- (5.49%), hexa- (2.90%), and penta- (1.00%) nucleotide repeat type. These characteristics suggest that Gp. lemaneiformis is a model organism for genetic study. This is the first report of genome-wide characterization within this taxon. PMID:23875008

  7. A Long-Read Transcriptome Assembly of Cotton (Gossypium hirsutum L. and Intraspecific Single Nucleotide Polymorphism Discovery

    Directory of Open Access Journals (Sweden)

    Hamid Ashrafi

    2015-07-01

    Full Text Available Upland cotton ( L. has a narrow germplasm base, which constrains marker development and hampers intraspecific breeding. A pressing need exists for high-throughput single nucleotide polymorphism (SNP markers that can be readily applied to germplasm in breeding and breeding-related research programs. Despite progress made in developing new sequencing technologies during the past decade, the cost of sequencing remains substantial when one is dealing with numerous samples and large genomes. Several strategies have been proposed to lower the cost of sequencing for multiple genotypes of large-genome species like cotton, such as transcriptome sequencing and reduced-representation DNA sequencing. This paper reports the development of a transcriptome assembly of the inbred line Texas Marker-1 (TM-1, a genetic standard for cotton, its usefulness as a reference for RNA sequencing (RNA-seq-based SNP identification, and the availability of transcriptome sequences of four other cotton cultivars. An assembly of TM-1 was made using Roche 454 transcriptome reads combined with an assembly of all available public expressed sequence tag (EST sequences of TM-1. The TM-1 assembly consists of 72,450 contigs with a total of 70 million bp. Functional predictions of the transcripts were estimated by alignment to selected protein databases. Transcriptome sequences of the five lines, including TM-1, were obtained using an Illumina Genome Analyzer-II, and the short reads were mapped to the TM-1 assembly to discover SNPs among the five lines. We identified >14,000 unfiltered allelic SNPs, of which ∼3,700 SNPs were retained for assay development after applying several rigorous filters. This paper reports availability of the reference transcriptome assembly and shows its utility in developing intraspecific SNP markers in upland cotton.

  8. Radical probing of spliceosome assembly.

    Science.gov (United States)

    Grewal, Charnpal S; Kent, Oliver A; MacMillan, Andrew M

    2017-08-01

    Here we describe the synthesis and use of a directed hydroxyl radical probe, tethered to a pre-mRNA substrate, to map the structure of this substrate during the spliceosome assembly process. These studies indicate an early organization and proximation of conserved pre-mRNA sequences during spliceosome assembly. This methodology may be adapted to the synthesis of a wide variety of modified RNAs for use as probes of RNA structure and RNA-protein interaction. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. A computational genomics pipeline for prokaryotic sequencing projects.

    Science.gov (United States)

    Kislyuk, Andrey O; Katz, Lee S; Agrawal, Sonia; Hagen, Matthew S; Conley, Andrew B; Jayaraman, Pushkala; Nelakuditi, Viswateja; Humphrey, Jay C; Sammons, Scott A; Govil, Dhwani; Mair, Raydel D; Tatti, Kathleen M; Tondella, Maria L; Harcourt, Brian H; Mayer, Leonard W; Jordan, I King

    2010-08-01

    New sequencing technologies have accelerated research on prokaryotic genomes and have made genome sequencing operations outside major genome sequencing centers routine. However, no off-the-shelf solution exists for the combined assembly, gene prediction, genome annotation and data presentation necessary to interpret sequencing data. The resulting requirement to invest significant resources into custom informatics support for genome sequencing projects remains a major impediment to the accessibility of high-throughput sequence data. We present a self-contained, automated high-throughput open source genome sequencing and computational genomics pipeline suitable for prokaryotic sequencing projects. The pipeline has been used at the Georgia Institute of Technology and the Centers for Disease Control and Prevention for the analysis of Neisseria meningitidis and Bordetella bronchiseptica genomes. The pipeline is capable of enhanced or manually assisted reference-based assembly using multiple assemblers and modes; gene predictor combining; and functional annotation of genes and gene products. Because every component of the pipeline is executed on a local machine with no need to access resources over the Internet, the pipeline is suitable for projects of a sensitive nature. Annotation of virulence-related features makes the pipeline particularly useful for projects working with pathogenic prokaryotes. The pipeline is licensed under the open-source GNU General Public License and available at the Georgia Tech Neisseria Base (http://nbase.biology.gatech.edu/). The pipeline is implemented with a combination of Perl, Bourne Shell and MySQL and is compatible with Linux and other Unix systems.

  10. Haplotype assembly in polyploid genomes and identical by descent shared tracts.

    Science.gov (United States)

    Aguiar, Derek; Istrail, Sorin

    2013-07-01

    Genome-wide haplotype reconstruction from sequence data, or haplotype assembly, is at the center of major challenges in molecular biology and life sciences. For complex eukaryotic organisms like humans, the genome is vast and the population samples are growing so rapidly that algorithms processing high-throughput sequencing data must scale favorably in terms of both accuracy and computational efficiency. Furthermore, current models and methodologies for haplotype assembly (i) do not consider individuals sharing haplotypes jointly, which reduces the size and accuracy of assembled haplotypes, and (ii) are unable to model genomes having more than two sets of homologous chromosomes (polyploidy). Polyploid organisms are increasingly becoming the target of many research groups interested in the genomics of disease, phylogenetics, botany and evolution but there is an absence of theory and methods for polyploid haplotype reconstruction. In this work, we present a number of results, extensions and generalizations of compass graphs and our HapCompass framework. We prove the theoretical complexity of two haplotype assembly optimizations, thereby motivating the use of heuristics. Furthermore, we present graph theory-based algorithms for the problem of haplotype assembly using our previously developed HapCompass framework for (i) novel implementations of haplotype assembly optimizations (minimum error correction), (ii) assembly of a pair of individuals sharing a haplotype tract identical by descent and (iii) assembly of polyploid genomes. We evaluate our methods on 1000 Genomes Project, Pacific Biosciences and simulated sequence data. HapCompass is available for download at http://www.brown.edu/Research/Istrail_Lab/. Supplementary data are available at Bioinformatics online.

  11. Preliminary High-Throughput Metagenome Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Dusheyko, Serge; Furman, Craig; Pangilinan, Jasmyn; Shapiro, Harris; Tu, Hank

    2007-03-26

    Metagenome data sets present a qualitatively different assembly problem than traditional single-organism whole-genome shotgun (WGS) assembly. The unique aspects of such projects include the presence of a potentially large number of distinct organisms and their representation in the data set at widely different fractions. In addition, multiple closely related strains could be present, which would be difficult to assemble separately. Failure to take these issues into account can result in poor assemblies that either jumble together different strains or which fail to yield useful results. The DOE Joint Genome Institute has sequenced a number of metagenomic projects and plans to considerably increase this number in the coming year. As a result, the JGI has a need for high-throughput tools and techniques for handling metagenome projects. We present the techniques developed to handle metagenome assemblies in a high-throughput environment. This includes a streamlined assembly wrapper, based on the JGI?s in-house WGS assembler, Jazz. It also includes the selection of sensible defaults targeted for metagenome data sets, as well as quality control automation for cleaning up the raw results. While analysis is ongoing, we will discuss preliminary assessments of the quality of the assembly results (http://fames.jgi-psf.org).

  12. Genome Sequence of the Freshwater Yangtze Finless Porpoise.

    Science.gov (United States)

    Yuan, Yuan; Zhang, Peijun; Wang, Kun; Liu, Mingzhong; Li, Jing; Zheng, Jingsong; Wang, Ding; Xu, Wenjie; Lin, Mingli; Dong, Lijun; Zhu, Chenglong; Qiu, Qiang; Li, Songhai

    2018-04-16

    The Yangtze finless porpoise ( Neophocaena asiaeorientalis ssp. asiaeorientalis ) is a subspecies of the narrow-ridged finless porpoise ( N. asiaeorientalis ). In total, 714.28 gigabases (Gb) of raw reads were generated by whole-genome sequencing of the Yangtze finless porpoise, using an Illumina HiSeq 2000 platform. After filtering the low-quality and duplicated reads, we assembled a draft genome of 2.22 Gb, with contig N50 and scaffold N50 values of 46.69 kilobases (kb) and 1.71 megabases (Mb), respectively. We identified 887.63 Mb of repetitive sequences and predicted 18,479 protein-coding genes in the assembled genome. The phylogenetic tree showed a relationship between the Yangtze finless porpoise and the Yangtze River dolphin, which diverged approximately 20.84 million years ago. In comparisons with the genomes of 10 other mammals, we detected 44 species-specific gene families, 164 expanded gene families, and 313 positively selected genes in the Yangtze finless porpoise genome. The assembled genome sequence and underlying sequence data are available at the National Center for Biotechnology Information under BioProject accession number PRJNA433603.

  13. Genomes correction and assembling: present methods and tools

    Science.gov (United States)

    Wojcieszek, Michał; Pawełkowicz, Magdalena; Nowak, Robert; Przybecki, Zbigniew

    2014-11-01

    Recent rapid development of next generation sequencing (NGS) technologies provided significant impact into genomics field of study enabling implementation of many de novo sequencing projects of new species which was previously confined by technological costs. Along with advancement of NGS there was need for adjustment in assembly programs. New algorithms must cope with massive amounts of data computation in reasonable time limits and processing power and hardware is also an important factor. In this paper, we address the issue of assembly pipeline for de novo genome assembly provided by programs presently available for scientist both as commercial and as open - source software. The implementation of four different approaches - Greedy, Overlap - Layout - Consensus (OLC), De Bruijn and Integrated resulting in variation of performance is the main focus of our discussion with additional insight into issue of short and long reads correction.

  14. Interactive domains in the molecular chaperone human alphaB crystallin modulate microtubule assembly and disassembly.

    Directory of Open Access Journals (Sweden)

    Joy G Ghosh

    2007-06-01

    Full Text Available Small heat shock proteins regulate microtubule assembly during cell proliferation and in response to stress through interactions that are poorly understood.Novel functions for five interactive sequences in the small heat shock protein and molecular chaperone, human alphaB crystallin, were investigated in the assembly/disassembly of microtubules and aggregation of tubulin using synthetic peptides and mutants of human alphaB crystallin.The interactive sequence (113FISREFHR(120 exposed on the surface of alphaB crystallin decreased microtubule assembly by approximately 45%. In contrast, the interactive sequences, (131LTITSSLSSDGV(142 and (156ERTIPITRE(164, corresponding to the beta8 strand and the C-terminal extension respectively, which are involved in complex formation, increased microtubule assembly by approximately 34-45%. The alphaB crystallin peptides, (113FISREFHR(120 and (156ERTIPITRE(164, inhibited microtubule disassembly by approximately 26-36%, and the peptides (113FISREFHR(120 and (131LTITSSLSSDGV(142 decreased the thermal aggregation of tubulin by approximately 42-44%. The (131LTITSSLSSDGV(142 and (156ERTIPITRE(164 peptides were more effective than the widely used anti-cancer drug, Paclitaxel, in modulating tubulinmicrotubule dynamics. Mutagenesis of these interactive sequences in wt human alphaB crystallin confirmed the effects of the alphaB crystallin peptides on microtubule assembly/disassembly and tubulin aggregation. The regulation of microtubule assembly by alphaB crystallin varied over a narrow range of concentrations. The assembly of microtubules was maximal at alphaB crystallin to tubulin molar ratios between 1:4 and 2:1, while molar ratios >2:1 inhibited microtubule assembly.Interactive sequences on the surface of human alphaB crystallin collectively modulate microtubule assembly through a dynamic subunit exchange mechanism that depends on the concentration and ratio of alphaB crystallin to tubulin. These are the first

  15. Self-assembly of proglycinin and hybrid proglycinin synthesized in vitro from cDNA

    Science.gov (United States)

    Dickinson, Craig D.; Floener, Liliane A.; Lilley, Glenn G.; Nielsen, Niels C.

    1987-01-01

    An in vitro system was developed that results in the self-assembly of subunit precursors into complexes that resemble those found naturally in the endoplasmic reticulum. Subunits of glycinin, the predominant seed protein of soybeans, were synthesized from modified cDNAs using a combination of the SP6 transcription and the rabbit reticulocyte translation systems. Subunits produced from plasmid constructions that encoded either Gy4 or Gy5 gene products, but modified such that their signal sequences were absent, self-assembled into trimers equivalent in size to those precursors found in the endoplasmic reticulum. In contrast, proteins synthesized in vitro from Gy4 constructs failed to self-assemble when the signal sequence was left intact (e.g., preproglycinin) or when the coding sequence was modified to remove 27 amino acids from an internal hydrophobic region, which is highly conserved among the glycinin subunits. Various hybrid subunits were also produced by trading portions of Gy4 and Gy5 cDNAs and all self-assembled in our system. The in vitro assembly system provides an opportunity to study the self-assembly of precursors and to probe for regions important for assembly. It will also be helpful in attempts to engineer beneficial nutritional changes into this important food protein. Images PMID:16593868

  16. QUAST: quality assessment tool for genome assemblies.

    Science.gov (United States)

    Gurevich, Alexey; Saveliev, Vladislav; Vyahhi, Nikolay; Tesler, Glenn

    2013-04-15

    Limitations of genome sequencing techniques have led to dozens of assembly algorithms, none of which is perfect. A number of methods for comparing assemblers have been developed, but none is yet a recognized benchmark. Further, most existing methods for comparing assemblies are only applicable to new assemblies of finished genomes; the problem of evaluating assemblies of previously unsequenced species has not been adequately considered. Here, we present QUAST-a quality assessment tool for evaluating and comparing genome assemblies. This tool improves on leading assembly comparison software with new ideas and quality metrics. QUAST can evaluate assemblies both with a reference genome, as well as without a reference. QUAST produces many reports, summary tables and plots to help scientists in their research and in their publications. In this study, we used QUAST to compare several genome assemblers on three datasets. QUAST tables and plots for all of them are available in the Supplementary Material, and interactive versions of these reports are on the QUAST website. http://bioinf.spbau.ru/quast . Supplementary data are available at Bioinformatics online.

  17. Lageos assembly operation plan

    Science.gov (United States)

    Brueger, J.

    1975-01-01

    Guidelines and constraints procedures for LAGEOS assembly, operation, and design performance are given. Special attention was given to thermal, optical, and dynamic analysis and testing. The operation procedures illustrate the interrelation and sequence of tasks in a flow diagram. The diagram also includes quality assurance functions for verification of operation tasks.

  18. Draft Genome Sequence of "Terrisporobacter othiniensis" Isolated from a Blood Culture from a Human Patient

    DEFF Research Database (Denmark)

    Lund, Lars Christian; Sydenham, Thomas Vognbjerg; Høgh, Silje Vermedal

    2015-01-01

    "Terrisporobacter othiniensis" (proposed species) was isolated from a blood culture. Genomic DNA was sequenced using a MiSeq benchtop sequencer (Illumina) and assembled using the SPAdes genome assembler. This resulted in a draft genome sequence comprising 3,980,019 bp in 167 contigs containing 3...

  19. Mapsembler, targeted and micro assembly of large NGS datasets on a desktop computer

    Directory of Open Access Journals (Sweden)

    Peterlongo Pierre

    2012-03-01

    Full Text Available Abstract Background The analysis of next-generation sequencing data from large genomes is a timely research topic. Sequencers are producing billions of short sequence fragments from newly sequenced organisms. Computational methods for reconstructing whole genomes/transcriptomes (de novo assemblers are typically employed to process such data. However, these methods require large memory resources and computation time. Many basic biological questions could be answered targeting specific information in the reads, thus avoiding complete assembly. Results We present Mapsembler, an iterative micro and targeted assembler which processes large datasets of reads on commodity hardware. Mapsembler checks for the presence of given regions of interest that can be constructed from reads and builds a short assembly around it, either as a plain sequence or as a graph, showing contextual structure. We introduce new algorithms to retrieve approximate occurrences of a sequence from reads and construct an extension graph. Among other results presented in this paper, Mapsembler enabled to retrieve previously described human breast cancer candidate fusion genes, and to detect new ones not previously known. Conclusions Mapsembler is the first software that enables de novo discovery around a region of interest of repeats, SNPs, exon skipping, gene fusion, as well as other structural events, directly from raw sequencing reads. As indexing is localized, the memory footprint of Mapsembler is negligible. Mapsembler is released under the CeCILL license and can be freely downloaded from http://alcovna.genouest.org/mapsembler/.

  20. Improvement of the Threespine Stickleback Genome Using a Hi-C-Based Proximity-Guided Assembly.

    Science.gov (United States)

    Peichel, Catherine L; Sullivan, Shawn T; Liachko, Ivan; White, Michael A

    2017-09-01

    Scaffolding genomes into complete chromosome assemblies remains challenging even with the rapidly increasing sequence coverage generated by current next-generation sequence technologies. Even with scaffolding information, many genome assemblies remain incomplete. The genome of the threespine stickleback (Gasterosteus aculeatus), a fish model system in evolutionary genetics and genomics, is not completely assembled despite scaffolding with high-density linkage maps. Here, we first test the ability of a Hi-C based proximity-guided assembly (PGA) to perform a de novo genome assembly from relatively short contigs. Using Hi-C based PGA, we generated complete chromosome assemblies from a distribution of short contigs (20-100 kb). We found that 96.40% of contigs were correctly assigned to linkage groups (LGs), with ordering nearly identical to the previous genome assembly. Using available bacterial artificial chromosome (BAC) end sequences, we provide evidence that some of the few discrepancies between the Hi-C assembly and the existing assembly are due to structural variation between the populations used for the 2 assemblies or errors in the existing assembly. This Hi-C assembly also allowed us to improve the existing assembly, assigning over 60% (13.35 Mb) of the previously unassigned (~21.7 Mb) contigs to LGs. Together, our results highlight the potential of the Hi-C based PGA method to be used in combination with short read data to perform relatively inexpensive de novo genome assemblies. This approach will be particularly useful in organisms in which it is difficult to perform linkage mapping or to obtain high molecular weight DNA required for other scaffolding methods. © The American Genetic Association 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  1. Assembly of highly repetitive genomes using short reads: the genome of discrete typing unit III Trypanosoma cruzi strain 231.

    Science.gov (United States)

    Baptista, Rodrigo P; Reis-Cunha, Joao Luis; DeBarry, Jeremy D; Chiari, Egler; Kissinger, Jessica C; Bartholomeu, Daniella C; Macedo, Andrea M

    2018-02-14

    Next-generation sequencing (NGS) methods are low-cost high-throughput technologies that produce thousands to millions of sequence reads. Despite the high number of raw sequence reads, their short length, relative to Sanger, PacBio or Nanopore reads, complicates the assembly of genomic repeats. Many genome tools are available, but the assembly of highly repetitive genome sequences using only NGS short reads remains challenging. Genome assembly of organisms responsible for important neglected diseases such as Trypanosoma cruzi, the aetiological agent of Chagas disease, is known to be challenging because of their repetitive nature. Only three of six recognized discrete typing units (DTUs) of the parasite have their draft genomes published and therefore genome evolution analyses in the taxon are limited. In this study, we developed a computational workflow to assemble highly repetitive genomes via a combination of de novo and reference-based assembly strategies to better overcome the intrinsic limitations of each, based on Illumina reads. The highly repetitive genome of the human-infecting parasite T. cruzi 231 strain was used as a test subject. The combined-assembly approach shown in this study benefits from the reference-based assembly ability to resolve highly repetitive sequences and from the de novo capacity to assemble genome-specific regions, improving the quality of the assembly. The acceptable confidence obtained by analyzing our results showed that our combined approach is an attractive option to assemble highly repetitive genomes with NGS short reads. Phylogenomic analysis including the 231 strain, the first representative of DTU III whose genome was sequenced, was also performed and provides new insights into T. cruzi genome evolution.

  2. Information-optimal genome assembly via sparse read-overlap graphs.

    Science.gov (United States)

    Shomorony, Ilan; Kim, Samuel H; Courtade, Thomas A; Tse, David N C

    2016-09-01

    In the context of third-generation long-read sequencing technologies, read-overlap-based approaches are expected to play a central role in the assembly step. A fundamental challenge in assembling from a read-overlap graph is that the true sequence corresponds to a Hamiltonian path on the graph, and, under most formulations, the assembly problem becomes NP-hard, restricting practical approaches to heuristics. In this work, we avoid this seemingly fundamental barrier by first setting the computational complexity issue aside, and seeking an algorithm that targets information limits In particular, we consider a basic feasibility question: when does the set of reads contain enough information to allow unambiguous reconstruction of the true sequence? Based on insights from this information feasibility question, we present an algorithm-the Not-So-Greedy algorithm-to construct a sparse read-overlap graph. Unlike most other assembly algorithms, Not-So-Greedy comes with a performance guarantee: whenever information feasibility conditions are satisfied, the algorithm reduces the assembly problem to an Eulerian path problem on the resulting graph, and can thus be solved in linear time. In practice, this theoretical guarantee translates into assemblies of higher quality. Evaluations on both simulated reads from real genomes and a PacBio Escherichia coli K12 dataset demonstrate that Not-So-Greedy compares favorably with standard string graph approaches in terms of accuracy of the resulting read-overlap graph and contig N50. Available at github.com/samhykim/nsg courtade@eecs.berkeley.edu or dntse@stanford.edu Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  3. Getting complete genomes from complex samples using nanopore sequencing

    DEFF Research Database (Denmark)

    Kirkegaard, Rasmus Hansen; Karst, Søren Michael; Albertsen, Mads

    Background Short read DNA sequencing and metagenomic binning workflows have made it possible to extract bacterial genome bins from environmental microbial samples containing hundreds to thousands of different species. However, these genome bins often do not represent complete genomes......, as they are mostly fragmented, incomplete and often contaminated with foreign DNA. The value of these `draft genomes` have limited, lasting value to the scientific community, as gene synteny is broken and there is some uncertainty of what is missing1. The genetic material most often missed is important multi......-copy and/or conserved marker genes such as the 16S rRNA gene, as sequence micro-heterogeneity prevents assembly of these genes in the de novo assembly. However, long read sequencing technologies are emerging promising an end to fragmented genome assemblies2. Experimental design We extracted DNA from a full...

  4. GRAbB: Selective Assembly of Genomic Regions, a New Niche for Genomic Research.

    Directory of Open Access Journals (Sweden)

    Balázs Brankovics

    2016-06-01

    Full Text Available GRAbB (Genomic Region Assembly by Baiting is a new program that is dedicated to assemble specific genomic regions from NGS data. This approach is especially useful when dealing with multi copy regions, such as mitochondrial genome and the rDNA repeat region, parts of the genome that are often neglected or poorly assembled, although they contain interesting information from phylogenetic or epidemiologic perspectives, but also single copy regions can be assembled. The program is capable of targeting multiple regions within a single run. Furthermore, GRAbB can be used to extract specific loci from NGS data, based on homology, like sequences that are used for barcoding. To make the assembly specific, a known part of the region, such as the sequence of a PCR amplicon or a homologous sequence from a related species must be specified. By assembling only the region of interest, the assembly process is computationally much less demanding and may lead to assemblies of better quality. In this study the different applications and functionalities of the program are demonstrated such as: exhaustive assembly (rDNA region and mitochondrial genome, extracting homologous regions or genes (IGS, RPB1, RPB2 and TEF1a, as well as extracting multiple regions within a single run. The program is also compared with MITObim, which is meant for the exhaustive assembly of a single target based on a similar query sequence. GRAbB is shown to be more efficient than MITObim in terms of speed, memory and disk usage. The other functionalities (handling multiple targets simultaneously and extracting homologous regions of the new program are not matched by other programs. The program is available with explanatory documentation at https://github.com/b-brankovics/grabb. GRAbB has been tested on Ubuntu (12.04 and 14.04, Fedora (23, CentOS (7.1.1503 and Mac OS X (10.7. Furthermore, GRAbB is available as a docker repository: brankovics/grabb (https://hub.docker.com/r/brankovics/grabb/.

  5. Extensive error in the number of genes inferred from draft genome assemblies.

    Directory of Open Access Journals (Sweden)

    James F Denton

    2014-12-01

    Full Text Available Current sequencing methods produce large amounts of data, but genome assemblies based on these data are often woefully incomplete. These incomplete and error-filled assemblies result in many annotation errors, especially in the number of genes present in a genome. In this paper we investigate the magnitude of the problem, both in terms of total gene number and the number of copies of genes in specific families. To do this, we compare multiple draft assemblies against higher-quality versions of the same genomes, using several new assemblies of the chicken genome based on both traditional and next-generation sequencing technologies, as well as published draft assemblies of chimpanzee. We find that upwards of 40% of all gene families are inferred to have the wrong number of genes in draft assemblies, and that these incorrect assemblies both add and subtract genes. Using simulated genome assemblies of Drosophila melanogaster, we find that the major cause of increased gene numbers in draft genomes is the fragmentation of genes onto multiple individual contigs. Finally, we demonstrate the usefulness of RNA-Seq in improving the gene annotation of draft assemblies, largely by connecting genes that have been fragmented in the assembly process.

  6. Optimal production planning for PCB assembly

    CERN Document Server

    Ho, William

    2006-01-01

    Focuses on the optimization of the Printed circuit board (PCB) assembly lines' efficiency. This book integrates the component sequencing and the feeder arrangement problems together for the pick-and-place machine and the chip shooter machines.

  7. Rapid hybrid de novo assembly of a microbial genome using only short reads: Corynebacterium pseudotuberculosis I19 as a case study.

    Science.gov (United States)

    Cerdeira, Louise Teixeira; Carneiro, Adriana Ribeiro; Ramos, Rommel Thiago Jucá; de Almeida, Sintia Silva; D'Afonseca, Vivian; Schneider, Maria Paula Cruz; Baumbach, Jan; Tauch, Andreas; McCulloch, John Anthony; Azevedo, Vasco Ariston Carvalho; Silva, Artur

    2011-08-01

    Due to the advent of the so-called Next-Generation Sequencing (NGS) technologies the amount of monetary and temporal resources for whole-genome sequencing has been reduced by several orders of magnitude. Sequence reads can be assembled either by anchoring them directly onto an available reference genome (classical reference assembly), or can be concatenated by overlap (de novo assembly). The latter strategy is preferable because it tends to maintain the architecture of the genome sequence the however, depending on the NGS platform used, the shortness of read lengths cause tremendous problems the in the subsequent genome assembly phase, impeding closing of the entire genome sequence. To address the problem, we developed a multi-pronged hybrid de novo strategy combining De Bruijn graph and Overlap-Layout-Consensus methods, which was used to assemble from short reads the entire genome of Corynebacterium pseudotuberculosis strain I19, a bacterium with immense importance in veterinary medicine that causes Caseous Lymphadenitis in ruminants, principally ovines and caprines. Briefly, contigs were assembled de novo from the short reads and were only oriented using a reference genome by anchoring. Remaining gaps were closed using iterative anchoring of short reads by craning to gap flanks. Finally, we compare the genome sequence assembled using our hybrid strategy to a classical reference assembly using the same data as input and show that with the availability of a reference genome, it pays off to use the hybrid de novo strategy, rather than a classical reference assembly, because more genome sequences are preserved using the former. Copyright © 2011 Elsevier B.V. All rights reserved.

  8. Genome Sequence of a Novel Archaeal Rudivirus Recovered from a Mexican Hot Spring

    DEFF Research Database (Denmark)

    Servín-Garcidueñas, L; Peng, X; Garrett, R

    2013-01-01

    We report the consensus genome sequence of a novel GC-rich rudivirus, designated SMR1 (Sulfolobales Mexican rudivirus 1), assembled from a high-throughput sequenced environmental sample from a hot spring in Los Azufres National Park in western Mexico.......We report the consensus genome sequence of a novel GC-rich rudivirus, designated SMR1 (Sulfolobales Mexican rudivirus 1), assembled from a high-throughput sequenced environmental sample from a hot spring in Los Azufres National Park in western Mexico....

  9. Detection and correction of false segmental duplications caused by genome mis-assembly

    Science.gov (United States)

    2010-01-01

    Diploid genomes with divergent chromosomes present special problems for assembly software as two copies of especially polymorphic regions may be mistakenly constructed, creating the appearance of a recent segmental duplication. We developed a method for identifying such false duplications and applied it to four vertebrate genomes. For each genome, we corrected mis-assemblies, improved estimates of the amount of duplicated sequence, and recovered polymorphisms between the sequenced chromosomes. PMID:20219098

  10. Sequencing of individual chromosomes of plant pathogenic Fusarium oxysporum.

    Science.gov (United States)

    Kashiwa, Takeshi; Kozaki, Toshinori; Ishii, Kazuo; Turgeon, B Gillian; Teraoka, Tohru; Komatsu, Ken; Arie, Tsutomu

    2017-01-01

    A small chromosome in reference isolate 4287 of F. oxysporum f. sp. lycopersici (Fol) has been designated as a 'pathogenicity chromosome' because it carries several pathogenicity related genes such as the Secreted In Xylem (SIX) genes. Sequence assembly of small chromosomes in other isolates, based on a reference genome template, is difficult because of karyotype variation among isolates and a high number of sequences associated with transposable elements. These factors often result in misassembly of sequences, making it unclear whether other isolates possess the same pathogenicity chromosome harboring SIX genes as in the reference isolate. To overcome this difficulty, single chromosome sequencing after Contour-clamped Homogeneous Electric Field (CHEF) separation of chromosomes was performed, followed by de novo assembly of sequences. The assembled sequences of individual chromosomes were consistent with results of probing gels of CHEF separated chromosomes with SIX genes. Individual chromosome sequencing revealed that several SIX genes are located on a single small chromosome in two pathogenic forms of F. oxysporum, beyond the reference isolate 4287, and in the cabbage yellows fungus F. oxysporum f. sp. conglutinans. The particular combination of SIX genes on each small chromosome varied. Moreover, not all SIX genes were found on small chromosomes; depending on the isolate, some were on big chromosomes. This suggests that recombination of chromosomes and/or translocation of SIX genes may occur frequently. Our method improves sequence comparison of small chromosomes among isolates. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. DNA-guided nanoparticle assemblies

    Science.gov (United States)

    Gang, Oleg; Nykypanchuk, Dmytro; Maye, Mathew; van der Lelie, Daniel

    2013-07-16

    In some embodiments, DNA-capped nanoparticles are used to define a degree of crystalline order in assemblies thereof. In some embodiments, thermodynamically reversible and stable body-centered cubic (bcc) structures, with particles occupying <.about.10% of the unit cell, are formed. Designs and pathways amenable to the crystallization of particle assemblies are identified. In some embodiments, a plasmonic crystal is provided. In some aspects, a method for controlling the properties of particle assemblages is provided. In some embodiments a catalyst is formed from nanoparticles linked by nucleic acid sequences and forming an open crystal structure with catalytically active agents attached to the crystal on its surface or in interstices.

  12. Alignment of 1000 Genomes Project reads to reference assembly GRCh38.

    Science.gov (United States)

    Zheng-Bradley, Xiangqun; Streeter, Ian; Fairley, Susan; Richardson, David; Clarke, Laura; Flicek, Paul

    2017-07-01

    The 1000 Genomes Project produced more than 100 trillion basepairs of short read sequence from more than 2600 samples in 26 populations over a period of five years. In its final phase, the project released over 85 million genotyped and phased variants on human reference genome assembly GRCh37. An updated reference assembly, GRCh38, was released in late 2013, but there was insufficient time for the final phase of the project analysis to change to the new assembly. Although it is possible to lift the coordinates of the 1000 Genomes Project variants to the new assembly, this is a potentially error-prone process as coordinate remapping is most appropriate only for non-repetitive regions of the genome and those that did not see significant change between the two assemblies. It will also miss variants in any region that was newly added to GRCh38. Thus, to produce the highest quality variants and genotypes on GRCh38, the best strategy is to realign the reads and recall the variants based on the new alignment. As the first step of variant calling for the 1000 Genomes Project data, we have finished remapping all of the 1000 Genomes sequence reads to GRCh38 with alternative scaffold-aware BWA-MEM. The resulting alignments are available as CRAM, a reference-based sequence compression format. The data have been released on our FTP site and are also available from European Nucleotide Archive to facilitate researchers discovering variants on the primary sequences and alternative contigs of GRCh38. © The Authors 2017. Published by Oxford University Press.

  13. SWAP-Assembler 2: Optimization of De Novo Genome Assembler at Large Scale

    Energy Technology Data Exchange (ETDEWEB)

    Meng, Jintao; Seo, Sangmin; Balaji, Pavan; Wei, Yanjie; Wang, Bingqiang; Feng, Shengzhong

    2016-08-16

    In this paper, we analyze and optimize the most time-consuming steps of the SWAP-Assembler, a parallel genome assembler, so that it can scale to a large number of cores for huge genomes with the size of sequencing data ranging from terabyes to petabytes. According to the performance analysis results, the most time-consuming steps are input parallelization, k-mer graph construction, and graph simplification (edge merging). For the input parallelization, the input data is divided into virtual fragments with nearly equal size, and the start position and end position of each fragment are automatically separated at the beginning of the reads. In k-mer graph construction, in order to improve the communication efficiency, the message size is kept constant between any two processes by proportionally increasing the number of nucleotides to the number of processes in the input parallelization step for each round. The memory usage is also decreased because only a small part of the input data is processed in each round. With graph simplification, the communication protocol reduces the number of communication loops from four to two loops and decreases the idle communication time. The optimized assembler is denoted as SWAP-Assembler 2 (SWAP2). In our experiments using a 1000 Genomes project dataset of 4 terabytes (the largest dataset ever used for assembling) on the supercomputer Mira, the results show that SWAP2 scales to 131,072 cores with an efficiency of 40%. We also compared our work with both the HipMER assembler and the SWAP-Assembler. On the Yanhuang dataset of 300 gigabytes, SWAP2 shows a 3X speedup and 4X better scalability compared with the HipMer assembler and is 45 times faster than the SWAP-Assembler. The SWAP2 software is available at https://sourceforge.net/projects/swapassembler.

  14. GAViT: Genome Assembly Visualization Tool for Short Read Data

    Energy Technology Data Exchange (ETDEWEB)

    Syed, Aijazuddin; Shapiro, Harris; Tu, Hank; Pangilinan, Jasmyn; Trong, Stephan

    2008-03-14

    It is a challenging job for genome analysts to accurately debug, troubleshoot, and validate genome assembly results. Genome analysts rely on visualization tools to help validate and troubleshoot assembly results, including such problems as mis-assemblies, low-quality regions, and repeats. Short read data adds further complexity and makes it extremely challenging for the visualization tools to scale and to view all needed assembly information. As a result, there is a need for a visualization tool that can scale to display assembly data from the new sequencing technologies. We present Genome Assembly Visualization Tool (GAViT), a highly scalable and interactive assembly visualization tool developed at the DOE Joint Genome Institute (JGI).

  15. De novo Assembly and Characterization of Cajanus scarabaeoides (L. Thouars Transcriptome by Paired-End Sequencing

    Directory of Open Access Journals (Sweden)

    Deepti Nigam

    2017-07-01

    Full Text Available Pigeonpea [Cajanus cajan (L. Millsp.] is a heat and drought resilient legume crop grown mostly in Asia and Africa. Pigeonpea is affected by various biotic (diseases and insect pests and abiotic stresses (salinity and water logging which limit the yield potential of this crop. However, resistance to all these constraints is not readily available in the cultivated genotypes and some of the wild relatives have been found to withstand these resistances. Thus, the utilization of crop wild relatives (CWR in pigeonpea breeding has been effective in conferring resistance, quality and breeding efficiency traits to this crop. Bud and leaf tissue of Cajanus scarabaeoides, a wild relative of pigeon pea were used for transcriptome profiling. Approximately 30 million clean reads filtered from raw reads by removal of adaptors, ambiguous reads and low-quality reads (3.02 gigabase pairs were generated by Illumina paired-end RNA-seq technology. All of these clean reads were pooled and assembled de novo into 1,17,007 transcripts using the Trinity. Finally, a total of 98,664 unigenes were derived with mean length of 396 bp and N50 values of 1393. The assembly produced significant mapping results (73.68% in BLASTN searches of the Glycine max CDS sequence database (Ensembl. Further, uniprot database of Viridiplantae was used for unigene annotation; 81,799 of 98,664 (82.90% unigenes were finally annotated with gene descriptions or conserved protein domains. Further, a total of 23,475 SSRs were identified in 27,321 unigenes. This data will provide useful information for mining of functionally important genes and SSR markers for pigeonpea improvement.

  16. ReRep: Computational detection of repetitive sequences in genome survey sequences (GSS

    Directory of Open Access Journals (Sweden)

    Alves-Ferreira Marcelo

    2008-09-01

    Full Text Available Abstract Background Genome survey sequences (GSS offer a preliminary global view of a genome since, unlike ESTs, they cover coding as well as non-coding DNA and include repetitive regions of the genome. A more precise estimation of the nature, quantity and variability of repetitive sequences very early in a genome sequencing project is of considerable importance, as such data strongly influence the estimation of genome coverage, library quality and progress in scaffold construction. Also, the elimination of repetitive sequences from the initial assembly process is important to avoid errors and unnecessary complexity. Repetitive sequences are also of interest in a variety of other studies, for instance as molecular markers. Results We designed and implemented a straightforward pipeline called ReRep, which combines bioinformatics tools for identifying repetitive structures in a GSS dataset. In a case study, we first applied the pipeline to a set of 970 GSSs, sequenced in our laboratory from the human pathogen Leishmania braziliensis, the causative agent of leishmaniosis, an important public health problem in Brazil. We also verified the applicability of ReRep to new sequencing technologies using a set of 454-reads of an Escheria coli. The behaviour of several parameters in the algorithm is evaluated and suggestions are made for tuning of the analysis. Conclusion The ReRep approach for identification of repetitive elements in GSS datasets proved to be straightforward and efficient. Several potential repetitive sequences were found in a L. braziliensis GSS dataset generated in our laboratory, and further validated by the analysis of a more complete genomic dataset from the EMBL and Sanger Centre databases. ReRep also identified most of the E. coli K12 repeats prior to assembly in an example dataset obtained by automated sequencing using 454 technology. The parameters controlling the algorithm behaved consistently and may be tuned to the properties

  17. Sequencing, De Novo Assembly, and Annotation of the Transcriptome of the Endangered Freshwater Pearl Bivalve, Cristaria plicata, Provides Novel Insights into Functional Genes and Marker Discovery.

    Directory of Open Access Journals (Sweden)

    Bharat Bhusan Patnaik

    Full Text Available The freshwater mussel Cristaria plicata (Bivalvia: Eulamellibranchia: Unionidae, is an economically important species in molluscan aquaculture due to its use in pearl farming. The species have been listed as endangered in South Korea due to the loss of natural habitats caused by anthropogenic activities. The decreasing population and a lack of genomic information on the species is concerning for environmentalists and conservationists. In this study, we conducted a de novo transcriptome sequencing and annotation analysis of C. plicata using Illumina HiSeq 2500 next-generation sequencing (NGS technology, the Trinity assembler, and bioinformatics databases to prepare a sustainable resource for the identification of candidate genes involved in immunity, defense, and reproduction.The C. plicata transcriptome analysis included a total of 286,152,584 raw reads and 281,322,837 clean reads. The de novo assembly identified a total of 453,931 contigs and 374,794 non-redundant unigenes with average lengths of 731.2 and 737.1 bp, respectively. Furthermore, 100% coverage of C. plicata mitochondrial genes within two unigenes supported the quality of the assembler. In total, 84,274 unigenes showed homology to entries in at least one database, and 23,246 unigenes were allocated to one or more Gene Ontology (GO terms. The most prominent GO biological process, cellular component, and molecular function categories (level 2 were cellular process, membrane, and binding, respectively. A total of 4,776 unigenes were mapped to 123 biological pathways in the KEGG database. Based on the GO terms and KEGG annotation, the unigenes were suggested to be involved in immunity, stress responses, sex-determination, and reproduction. A total of 17,251 cDNA simple sequence repeats (cSSRs were identified from 61,141 unigenes (size of >1 kb with the most abundant being dinucleotide repeats.This dataset represents the first transcriptome analysis of the endangered mollusc, C. plicata

  18. Dramatic improvement in genome assembly achieved using doubled-haploid genomes.

    Science.gov (United States)

    Zhang, Hong; Tan, Engkong; Suzuki, Yutaka; Hirose, Yusuke; Kinoshita, Shigeharu; Okano, Hideyuki; Kudoh, Jun; Shimizu, Atsushi; Saito, Kazuyoshi; Watabe, Shugo; Asakawa, Shuichi

    2014-10-27

    Improvement in de novo assembly of large genomes is still to be desired. Here, we improved draft genome sequence quality by employing doubled-haploid individuals. We sequenced wildtype and doubled-haploid Takifugu rubripes genomes, under the same conditions, using the Illumina platform and assembled contigs with SOAPdenovo2. We observed 5.4-fold and 2.6-fold improvement in the sizes of the N50 contig and scaffold of doubled-haploid individuals, respectively, compared to the wildtype, indicating that the use of a doubled-haploid genome aids in accurate genome analysis.

  19. De novo transcriptome sequence assembly and identification of AP2/ERF transcription factor related to abiotic stress in parsley (Petroselinum crispum.

    Directory of Open Access Journals (Sweden)

    Meng-Yao Li

    Full Text Available Parsley is an important biennial Apiaceae species that is widely cultivated as herb, spice, and vegetable. Previous studies on parsley principally focused on its physiological and biochemical properties, including phenolic compound and volatile oil contents. However, little is known about the molecular and genetic properties of parsley. In this study, 23,686,707 high-quality reads were obtained and assembled into 81,852 transcripts and 50,161 unigenes for the first time. Functional annotation showed that 30,516 unigenes had sequence similarity to known genes. In addition, 3,244 putative simple sequence repeats were detected in curly parsley. Finally, 1,569 of the identified unigenes belonged to 58 transcription factor families. Various abiotic stresses have a strong detrimental effect on the yield and quality of parsley. AP2/ERF transcription factors have important functions in plant development, hormonal regulation, and abiotic response. A total of 88 putative AP2/ERF factors were identified from the transcriptome sequence of parsley. Seven AP2/ERF transcription factors were selected in this study to analyze the expression profiles of parsley under different abiotic stresses. Our data provide a potentially valuable resource that can be used for intensive parsley research.

  20. De novo transcriptome sequence assembly and identification of AP2/ERF transcription factor related to abiotic stress in parsley (Petroselinum crispum).

    Science.gov (United States)

    Li, Meng-Yao; Tan, Hua-Wei; Wang, Feng; Jiang, Qian; Xu, Zhi-Sheng; Tian, Chang; Xiong, Ai-Sheng

    2014-01-01

    Parsley is an important biennial Apiaceae species that is widely cultivated as herb, spice, and vegetable. Previous studies on parsley principally focused on its physiological and biochemical properties, including phenolic compound and volatile oil contents. However, little is known about the molecular and genetic properties of parsley. In this study, 23,686,707 high-quality reads were obtained and assembled into 81,852 transcripts and 50,161 unigenes for the first time. Functional annotation showed that 30,516 unigenes had sequence similarity to known genes. In addition, 3,244 putative simple sequence repeats were detected in curly parsley. Finally, 1,569 of the identified unigenes belonged to 58 transcription factor families. Various abiotic stresses have a strong detrimental effect on the yield and quality of parsley. AP2/ERF transcription factors have important functions in plant development, hormonal regulation, and abiotic response. A total of 88 putative AP2/ERF factors were identified from the transcriptome sequence of parsley. Seven AP2/ERF transcription factors were selected in this study to analyze the expression profiles of parsley under different abiotic stresses. Our data provide a potentially valuable resource that can be used for intensive parsley research.

  1. Next generation sequencing (NGS)technologies and applications

    Energy Technology Data Exchange (ETDEWEB)

    Vuyisich, Momchilo [Los Alamos National Laboratory

    2012-09-11

    NGS technology overview: (1) NGS library preparation - Nucleic acids extraction, Sample quality control, RNA conversion to cDNA, Addition of sequencing adapters, Quality control of library; (2) Sequencing - Clonal amplification of library fragments, (except PacBio), Sequencing by synthesis, Data output (reads and quality); and (3) Data analysis - Read mapping, Genome assembly, Gene expression, Operon structure, sRNA discovery, and Epigenetic analyses.

  2. A divide-and-conquer algorithm for large-scale de novo transcriptome assembly through combining small assemblies from existing algorithms.

    Science.gov (United States)

    Sze, Sing-Hoi; Parrott, Jonathan J; Tarone, Aaron M

    2017-12-06

    While the continued development of high-throughput sequencing has facilitated studies of entire transcriptomes in non-model organisms, the incorporation of an increasing amount of RNA-Seq libraries has made de novo transcriptome assembly difficult. Although algorithms that can assemble a large amount of RNA-Seq data are available, they are generally very memory-intensive and can only be used to construct small assemblies. We develop a divide-and-conquer strategy that allows these algorithms to be utilized, by subdividing a large RNA-Seq data set into small libraries. Each individual library is assembled independently by an existing algorithm, and a merging algorithm is developed to combine these assemblies by picking a subset of high quality transcripts to form a large transcriptome. When compared to existing algorithms that return a single assembly directly, this strategy achieves comparable or increased accuracy as memory-efficient algorithms that can be used to process a large amount of RNA-Seq data, and comparable or decreased accuracy as memory-intensive algorithms that can only be used to construct small assemblies. Our divide-and-conquer strategy allows memory-intensive de novo transcriptome assembly algorithms to be utilized to construct large assemblies.

  3. Analysis of dismantling possibility and unloading efforts of fuel assemblies from core of WWER

    International Nuclear Information System (INIS)

    Danilov, V.; Dobrov, V.; Semishkin, V.; Vasilchenko, I.

    2006-01-01

    The computation methods of optimal dismantling sequence of fuel assemblies (FA) from core of WWER after different operating periods and accident conditions are considered. The algorithms of fuel dismantling sequence are constructed both on the basis of analysis of mutual spacer grid overlaps of adjacent fuel assemblies and numerical structure analysis of efforts required for FA removal as FA heaving from the core. Computation results for core dismantling sequence after 3-year operating period and LB LOCA are presented in the paper

  4. Evaluation and Validation of Assembling Corrected PacBio Long Reads for Microbial Genome Completion via Hybrid Approaches.

    Science.gov (United States)

    Lin, Hsin-Hung; Liao, Yu-Chieh

    2015-01-01

    Despite the ever-increasing output of next-generation sequencing data along with developing assemblers, dozens to hundreds of gaps still exist in de novo microbial assemblies due to uneven coverage and large genomic repeats. Third-generation single-molecule, real-time (SMRT) sequencing technology avoids amplification artifacts and generates kilobase-long reads with the potential to complete microbial genome assembly. However, due to the low accuracy (~85%) of third-generation sequences, a considerable amount of long reads (>50X) are required for self-correction and for subsequent de novo assembly. Recently-developed hybrid approaches, using next-generation sequencing data and as few as 5X long reads, have been proposed to improve the completeness of microbial assembly. In this study we have evaluated the contemporary hybrid approaches and demonstrated that assembling corrected long reads (by runCA) produced the best assembly compared to long-read scaffolding (e.g., AHA, Cerulean and SSPACE-LongRead) and gap-filling (SPAdes). For generating corrected long reads, we further examined long-read correction tools, such as ECTools, LSC, LoRDEC, PBcR pipeline and proovread. We have demonstrated that three microbial genomes including Escherichia coli K12 MG1655, Meiothermus ruber DSM1279 and Pdeobacter heparinus DSM2366 were successfully hybrid assembled by runCA into near-perfect assemblies using ECTools-corrected long reads. In addition, we developed a tool, Patch, which implements corrected long reads and pre-assembled contigs as inputs, to enhance microbial genome assemblies. With the additional 20X long reads, short reads of S. cerevisiae W303 were hybrid assembled into 115 contigs using the verified strategy, ECTools + runCA. Patch was subsequently applied to upgrade the assembly to a 35-contig draft genome. Our evaluation of the hybrid approaches shows that assembling the ECTools-corrected long reads via runCA generates near complete microbial genomes, suggesting

  5. The Second Garnet Compendium: Collected Papers 1990-1992.

    Science.gov (United States)

    1993-02-01

    BELIEF Robert Goldman Russell Almond 33. Tulane University, CS 12. The MITRE Corporation THESEUS AIMI (An Intelligent Multimedia Interface) Raymond Lang...These are images of windows from the THESEUS application used by the Tulanc University Computer Science Department on guided tours of the department given...to visiting high school seniors and other interested paries. THESEUS is intended to be used as part of a presentation on what the study of computcr

  6. Collaboration for Land, Air, Sea, and Space Vehicles: Developing the Common Ground in Vehicle Dynamics, System Identification, Control, and Handling Qualities (La collaboration dans le domaine des vehicules terrestres, aeriens, maritimes et spatiaux: L’etablissement d’une approche commune de la dynamique des vehicules, l’identification des systemes, et les qualite’s de controle et de pilotage)

    Science.gov (United States)

    2002-11-01

    Sailplanes and X-Rudders, and Type 214 (right) with 79 Bowplanes and a Cruciform Tail Arrangement Figure 4.15 Theseus UUV 80 Figure 4.16 Some...for mine countermeasures and other purposes. A notable example was the 180 km out and back mission performed in 1996 by the cable-laying UUV Theseus ...predetermined response table. 80 Figure 4.15 - Theseus UUV Although autonomous UUVs are finding increasing application, Remotely Operated

  7. FOLDNA, a Web Server for Self-Assembled DNA Nanostructure Autoscaffolds and Autostaples

    Directory of Open Access Journals (Sweden)

    Chensheng Zhou

    2012-01-01

    Full Text Available DNA self-assembly is a nanotechnology that folds DNA into desired shapes. Self-assembled DNA nanostructures, also known as origami, are increasingly valuable in nanomaterial and biosensing applications. Two ways to use DNA nanostructures in medicine are to form nanoarrays, and to work as vehicles in drug delivery. The DNA nanostructures perform well as a biomaterial in these areas because they have spatially addressable and size controllable properties. However, manually designing complementary DNA sequences for self-assembly is a technically demanding and time consuming task, which makes it advantageous for computers to do this job instead. We have developed a web server, FOLDNA, which can automatically design 2D self-assembled DNA nanostructures according to custom pictures and scaffold sequences provided by the users. It is the first web server to provide an entirely automatic design of self-assembled DNA nanostructure, and it takes merely a second to generate comprehensive information for molecular experiments including: scaffold DNA pathways, staple DNA directions, and staple DNA sequences. This program could save as much as several hours in the designing step for each DNA nanostructure. We randomly selected some shapes and corresponding outputs from our server and validated its performance in molecular experiments.

  8. Analysis Of Transcriptomes In A Porcine Tissue Collection Using RNA-Seq And Genome Assembly 10

    DEFF Research Database (Denmark)

    Hornshøj, Henrik; Thomsen, Bo; Hedegaard, Jakob

    2011-01-01

    The release of Sus scrofa genome assembly 10 supports improvement of the pig genome annotation and in depth transcriptome analyses using next-generation sequencing technologies. In this study we analyze RNA-seq reads from a tissue collection, including 10 separate tissues from Duroc boars and 10...... short read alignment software we mapped the reads to the genome assembly 10. We extracted contig sequences of gene transcripts using the Cufflinks software. Based on this information we identified expressed genes that are present in the genome assembly. The portion of these genes being previously known...... was roughly estimated by sequence comparison to known genes. Similarly, we searched for genes that are expressed in the tissues but not present in the genome assembly by aligning the non-genome-mapped reads to known gene transcripts. For the genes predicted to have alternative transcript variants by Cufflinks...

  9. Navigating the tip of the genomic iceberg: Next-generation sequencing for plant systematics.

    Science.gov (United States)

    Straub, Shannon C K; Parks, Matthew; Weitemier, Kevin; Fishbein, Mark; Cronn, Richard C; Liston, Aaron

    2012-02-01

    Just as Sanger sequencing did more than 20 years ago, next-generation sequencing (NGS) is poised to revolutionize plant systematics. By combining multiplexing approaches with NGS throughput, systematists may no longer need to choose between more taxa or more characters. Here we describe a genome skimming (shallow sequencing) approach for plant systematics. Through simulations, we evaluated optimal sequencing depth and performance of single-end and paired-end short read sequences for assembly of nuclear ribosomal DNA (rDNA) and plastomes and addressed the effect of divergence on reference-guided plastome assembly. We also used simulations to identify potential phylogenetic markers from low-copy nuclear loci at different sequencing depths. We demonstrated the utility of genome skimming through phylogenetic analysis of the Sonoran Desert clade (SDC) of Asclepias (Apocynaceae). Paired-end reads performed better than single-end reads. Minimum sequencing depths for high quality rDNA and plastome assemblies were 40× and 30×, respectively. Divergence from the reference significantly affected plastome assembly, but relatively similar references are available for most seed plants. Deeper rDNA sequencing is necessary to characterize intragenomic polymorphism. The low-copy fraction of the nuclear genome was readily surveyed, even at low sequencing depths. Nearly 160000 bp of sequence from three organelles provided evidence of phylogenetic incongruence in the SDC. Adoption of NGS will facilitate progress in plant systematics, as whole plastome and rDNA cistrons, partial mitochondrial genomes, and low-copy nuclear markers can now be efficiently obtained for molecular phylogenetics studies.

  10. Radiation hybrid maps of the D-genome of Aegilops tauschii and their application in sequence assembly of large and complex plant genomes.

    Science.gov (United States)

    Kumar, Ajay; Seetan, Raed; Mergoum, Mohamed; Tiwari, Vijay K; Iqbal, Muhammad J; Wang, Yi; Al-Azzam, Omar; Šimková, Hana; Luo, Ming-Cheng; Dvorak, Jan; Gu, Yong Q; Denton, Anne; Kilian, Andrzej; Lazo, Gerard R; Kianian, Shahryar F

    2015-10-16

    The large and complex genome of bread wheat (Triticum aestivum L., ~17 Gb) requires high resolution genome maps with saturated marker scaffolds to anchor and orient BAC contigs/ sequence scaffolds for whole genome assembly. Radiation hybrid (RH) mapping has proven to be an excellent tool for the development of such maps for it offers much higher and more uniform marker resolution across the length of the chromosome compared to genetic mapping and does not require marker polymorphism per se, as it is based on presence (retention) vs. absence (deletion) marker assay. In this study, a 178 line RH panel was genotyped with SSRs and DArT markers to develop the first high resolution RH maps of the entire D-genome of Ae. tauschii accession AL8/78. To confirm map order accuracy, the AL8/78-RH maps were compared with:1) a DArT consensus genetic map constructed using more than 100 bi-parental populations, 2) a RH map of the D-genome of reference hexaploid wheat 'Chinese Spring', and 3) two SNP-based genetic maps, one with anchored D-genome BAC contigs and another with anchored D-genome sequence scaffolds. Using marker sequences, the RH maps were also anchored with a BAC contig based physical map and draft sequence of the D-genome of Ae. tauschii. A total of 609 markers were mapped to 503 unique positions on the seven D-genome chromosomes, with a total map length of 14,706.7 cR. The average distance between any two marker loci was 29.2 cR which corresponds to 2.1 cM or 9.8 Mb. The average mapping resolution across the D-genome was estimated to be 0.34 Mb (Mb/cR) or 0.07 cM (cM/cR). The RH maps showed almost perfect agreement with several published maps with regard to chromosome assignments of markers. The mean rank correlations between the position of markers on AL8/78 maps and the four published maps, ranged from 0.75 to 0.92, suggesting a good agreement in marker order. With 609 mapped markers, a total of 2481 deletions for the whole D-genome were detected with an average

  11. The Release 6 reference sequence of the Drosophila melanogaster genome.

    Science.gov (United States)

    Hoskins, Roger A; Carlson, Joseph W; Wan, Kenneth H; Park, Soo; Mendez, Ivonne; Galle, Samuel E; Booth, Benjamin W; Pfeiffer, Barret D; George, Reed A; Svirskas, Robert; Krzywinski, Martin; Schein, Jacqueline; Accardo, Maria Carmela; Damia, Elisabetta; Messina, Giovanni; Méndez-Lago, María; de Pablos, Beatriz; Demakova, Olga V; Andreyeva, Evgeniya N; Boldyreva, Lidiya V; Marra, Marco; Carvalho, A Bernardo; Dimitri, Patrizio; Villasante, Alfredo; Zhimulev, Igor F; Rubin, Gerald M; Karpen, Gary H; Celniker, Susan E

    2015-03-01

    Drosophila melanogaster plays an important role in molecular, genetic, and genomic studies of heredity, development, metabolism, behavior, and human disease. The initial reference genome sequence reported more than a decade ago had a profound impact on progress in Drosophila research, and improving the accuracy and completeness of this sequence continues to be important to further progress. We previously described improvement of the 117-Mb sequence in the euchromatic portion of the genome and 21 Mb in the heterochromatic portion, using a whole-genome shotgun assembly, BAC physical mapping, and clone-based finishing. Here, we report an improved reference sequence of the single-copy and middle-repetitive regions of the genome, produced using cytogenetic mapping to mitotic and polytene chromosomes, clone-based finishing and BAC fingerprint verification, ordering of scaffolds by alignment to cDNA sequences, incorporation of other map and sequence data, and validation by whole-genome optical restriction mapping. These data substantially improve the accuracy and completeness of the reference sequence and the order and orientation of sequence scaffolds into chromosome arm assemblies. Representation of the Y chromosome and other heterochromatic regions is particularly improved. The new 143.9-Mb reference sequence, designated Release 6, effectively exhausts clone-based technologies for mapping and sequencing. Highly repeat-rich regions, including large satellite blocks and functional elements such as the ribosomal RNA genes and the centromeres, are largely inaccessible to current sequencing and assembly methods and remain poorly represented. Further significant improvements will require sequencing technologies that do not depend on molecular cloning and that produce very long reads. © 2015 Hoskins et al.; Published by Cold Spring Harbor Laboratory Press.

  12. Whole genome assembly of a natto production strain Bacillus subtilis natto from very short read data.

    Science.gov (United States)

    Nishito, Yukari; Osana, Yasunori; Hachiya, Tsuyoshi; Popendorf, Kris; Toyoda, Atsushi; Fujiyama, Asao; Itaya, Mitsuhiro; Sakakibara, Yasubumi

    2010-04-16

    Bacillus subtilis natto is closely related to the laboratory standard strain B. subtilis Marburg 168, and functions as a starter for the production of the traditional Japanese food "natto" made from soybeans. Although re-sequencing whole genomes of several laboratory domesticated B. subtilis 168 derivatives has already been attempted using short read sequencing data, the assembly of the whole genome sequence of a closely related strain, B. subtilis natto, from very short read data is more challenging, particularly with our aim to assemble one fully connected scaffold from short reads around 35 bp in length. We applied a comparative genome assembly method, which combines de novo assembly and reference guided assembly, to one of the B. subtilis natto strains. We successfully assembled 28 scaffolds and managed to avoid substantial fragmentation. Completion of the assembly through long PCR experiments resulted in one connected scaffold for B. subtilis natto. Based on the assembled genome sequence, our orthologous gene analysis between natto BEST195 and Marburg 168 revealed that 82.4% of 4375 predicted genes in BEST195 are one-to-one orthologous to genes in 168, with two genes in-paralog, 3.2% are deleted in 168, 14.3% are inserted in BEST195, and 5.9% of genes present in 168 are deleted in BEST195. The natto genome contains the same alleles in the promoter region of degQ and the coding region of swrAA as the wild strain, RO-FF-1. These are specific for gamma-PGA production ability, which is related to natto production. Further, the B. subtilis natto strain completely lacked a polyketide synthesis operon, disrupted the plipastatin production operon, and possesses previously unidentified transposases. The determination of the whole genome sequence of Bacillus subtilis natto provided detailed analyses of a set of genes related to natto production, demonstrating the number and locations of insertion sequences that B. subtilis natto harbors but B. subtilis 168 lacks

  13. Whole genome assembly of a natto production strain Bacillus subtilis natto from very short read data

    Directory of Open Access Journals (Sweden)

    Fujiyama Asao

    2010-04-01

    Full Text Available Abstract Background Bacillus subtilis natto is closely related to the laboratory standard strain B. subtilis Marburg 168, and functions as a starter for the production of the traditional Japanese food "natto" made from soybeans. Although re-sequencing whole genomes of several laboratory domesticated B. subtilis 168 derivatives has already been attempted using short read sequencing data, the assembly of the whole genome sequence of a closely related strain, B. subtilis natto, from very short read data is more challenging, particularly with our aim to assemble one fully connected scaffold from short reads around 35 bp in length. Results We applied a comparative genome assembly method, which combines de novo assembly and reference guided assembly, to one of the B. subtilis natto strains. We successfully assembled 28 scaffolds and managed to avoid substantial fragmentation. Completion of the assembly through long PCR experiments resulted in one connected scaffold for B. subtilis natto. Based on the assembled genome sequence, our orthologous gene analysis between natto BEST195 and Marburg 168 revealed that 82.4% of 4375 predicted genes in BEST195 are one-to-one orthologous to genes in 168, with two genes in-paralog, 3.2% are deleted in 168, 14.3% are inserted in BEST195, and 5.9% of genes present in 168 are deleted in BEST195. The natto genome contains the same alleles in the promoter region of degQ and the coding region of swrAA as the wild strain, RO-FF-1. These are specific for γ-PGA production ability, which is related to natto production. Further, the B. subtilis natto strain completely lacked a polyketide synthesis operon, disrupted the plipastatin production operon, and possesses previously unidentified transposases. Conclusions The determination of the whole genome sequence of Bacillus subtilis natto provided detailed analyses of a set of genes related to natto production, demonstrating the number and locations of insertion sequences that B

  14. Enabling Graph Appliance for Genome Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Rina [ORNL; Graves, Jeffrey A [ORNL; Lee, Sangkeun (Matt) [ORNL; Sukumar, Sreenivas R [ORNL; Shankar, Mallikarjun [ORNL

    2015-01-01

    In recent years, there has been a huge growth in the amount of genomic data available as reads generated from various genome sequencers. The number of reads generated can be huge, ranging from hundreds to billions of nucleotide, each varying in size. Assembling such large amounts of data is one of the challenging computational problems for both biomedical and data scientists. Most of the genome assemblers developed have used de Bruijn graph techniques. A de Bruijn graph represents a collection of read sequences by billions of vertices and edges, which require large amounts of memory and computational power to store and process. This is the major drawback to de Bruijn graph assembly. Massively parallel, multi-threaded, shared memory systems can be leveraged to overcome some of these issues. The objective of our research is to investigate the feasibility and scalability issues of de Bruijn graph assembly on Cray s Urika-GD system; Urika-GD is a high performance graph appliance with a large shared memory and massively multithreaded custom processor designed for executing SPARQL queries over large-scale RDF data sets. However, to the best of our knowledge, there is no research on representing a de Bruijn graph as an RDF graph or finding Eulerian paths in RDF graphs using SPARQL for potential genome discovery. In this paper, we address the issues involved in representing a de Bruin graphs as RDF graphs and propose an iterative querying approach for finding Eulerian paths in large RDF graphs. We evaluate the performance of our implementation on real world ebola genome datasets and illustrate how genome assembly can be accomplished with Urika-GD using iterative SPARQL queries.

  15. Hybrid De Novo Genome Assembly Using MiSeq and SOLiD Short Read Data.

    Directory of Open Access Journals (Sweden)

    Tsutomu Ikegami

    Full Text Available A hybrid de novo assembly pipeline was constructed to utilize both MiSeq and SOLiD short read data in combination in the assembly. The short read data were converted to a standard format of the pipeline, and were supplied to the pipeline components such as ABySS and SOAPdenovo. The assembly pipeline proceeded through several stages, and either MiSeq paired-end data, SOLiD mate-paired data, or both of them could be specified as input data at each stage separately. The pipeline was examined on the filamentous fungus Aspergillus oryzae RIB40, by aligning the assembly results against the reference sequences. Using both the MiSeq and the SOLiD data in the hybrid assembly, the alignment length was improved by a factor of 3 to 8, compared with the assemblies using either one of the data types. The number of the reproduced gene cluster regions encoding secondary metabolite biosyntheses (SMB was also improved by the hybrid assemblies. These results imply that the MiSeq data with long read length are essential to construct accurate nucleotide sequences, while the SOLiD mate-paired reads with long insertion length enhance long-range arrangements of the sequences. The pipeline was also tested on the actinomycete Streptomyces avermitilis MA-4680, whose gene is known to have high-GC content. Although the quality of the SOLiD reads was too low to perform any meaningful assemblies by themselves, the alignment length to the reference was improved by a factor of 2, compared with the assembly using only the MiSeq data.

  16. Evaluation of the impact of RNA preservation methods of spiders for de novo transcriptome assembly.

    Science.gov (United States)

    Kono, Nobuaki; Nakamura, Hiroyuki; Ito, Yusuke; Tomita, Masaru; Arakawa, Kazuharu

    2016-05-01

    With advances in high-throughput sequencing technologies, de novo transcriptome sequencing and assembly has become a cost-effective method to obtain comprehensive genetic information of a species of interest, especially in nonmodel species with large genomes such as spiders. However, high-quality RNA is essential for successful sequencing, and sample preservation conditions require careful consideration for the effective storage of field-collected samples. To this end, we report a streamlined feasibility study of various storage conditions and their effects on de novo transcriptome assembly results. The storage parameters considered include temperatures ranging from room temperature to -80°C; preservatives, including ethanol, RNAlater, TRIzol and RNAlater-ICE; and sample submersion states. As a result, intact RNA was extracted and assembly was successful when samples were preserved at low temperatures regardless of the type of preservative used. The assemblies as well as the gene expression profiles were shown to be robust to RNA degradation, when 30 million 150-bp paired-end reads are obtained. The parameters for sample storage, RNA extraction, library preparation, sequencing and in silico assembly considered in this work provide a guideline for the study of field-collected samples of spiders. © 2015 John Wiley & Sons Ltd.

  17. AUV Commercialization - Who’s Leading the Pack?

    Science.gov (United States)

    2000-09-01

    the Theseus and ARCS, is designing a deep water commercial site survey AUV for Fugro GeoServices Inc. Called the Explorer, the vehicle will conduct...ISE has the ARCS and the Theseus vehicles and Perry Technologies has the MUST. These vehicles have each performed some dramatic operations including the...deployment of fiber optic cables. In the case of Theseus , the fiber optic cable was deployed under the ice pack. Mid-size vehicles include those from

  18. Multiple hybrid de novo genome assembly of finger millet, an orphan allotetraploid crop.

    Science.gov (United States)

    Hatakeyama, Masaomi; Aluri, Sirisha; Balachadran, Mathi Thumilan; Sivarajan, Sajeevan Radha; Patrignani, Andrea; Grüter, Simon; Poveda, Lucy; Shimizu-Inatsugi, Rie; Baeten, John; Francoijs, Kees-Jan; Nataraja, Karaba N; Reddy, Yellodu A Nanja; Phadnis, Shamprasad; Ravikumar, Ramapura L; Schlapbach, Ralph; Sreeman, Sheshshayee M; Shimizu, Kentaro K

    2017-09-05

    Finger millet (Eleusine coracana (L.) Gaertn) is an important crop for food security because of its tolerance to drought, which is expected to be exacerbated by global climate changes. Nevertheless, it is often classified as an orphan/underutilized crop because of the paucity of scientific attention. Among several small millets, finger millet is considered as an excellent source of essential nutrient elements, such as iron and zinc; hence, it has potential as an alternate coarse cereal. However, high-quality genome sequence data of finger millet are currently not available. One of the major problems encountered in the genome assembly of this species was its polyploidy, which hampers genome assembly compared with a diploid genome. To overcome this problem, we sequenced its genome using diverse technologies with sufficient coverage and assembled it via a novel multiple hybrid assembly workflow that combines next-generation with single-molecule sequencing, followed by whole-genome optical mapping using the Bionano Irys® system. The total number of scaffolds was 1,897 with an N50 length >2.6 Mb and detection of 96% of the universal single-copy orthologs. The majority of the homeologs were assembled separately. This indicates that the proposed workflow is applicable to the assembly of other allotetraploid genomes. © The Author 2017. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

  19. Characterization of transcriptome dynamics during watermelon fruit development: sequencing, assembly, annotation and gene expression profiles.

    Science.gov (United States)

    Guo, Shaogui; Liu, Jingan; Zheng, Yi; Huang, Mingyun; Zhang, Haiying; Gong, Guoyi; He, Hongju; Ren, Yi; Zhong, Silin; Fei, Zhangjun; Xu, Yong

    2011-09-21

    Cultivated watermelon [Citrullus lanatus (Thunb.) Matsum. & Nakai var. lanatus] is an important agriculture crop world-wide. The fruit of watermelon undergoes distinct stages of development with dramatic changes in its size, color, sweetness, texture and aroma. In order to better understand the genetic and molecular basis of these changes and significantly expand the watermelon transcript catalog, we have selected four critical stages of watermelon fruit development and used Roche/454 next-generation sequencing technology to generate a large expressed sequence tag (EST) dataset and a comprehensive transcriptome profile for watermelon fruit flesh tissues. We performed half Roche/454 GS-FLX run for each of the four watermelon fruit developmental stages (immature white, white-pink flesh, red flesh and over-ripe) and obtained 577,023 high quality ESTs with an average length of 302.8 bp. De novo assembly of these ESTs together with 11,786 watermelon ESTs collected from GenBank produced 75,068 unigenes with a total length of approximately 31.8 Mb. Overall 54.9% of the unigenes showed significant similarities to known sequences in GenBank non-redundant (nr) protein database and around two-thirds of them matched proteins of cucumber, the most closely-related species with a sequenced genome. The unigenes were further assigned with gene ontology (GO) terms and mapped to biochemical pathways. More than 5,000 SSRs were identified from the EST collection. Furthermore we carried out digital gene expression analysis of these ESTs and identified 3,023 genes that were differentially expressed during watermelon fruit development and ripening, which provided novel insights into watermelon fruit biology and a comprehensive resource of candidate genes for future functional analysis. We then generated profiles of several interesting metabolites that are important to fruit quality including pigmentation and sweetness. Integrative analysis of metabolite and digital gene expression

  20. Discovery of human inversion polymorphisms by comparative analysis of human and chimpanzee DNA sequence assemblies.

    Directory of Open Access Journals (Sweden)

    2005-10-01

    Full Text Available With a draft genome-sequence assembly for the chimpanzee available, it is now possible to perform genome-wide analyses to identify, at a submicroscopic level, structural rearrangements that have occurred between chimpanzees and humans. The goal of this study was to investigate chromosomal regions that are inverted between the chimpanzee and human genomes. Using the net alignments for the builds of the human and chimpanzee genome assemblies, we identified a total of 1,576 putative regions of inverted orientation, covering more than 154 mega-bases of DNA. The DNA segments are distributed throughout the genome and range from 23 base pairs to 62 mega-bases in length. For the 66 inversions more than 25 kilobases (kb in length, 75% were flanked on one or both sides by (often unrelated segmental duplications. Using PCR and fluorescence in situ hybridization we experimentally validated 23 of 27 (85% semi-randomly chosen regions; the largest novel inversion confirmed was 4.3 mega-bases at human Chromosome 7p14. Gorilla was used as an out-group to assign ancestral status to the variants. All experimentally validated inversion regions were then assayed against a panel of human samples and three of the 23 (13% regions were found to be polymorphic in the human genome. These polymorphic inversions include 730 kb (at 7p22, 13 kb (at 7q11, and 1 kb (at 16q24 fragments with a 5%, 30%, and 48% minor allele frequency, respectively. Our results suggest that inversions are an important source of variation in primate genome evolution. The finding of at least three novel inversion polymorphisms in humans indicates this type of structural variation may be a more common feature of our genome than previously realized.

  1. SAGE: String-overlap Assembly of GEnomes.

    Science.gov (United States)

    Ilie, Lucian; Haider, Bahlul; Molnar, Michael; Solis-Oba, Roberto

    2014-09-15

    De novo genome assembly of next-generation sequencing data is one of the most important current problems in bioinformatics, essential in many biological applications. In spite of significant amount of work in this area, better solutions are still very much needed. We present a new program, SAGE, for de novo genome assembly. As opposed to most assemblers, which are de Bruijn graph based, SAGE uses the string-overlap graph. SAGE builds upon great existing work on string-overlap graph and maximum likelihood assembly, bringing an important number of new ideas, such as the efficient computation of the transitive reduction of the string overlap graph, the use of (generalized) edge multiplicity statistics for more accurate estimation of read copy counts, and the improved use of mate pairs and min-cost flow for supporting edge merging. The assemblies produced by SAGE for several short and medium-size genomes compared favourably with those of existing leading assemblers. SAGE benefits from innovations in almost every aspect of the assembly process: error correction of input reads, string-overlap graph construction, read copy counts estimation, overlap graph analysis and reduction, contig extraction, and scaffolding. We hope that these new ideas will help advance the current state-of-the-art in an essential area of research in genomics.

  2. Self-assembled software and method of overriding software execution

    Science.gov (United States)

    Bouchard, Ann M.; Osbourn, Gordon C.

    2013-01-08

    A computer-implemented software self-assembled system and method for providing an external override and monitoring capability to dynamically self-assembling software containing machines that self-assemble execution sequences and data structures. The method provides an external override machine that can be introduced into a system of self-assembling machines while the machines are executing such that the functionality of the executing software can be changed or paused without stopping the code execution and modifying the existing code. Additionally, a monitoring machine can be introduced without stopping code execution that can monitor specified code execution functions by designated machines and communicate the status to an output device.

  3. De novo 454 sequencing of barcoded BAC pools for comprehensive gene survey and genome analysis in the complex genome of barley

    Directory of Open Access Journals (Sweden)

    Scholz Uwe

    2009-11-01

    Full Text Available Abstract Background De novo sequencing the entire genome of a large complex plant genome like the one of barley (Hordeum vulgare L. is a major challenge both in terms of experimental feasibility and costs. The emergence and breathtaking progress of next generation sequencing technologies has put this goal into focus and a clone based strategy combined with the 454/Roche technology is conceivable. Results To test the feasibility, we sequenced 91 barcoded, pooled, gene containing barley BACs using the GS FLX platform and assembled the sequences under iterative change of parameters. The BAC assemblies were characterized by N50 of ~50 kb (N80 ~31 kb, N90 ~21 kb and a Q40 of 94%. For ~80% of the clones, the best assemblies consisted of less than 10 contigs at 24-fold mean sequence coverage. Moreover we show that gene containing regions seem to assemble completely and uninterrupted thus making the approach suitable for detecting complete and positionally anchored genes. By comparing the assemblies of four clones to their complete reference sequences generated by the Sanger method, we evaluated the distribution, quality and representativeness of the 454 sequences as well as the consistency and reliability of the assemblies. Conclusion The described multiplex 454 sequencing of barcoded BACs leads to sequence consensi highly representative for the clones. Assemblies are correct for the majority of contigs. Though the resolution of complex repetitive structures requires additional experimental efforts, our approach paves the way for a clone based strategy of sequencing the barley genome.

  4. A conceptual design and structural stabilities of in-pit assembly tools for the completion of final sector assembly at tokamak hall

    International Nuclear Information System (INIS)

    Nam, K.O.; Park, H.K.; Kim, D.J.; Ahn, H.J.; Kim, K.K.; Im, K.; Shaw, R.

    2010-01-01

    The final assembly of main components of the International Thermonuclear Experimental Reactor (ITER) tokamak, Vacuum Vessel (VV) and Toroidal Field Coils (TFCs), is achieved by the sequential assembly of the nine sub-assembled 40 o sectors in tokamak pit. Each sub-assembled 40 o sector is composed of one VV 40 o sector, two TFCs, and in-between Vacuum Vessel Thermal Shield (VVTS) segments. Sub-assembly is carried out in the assembly building and then the sub-assembled sectors are transferred into tokamak pit, in sequence, to complete sector assembly. The role of in-pit assembly tool is to support and align the sub-assembled sectors in tokamak pit. It also plays the role of reference datum during assembly until the completion of main components assembly. Korea Domestic Agency (KO DA) has developed the conceptual design of most ITER purpose-built assembly tools under the collaboration with the ITER Organization. Among the conceptual designs carried out, this paper describes the function, the structure, the selected material and the design results of the in-pit assembly tools comprising central column, radial beams and their supports, TF inner supports and in-pit working floor. The results of structural analysis using ANSYS for the various loading cases are given as well. The resultant stresses and deflections turned out to fall within the allowable ranges.

  5. [Complete genome sequencing and sequence analysis of BCG Tice].

    Science.gov (United States)

    Wang, Zhiming; Pan, Yuanlong; Wu, Jun; Zhu, Baoli

    2012-10-04

    The objective of this study is to obtain the complete genome sequence of Bacillus Calmette-Guerin Tice (BCG Tice), in order to provide more information about the molecular biology of BCG Tice and design more reasonable vaccines to prevent tuberculosis. We assembled the data from high-throughput sequencing with SOAPdenovo software, with many contigs and scaffolds obtained. There are many sequence gaps and physical gaps remained as a result of regional low coverage and low quality. We designed primers at the end of contigs and performed PCR amplification in order to link these contigs and scaffolds. With various enzymes to perform PCR amplification, adjustment of PCR reaction conditions, and combined with clone construction to sequence, all the gaps were finished. We obtained the complete genome sequence of BCG Tice and submitted it to GenBank of National Center for Biotechnology Information (NCBI). The genome of BCG Tice is 4334064 base pairs in length, with GC content 65.65%. The problems and strategies during the finishing step of BCG Tice sequencing are illuminated here, with the hope of affording some experience to those who are involved in the finishing step of genome sequencing. The microarray data were verified by our results.

  6. GAAP: Genome-organization-framework-Assisted Assembly Pipeline for prokaryotic genomes.

    Science.gov (United States)

    Yuan, Lina; Yu, Yang; Zhu, Yanmin; Li, Yulai; Li, Changqing; Li, Rujiao; Ma, Qin; Siu, Gilman Kit-Hang; Yu, Jun; Jiang, Taijiao; Xiao, Jingfa; Kang, Yu

    2017-01-25

    Next-generation sequencing (NGS) technologies have greatly promoted the genomic study of prokaryotes. However, highly fragmented assemblies due to short reads from NGS are still a limiting factor in gaining insights into the genome biology. Reference-assisted tools are promising in genome assembly, but tend to result in false assembly when the assigned reference has extensive rearrangements. Herein, we present GAAP, a genome assembly pipeline for scaffolding based on core-gene-defined Genome Organizational Framework (cGOF) described in our previous study. Instead of assigning references, we use the multiple-reference-derived cGOFs as indexes to assist in order and orientation of the scaffolds and build a skeleton structure, and then use read pairs to extend scaffolds, called local scaffolding, and distinguish between true and chimeric adjacencies in the scaffolds. In our performance tests using both empirical and simulated data of 15 genomes in six species with diverse genome size, complexity, and all three categories of cGOFs, GAAP outcompetes or achieves comparable results when compared to three other reference-assisted programs, AlignGraph, Ragout and MeDuSa. GAAP uses both cGOF and pair-end reads to create assemblies in genomic scale, and performs better than the currently available reference-assisted assembly tools as it recovers more assemblies and makes fewer false locations, especially for species with extensive rearranged genomes. Our method is a promising solution for reconstruction of genome sequence from short reads of NGS.

  7. Over the Beach. US Army Amphibious Operations in the Korean War

    Science.gov (United States)

    2008-01-01

    steaming toward the west coast with British aircraft carrier Theseus and four destroyers.59 At 0930 on 4 December, when Kelly and his transports...throughout 5 December, with Royal Navy aircraft of the Theseus arriving overhead around noon, the Foss continuing to provide power, and the ROK Navy shore...carrier H.M.S. Theseus , cruis­ ers, and destroyers. Kelly’s small amphibious force arrived off Incheon on 8 February and the next day the Missouri began

  8. Building block synthesis using the polymerase chain assembly method.

    Science.gov (United States)

    Marchand, Julie A; Peccoud, Jean

    2012-01-01

    De novo gene synthesis allows the creation of custom DNA molecules without the typical constraints of traditional cloning assembly: scars, restriction site incompatibility, and the quest to find all the desired parts to name a few. Moreover, with the help of computer-assisted design, the perfect DNA molecule can be created along with its matching sequence ready to download. The challenge is to build the physical DNA molecules that have been designed with the software. Although there are several DNA assembly methods, this section presents and describes a method using the polymerase chain assembly (PCA).

  9. Noncanonical self-assembly of multifunctional DNA nanoflowers for biomedical applications.

    Science.gov (United States)

    Zhu, Guizhi; Hu, Rong; Zhao, Zilong; Chen, Zhuo; Zhang, Xiaobing; Tan, Weihong

    2013-11-06

    DNA nanotechnology has been extensively explored to assemble various functional nanostructures for versatile applications. Mediated by Watson-Crick base-pairing, these DNA nanostructures have been conventionally assembled through hybridization of many short DNA building blocks. Here we report the noncanonical self-assembly of multifunctional DNA nanostructures, termed as nanoflowers (NFs), and the versatile biomedical applications. These NFs were assembled from long DNA building blocks generated via rolling circle replication (RCR) of a designer template. NF assembly was driven by liquid crystallization and dense packaging of building blocks, without relying on Watson-Crick base-pairing between DNA strands, thereby avoiding the otherwise conventional complicated DNA sequence design. NF sizes were readily tunable in a wide range, by simply adjusting such parameters as assembly time and template sequences. NFs were exceptionally resistant to nuclease degradation, denaturation, or dissociation at extremely low concentration, presumably resulting from the dense DNA packaging in NFs. The exceptional biostability is critical for biomedical applications. By rational design, NFs can be readily incorporated with myriad functional moieties. All these properties make NFs promising for versatile applications. As a proof-of-principle demonstration, in this study, NFs were integrated with aptamers, bioimaging agents, and drug loading sites, and the resultant multifunctional NFs were demonstrated for selective cancer cell recognition, bioimaging, and targeted anticancer drug delivery.

  10. Three invariant Hi-C interaction patterns: Applications to genome assembly.

    Science.gov (United States)

    Oddes, Sivan; Zelig, Aviv; Kaplan, Noam

    2018-06-01

    Assembly of reference-quality genomes from next-generation sequencing data is a key challenge in genomics. Recently, we and others have shown that Hi-C data can be used to address several outstanding challenges in the field of genome assembly. This principle has since been developed in academia and industry, and has been used in the assembly of several major genomes. In this paper, we explore the central principles underlying Hi-C-based assembly approaches, by quantitatively defining and characterizing three invariant Hi-C interaction patterns on which these approaches can build: Intrachromosomal interaction enrichment, distance-dependent interaction decay and local interaction smoothness. Specifically, we evaluate to what degree each invariant pattern holds on a single locus level in different species, cell types and Hi-C map resolutions. We find that these patterns are generally consistent across species and cell types but are affected by sequencing depth, and that matrix balancing improves consistency of loci with all three invariant patterns. Finally, we overview current Hi-C-based assembly approaches in light of these invariant patterns and demonstrate how local interaction smoothness can be used to easily detect scaffolding errors in extremely sparse Hi-C maps. We suggest that simultaneously considering all three invariant patterns may lead to better Hi-C-based genome assembly methods. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Functionalization and Self-Assembly of DNA Bidimensional Arrays

    Directory of Open Access Journals (Sweden)

    Ramon Eritja

    2011-09-01

    Full Text Available Oligonucleotides carrying amino, thiol groups, as well as fluorescein, c-myc peptide sequence and nanogold at internal positions were prepared and used for the assembly of bidimensional DNA arrays.

  12. Comparison of next generation sequencing technologies for transcriptome characterization

    Directory of Open Access Journals (Sweden)

    Soltis Douglas E

    2009-08-01

    Full Text Available Abstract Background We have developed a simulation approach to help determine the optimal mixture of sequencing methods for most complete and cost effective transcriptome sequencing. We compared simulation results for traditional capillary sequencing with "Next Generation" (NG ultra high-throughput technologies. The simulation model was parameterized using mappings of 130,000 cDNA sequence reads to the Arabidopsis genome (NCBI Accession SRA008180.19. We also generated 454-GS20 sequences and de novo assemblies for the basal eudicot California poppy (Eschscholzia californica and the magnoliid avocado (Persea americana using a variety of methods for cDNA synthesis. Results The Arabidopsis reads tagged more than 15,000 genes, including new splice variants and extended UTR regions. Of the total 134,791 reads (13.8 MB, 119,518 (88.7% mapped exactly to known exons, while 1,117 (0.8% mapped to introns, 11,524 (8.6% spanned annotated intron/exon boundaries, and 3,066 (2.3% extended beyond the end of annotated UTRs. Sequence-based inference of relative gene expression levels correlated significantly with microarray data. As expected, NG sequencing of normalized libraries tagged more genes than non-normalized libraries, although non-normalized libraries yielded more full-length cDNA sequences. The Arabidopsis data were used to simulate additional rounds of NG and traditional EST sequencing, and various combinations of each. Our simulations suggest a combination of FLX and Solexa sequencing for optimal transcriptome coverage at modest cost. We have also developed ESTcalc http://fgp.huck.psu.edu/NG_Sims/ngsim.pl, an online webtool, which allows users to explore the results of this study by specifying individualized costs and sequencing characteristics. Conclusion NG sequencing technologies are a highly flexible set of platforms that can be scaled to suit different project goals. In terms of sequence coverage alone, the NG sequencing is a dramatic advance

  13. MetaQUAST: evaluation of metagenome assemblies.

    Science.gov (United States)

    Mikheenko, Alla; Saveliev, Vladislav; Gurevich, Alexey

    2016-04-01

    During the past years we have witnessed the rapid development of new metagenome assembly methods. Although there are many benchmark utilities designed for single-genome assemblies, there is no well-recognized evaluation and comparison tool for metagenomic-specific analogues. In this article, we present MetaQUAST, a modification of QUAST, the state-of-the-art tool for genome assembly evaluation based on alignment of contigs to a reference. MetaQUAST addresses such metagenome datasets features as (i) unknown species content by detecting and downloading reference sequences, (ii) huge diversity by giving comprehensive reports for multiple genomes and (iii) presence of highly relative species by detecting chimeric contigs. We demonstrate MetaQUAST performance by comparing several leading assemblers on one simulated and two real datasets. http://bioinf.spbau.ru/metaquast aleksey.gurevich@spbu.ru Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  14. Whole-Genome Sequences of Three Symbiotic Endozoicomonas Bacteria

    KAUST Repository

    Neave, Matthew J.

    2014-08-14

    Members of the genus Endozoicomonas associate with a wide range of marine organisms. Here, we report on the whole-genome sequencing, assembly, and annotation of three Endozoicomonas type strains. These data will assist in exploring interactions between Endozoicomonas organisms and their hosts, and it will aid in the assembly of genomes from uncultivated Endozoicomonas spp.

  15. Whole-Genome Sequences of Three Symbiotic Endozoicomonas Bacteria

    KAUST Repository

    Neave, Matthew J.; Michell, Craig; Apprill, Amy; Voolstra, Christian R.

    2014-01-01

    Members of the genus Endozoicomonas associate with a wide range of marine organisms. Here, we report on the whole-genome sequencing, assembly, and annotation of three Endozoicomonas type strains. These data will assist in exploring interactions between Endozoicomonas organisms and their hosts, and it will aid in the assembly of genomes from uncultivated Endozoicomonas spp.

  16. Challenges and opportunities in understanding microbial communities with metagenome assembly (accompanied by IPython Notebook tutorial)

    Science.gov (United States)

    Howe, Adina; Chain, Patrick S. G.

    2015-01-01

    Metagenomic investigations hold great promise for informing the genetics, physiology, and ecology of environmental microorganisms. Current challenges for metagenomic analysis are related to our ability to connect the dots between sequencing reads, their population of origin, and their encoding functions. Assembly-based methods reduce dataset size by extending overlapping reads into larger contiguous sequences (contigs), providing contextual information for genetic sequences that does not rely on existing references. These methods, however, tend to be computationally intensive and are again challenged by sequencing errors as well as by genomic repeats While numerous tools have been developed based on these methodological concepts, they present confounding choices and training requirements to metagenomic investigators. To help with accessibility to assembly tools, this review also includes an IPython Notebook metagenomic assembly tutorial. This tutorial has instructions for execution any operating system using Amazon Elastic Cloud Compute and guides users through downloading, assembly, and mapping reads to contigs of a mock microbiome metagenome. Despite its challenges, metagenomic analysis has already revealed novel insights into many environments on Earth. As software, training, and data continue to emerge, metagenomic data access and its discoveries will to grow. PMID:26217314

  17. Challenges and opportunities in understanding microbial communities with metagenome assembly (accompanied by IPython Notebook tutorial

    Directory of Open Access Journals (Sweden)

    Adina eHowe

    2015-07-01

    Full Text Available Metagenomic investigations hold great promise for informing the genetics, physiology, and ecology of environmental microorganisms. Current challenges for metagenomic analysis are related to our ability to connect the dots between sequencing reads, their population of origin, and their encoding functions. Assembly-based methods reduce dataset size by extending overlapping reads into larger contiguous sequences (contigs, providing contextual information for genetic sequences that does not rely on existing references. These methods, however, tend to be computationally intensive and are again challenged by sequencing errors as well as by genomic repeats While numerous tools have been developed based on these methodological concepts, they present confounding choices and training requirements to metagenomic investigators. To help with accessibility to assembly tools, this review also includes an IPython Notebook metagenomic assembly tutorial. This tutorial has instructions for execution any operating system using Amazon Elastic Cloud Compute and guides users through downloading, assembly, and mapping reads to contigs of a mock microbiome metagenome. Despite its challenges, metagenomic analysis has already revealed novel insights into many environments on Earth. As software, training, and data continue to emerge, metagenomic data access and its discoveries will to grow.

  18. TAGUCHI METHOD FOR THREE-STAGE ASSEMBLY FLOW SHOP SCHEDULING PROBLEM WITH BLOCKING AND SEQUENCE-DEPENDENT SET UP TIMES

    Directory of Open Access Journals (Sweden)

    AREF MALEKI-DARONKOLAEI

    2013-10-01

    Full Text Available This article considers a three-stage assembly flowshop scheduling problem minimizing the weighted sum of mean completion time and makespan with sequence-dependent setup times at the first stage and blocking times between each stage. To tackle such an NP-hard, two meta-heuristic algorithms are presented. The novelty of our approach is to develop a variable neighborhood search algorithm (VNS and a well-known simulated annealing (SA for the problem. Furthermore, to enhance the performance of the (SA, its parameters are optimized by the use of Taguchi method, but to setting parameters of VNS just one parameter has been used without Taguchi. The computational results show that the proposed VNS is better in mean and standard deviation for all sizes of the problem than SA, but on the contrary about CPU Time SA outperforms VNS.

  19. RAMBO-K: Rapid and Sensitive Removal of Background Sequences from Next Generation Sequencing Data.

    Directory of Open Access Journals (Sweden)

    Simon H Tausch

    Full Text Available The assembly of viral or endosymbiont genomes from Next Generation Sequencing (NGS data is often hampered by the predominant abundance of reads originating from the host organism. These reads increase the memory and CPU time usage of the assembler and can lead to misassemblies.We developed RAMBO-K (Read Assignment Method Based On K-mers, a tool which allows rapid and sensitive removal of unwanted host sequences from NGS datasets. Reaching a speed of 10 Megabases/s on 4 CPU cores and a standard hard drive, RAMBO-K is faster than any tool we tested, while showing a consistently high sensitivity and specificity across different datasets.RAMBO-K rapidly and reliably separates reads from different species without data preprocessing. It is suitable as a straightforward standard solution for workflows dealing with mixed datasets. Binaries and source code (java and python are available from http://sourceforge.net/projects/rambok/.

  20. Sequencing of a QTL-rich region of the Theobroma cacao genome using pooled BACs and the identification of trait specific candidate genes

    Directory of Open Access Journals (Sweden)

    Blackmon Barbara P

    2011-07-01

    Full Text Available Abstract Background BAC-based physical maps provide for sequencing across an entire genome or a selected sub-genomic region of biological interest. Such a region can be approached with next-generation whole-genome sequencing and assembly as if it were an independent small genome. Using the minimum tiling path as a guide, specific BAC clones representing the prioritized genomic interval are selected, pooled, and used to prepare a sequencing library. Results This pooled BAC approach was taken to sequence and assemble a QTL-rich region, of ~3 Mbp and represented by twenty-seven BACs, on linkage group 5 of the Theobroma cacao cv. Matina 1-6 genome. Using various mixtures of read coverages from paired-end and linear 454 libraries, multiple assemblies of varied quality were generated. Quality was assessed by comparing the assembly of 454 reads with a subset of ten BACs individually sequenced and assembled using Sanger reads. A mixture of reads optimal for assembly was identified. We found, furthermore, that a quality assembly suitable for serving as a reference genome template could be obtained even with a reduced depth of sequencing coverage. Annotation of the resulting assembly revealed several genes potentially responsible for three T. cacao traits: black pod disease resistance, bean shape index, and pod weight. Conclusions Our results, as with other pooled BAC sequencing reports, suggest that pooling portions of a minimum tiling path derived from a BAC-based physical map is an effective method to target sub-genomic regions for sequencing. While we focused on a single QTL region, other QTL regions of importance could be similarly sequenced allowing for biological discovery to take place before a high quality whole-genome assembly is completed.

  1. MEGGASENSE - The Metagenome/Genome Annotated Sequence Natural Language Search Engine: A Platform for 
the Construction of Sequence Data Warehouses.

    Science.gov (United States)

    Gacesa, Ranko; Zucko, Jurica; Petursdottir, Solveig K; Gudmundsdottir, Elisabet Eik; Fridjonsson, Olafur H; Diminic, Janko; Long, Paul F; Cullum, John; Hranueli, Daslav; Hreggvidsson, Gudmundur O; Starcevic, Antonio

    2017-06-01

    The MEGGASENSE platform constructs relational databases of DNA or protein sequences. The default functional analysis uses 14 106 hidden Markov model (HMM) profiles based on sequences in the KEGG database. The Solr search engine allows sophisticated queries and a BLAST search function is also incorporated. These standard capabilities were used to generate the SCATT database from the predicted proteome of Streptomyces cattleya . The implementation of a specialised metagenome database (AMYLOMICS) for bioprospecting of carbohydrate-modifying enzymes is described. In addition to standard assembly of reads, a novel 'functional' assembly was developed, in which screening of reads with the HMM profiles occurs before the assembly. The AMYLOMICS database incorporates additional HMM profiles for carbohydrate-modifying enzymes and it is illustrated how the combination of HMM and BLAST analyses helps identify interesting genes. A variety of different proteome and metagenome databases have been generated by MEGGASENSE.

  2. Evaluation of de novo assembly technique in the South African abalone Haliotis midae transcriptome: A comparison from Illumina and 454 systems

    Directory of Open Access Journals (Sweden)

    Barbara Picone

    2016-12-01

    Full Text Available Next generation sequencing platforms have recently been used to rapidly characterize transcriptome sequences from a number of non-model organisms. The present study compares two of the most frequently used platforms, the Roche 454-pyrosequencing and the Illumina sequencing-by-synthesis (SBS, on the same RNA sample obtained from an intertidal gastropod mollusc species, Haliotis midae. All the sequencing reads were deposited in the Short Read Archive (SRA database are retrievable under the accession number [SRR071314 (Illumina Genome Analyzer II] and [SRR1737738, SRR1737737, SRR1737735, SRR1737734 (454 GS FLX] in the SRA database of NCBI. Three transcriptomes, composed of either pure 454 or Illumina reads or a mixture of read types (Hybrid, were assembled using CLC Genomics Workbench software. Illumina assemblies performed the best de novo transcriptome characterization in terms of contig length, whereas the 454 assemblies tended to improve the complete assembly of gene transcripts. Both the Hybrid and Illumina assemblies produced longer contigs covering more of the transcriptome than 454 assemblies. However, the addition of 454 significantly increased the number of genes annotated.

  3. Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing

    Directory of Open Access Journals (Sweden)

    Yu-Chih Tsai

    2016-02-01

    Full Text Available Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant “genomes” are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation.

  4. Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing

    Science.gov (United States)

    Tsai, Yu-Chih; Deming, Clayton; Segre, Julia A.; Kong, Heidi H.; Korlach, Jonas

    2016-01-01

    ABSTRACT Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant “genomes” are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT) sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation. PMID:26861018

  5. Draft sequencing and assembly of the genome of the world's largest fish, the whale shark: Rhincodon typus Smith 1828.

    Science.gov (United States)

    Read, Timothy D; Petit, Robert A; Joseph, Sandeep J; Alam, Md Tauqeer; Weil, M Ryan; Ahmad, Maida; Bhimani, Ravila; Vuong, Jocelyn S; Haase, Chad P; Webb, D Harry; Tan, Milton; Dove, Alistair D M

    2017-07-14

    The whale shark (Rhincodon typus) has by far the largest body size of any elasmobranch (shark or ray) species. Therefore, it is also the largest extant species of the paraphyletic assemblage commonly referred to as fishes. As both a phenotypic extreme and a member of the group Chondrichthyes - the sister group to the remaining gnathostomes, which includes all tetrapods and therefore also humans - its genome is of substantial comparative interest. Whale sharks are also listed as an endangered species on the International Union for Conservation of Nature's Red List of threatened species and are of growing popularity as both a target of ecotourism and as a charismatic conservation ambassador for the pelagic ecosystem. A genome map for this species would aid in defining effective conservation units and understanding global population structure. We characterised the nuclear genome of the whale shark using next generation sequencing (454, Illumina) and de novo assembly and annotation methods, based on material collected from the Georgia Aquarium. The data set consisted of 878,654,233 reads, which yielded a draft assembly of 1,213,200 contigs and 997,976 scaffolds. The estimated genome size was 3.44Gb. As expected, the proteome of the whale shark was most closely related to the only other complete genome of a cartilaginous fish, the holocephalan elephant shark. The whale shark contained a novel Toll-like-receptor (TLR) protein with sequence similarity to both the TLR4 and TLR13 proteins of mammals and TLR21 of teleosts. The data are publicly available on GenBank, FigShare, and from the NCBI Short Read Archive under accession number SRP044374. This represents the first shotgun elasmobranch genome and will aid studies of molecular systematics, biogeography, genetic differentiation, and conservation genetics in this and other shark species, as well as providing comparative data for studies of evolutionary biology and immunology across the jawed vertebrate lineages.

  6. The diploid genome sequence of an individual human.

    Directory of Open Access Journals (Sweden)

    Samuel Levy

    2007-09-01

    Full Text Available Presented here is a genome sequence of an individual human. It was produced from approximately 32 million random DNA fragments, sequenced by Sanger dideoxy technology and assembled into 4,528 scaffolds, comprising 2,810 million bases (Mb of contiguous sequence with approximately 7.5-fold coverage for any given region. We developed a modified version of the Celera assembler to facilitate the identification and comparison of alternate alleles within this individual diploid genome. Comparison of this genome and the National Center for Biotechnology Information human reference assembly revealed more than 4.1 million DNA variants, encompassing 12.3 Mb. These variants (of which 1,288,319 were novel included 3,213,401 single nucleotide polymorphisms (SNPs, 53,823 block substitutions (2-206 bp, 292,102 heterozygous insertion/deletion events (indels(1-571 bp, 559,473 homozygous indels (1-82,711 bp, 90 inversions, as well as numerous segmental duplications and copy number variation regions. Non-SNP DNA variation accounts for 22% of all events identified in the donor, however they involve 74% of all variant bases. This suggests an important role for non-SNP genetic alterations in defining the diploid genome structure. Moreover, 44% of genes were heterozygous for one or more variants. Using a novel haplotype assembly strategy, we were able to span 1.5 Gb of genome sequence in segments >200 kb, providing further precision to the diploid nature of the genome. These data depict a definitive molecular portrait of a diploid human genome that provides a starting point for future genome comparisons and enables an era of individualized genomic information.

  7. Genome Assembly and Computational Analysis Pipelines for Bacterial Pathogens

    KAUST Repository

    Rangkuti, Farania Gama Ardhina

    2011-06-01

    Pathogens lie behind the deadliest pandemics in history. To date, AIDS pandemic has resulted in more than 25 million fatal cases, while tuberculosis and malaria annually claim more than 2 million lives. Comparative genomic analyses are needed to gain insights into the molecular mechanisms of pathogens, but the abundance of biological data dictates that such studies cannot be performed without the assistance of computational approaches. This explains the significant need for computational pipelines for genome assembly and analyses. The aim of this research is to develop such pipelines. This work utilizes various bioinformatics approaches to analyze the high-­throughput genomic sequence data that has been obtained from several strains of bacterial pathogens. A pipeline has been compiled for quality control for sequencing and assembly, and several protocols have been developed to detect contaminations. Visualization has been generated of genomic data in various formats, in addition to alignment, homology detection and sequence variant detection. We have also implemented a metaheuristic algorithm that significantly improves bacterial genome assemblies compared to other known methods. Experiments on Mycobacterium tuberculosis H37Rv data showed that our method resulted in improvement of N50 value of up to 9697% while consistently maintaining high accuracy, covering around 98% of the published reference genome. Other improvement efforts were also implemented, consisting of iterative local assemblies and iterative correction of contiguated bases. Our result expedites the genomic analysis of virulent genes up to single base pair resolution. It is also applicable to virtually every pathogenic microorganism, propelling further research in the control of and protection from pathogen-­associated diseases.

  8. Combinatorial Pooling Enables Selective Sequencing of the Barley Gene Space

    Science.gov (United States)

    Lonardi, Stefano; Duma, Denisa; Alpert, Matthew; Cordero, Francesca; Beccuti, Marco; Bhat, Prasanna R.; Wu, Yonghui; Ciardo, Gianfranco; Alsaihati, Burair; Ma, Yaqin; Wanamaker, Steve; Resnik, Josh; Bozdag, Serdar; Luo, Ming-Cheng; Close, Timothy J.

    2013-01-01

    For the vast majority of species – including many economically or ecologically important organisms, progress in biological research is hampered due to the lack of a reference genome sequence. Despite recent advances in sequencing technologies, several factors still limit the availability of such a critical resource. At the same time, many research groups and international consortia have already produced BAC libraries and physical maps and now are in a position to proceed with the development of whole-genome sequences organized around a physical map anchored to a genetic map. We propose a BAC-by-BAC sequencing protocol that combines combinatorial pooling design and second-generation sequencing technology to efficiently approach denovo selective genome sequencing. We show that combinatorial pooling is a cost-effective and practical alternative to exhaustive DNA barcoding when preparing sequencing libraries for hundreds or thousands of DNA samples, such as in this case gene-bearing minimum-tiling-path BAC clones. The novelty of the protocol hinges on the computational ability to efficiently compare hundred millions of short reads and assign them to the correct BAC clones (deconvolution) so that the assembly can be carried out clone-by-clone. Experimental results on simulated data for the rice genome show that the deconvolution is very accurate, and the resulting BAC assemblies have high quality. Results on real data for a gene-rich subset of the barley genome confirm that the deconvolution is accurate and the BAC assemblies have good quality. While our method cannot provide the level of completeness that one would achieve with a comprehensive whole-genome sequencing project, we show that it is quite successful in reconstructing the gene sequences within BACs. In the case of plants such as barley, this level of sequence knowledge is sufficient to support critical end-point objectives such as map-based cloning and marker-assisted breeding. PMID:23592960

  9. Combinatorial pooling enables selective sequencing of the barley gene space.

    Directory of Open Access Journals (Sweden)

    Stefano Lonardi

    2013-04-01

    Full Text Available For the vast majority of species - including many economically or ecologically important organisms, progress in biological research is hampered due to the lack of a reference genome sequence. Despite recent advances in sequencing technologies, several factors still limit the availability of such a critical resource. At the same time, many research groups and international consortia have already produced BAC libraries and physical maps and now are in a position to proceed with the development of whole-genome sequences organized around a physical map anchored to a genetic map. We propose a BAC-by-BAC sequencing protocol that combines combinatorial pooling design and second-generation sequencing technology to efficiently approach denovo selective genome sequencing. We show that combinatorial pooling is a cost-effective and practical alternative to exhaustive DNA barcoding when preparing sequencing libraries for hundreds or thousands of DNA samples, such as in this case gene-bearing minimum-tiling-path BAC clones. The novelty of the protocol hinges on the computational ability to efficiently compare hundred millions of short reads and assign them to the correct BAC clones (deconvolution so that the assembly can be carried out clone-by-clone. Experimental results on simulated data for the rice genome show that the deconvolution is very accurate, and the resulting BAC assemblies have high quality. Results on real data for a gene-rich subset of the barley genome confirm that the deconvolution is accurate and the BAC assemblies have good quality. While our method cannot provide the level of completeness that one would achieve with a comprehensive whole-genome sequencing project, we show that it is quite successful in reconstructing the gene sequences within BACs. In the case of plants such as barley, this level of sequence knowledge is sufficient to support critical end-point objectives such as map-based cloning and marker-assisted breeding.

  10. Combinatorial pooling enables selective sequencing of the barley gene space.

    Science.gov (United States)

    Lonardi, Stefano; Duma, Denisa; Alpert, Matthew; Cordero, Francesca; Beccuti, Marco; Bhat, Prasanna R; Wu, Yonghui; Ciardo, Gianfranco; Alsaihati, Burair; Ma, Yaqin; Wanamaker, Steve; Resnik, Josh; Bozdag, Serdar; Luo, Ming-Cheng; Close, Timothy J

    2013-04-01

    For the vast majority of species - including many economically or ecologically important organisms, progress in biological research is hampered due to the lack of a reference genome sequence. Despite recent advances in sequencing technologies, several factors still limit the availability of such a critical resource. At the same time, many research groups and international consortia have already produced BAC libraries and physical maps and now are in a position to proceed with the development of whole-genome sequences organized around a physical map anchored to a genetic map. We propose a BAC-by-BAC sequencing protocol that combines combinatorial pooling design and second-generation sequencing technology to efficiently approach denovo selective genome sequencing. We show that combinatorial pooling is a cost-effective and practical alternative to exhaustive DNA barcoding when preparing sequencing libraries for hundreds or thousands of DNA samples, such as in this case gene-bearing minimum-tiling-path BAC clones. The novelty of the protocol hinges on the computational ability to efficiently compare hundred millions of short reads and assign them to the correct BAC clones (deconvolution) so that the assembly can be carried out clone-by-clone. Experimental results on simulated data for the rice genome show that the deconvolution is very accurate, and the resulting BAC assemblies have high quality. Results on real data for a gene-rich subset of the barley genome confirm that the deconvolution is accurate and the BAC assemblies have good quality. While our method cannot provide the level of completeness that one would achieve with a comprehensive whole-genome sequencing project, we show that it is quite successful in reconstructing the gene sequences within BACs. In the case of plants such as barley, this level of sequence knowledge is sufficient to support critical end-point objectives such as map-based cloning and marker-assisted breeding.

  11. An advanced draft genome assembly of a desi type chickpea (Cicer arietinum L.).

    Science.gov (United States)

    Parween, Sabiha; Nawaz, Kashif; Roy, Riti; Pole, Anil K; Venkata Suresh, B; Misra, Gopal; Jain, Mukesh; Yadav, Gitanjali; Parida, Swarup K; Tyagi, Akhilesh K; Bhatia, Sabhyata; Chattopadhyay, Debasis

    2015-08-11

    Chickpea (Cicer arietinum L.) is an important pulse legume crop. We previously reported a draft genome assembly of the desi chickpea cultivar ICC 4958. Here we report an advanced version of the ICC 4958 genome assembly (version 2.0) generated using additional sequence data and an improved genetic map. This resulted in 2.7-fold increase in the length of the pseudomolecules and substantial reduction of sequence gaps. The genome assembly covered more than 94% of the estimated gene space and predicted the presence of 30,257 protein-coding genes including 2230 and 133 genes encoding potential transcription factors (TF) and resistance gene homologs, respectively. Gene expression analysis identified several TF and chickpea-specific genes with tissue-specific expression and displayed functional diversification of the paralogous genes. Pairwise comparison of pseudomolecules in the desi (ICC 4958) and the earlier reported kabuli (CDC Frontier) chickpea assemblies showed an extensive local collinearity with incongruity in the placement of large sequence blocks along the linkage groups, apparently due to use of different genetic maps. Single nucleotide polymorphism (SNP)-based mining of intra-specific polymorphism identified more than four thousand SNPs differentiating a desi group and a kabuli group of chickpea genotypes.

  12. A Framework for the Development of Automatic DFA Method to Minimize the Number of Components and Assembly Reorientations

    Science.gov (United States)

    Alfadhlani; Samadhi, T. M. A. Ari; Ma’ruf, Anas; Setiasyah Toha, Isa

    2018-03-01

    Assembly is a part of manufacturing processes that must be considered at the product design stage. Design for Assembly (DFA) is a method to evaluate product design in order to make it simpler, easier and quicker to assemble, so that assembly cost is reduced. This article discusses a framework for developing a computer-based DFA method. The method is expected to aid product designer to extract data, evaluate assembly process, and provide recommendation for the product design improvement. These three things are desirable to be performed without interactive process or user intervention, so product design evaluation process could be done automatically. Input for the proposed framework is a 3D solid engineering drawing. Product design evaluation is performed by: minimizing the number of components; generating assembly sequence alternatives; selecting the best assembly sequence based on the minimum number of assembly reorientations; and providing suggestion for design improvement.

  13. Hierarchical assembly of viral nanotemplates with encoded microparticles via nucleic acid hybridization.

    Science.gov (United States)

    Tan, Wui Siew; Lewis, Christina L; Horelik, Nicholas E; Pregibon, Daniel C; Doyle, Patrick S; Yi, Hyunmin

    2008-11-04

    We demonstrate hierarchical assembly of tobacco mosaic virus (TMV)-based nanotemplates with hydrogel-based encoded microparticles via nucleic acid hybridization. TMV nanotemplates possess a highly defined structure and a genetically engineered high density thiol functionality. The encoded microparticles are produced in a high throughput microfluidic device via stop-flow lithography (SFL) and consist of spatially discrete regions containing encoded identity information, an internal control, and capture DNAs. For the hybridization-based assembly, partially disassembled TMVs were programmed with linker DNAs that contain sequences complementary to both the virus 5' end and a selected capture DNA. Fluorescence microscopy, atomic force microscopy (AFM), and confocal microscopy results clearly indicate facile assembly of TMV nanotemplates onto microparticles with high spatial and sequence selectivity. We anticipate that our hybridization-based assembly strategy could be employed to create multifunctional viral-synthetic hybrid materials in a rapid and high-throughput manner. Additionally, we believe that these viral-synthetic hybrid microparticles may find broad applications in high capacity, multiplexed target sensing.

  14. De novo assembly, gene annotation, and marker discovery in stored-product pest Liposcelis entomophila (Enderlein using transcriptome sequences.

    Directory of Open Access Journals (Sweden)

    Dan-Dan Wei

    Full Text Available BACKGROUND: As a major stored-product pest insect, Liposcelis entomophila has developed high levels of resistance to various insecticides in grain storage systems. However, the molecular mechanisms underlying resistance and environmental stress have not been characterized. To date, there is a lack of genomic information for this species. Therefore, studies aimed at profiling the L. entomophila transcriptome would provide a better understanding of the biological functions at the molecular levels. METHODOLOGY/PRINCIPAL FINDINGS: We applied Illumina sequencing technology to sequence the transcriptome of L. entomophila. A total of 54,406,328 clean reads were obtained and that de novo assembled into 54,220 unigenes, with an average length of 571 bp. Through a similarity search, 33,404 (61.61% unigenes were matched to known proteins in the NCBI non-redundant (Nr protein database. These unigenes were further functionally annotated with gene ontology (GO, cluster of orthologous groups of proteins (COG, and Kyoto Encyclopedia of Genes and Genomes (KEGG databases. A large number of genes potentially involved in insecticide resistance were manually curated, including 68 putative cytochrome P450 genes, 37 putative glutathione S-transferase (GST genes, 19 putative carboxyl/cholinesterase (CCE genes, and other 126 transcripts to contain target site sequences or encoding detoxification genes representing eight types of resistance enzymes. Furthermore, to gain insight into the molecular basis of the L. entomophila toward thermal stresses, 25 heat shock protein (Hsp genes were identified. In addition, 1,100 SSRs and 57,757 SNPs were detected and 231 pairs of SSR primes were designed for investigating the genetic diversity in future. CONCLUSIONS/SIGNIFICANCE: We developed a comprehensive transcriptomic database for L. entomophila. These sequences and putative molecular markers would further promote our understanding of the molecular mechanisms underlying

  15. De novo assembly of the pepper transcriptome (Capsicum annuum): a benchmark for in silico discovery of SNPs, SSRs and candidate genes

    Science.gov (United States)

    2012-01-01

    Background Molecular breeding of pepper (Capsicum spp.) can be accelerated by developing DNA markers associated with transcriptomes in breeding germplasm. Before the advent of next generation sequencing (NGS) technologies, the majority of sequencing data were generated by the Sanger sequencing method. By leveraging Sanger EST data, we have generated a wealth of genetic information for pepper including thousands of SNPs and Single Position Polymorphic (SPP) markers. To complement and enhance these resources, we applied NGS to three pepper genotypes: Maor, Early Jalapeño and Criollo de Morelos-334 (CM334) to identify SNPs and SSRs in the assembly of these three genotypes. Results Two pepper transcriptome assemblies were developed with different purposes. The first reference sequence, assembled by CAP3 software, comprises 31,196 contigs from >125,000 Sanger-EST sequences that were mainly derived from a Korean F1-hybrid line, Bukang. Overlapping probes were designed for 30,815 unigenes to construct a pepper Affymetrix GeneChip® microarray for whole genome analyses. In addition, custom Python scripts were used to identify 4,236 SNPs in contigs of the assembly. A total of 2,489 simple sequence repeats (SSRs) were identified from the assembly, and primers were designed for the SSRs. Annotation of contigs using Blast2GO software resulted in information for 60% of the unigenes in the assembly. The second transcriptome assembly was constructed from more than 200 million Illumina Genome Analyzer II reads (80–120 nt) using a combination of Velvet, CLC workbench and CAP3 software packages. BWA, SAMtools and in-house Perl scripts were used to identify SNPs among three pepper genotypes. The SNPs were filtered to be at least 50 bp from any intron-exon junctions as well as flanking SNPs. More than 22,000 high-quality putative SNPs were identified. Using the MISA software, 10,398 SSR markers were also identified within the Illumina transcriptome assembly and primers were

  16. Advantages of genome sequencing by long-read sequencer using SMRT technology in medical area.

    Science.gov (United States)

    Nakano, Kazuma; Shiroma, Akino; Shimoji, Makiko; Tamotsu, Hinako; Ashimine, Noriko; Ohki, Shun; Shinzato, Misuzu; Minami, Maiko; Nakanishi, Tetsuhiro; Teruya, Kuniko; Satou, Kazuhito; Hirano, Takashi

    2017-07-01

    PacBio RS II is the first commercialized third-generation DNA sequencer able to sequence a single molecule DNA in real-time without amplification. PacBio RS II's sequencing technology is novel and unique, enabling the direct observation of DNA synthesis by DNA polymerase. PacBio RS II confers four major advantages compared to other sequencing technologies: long read lengths, high consensus accuracy, a low degree of bias, and simultaneous capability of epigenetic characterization. These advantages surmount the obstacle of sequencing genomic regions such as high/low G+C, tandem repeat, and interspersed repeat regions. Moreover, PacBio RS II is ideal for whole genome sequencing, targeted sequencing, complex population analysis, RNA sequencing, and epigenetics characterization. With PacBio RS II, we have sequenced and analyzed the genomes of many species, from viruses to humans. Herein, we summarize and review some of our key genome sequencing projects, including full-length viral sequencing, complete bacterial genome and almost-complete plant genome assemblies, and long amplicon sequencing of a disease-associated gene region. We believe that PacBio RS II is not only an effective tool for use in the basic biological sciences but also in the medical/clinical setting.

  17. Impact of genome assembly status on ChIP-Seq and ChIP-PET data mapping

    Directory of Open Access Journals (Sweden)

    Sachs Laurent

    2009-12-01

    Full Text Available Abstract Background ChIP-Seq and ChIP-PET can potentially be used with any genome for genome wide profiling of protein-DNA interaction sites. Unfortunately, it is probable that most genome assemblies will never reach the quality of the human genome assembly. Therefore, it remains to be determined whether ChIP-Seq and ChIP-PET are practicable with genome sequences other than a few (e.g. human and mouse. Findings Here, we used in silico simulations to assess the impact of completeness or fragmentation of genome assemblies on ChIP-Seq and ChIP-PET data mapping. Conclusions Most currently published genome assemblies are suitable for mapping the short sequence tags produced by ChIP-Seq or ChIP-PET.

  18. A mathematical model of an automatic assembler to stack fuel pellets

    International Nuclear Information System (INIS)

    Jarvis, R.G.; Joynes, R.; Bretzlaff, C.I.

    1980-11-01

    Fuel elements for CANDU reactors are assembled from stacks of cylindrical UO 2 pellets, with close tolerances on lengths and diameters. Present stacking techniques involve extensive manual operations and they can be speeded up and reduced in cost by an automated device. If gamma-active fuel is handled such a device is essential. An automatic fuel pellet assembly process was modelled mathematically. The model indicated a suitable sequence of pellet manipulations to arrive at a stack length that was always within tolerance. This sequence was used as the inital input for the design of mechanical hardware. The mechanical design and the refinement of the mathematical model proceeded simultaneously. Mechanical constraints were allowed for in the model, and its optimized sequence of operations was incorporated in a microcomputer program to control the mechanical hardware. (auth)

  19. Integrating sequencing technologies in personal genomics: optimal low cost reconstruction of structural variants.

    Directory of Open Access Journals (Sweden)

    Jiang Du

    2009-07-01

    Full Text Available The goal of human genome re-sequencing is obtaining an accurate assembly of an individual's genome. Recently, there has been great excitement in the development of many technologies for this (e.g. medium and short read sequencing from companies such as 454 and SOLiD, and high-density oligo-arrays from Affymetrix and NimbelGen, with even more expected to appear. The costs and sensitivities of these technologies differ considerably from each other. As an important goal of personal genomics is to reduce the cost of re-sequencing to an affordable point, it is worthwhile to consider optimally integrating technologies. Here, we build a simulation toolbox that will help us optimally combine different technologies for genome re-sequencing, especially in reconstructing large structural variants (SVs. SV reconstruction is considered the most challenging step in human genome re-sequencing. (It is sometimes even harder than de novo assembly of small genomes because of the duplications and repetitive sequences in the human genome. To this end, we formulate canonical problems that are representative of issues in reconstruction and are of small enough scale to be computationally tractable and simulatable. Using semi-realistic simulations, we show how we can combine different technologies to optimally solve the assembly at low cost. With mapability maps, our simulations efficiently handle the inhomogeneous repeat-containing structure of the human genome and the computational complexity of practical assembly algorithms. They quantitatively show how combining different read lengths is more cost-effective than using one length, how an optimal mixed sequencing strategy for reconstructing large novel SVs usually also gives accurate detection of SNPs/indels, how paired-end reads can improve reconstruction efficiency, and how adding in arrays is more efficient than just sequencing for disentangling some complex SVs. Our strategy should facilitate the sequencing of

  20. Improvement of the banana "Musa acuminata" reference sequence using NGS data and semi-automated bioinformatics methods.

    Science.gov (United States)

    Martin, Guillaume; Baurens, Franc-Christophe; Droc, Gaëtan; Rouard, Mathieu; Cenci, Alberto; Kilian, Andrzej; Hastie, Alex; Doležel, Jaroslav; Aury, Jean-Marc; Alberti, Adriana; Carreel, Françoise; D'Hont, Angélique

    2016-03-16

    Recent advances in genomics indicate functional significance of a majority of genome sequences and their long range interactions. As a detailed examination of genome organization and function requires very high quality genome sequence, the objective of this study was to improve reference genome assembly of banana (Musa acuminata). We have developed a modular bioinformatics pipeline to improve genome sequence assemblies, which can handle various types of data. The pipeline comprises several semi-automated tools. However, unlike classical automated tools that are based on global parameters, the semi-automated tools proposed an expert mode for a user who can decide on suggested improvements through local compromises. The pipeline was used to improve the draft genome sequence of Musa acuminata. Genotyping by sequencing (GBS) of a segregating population and paired-end sequencing were used to detect and correct scaffold misassemblies. Long insert size paired-end reads identified scaffold junctions and fusions missed by automated assembly methods. GBS markers were used to anchor scaffolds to pseudo-molecules with a new bioinformatics approach that avoids the tedious step of marker ordering during genetic map construction. Furthermore, a genome map was constructed and used to assemble scaffolds into super scaffolds. Finally, a consensus gene annotation was projected on the new assembly from two pre-existing annotations. This approach reduced the total Musa scaffold number from 7513 to 1532 (i.e. by 80%), with an N50 that increased from 1.3 Mb (65 scaffolds) to 3.0 Mb (26 scaffolds). 89.5% of the assembly was anchored to the 11 Musa chromosomes compared to the previous 70%. Unknown sites (N) were reduced from 17.3 to 10.0%. The release of the Musa acuminata reference genome version 2 provides a platform for detailed analysis of banana genome variation, function and evolution. Bioinformatics tools developed in this work can be used to improve genome sequence assemblies in

  1. A pipeline for the de novo assembly of the Themira biloba (Sepsidae: Diptera) transcriptome using a multiple k-mer length approach.

    Science.gov (United States)

    Melicher, Dacotah; Torson, Alex S; Dworkin, Ian; Bowsher, Julia H

    2014-03-12

    The Sepsidae family of flies is a model for investigating how sexual selection shapes courtship and sexual dimorphism in a comparative framework. However, like many non-model systems, there are few molecular resources available. Large-scale sequencing and assembly have not been performed in any sepsid, and the lack of a closely related genome makes investigation of gene expression challenging. Our goal was to develop an automated pipeline for de novo transcriptome assembly, and to use that pipeline to assemble and analyze the transcriptome of the sepsid Themira biloba. Our bioinformatics pipeline uses cloud computing services to assemble and analyze the transcriptome with off-site data management, processing, and backup. It uses a multiple k-mer length approach combined with a second meta-assembly to extend transcripts and recover more bases of transcript sequences than standard single k-mer assembly. We used 454 sequencing to generate 1.48 million reads from cDNA generated from embryo, larva, and pupae of T. biloba and assembled a transcriptome consisting of 24,495 contigs. Annotation identified 16,705 transcripts, including those involved in embryogenesis and limb patterning. We assembled transcriptomes from an additional three non-model organisms to demonstrate that our pipeline assembled a higher-quality transcriptome than single k-mer approaches across multiple species. The pipeline we have developed for assembly and analysis increases contig length, recovers unique transcripts, and assembles more base pairs than other methods through the use of a meta-assembly. The T. biloba transcriptome is a critical resource for performing large-scale RNA-Seq investigations of gene expression patterns, and is the first transcriptome sequenced in this Dipteran family.

  2. Transcriptome survey of Patagonian southern beech Nothofagus nervosa (= N. Alpina: assembly, annotation and molecular marker discovery

    Directory of Open Access Journals (Sweden)

    Torales Susana L

    2012-07-01

    Full Text Available Abstract Background Nothofagus nervosa is one of the most emblematic native tree species of Patagonian temperate forests. Here, the shotgun RNA-sequencing (RNA-Seq of the transcriptome of N. nervosa, including de novo assembly, functional annotation, and in silico discovery of potential molecular markers to support population and associations genetic studies, are described. Results Pyrosequencing of a young leaf cDNA library generated a total of 111,814 high quality reads, with an average length of 447 bp. De novo assembly using Newbler resulted into 3,005 tentative isotigs (including alternative transcripts. The non-assembled sequences (singletons were clustered with CD-HIT-454 to identify natural and artificial duplicates from pyrosequencing reads, leading to 21,881 unique singletons. 15,497 out of 24,886 non-redundant sequences or unigenes, were successfully annotated against a plant protein database. A substantial number of simple sequence repeat markers (SSRs were discovered in the assembled and annotated sequences. More than 40% of the SSR sequences were inside ORF sequences. To confirm the validity of these predicted markers, a subset of 73 SSRs selected through functional annotation evidences were successfully amplified from six seedlings DNA samples, being 14 polymorphic. Conclusions This paper is the first report that shows a highly precise representation of the mRNAs diversity present in young leaves of a native South American tree, N. nervosa, as well as its in silico deduced putative functionality. The reported Nothofagus transcriptome sequences represent a unique resource for genetic studies and provide a tool to discover genes of interest and genetic markers that will greatly aid questions involving evolution, ecology, and conservation using genetic and genomic approaches in the genus.

  3. Evaluating de Bruijn graph assemblers on 454 transcriptomic data.

    Directory of Open Access Journals (Sweden)

    Xianwen Ren

    Full Text Available Next generation sequencing (NGS technologies have greatly changed the landscape of transcriptomic studies of non-model organisms. Since there is no reference genome available, de novo assembly methods play key roles in the analysis of these data sets. Because of the huge amount of data generated by NGS technologies for each run, many assemblers, e.g., ABySS, Velvet and Trinity, are developed based on a de Bruijn graph due to its time- and space-efficiency. However, most of these assemblers were developed initially for the Illumina/Solexa platform. The performance of these assemblers on 454 transcriptomic data is unknown. In this study, we evaluated and compared the relative performance of these de Bruijn graph based assemblers on both simulated and real 454 transcriptomic data. The results suggest that Trinity, the Illumina/Solexa-specialized transcriptomic assembler, performs the best among the multiple de Bruijn graph assemblers, comparable to or even outperforming the standard 454 assembler Newbler which is based on the overlap-layout-consensus algorithm. Our evaluation is expected to provide helpful guidance for researchers to choose assemblers when analyzing 454 transcriptomic data.

  4. SeqEntropy: genome-wide assessment of repeats for short read sequencing.

    Directory of Open Access Journals (Sweden)

    Hsueh-Ting Chu

    Full Text Available BACKGROUND: Recent studies on genome assembly from short-read sequencing data reported the limitation of this technology to reconstruct the entire genome even at very high depth coverage. We investigated the limitation from the perspective of information theory to evaluate the effect of repeats on short-read genome assembly using idealized (error-free reads at different lengths. METHODOLOGY/PRINCIPAL FINDINGS: We define a metric H(k to be the entropy of sequencing reads at a read length k and use the relative loss of entropy ΔH(k to measure the impact of repeats for the reconstruction of whole-genome from sequences of length k. In our experiments, we found that entropy loss correlates well with de-novo assembly coverage of a genome, and a score of ΔH(k>1% indicates a severe loss in genome reconstruction fidelity. The minimal read lengths to achieve ΔH(k<1% are different for various organisms and are independent of the genome size. For example, in order to meet the threshold of ΔH(k<1%, a read length of 60 bp is needed for the sequencing of human genome (3.2 10(9 bp and 320 bp for the sequencing of fruit fly (1.8×10(8 bp. We also calculated the ΔH(k scores for 2725 prokaryotic chromosomes and plasmids at several read lengths. Our results indicate that the levels of repeats in different genomes are diverse and the entropy of sequencing reads provides a measurement for the repeat structures. CONCLUSIONS/SIGNIFICANCE: The proposed entropy-based measurement, which can be calculated in seconds to minutes in most cases, provides a rapid quantitative evaluation on the limitation of idealized short-read genome sequencing. Moreover, the calculation can be parallelized to scale up to large euakryotic genomes. This approach may be useful to tune the sequencing parameters to achieve better genome assemblies when a closely related genome is already available.

  5. Cholera toxin B subunits assemble into pentamers--proposition of a fly-casting mechanism.

    Directory of Open Access Journals (Sweden)

    Jihad Zrimi

    Full Text Available The cholera toxin B pentamer (CtxB(5, which belongs to the AB(5 toxin family, is used as a model study for protein assembly. The effect of the pH on the reassembly of the toxin was investigated using immunochemical, electrophoretic and spectroscopic methods. Three pH-dependent steps were identified during the toxin reassembly: (i acquisition of a fully assembly-competent fold by the CtxB monomer, (ii association of CtxB monomer into oligomers, (iii acquisition of the native fold by the CtxB pentamer. The results show that CtxB(5 and the related heat labile enterotoxin LTB(5 have distinct mechanisms of assembly despite sharing high sequence identity (84% and almost identical atomic structures. The difference can be pinpointed to four histidines which are spread along the protein sequence and may act together. Thus, most of the toxin B amino acids appear negligible for the assembly, raising the possibility that assembly is driven by a small network of amino acids instead of involving all of them.

  6. Assembly and diploid architecture of an individual human genome via single-molecule technologies.

    Science.gov (United States)

    Pendleton, Matthew; Sebra, Robert; Pang, Andy Wing Chun; Ummat, Ajay; Franzen, Oscar; Rausch, Tobias; Stütz, Adrian M; Stedman, William; Anantharaman, Thomas; Hastie, Alex; Dai, Heng; Fritz, Markus Hsi-Yang; Cao, Han; Cohain, Ariella; Deikus, Gintaras; Durrett, Russell E; Blanchard, Scott C; Altman, Roger; Chin, Chen-Shan; Guo, Yan; Paxinos, Ellen E; Korbel, Jan O; Darnell, Robert B; McCombie, W Richard; Kwok, Pui-Yan; Mason, Christopher E; Schadt, Eric E; Bashir, Ali

    2015-08-01

    We present the first comprehensive analysis of a diploid human genome that combines single-molecule sequencing with single-molecule genome maps. Our hybrid assembly markedly improves upon the contiguity observed from traditional shotgun sequencing approaches, with scaffold N50 values approaching 30 Mb, and we identified complex structural variants (SVs) missed by other high-throughput approaches. Furthermore, by combining Illumina short-read data with long reads, we phased both single-nucleotide variants and SVs, generating haplotypes with over 99% consistency with previous trio-based studies. Our work shows that it is now possible to integrate single-molecule and high-throughput sequence data to generate de novo assembled genomes that approach reference quality.

  7. Comparison of de novo assembly statistics of Cucumis sativus L.

    Science.gov (United States)

    Wojcieszek, Michał; Kuśmirek, Wiktor; Pawełkowicz, Magdalena; PlÄ der, Wojciech; Nowak, Robert M.

    2017-08-01

    Genome sequencing is the core of genomic research. With the development of NGS and lowering the cost of procedure there is another tight gap - genome assembly. Developing the proper tool for this task is essential as quality of genome has important impact on further research. Here we present comparison of several de Bruijn assemblers tested on C. sativus genomic reads. The assessment shows that newly developed software - dnaasm provides better results in terms of quantity and quality. The number of generated sequences is lower by 5 - 33% with even two fold higher N50. Quality check showed reliable results were generated by dnaasm. This provides us with very strong base for future genomic analysis.

  8. Matlab enhanced multi-threaded tomography optimization sequence (MEMTOS)

    International Nuclear Information System (INIS)

    Lum, Edward S.; Pope, Chad L.

    2016-01-01

    Highlights: • Monte Carlo simulation of spent nuclear fuel assembly neutron computed tomography. • Optimized parallel calculations conducted from within the MATLAB environment. • Projection difference technique used to identify anomalies in spent nuclear fuel assemblies. - Abstract: One challenge associated with spent nuclear fuel assemblies is the lack of non-destructive analysis techniques to determine if fuel pins have been removed or replaced or if there are significant defects associated with fuel pins deep within a fuel assembly. Neutron computed tomography is a promising technique for addressing these qualitative issues. Monte Carlo simulation of spent nuclear fuel neutron computed tomography allows inexpensive process investigation and optimization. The main purpose of this work is to provide a fully automated advanced simulation framework for the analysis of spent nuclear fuel inspection using neutron computed tomography. The simulation framework, called Matlab Enhanced Multi-Threaded Tomography Optimization Sequence (MEMTOS) not only automates the simulation process, but also generates superior tomography image results. MEMTOS is written in the MATLAB scripting language and addresses file management, parallel Monte Carlo execution, results extraction, and tomography image generation. This paper describes the mathematical basis for neutron computed tomography, the Monte Carlo technique used to simulate neutron computed tomography, and the overall tomography simulation optimization algorithm. Sequence results presented include overall simulation speed enhancement, tomography and image results obtained for Experimental Breeder Reactor II spent fuel assemblies and light water reactor fuel assemblies. Optimization using a projection difference technique are also described.

  9. Sensitivity Analysis of Deviation Source for Fast Assembly Precision Optimization

    Directory of Open Access Journals (Sweden)

    Jianjun Tang

    2014-01-01

    Full Text Available Assembly precision optimization of complex product has a huge benefit in improving the quality of our products. Due to the impact of a variety of deviation source coupling phenomena, the goal of assembly precision optimization is difficult to be confirmed accurately. In order to achieve optimization of assembly precision accurately and rapidly, sensitivity analysis of deviation source is proposed. First, deviation source sensitivity is defined as the ratio of assembly dimension variation and deviation source dimension variation. Second, according to assembly constraint relations, assembly sequences and locating, deviation transmission paths are established by locating the joints between the adjacent parts, and establishing each part’s datum reference frame. Third, assembly multidimensional vector loops are created using deviation transmission paths, and the corresponding scalar equations of each dimension are established. Then, assembly deviation source sensitivity is calculated by using a first-order Taylor expansion and matrix transformation method. Finally, taking assembly precision optimization of wing flap rocker as an example, the effectiveness and efficiency of the deviation source sensitivity analysis method are verified.

  10. Genomic characterization of large heterochromatic gaps in the human genome assembly.

    Directory of Open Access Journals (Sweden)

    Nicolas Altemose

    2014-05-01

    Full Text Available The largest gaps in the human genome assembly correspond to multi-megabase heterochromatic regions composed primarily of two related families of tandem repeats, Human Satellites 2 and 3 (HSat2,3. The abundance of repetitive DNA in these regions challenges standard mapping and assembly algorithms, and as a result, the sequence composition and potential biological functions of these regions remain largely unexplored. Furthermore, existing genomic tools designed to predict consensus-based descriptions of repeat families cannot be readily applied to complex satellite repeats such as HSat2,3, which lack a consistent repeat unit reference sequence. Here we present an alignment-free method to characterize complex satellites using whole-genome shotgun read datasets. Utilizing this approach, we classify HSat2,3 sequences into fourteen subfamilies and predict their chromosomal distributions, resulting in a comprehensive satellite reference database to further enable genomic studies of heterochromatic regions. We also identify 1.3 Mb of non-repetitive sequence interspersed with HSat2,3 across 17 unmapped assembly scaffolds, including eight annotated gene predictions. Finally, we apply our satellite reference database to high-throughput sequence data from 396 males to estimate array size variation of the predominant HSat3 array on the Y chromosome, confirming that satellite array sizes can vary between individuals over an order of magnitude (7 to 98 Mb and further demonstrating that array sizes are distributed differently within distinct Y haplogroups. In summary, we present a novel framework for generating initial reference databases for unassembled genomic regions enriched with complex satellite DNA, and we further demonstrate the utility of these reference databases for studying patterns of sequence variation within human populations.

  11. Identification of human chromosome 22 transcribed sequences with ORF expressed sequence tags

    Science.gov (United States)

    de Souza, Sandro J.; Camargo, Anamaria A.; Briones, Marcelo R. S.; Costa, Fernando F.; Nagai, Maria Aparecida; Verjovski-Almeida, Sergio; Zago, Marco A.; Andrade, Luis Eduardo C.; Carrer, Helaine; El-Dorry, Hamza F. A.; Espreafico, Enilza M.; Habr-Gama, Angelita; Giannella-Neto, Daniel; Goldman, Gustavo H.; Gruber, Arthur; Hackel, Christine; Kimura, Edna T.; Maciel, Rui M. B.; Marie, Suely K. N.; Martins, Elizabeth A. L.; Nóbrega, Marina P.; Paçó-Larson, Maria Luisa; Pardini, Maria Inês M. C.; Pereira, Gonçalo G.; Pesquero, João Bosco; Rodrigues, Vanderlei; Rogatto, Silvia R.; da Silva, Ismael D. C. G.; Sogayar, Mari C.; de Fátima Sonati, Maria; Tajara, Eloiza H.; Valentini, Sandro R.; Acencio, Marcio; Alberto, Fernando L.; Amaral, Maria Elisabete J.; Aneas, Ivy; Bengtson, Mário Henrique; Carraro, Dirce M.; Carvalho, Alex F.; Carvalho, Lúcia Helena; Cerutti, Janete M.; Corrêa, Maria Lucia C.; Costa, Maria Cristina R.; Curcio, Cyntia; Gushiken, Tsieko; Ho, Paulo L.; Kimura, Elza; Leite, Luciana C. C.; Maia, Gustavo; Majumder, Paromita; Marins, Mozart; Matsukuma, Adriana; Melo, Analy S. A.; Mestriner, Carlos Alberto; Miracca, Elisabete C.; Miranda, Daniela C.; Nascimento, Ana Lucia T. O.; Nóbrega, Francisco G.; Ojopi, Élida P. B.; Pandolfi, José Rodrigo C.; Pessoa, Luciana Gilbert; Rahal, Paula; Rainho, Claudia A.; da Ro's, Nancy; de Sá, Renata G.; Sales, Magaly M.; da Silva, Neusa P.; Silva, Tereza C.; da Silva, Wilson; Simão, Daniel F.; Sousa, Josane F.; Stecconi, Daniella; Tsukumo, Fernando; Valente, Valéria; Zalcberg, Heloisa; Brentani, Ricardo R.; Reis, Luis F. L.; Dias-Neto, Emmanuel; Simpson, Andrew J. G.

    2000-01-01

    Transcribed sequences in the human genome can be identified with confidence only by alignment with sequences derived from cDNAs synthesized from naturally occurring mRNAs. We constructed a set of 250,000 cDNAs that represent partial expressed gene sequences and that are biased toward the central coding regions of the resulting transcripts. They are termed ORF expressed sequence tags (ORESTES). The 250,000 ORESTES were assembled into 81,429 contigs. Of these, 1,181 (1.45%) were found to match sequences in chromosome 22 with at least one ORESTES contig for 162 (65.6%) of the 247 known genes, for 67 (44.6%) of the 150 related genes, and for 45 of the 148 (30.4%) EST-predicted genes on this chromosome. Using a set of stringent criteria to validate our sequences, we identified a further 219 previously unannotated transcribed sequences on chromosome 22. Of these, 171 were in fact also defined by EST or full length cDNA sequences available in GenBank but not utilized in the initial annotation of the first human chromosome sequence. Thus despite representing less than 15% of all expressed human sequences in the public databases at the time of the present analysis, ORESTES sequences defined 48 transcribed sequences on chromosome 22 not defined by other sequences. All of the transcribed sequences defined by ORESTES coincided with DNA regions predicted as encoding exons by genscan. (http://genes.mit.edu/GENSCAN.html). PMID:11070084

  12. Genomic treasure troves: complete genome sequencing of herbarium and insect museum specimens.

    Science.gov (United States)

    Staats, Martijn; Erkens, Roy H J; van de Vossenberg, Bart; Wieringa, Jan J; Kraaijeveld, Ken; Stielow, Benjamin; Geml, József; Richardson, James E; Bakker, Freek T

    2013-01-01

    Unlocking the vast genomic diversity stored in natural history collections would create unprecedented opportunities for genome-scale evolutionary, phylogenetic, domestication and population genomic studies. Many researchers have been discouraged from using historical specimens in molecular studies because of both generally limited success of DNA extraction and the challenges associated with PCR-amplifying highly degraded DNA. In today's next-generation sequencing (NGS) world, opportunities and prospects for historical DNA have changed dramatically, as most NGS methods are actually designed for taking short fragmented DNA molecules as templates. Here we show that using a standard multiplex and paired-end Illumina sequencing approach, genome-scale sequence data can be generated reliably from dry-preserved plant, fungal and insect specimens collected up to 115 years ago, and with minimal destructive sampling. Using a reference-based assembly approach, we were able to produce the entire nuclear genome of a 43-year-old Arabidopsis thaliana (Brassicaceae) herbarium specimen with high and uniform sequence coverage. Nuclear genome sequences of three fungal specimens of 22-82 years of age (Agaricus bisporus, Laccaria bicolor, Pleurotus ostreatus) were generated with 81.4-97.9% exome coverage. Complete organellar genome sequences were assembled for all specimens. Using de novo assembly we retrieved between 16.2-71.0% of coding sequence regions, and hence remain somewhat cautious about prospects for de novo genome assembly from historical specimens. Non-target sequence contaminations were observed in 2 of our insect museum specimens. We anticipate that future museum genomics projects will perhaps not generate entire genome sequences in all cases (our specimens contained relatively small and low-complexity genomes), but at least generating vital comparative genomic data for testing (phylo)genetic, demographic and genetic hypotheses, that become increasingly more horizontal

  13. Direct chloroplast sequencing: comparison of sequencing platforms and analysis tools for whole chloroplast barcoding.

    Directory of Open Access Journals (Sweden)

    Marta Brozynska

    Full Text Available Direct sequencing of total plant DNA using next generation sequencing technologies generates a whole chloroplast genome sequence that has the potential to provide a barcode for use in plant and food identification. Advances in DNA sequencing platforms may make this an attractive approach for routine plant identification. The HiSeq (Illumina and Ion Torrent (Life Technology sequencing platforms were used to sequence total DNA from rice to identify polymorphisms in the whole chloroplast genome sequence of a wild rice plant relative to cultivated rice (cv. Nipponbare. Consensus chloroplast sequences were produced by mapping sequence reads to the reference rice chloroplast genome or by de novo assembly and mapping of the resulting contigs to the reference sequence. A total of 122 polymorphisms (SNPs and indels between the wild and cultivated rice chloroplasts were predicted by these different sequencing and analysis methods. Of these, a total of 102 polymorphisms including 90 SNPs were predicted by both platforms. Indels were more variable with different sequencing methods, with almost all discrepancies found in homopolymers. The Ion Torrent platform gave no apparent false SNP but was less reliable for indels. The methods should be suitable for routine barcoding using appropriate combinations of sequencing platform and data analysis.

  14. Design strategies for self-assembly of discrete targets

    International Nuclear Information System (INIS)

    Madge, Jim; Miller, Mark A.

    2015-01-01

    Both biological and artificial self-assembly processes can take place by a range of different schemes, from the successive addition of identical building blocks to hierarchical sequences of intermediates, all the way to the fully addressable limit in which each component is unique. In this paper, we introduce an idealized model of cubic particles with patterned faces that allows self-assembly strategies to be compared and tested. We consider a simple octameric target, starting with the minimal requirements for successful self-assembly and comparing the benefits and limitations of more sophisticated hierarchical and addressable schemes. Simulations are performed using a hybrid dynamical Monte Carlo protocol that allows self-assembling clusters to rearrange internally while still providing Stokes-Einstein-like diffusion of aggregates of different sizes. Our simulations explicitly capture the thermodynamic, dynamic, and steric challenges typically faced by self-assembly processes, including competition between multiple partially completed structures. Self-assembly pathways are extracted from the simulation trajectories by a fully extendable scheme for identifying structural fragments, which are then assembled into history diagrams for successfully completed target structures. For the simple target, a one-component assembly scheme is most efficient and robust overall, but hierarchical and addressable strategies can have an advantage under some conditions if high yield is a priority

  15. DNA-PK dependent targeting of DNA-ends to a protein complex assembled on matrix attachment region DNA sequences

    International Nuclear Information System (INIS)

    Mauldin, S.K.; Getts, R.C.; Perez, M.L.; DiRienzo, S.; Stamato, T.D.

    2003-01-01

    Full text: We find that nuclear protein extracts from mammalian cells contain an activity that allows DNA ends to associate with circular pUC18 plasmid DNA. This activity requires the catalytic subunit of DNA-PK (DNA-PKcs) and Ku since it was not observed in mutants lacking Ku or DNA-PKcs but was observed when purified Ku/DNA-PKcs was added to these mutant extracts. Competition experiments between pUC18 and pUC18 plasmids containing various nuclear matrix attachment region (MAR) sequences suggest that DNA ends preferentially associate with plasmids containing MAR DNA sequences. At a 1:5 mass ratio of MAR to pUC18, approximately equal amounts of DNA end binding to the two plasmids were observed, while at a 1:1 ratio no pUC18 end-binding was observed. Calculation of relative binding activities indicates that DNA-end binding activities to MAR sequences was 7 to 21 fold higher than pUC18. Western analysis of proteins bound to pUC18 and MAR plasmids indicates that XRCC4, DNA ligase IV, scaffold attachment factor A, topoisomerase II, and poly(ADP-ribose) polymerase preferentially associate with the MAR plasmid in the absence or presence of DNA ends. In contrast, Ku and DNA-PKcs were found on the MAR plasmid only in the presence of DNA ends. After electroporation of a 32P-labeled DNA probe into human cells and cell fractionation, 87% of the total intercellular radioactivity remained in nuclei after a 0.5M NaCl extraction suggesting the probe was strongly bound in the nucleus. The above observations raise the possibility that DNA-PK targets DNA-ends to a repair and/or DNA damage signaling complex which is assembled on MAR sites in the nucleus

  16. Assembly of the Lactuca sativa, L. cv. Tizian draft genome sequence reveals differences within major resistance complex 1 as compared to the cv. Salinas reference genome.

    Science.gov (United States)

    Verwaaijen, Bart; Wibberg, Daniel; Nelkner, Johanna; Gordin, Miriam; Rupp, Oliver; Winkler, Anika; Bremges, Andreas; Blom, Jochen; Grosch, Rita; Pühler, Alfred; Schlüter, Andreas

    2018-02-10

    Lettuce (Lactuca sativa, L.) is an important annual plant of the family Asteraceae (Compositae). The commercial lettuce cultivar Tizian has been used in various scientific studies investigating the interaction of the plant with phytopathogens or biological control agents. Here, we present the de novo draft genome sequencing and gene prediction for this specific cultivar derived from transcriptome sequence data. The assembled scaffolds amount to a size of 2.22 Gb. Based on RNAseq data, 31,112 transcript isoforms were identified. Functional predictions for these transcripts were determined within the GenDBE annotation platform. Comparison with the cv. Salinas reference genome revealed a high degree of sequence similarity on genome and transcriptome levels, with an average amino acid identity of 99%. Furthermore, it was observed that two large regions are either missing or are highly divergent within the cv. Tizian genome compared to cv. Salinas. One of these regions covers the major resistance complex 1 region of cv. Salinas. The cv. Tizian draft genome sequence provides a valuable resource for future functional and transcriptome analyses focused on this lettuce cultivar. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Draft genome sequence of ramie, Boehmeria nivea (L.) Gaudich.

    Science.gov (United States)

    Luan, Ming-Bao; Jian, Jian-Bo; Chen, Ping; Chen, Jun-Hui; Chen, Jian-Hua; Gao, Qiang; Gao, Gang; Zhou, Ju-Hong; Chen, Kun-Mei; Guang, Xuan-Min; Chen, Ji-Kang; Zhang, Qian-Qian; Wang, Xiao-Fei; Fang, Long; Sun, Zhi-Min; Bai, Ming-Zhou; Fang, Xiao-Dong; Zhao, Shan-Cen; Xiong, He-Ping; Yu, Chun-Ming; Zhu, Ai-Guo

    2018-05-01

    Ramie, Boehmeria nivea (L.) Gaudich, family Urticaceae, is a plant native to eastern Asia, and one of the world's oldest fibre crops. It is also used as animal feed and for the phytoremediation of heavy metal-contaminated farmlands. Thus, the genome sequence of ramie was determined to explore the molecular basis of its fibre quality, protein content and phytoremediation. For further understanding ramie genome, different paired-end and mate-pair libraries were combined to generate 134.31 Gb of raw DNA sequences using the Illumina whole-genome shotgun sequencing approach. The highly heterozygous B. nivea genome was assembled using the Platanus Genome Assembler, which is an effective tool for the assembly of highly heterozygous genome sequences. The final length of the draft genome of this species was approximately 341.9 Mb (contig N50 = 22.62 kb, scaffold N50 = 1,126.36 kb). Based on ramie genome annotations, 30,237 protein-coding genes were predicted, and the repetitive element content was 46.3%. The completeness of the final assembly was evaluated by benchmarking universal single-copy orthologous genes (BUSCO); 90.5% of the 1,440 expected embryophytic genes were identified as complete, and 4.9% were identified as fragmented. Phylogenetic analysis based on single-copy gene families and one-to-one orthologous genes placed ramie with mulberry and cannabis, within the clade of urticalean rosids. Genome information of ramie will be a valuable resource for the conservation of endangered Boehmeria species and for future studies on the biogeography and characteristic evolution of members of Urticaceae. © 2018 John Wiley & Sons Ltd.

  18. CBrowse: a SAM/BAM-based contig browser for transcriptome assembly visualization and analysis.

    Science.gov (United States)

    Li, Pei; Ji, Guoli; Dong, Min; Schmidt, Emily; Lenox, Douglas; Chen, Liangliang; Liu, Qi; Liu, Lin; Zhang, Jie; Liang, Chun

    2012-09-15

    To address the impending need for exploring rapidly increased transcriptomics data generated for non-model organisms, we developed CBrowse, an AJAX-based web browser for visualizing and analyzing transcriptome assemblies and contigs. Designed in a standard three-tier architecture with a data pre-processing pipeline, CBrowse is essentially a Rich Internet Application that offers many seamlessly integrated web interfaces and allows users to navigate, sort, filter, search and visualize data smoothly. The pre-processing pipeline takes the contig sequence file in FASTA format and its relevant SAM/BAM file as the input; detects putative polymorphisms, simple sequence repeats and sequencing errors in contigs and generates image, JSON and database-compatible CSV text files that are directly utilized by different web interfaces. CBowse is a generic visualization and analysis tool that facilitates close examination of assembly quality, genetic polymorphisms, sequence repeats and/or sequencing errors in transcriptome sequencing projects. CBrowse is distributed under the GNU General Public License, available at http://bioinfolab.muohio.edu/CBrowse/ liangc@muohio.edu or liangc.mu@gmail.com; glji@xmu.edu.cn Supplementary data are available at Bioinformatics online.

  19. An improved filtering algorithm for big read datasets and its application to single-cell assembly.

    Science.gov (United States)

    Wedemeyer, Axel; Kliemann, Lasse; Srivastav, Anand; Schielke, Christian; Reusch, Thorsten B; Rosenstiel, Philip

    2017-07-03

    For single-cell or metagenomic sequencing projects, it is necessary to sequence with a very high mean coverage in order to make sure that all parts of the sample DNA get covered by the reads produced. This leads to huge datasets with lots of redundant data. A filtering of this data prior to assembly is advisable. Brown et al. (2012) presented the algorithm Diginorm for this purpose, which filters reads based on the abundance of their k-mers. We present Bignorm, a faster and quality-conscious read filtering algorithm. An important new algorithmic feature is the use of phred quality scores together with a detailed analysis of the k-mer counts to decide which reads to keep. We qualify and recommend parameters for our new read filtering algorithm. Guided by these parameters, we remove in terms of median 97.15% of the reads while keeping the mean phred score of the filtered dataset high. Using the SDAdes assembler, we produce assemblies of high quality from these filtered datasets in a fraction of the time needed for an assembly from the datasets filtered with Diginorm. We conclude that read filtering is a practical and efficient method for reducing read data and for speeding up the assembly process. This applies not only for single cell assembly, as shown in this paper, but also to other projects with high mean coverage datasets like metagenomic sequencing projects. Our Bignorm algorithm allows assemblies of competitive quality in comparison to Diginorm, while being much faster. Bignorm is available for download at https://git.informatik.uni-kiel.de/axw/Bignorm .

  20. A simple and versatile system for the ATP-dependent assembly of chromatin.

    Science.gov (United States)

    Khuong, Mai T; Fei, Jia; Cruz-Becerra, Grisel; Kadonaga, James T

    2017-11-24

    Chromatin is the natural form of DNA in the eukaryotic nucleus and is the substrate for diverse biological phenomena. The functional analysis of these processes ideally would be carried out with nucleosomal templates that are assembled with customized core histones, DNA sequences, and chromosomal proteins. Here we report a simple, reliable, and versatile method for the ATP-dependent assembly of evenly spaced nucleosome arrays. This minimal chromatin assembly system comprises the Drosophila nucleoplasmin-like protein (dNLP) histone chaperone, the imitation switch (ISWI) ATP-driven motor protein, core histones, template DNA, and ATP. The dNLP and ISWI components were synthesized in bacteria, and each protein could be purified in a single step by affinity chromatography. We show that the dNLP-ISWI system can be used with different DNA sequences, linear or circular DNA, bulk genomic DNA, recombinant or native Drosophila core histones, native human histones, the linker histone H1, the non-histone chromosomal protein HMGN2, and the core histone variants H3.3 and H2A.V. The dNLP-ISWI system should be accessible to a wide range of researchers and enable the assembly of customized chromatin with specifically desired DNA sequences, core histones, and other chromosomal proteins. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Divide and conquer: enriching environmental sequencing data.

    Directory of Open Access Journals (Sweden)

    Anne Bergeron

    2007-09-01

    Full Text Available In environmental sequencing projects, a mix of DNA from a whole microbial community is fragmented and sequenced, with one of the possible goals being to reconstruct partial or complete genomes of members of the community. In communities with high diversity of species, a significant proportion of the sequences do not overlap any other fragment in the sample. This problem will arise not only in situations with a relatively even distribution of many species, but also when the community in a particular environment is routinely dominated by the same few species. In the former case, no genomes may be assembled at all, while in the latter case a few dominant species in an environment will always be sequenced at high coverage to the detriment of coverage of the greater number of sparse species.Here we show that, with the same global sequencing effort, separating the species into two or more sub-communities prior to sequencing can yield a much higher proportion of sequences that can be assembled. We first use the Lander-Waterman model to show that, if the expected percentage of singleton sequences is higher than 25%, then, under the uniform distribution hypothesis, splitting the community is always a wise choice. We then construct simulated microbial communities to show that the results hold for highly non-uniform distributions. We also show that, for the distributions considered in the experiments, it is possible to estimate quite accurately the relative diversity of the two sub-communities.Given the fact that several methods exist to split microbial communities based on physical properties such as size, density, surface biochemistry, or optical properties, we strongly suggest that groups involved in environmental sequencing, and expecting high diversity, consider splitting their communities in order to maximize the information content of their sequencing effort.

  2. Sequencing and comparing whole mitochondrial genomes ofanimals

    Energy Technology Data Exchange (ETDEWEB)

    Boore, Jeffrey L.; Macey, J. Robert; Medina, Monica

    2005-04-22

    Comparing complete animal mitochondrial genome sequences is becoming increasingly common for phylogenetic reconstruction and as a model for genome evolution. Not only are they much more informative than shorter sequences of individual genes for inferring evolutionary relatedness, but these data also provide sets of genome-level characters, such as the relative arrangements of genes, that can be especially powerful. We describe here the protocols commonly used for physically isolating mtDNA, for amplifying these by PCR or RCA, for cloning,sequencing, assembly, validation, and gene annotation, and for comparing both sequences and gene arrangements. On several topics, we offer general observations based on our experiences to date with determining and comparing complete mtDNA sequences.

  3. Genomics of Compositae crops: reference transcriptome assemblies and evidence of hybridization with wild relatives.

    Science.gov (United States)

    Hodgins, Kathryn A; Lai, Zhao; Oliveira, Luiz O; Still, David W; Scascitelli, Moira; Barker, Michael S; Kane, Nolan C; Dempewolf, Hannes; Kozik, Alex; Kesseli, Richard V; Burke, John M; Michelmore, Richard W; Rieseberg, Loren H

    2014-01-01

    Although the Compositae harbours only two major food crops, sunflower and lettuce, many other species in this family are utilized by humans and have experienced various levels of domestication. Here, we have used next-generation sequencing technology to develop 15 reference transcriptome assemblies for Compositae crops or their wild relatives. These data allow us to gain insight into the evolutionary and genomic consequences of plant domestication. Specifically, we performed Illumina sequencing of Cichorium endivia, Cichorium intybus, Echinacea angustifolia, Iva annua, Helianthus tuberosus, Dahlia hybrida, Leontodon taraxacoides and Glebionis segetum, as well 454 sequencing of Guizotia scabra, Stevia rebaudiana, Parthenium argentatum and Smallanthus sonchifolius. Illumina reads were assembled using Trinity, and 454 reads were assembled using MIRA and CAP3. We evaluated the coverage of the transcriptomes using BLASTX analysis of a set of ultra-conserved orthologs (UCOs) and recovered most of these genes (88-98%). We found a correlation between contig length and read length for the 454 assemblies, and greater contig lengths for the 454 compared with the Illumina assemblies. This suggests that longer reads can aid in the assembly of more complete transcripts. Finally, we compared the divergence of orthologs at synonymous sites (Ks) between Compositae crops and their wild relatives and found greater divergence when the progenitors were self-incompatible. We also found greater divergence between pairs of taxa that had some evidence of postzygotic isolation. For several more distantly related congeners, such as chicory and endive, we identified a signature of introgression in the distribution of Ks values. © 2013 John Wiley & Sons Ltd.

  4. De novo transcriptome assembly of shrimp Palaemon serratus

    Directory of Open Access Journals (Sweden)

    Alejandra Perina

    2017-03-01

    Full Text Available The shrimp Palaemon serratus is a coastal decapod crustacean with a high commercial value. It is harvested for human consumption. In this study, we used Illumina sequencing technology (HiSeq 2000 to sequence, assemble and annotate the transcriptome of P. serratus. RNA was isolated from muscle of adults individuals and, from a pool of larvae. A total number of 4 cDNA libraries were constructed, using the TruSeq RNA Sample Preparation Kit v2. The raw data in this study was deposited in NCBI SRA database with study accession number of SRP090769. The obtained data were subjected to de novo transcriptome assembly using Trinity software, and coding regions were predicted by TransDecoder. We used Blastp and Sma3s to annotate the identified proteins. The transcriptome data could provide some insight into the understanding of genes involved in the larval development and metamorphosis.

  5. Simultaneous identification of long similar substrings in large sets of sequences

    Directory of Open Access Journals (Sweden)

    Wittig Burghardt

    2007-05-01

    Full Text Available Abstract Background Sequence comparison faces new challenges today, with many complete genomes and large libraries of transcripts known. Gene annotation pipelines match these sequences in order to identify genes and their alternative splice forms. However, the software currently available cannot simultaneously compare sets of sequences as large as necessary especially if errors must be considered. Results We therefore present a new algorithm for the identification of almost perfectly matching substrings in very large sets of sequences. Its implementation, called ClustDB, is considerably faster and can handle 16 times more data than VMATCH, the most memory efficient exact program known today. ClustDB simultaneously generates large sets of exactly matching substrings of a given minimum length as seeds for a novel method of match extension with errors. It generates alignments of maximum length with a considered maximum number of errors within each overlapping window of a given size. Such alignments are not optimal in the usual sense but faster to calculate and often more appropriate than traditional alignments for genomic sequence comparisons, EST and full-length cDNA matching, and genomic sequence assembly. The method is used to check the overlaps and to reveal possible assembly errors for 1377 Medicago truncatula BAC-size sequences published at http://www.medicago.org/genome/assembly_table.php?chr=1. Conclusion The program ClustDB proves that window alignment is an efficient way to find long sequence sections of homogenous alignment quality, as expected in case of random errors, and to detect systematic errors resulting from sequence contaminations. Such inserts are systematically overlooked in long alignments controlled by only tuning penalties for mismatches and gaps. ClustDB is freely available for academic use.

  6. Long-read sequencing data analysis for yeasts.

    Science.gov (United States)

    Yue, Jia-Xing; Liti, Gianni

    2018-06-01

    Long-read sequencing technologies have become increasingly popular due to their strengths in resolving complex genomic regions. As a leading model organism with small genome size and great biotechnological importance, the budding yeast Saccharomyces cerevisiae has many isolates currently being sequenced with long reads. However, analyzing long-read sequencing data to produce high-quality genome assembly and annotation remains challenging. Here, we present a modular computational framework named long-read sequencing data analysis for yeasts (LRSDAY), the first one-stop solution that streamlines this process. Starting from the raw sequencing reads, LRSDAY can produce chromosome-level genome assembly and comprehensive genome annotation in a highly automated manner with minimal manual intervention, which is not possible using any alternative tool available to date. The annotated genomic features include centromeres, protein-coding genes, tRNAs, transposable elements (TEs), and telomere-associated elements. Although tailored for S. cerevisiae, we designed LRSDAY to be highly modular and customizable, making it adaptable to virtually any eukaryotic organism. When applying LRSDAY to an S. cerevisiae strain, it takes ∼41 h to generate a complete and well-annotated genome from ∼100× Pacific Biosciences (PacBio) running the basic workflow with four threads. Basic experience working within the Linux command-line environment is recommended for carrying out the analysis using LRSDAY.

  7. Gigadalton-scale shape-programmable DNA assemblies

    Science.gov (United States)

    Wagenbauer, Klaus F.; Sigl, Christian; Dietz, Hendrik

    2017-12-01

    Natural biomolecular assemblies such as molecular motors, enzymes, viruses and subcellular structures often form by self-limiting hierarchical oligomerization of multiple subunits. Large structures can also assemble efficiently from a few components by combining hierarchical assembly and symmetry, a strategy exemplified by viral capsids. De novo protein design and RNA and DNA nanotechnology aim to mimic these capabilities, but the bottom-up construction of artificial structures with the dimensions and complexity of viruses and other subcellular components remains challenging. Here we show that natural assembly principles can be combined with the methods of DNA origami to produce gigadalton-scale structures with controlled sizes. DNA sequence information is used to encode the shapes of individual DNA origami building blocks, and the geometry and details of the interactions between these building blocks then control their copy numbers, positions and orientations within higher-order assemblies. We illustrate this strategy by creating planar rings of up to 350 nanometres in diameter and with atomic masses of up to 330 megadaltons, micrometre-long, thick tubes commensurate in size to some bacilli, and three-dimensional polyhedral assemblies with sizes of up to 1.2 gigadaltons and 450 nanometres in diameter. We achieve efficient assembly, with yields of up to 90 per cent, by using building blocks with validated structure and sufficient rigidity, and an accurate design with interaction motifs that ensure that hierarchical assembly is self-limiting and able to proceed in equilibrium to allow for error correction. We expect that our method, which enables the self-assembly of structures with sizes approaching that of viruses and cellular organelles, can readily be used to create a range of other complex structures with well defined sizes, by exploiting the modularity and high degree of addressability of the DNA origami building blocks used.

  8. De novo assembly of leaf transcriptome in the medicinal plant Andrographis paniculata

    Directory of Open Access Journals (Sweden)

    Neeraja Cherukupalli

    2016-08-01

    Full Text Available Andrographis paniculata is an important medicinal plant containing various bioactive terpenoids and flavonoids. Despite its importance in herbal medicine, no ready-to-use transcript sequence information of this plant is made available in the public data base, this study mainly deals with the sequencing of RNA from A. paniculata leaf using Illumina HiSeqTM 2000 platform followed by the de novo transcriptome assembly. A total of 189.22 million high quality paired reads were generated and 1,70,724 transcripts were predicted in the primary assembly. Secondary assembly generated a transcriptome size of ~88 Mb with 83,800 clustered transcripts. Based on the similarity searches against plant nonredundant protein database, gene ontology and eukaryotic orthologous groups, 49,363 transcripts were annotated constituting upto 58.91% of the identified unigenes. Annotation of transcripts − using kyoto encyclopedia of genes and genomes database − revealed 5,606 transcripts plausibly involved in 140 pathways including biosynthesis of terpenoids and other secondary metabolites. Transcription factor analysis showed 6,767 unique transcripts belonging to 97 different transcription factor families. A total number of 124 CYP450 transcripts belonging to seven divergent clans have been identified. Transcriptome revealed 146 different transcripts coding for enzymes involved in the biosynthesis of terpenoids of which 35 contained terpene synthase motifs. This study also revealed 32,341 simple sequence repeats (SSRs in 23,168 transcripts. Assembled sequences of transcriptome of A.paniculata generated in this study are made available, for the first time, in the TSA database, which provides useful information for functional and comparative genomic analyses besides identification of key enzymes involved in the various pathways of secondary metabolism.

  9. De novo Assembly of Leaf Transcriptome in the Medicinal Plant Andrographis paniculata

    Science.gov (United States)

    Cherukupalli, Neeraja; Divate, Mayur; Mittapelli, Suresh R.; Khareedu, Venkateswara R.; Vudem, Dashavantha R.

    2016-01-01

    Andrographis paniculata is an important medicinal plant containing various bioactive terpenoids and flavonoids. Despite its importance in herbal medicine, no ready-to-use transcript sequence information of this plant is made available in the public data base, this study mainly deals with the sequencing of RNA from A. paniculata leaf using Illumina HiSeq™ 2000 platform followed by the de novo transcriptome assembly. A total of 189.22 million high quality paired reads were generated and 1,70,724 transcripts were predicted in the primary assembly. Secondary assembly generated a transcriptome size of ~88 Mb with 83,800 clustered transcripts. Based on the similarity searches against plant non-redundant protein database, gene ontology, and eukaryotic orthologous groups, 49,363 transcripts were annotated constituting upto 58.91% of the identified unigenes. Annotation of transcripts—using kyoto encyclopedia of genes and genomes database—revealed 5606 transcripts plausibly involved in 140 pathways including biosynthesis of terpenoids and other secondary metabolites. Transcription factor analysis showed 6767 unique transcripts belonging to 97 different transcription factor families. A total number of 124 CYP450 transcripts belonging to seven divergent clans have been identified. Transcriptome revealed 146 different transcripts coding for enzymes involved in the biosynthesis of terpenoids of which 35 contained terpene synthase motifs. This study also revealed 32,341 simple sequence repeats (SSRs) in 23,168 transcripts. Assembled sequences of transcriptome of A. paniculata generated in this study are made available, for the first time, in the TSA database, which provides useful information for functional and comparative genomic analysis besides identification of key enzymes involved in the various pathways of secondary metabolism. PMID:27582746

  10. A human gut microbial gene catalogue established by metagenomic sequencing

    DEFF Research Database (Denmark)

    dos Santos, Marcelo Bertalan Quintanilha; Sicheritz-Pontén, Thomas; Nielsen, Henrik Bjørn

    2010-01-01

    To understand the impact of gut microbes on human health and well-being it is crucial to assess their genetic potential. Here we describe the Illumina-based metagenomic sequencing, assembly and characterization of 3.3 million non-redundant microbial genes, derived from 576.7 gigabases of sequence...

  11. Functional analysis of the accessory protein TapA in Bacillus subtilis amyloid fiber assembly.

    Science.gov (United States)

    Romero, Diego; Vlamakis, Hera; Losick, Richard; Kolter, Roberto

    2014-04-01

    Bacillus subtilis biofilm formation relies on the assembly of a fibrous scaffold formed by the protein TasA. TasA polymerizes into highly stable fibers with biochemical and morphological features of functional amyloids. Previously, we showed that assembly of TasA fibers requires the auxiliary protein TapA. In this study, we investigated the roles of TapA sequences from the C-terminal and N-terminal ends and TapA cysteine residues in its ability to promote the assembly of TasA amyloid-like fibers. We found that the cysteine residues are not essential for the formation of TasA fibers, as their replacement by alanine residues resulted in only minor defects in biofilm formation. Mutating sequences in the C-terminal half had no effect on biofilm formation. However, we identified a sequence of 8 amino acids in the N terminus that is key for TasA fiber formation. Strains expressing TapA lacking these 8 residues were completely defective in biofilm formation. In addition, this TapA mutant protein exhibited a dominant negative effect on TasA fiber formation. Even in the presence of wild-type TapA, the mutant protein inhibited fiber assembly in vitro and delayed biofilm formation in vivo. We propose that this 8-residue sequence is crucial for the formation of amyloid-like fibers on the cell surface, perhaps by mediating the interaction between TapA or TapA and TasA molecules.

  12. Establishing gene models from the Pinus pinaster genome using gene capture and BAC sequencing.

    Science.gov (United States)

    Seoane-Zonjic, Pedro; Cañas, Rafael A; Bautista, Rocío; Gómez-Maldonado, Josefa; Arrillaga, Isabel; Fernández-Pozo, Noé; Claros, M Gonzalo; Cánovas, Francisco M; Ávila, Concepción

    2016-02-27

    In the era of DNA throughput sequencing, assembling and understanding gymnosperm mega-genomes remains a challenge. Although drafts of three conifer genomes have recently been published, this number is too low to understand the full complexity of conifer genomes. Using techniques focused on specific genes, gene models can be established that can aid in the assembly of gene-rich regions, and this information can be used to compare genomes and understand functional evolution. In this study, gene capture technology combined with BAC isolation and sequencing was used as an experimental approach to establish de novo gene structures without a reference genome. Probes were designed for 866 maritime pine transcripts to sequence genes captured from genomic DNA. The gene models were constructed using GeneAssembler, a new bioinformatic pipeline, which reconstructed over 82% of the gene structures, and a high proportion (85%) of the captured gene models contained sequences from the promoter regulatory region. In a parallel experiment, the P. pinaster BAC library was screened to isolate clones containing genes whose cDNA sequence were already available. BAC clones containing the asparagine synthetase, sucrose synthase and xyloglucan endotransglycosylase gene sequences were isolated and used in this study. The gene models derived from the gene capture approach were compared with the genomic sequences derived from the BAC clones. This combined approach is a particularly efficient way to capture the genomic structures of gene families with a small number of members. The experimental approach used in this study is a valuable combined technique to study genomic gene structures in species for which a reference genome is unavailable. It can be used to establish exon/intron boundaries in unknown gene structures, to reconstruct incomplete genes and to obtain promoter sequences that can be used for transcriptional studies. A bioinformatics algorithm (GeneAssembler) is also provided as a

  13. Liaison concatenation – A method to obtain feasible assembly ...

    Indian Academy of Sciences (India)

    non-possible assembly sequences using liaison graph. In this paper a new ... Applications (CATIA) software is a user friendly CAD tool with the feasibility of .... to perform the contact analysis to check the possibility of interference. Table 5.

  14. Integration of Systems with Varying Levels of Autonomy (Integration de Systemes a Niveau d’Autonomie Variable)

    Science.gov (United States)

    2008-09-01

    2.2.2.1 Theseus AUV 2-15 2.2.3 Air and Space 2-18 2.2.3.1 F-22A Flying Qualities Development 2-18 2.2.3.2 Apollo and Space Shuttle 2-19 2.2.3.3...Figure 2.14 The Multi-Mission Spartan Vehicle and the Remote Minehunting Vehicle Dorado 2-15 Figure 2.15 Theseus Schematic 2-16 Figure 2.16 The...Disciplines 6-3 RTO-TR-SCI-144 ix List of Tables Table Page Table 2.1 Thrust and Pitch Attitude Before the Accident 2-6 Table 2.2 Theseus

  15. Yeast prions assembly and propagation: contributions of the prion and non-prion moieties and the nature of assemblies.

    Science.gov (United States)

    Kabani, Mehdi; Melki, Ronald

    2011-01-01

    Yeast prions are self-perpetuating protein aggregates that are at the origin of heritable and transmissible non-Mendelian phenotypic traits. Among these, [PSI+], [URE3] and [PIN+] are the most well documented prions and arise from the assembly of Sup35p, Ure2p and Rnq1p, respectively, into insoluble fibrillar assemblies. Fibril assembly depends on the presence of N- or C-terminal prion domains (PrDs) which are not homologous in sequence but share unusual amino-acid compositions, such as enrichment in polar residues (glutamines and asparagines) or the presence of oligopeptide repeats. Purified PrDs form amyloid fibrils that can convert prion-free cells to the prion state upon transformation. Nonetheless, isolated PrDs and full-length prion proteins have different aggregation, structural and infectious properties. In addition, mutations in the "non-prion" domains (non-PrDs) of Sup35p, Ure2p and Rnq1p were shown to affect their prion properties in vitro and in vivo. Despite these evidences, the implication of the functional non-PrDs in fibril assembly and prion propagation has been mostly overlooked. In this review, we discuss the contribution of non-PrDs to prion assemblies, and the structure-function relationship in prion infectivity in the light of recent findings on Sup35p and Ure2p assembly into infectious fibrils from our laboratory and others.

  16. Dataset of the first transcriptome assembly of the tree crop “yerba mate” (Ilex paraguariensis and systematic characterization of protein coding genes

    Directory of Open Access Journals (Sweden)

    Patricia M. Aguilera

    2018-04-01

    Full Text Available This contribution contains data associated to the research article entitled “Exploring the genes of yerba mate (Ilex paraguariensis A. St.-Hil. by NGS and de novo transcriptome assembly” (Debat et al., 2014 [1]. By means of a bioinformatic approach involving extensive NGS data analyses, we provide a resource encompassing the full transcriptome assembly of yerba mate, the first available reference for the Ilex L. genus. This dataset (Supplementary files 1 and 2 consolidates the transcriptome-wide assembled sequences of I. paraguariensis with further comprehensive annotation of the protein coding genes of yerba mate via the integration of Arabidopsis thaliana databases. The generated data is pivotal for the characterization of agronomical relevant genes in the tree crop yerba mate -a non-model species- and related taxa in Ilex. The raw sequencing data dissected here is available at DDBJ/ENA/GenBank (NCBI Resource Coordinators, 2016 [2] Sequence Read Archive (SRA under the accession SRP043293 and the assembled sequences have been deposited at the Transcriptome Shotgun Assembly Sequence Database (TSA under the accession GFHV00000000.

  17. De novo transcriptome assembly for a non-model species, the blood-sucking bug Triatoma brasiliensis, a vector of Chagas disease.

    Science.gov (United States)

    Marchant, A; Mougel, F; Almeida, C; Jacquin-Joly, E; Costa, J; Harry, M

    2015-04-01

    High throughput sequencing (HTS) provides new research opportunities for work on non-model organisms, such as differential expression studies between populations exposed to different environmental conditions. However, such transcriptomic studies first require the production of a reference assembly. The choice of sampling procedure, sequencing strategy and assembly workflow is crucial. To develop a reliable reference transcriptome for Triatoma brasiliensis, the major Chagas disease vector in Northeastern Brazil, different de novo assembly protocols were generated using various datasets and software. Both 454 and Illumina sequencing technologies were applied on RNA extracted from antennae and mouthparts from single or pooled individuals. The 454 library yielded 278 Mb. Fifteen Illumina libraries were constructed and yielded nearly 360 million RNA-seq single reads and 46 million RNA-seq paired-end reads for nearly 45 Gb. For the 454 reads, we used three assemblers, Newbler, CAP3 and/or MIRA and for the Illumina reads, the Trinity assembler. Ten assembly workflows were compared using these programs separately or in combination. To compare the assemblies obtained, quantitative and qualitative criteria were used, including contig length, N50, contig number and the percentage of chimeric contigs. Completeness of the assemblies was estimated using the CEGMA pipeline. The best assembly (57,657 contigs, completeness of 80 %, <1 % chimeric contigs) was a hybrid assembly leading to recommend the use of (1) a single individual with large representation of biological tissues, (2) merging both long reads and short paired-end Illumina reads, (3) several assemblers in order to combine the specific advantages of each.

  18. VAGUE: a graphical user interface for the Velvet assembler.

    Science.gov (United States)

    Powell, David R; Seemann, Torsten

    2013-01-15

    Velvet is a popular open-source de novo genome assembly software tool, which is run from the Unix command line. Most of the problems experienced by new users of Velvet revolve around constructing syntactically and semantically correct command lines, getting input files into acceptable formats and assessing the output. Here, we present Velvet Assembler Graphical User Environment (VAGUE), a multi-platform graphical front-end for Velvet. VAGUE aims to make sequence assembly accessible to a wider audience and to facilitate better usage amongst existing users of Velvet. VAGUE is implemented in JRuby and targets the Java Virtual Machine. It is available under an open-source GPLv2 licence from http://www.vicbioinformatics.com/. torsten.seemann@monash.edu.

  19. A consolidation process for spent burnable poison rod assemblies

    International Nuclear Information System (INIS)

    Yamamoto, Y.; Harada, M.; Komatsu, Y.

    1985-01-01

    A new consolidation system for the spent burnable poison assembly utilizing a sequence control robot operated under water was proposed. A credible accident in the system was analyzed mainly from the viewpoint of tritium release, based on the diffusion analysis of tritium in borosilicate glass. It was found that the amount of tritium released would be small even after the rupture of burnable poison rods. An experiment on a new consolidation system was performed using spent burnable poison assemblies. The volume of burnable poison assemblies was reduced safely and securely by a factor of 7 to 14 for burnable poison rods and by 22 for hold-down portions. It was proved that the consolidation system is collectively feasible

  20. Snake Genome Sequencing: Results and Future Prospects.

    Science.gov (United States)

    Kerkkamp, Harald M I; Kini, R Manjunatha; Pospelov, Alexey S; Vonk, Freek J; Henkel, Christiaan V; Richardson, Michael K

    2016-12-01

    Snake genome sequencing is in its infancy-very much behind the progress made in sequencing the genomes of humans, model organisms and pathogens relevant to biomedical research, and agricultural species. We provide here an overview of some of the snake genome projects in progress, and discuss the biological findings, with special emphasis on toxinology, from the small number of draft snake genomes already published. We discuss the future of snake genomics, pointing out that new sequencing technologies will help overcome the problem of repetitive sequences in assembling snake genomes. Genome sequences are also likely to be valuable in examining the clustering of toxin genes on the chromosomes, in designing recombinant antivenoms and in studying the epigenetic regulation of toxin gene expression.

  1. Snake Genome Sequencing: Results and Future Prospects

    Directory of Open Access Journals (Sweden)

    Harald M. I. Kerkkamp

    2016-12-01

    Full Text Available Snake genome sequencing is in its infancy—very much behind the progress made in sequencing the genomes of humans, model organisms and pathogens relevant to biomedical research, and agricultural species. We provide here an overview of some of the snake genome projects in progress, and discuss the biological findings, with special emphasis on toxinology, from the small number of draft snake genomes already published. We discuss the future of snake genomics, pointing out that new sequencing technologies will help overcome the problem of repetitive sequences in assembling snake genomes. Genome sequences are also likely to be valuable in examining the clustering of toxin genes on the chromosomes, in designing recombinant antivenoms and in studying the epigenetic regulation of toxin gene expression.

  2. Sequencing, mapping, and analysis of 27,455 maize full-length cDNAs.

    Directory of Open Access Journals (Sweden)

    Carol Soderlund

    2009-11-01

    Full Text Available Full-length cDNA (FLcDNA sequencing establishes the precise primary structure of individual gene transcripts. From two libraries representing 27 B73 tissues and abiotic stress treatments, 27,455 high-quality FLcDNAs were sequenced. The average transcript length was 1.44 kb including 218 bases and 321 bases of 5' and 3' UTR, respectively, with 8.6% of the FLcDNAs encoding predicted proteins of fewer than 100 amino acids. Approximately 94% of the FLcDNAs were stringently mapped to the maize genome. Although nearly two-thirds of this genome is composed of transposable elements (TEs, only 5.6% of the FLcDNAs contained TE sequences in coding or UTR regions. Approximately 7.2% of the FLcDNAs are putative transcription factors, suggesting that rare transcripts are well-enriched in our FLcDNA set. Protein similarity searching identified 1,737 maize transcripts not present in rice, sorghum, Arabidopsis, or poplar annotated genes. A strict FLcDNA assembly generated 24,467 non-redundant sequences, of which 88% have non-maize protein matches. The FLcDNAs were also assembled with 41,759 FLcDNAs in GenBank from other projects, where semi-strict parameters were used to identify 13,368 potentially unique non-redundant sequences from this project. The libraries, ESTs, and FLcDNA sequences produced from this project are publicly available. The annotated EST and FLcDNA assemblies are available through the maize FLcDNA web resource (www.maizecdna.org.

  3. Self-assembling peptide amphiphiles and related methods for growth factor delivery

    Science.gov (United States)

    Stupp, Samuel I [Chicago, IL; Donners, Jack J. J. M.; Silva, Gabriel A [Chicago, IL; Behanna, Heather A [Chicago, IL; Anthony, Shawn G [New Stanton, PA

    2009-06-09

    Amphiphilic peptide compounds comprising one or more epitope sequences for binding interaction with one or more corresponding growth factors, micellar assemblies of such compounds and related methods of use.

  4. Simulation of Assembly Tolerance and Characteristics of High Pressure Common Rail Injector

    Directory of Open Access Journals (Sweden)

    Jiping Lu

    2011-12-01

    Full Text Available Fuel injector is the key part of a high-pressure common rail fuel injection system. Its manufacturing precision and assembly quality affect system's property and performance. According to the characteristics and demands of assembly of the fuel injector, an intelligent optimization algorithm is proposed to resolve the problem of assembly sequence planning. Based on geometric modeling, assembly dimension chain of the injector control chamber is established, and the relationship between assembly tolerance and volume change of control chamber is analyzed. The optimization model of the assembly is established. The impact of assembly tolerance on injector's performance is simulated according to the optimization algorithm. The simulation result shows that quantity of injection fuel changes correspondingly with the change of assembly tolerance, while injection rate and pressure do not change significantly, and the response rate of needle considerably slow. Similarly, the leakage rate of fuel in control chamber is calculated, indicating that the assembly tolerance has obvious impact on fuel leakage and its rate. The study illuminates that injector's assembly tolerance has prominent effect on injection.

  5. Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea

    Energy Technology Data Exchange (ETDEWEB)

    Bowers, Robert M.; Kyrpides, Nikos C.; Stepanauskas, Ramunas; Harmon-Smith, Miranda; Doud, Devin; Reddy, T. B. K.; Schulz, Frederik; Jarett, Jessica; Rivers, Adam R.; Eloe-Fadrosh, Emiley A.; Tringe, Susannah G.; Ivanova, Natalia N.; Copeland, Alex; Clum, Alicia; Becraft, Eric D.; Malmstrom, Rex R.; Birren, Bruce; Podar, Mircea; Bork, Peer; Weinstock, George M.; Garrity, George M.; Dodsworth, Jeremy A.; Yooseph, Shibu; Sutton, Granger; Glöckner, Frank O.; Gilbert, Jack A.; Nelson, William C.; Hallam, Steven J.; Jungbluth, Sean P.; Ettema, Thijs J. G.; Tighe, Scott; Konstantinidis, Konstantinos T.; Liu, Wen-Tso; Baker, Brett J.; Rattei, Thomas; Eisen, Jonathan A.; Hedlund, Brian; McMahon, Katherine D.; Fierer, Noah; Knight, Rob; Finn, Rob; Cochrane, Guy; Karsch-Mizrachi, Ilene; Tyson, Gene W.; Rinke, Christian; Kyrpides, Nikos C.; Schriml, Lynn; Garrity, George M.; Hugenholtz, Philip; Sutton, Granger; Yilmaz, Pelin; Meyer, Folker; Glöckner, Frank O.; Gilbert, Jack A.; Knight, Rob; Finn, Rob; Cochrane, Guy; Karsch-Mizrachi, Ilene; Lapidus, Alla; Meyer, Folker; Yilmaz, Pelin; Parks, Donovan H.; Eren, A. M.; Schriml, Lynn; Banfield, Jillian F.; Hugenholtz, Philip; Woyke, Tanja

    2017-08-08

    We present two standards developed by the Genomic Standards Consortium (GSC) for reporting bacterial and archaeal genome sequences. Both are extensions of the Minimum Information about Any (x) Sequence (MIxS). The standards are the Minimum Information about a Single Amplified Genome (MISAG) and the Minimum Information about a Metagenome-Assembled Genome (MIMAG), including, but not limited to, assembly quality, and estimates of genome completeness and contamination. These standards can be used in combination with other GSC checklists, including the Minimum Information about a Genome Sequence (MIGS), Minimum Information about a Metagenomic Sequence (MIMS), and Minimum Information about a Marker Gene Sequence (MIMARKS). Community-wide adoption of MISAG and MIMAG will facilitate more robust comparative genomic analyses of bacterial and archaeal diversity.

  6. MIDAS: A Modular DNA Assembly System for Synthetic Biology.

    Science.gov (United States)

    van Dolleweerd, Craig J; Kessans, Sarah A; Van de Bittner, Kyle C; Bustamante, Leyla Y; Bundela, Rudranuj; Scott, Barry; Nicholson, Matthew J; Parker, Emily J

    2018-04-20

    A modular and hierarchical DNA assembly platform for synthetic biology based on Golden Gate (Type IIS restriction enzyme) cloning is described. This enabling technology, termed MIDAS (for Modular Idempotent DNA Assembly System), can be used to precisely assemble multiple DNA fragments in a single reaction using a standardized assembly design. It can be used to build genes from libraries of sequence-verified, reusable parts and to assemble multiple genes in a single vector, with full user control over gene order and orientation, as well as control of the direction of growth (polarity) of the multigene assembly, a feature that allows genes to be nested between other genes or genetic elements. We describe the detailed design and use of MIDAS, exemplified by the reconstruction, in the filamentous fungus Penicillium paxilli, of the metabolic pathway for production of paspaline and paxilline, key intermediates in the biosynthesis of a range of indole diterpenes-a class of secondary metabolites produced by several species of filamentous fungi. MIDAS was used to efficiently assemble a 25.2 kb plasmid from 21 different modules (seven genes, each composed of three basic parts). By using a parts library-based system for construction of complex assemblies, and a unique set of vectors, MIDAS can provide a flexible route to assembling tailored combinations of genes and other genetic elements, thereby supporting synthetic biology applications in a wide range of expression hosts.

  7. The LINKS motif zippers trans-acyltransferase polyketide synthase assembly lines into a biosynthetic megacomplex.

    Science.gov (United States)

    Gay, Darren C; Wagner, Drew T; Meinke, Jessica L; Zogzas, Charles E; Gay, Glen R; Keatinge-Clay, Adrian T

    2016-03-01

    Polyketides such as the clinically-valuable antibacterial agent mupirocin are constructed by architecturally-sophisticated assembly lines known as trans-acyltransferase polyketide synthases. Organelle-sized megacomplexes composed of several copies of trans-acyltransferase polyketide synthase assembly lines have been observed by others through transmission electron microscopy to be located at the Bacillus subtilis plasma membrane, where the synthesis and export of the antibacterial polyketide bacillaene takes place. In this work we analyze ten crystal structures of trans-acyltransferase polyketide synthases ketosynthase domains, seven of which are reported here for the first time, to characterize a motif capable of zippering assembly lines into a megacomplex. While each of the three-helix LINKS (Laterally-INteracting Ketosynthase Sequence) motifs is observed to similarly dock with a spatially-reversed copy of itself through hydrophobic and ionic interactions, the amino acid sequences of this motif are not conserved. Such a code is appropriate for mediating homotypic contacts between assembly lines to ensure the ordered self-assembly of a noncovalent, yet tightly-knit, enzymatic network. LINKS-mediated lateral interactions would also have the effect of bolstering the vertical association of the polypeptides that comprise a polyketide synthase assembly line. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. An Efficient Genome Fragment Assembling Using GA with Neighborhood Aware Fitness Function

    Directory of Open Access Journals (Sweden)

    Satoko Kikuchi

    2012-01-01

    Full Text Available To decode a long genome sequence, shotgun sequencing is the state-of-the-art technique. It needs to properly sequence a very large number, sometimes as large as millions, of short partially readable strings (fragments. Arranging those fragments in correct sequence is known as fragment assembling, which is an NP-problem. Presently used methods require enormous computational cost. In this work, we have shown how our modified genetic algorithm (GA could solve this problem efficiently. In the proposed GA, the length of the chromosome, which represents the volume of the search space, is reduced with advancing generations, and thereby improves search efficiency. We also introduced a greedy mutation, by swapping nearby fragments using some heuristics, to improve the fitness of chromosomes. We compared results with Parsons’ algorithm which is based on GA too. We used fragments with partial reads on both sides, mimicking fragments in real genome assembling process. In Parsons’ work base-pair array of the whole fragment is known. Even then, we could obtain much better results, and we succeeded in restructuring contigs covering 100% of the genome sequences.

  9. Comparative analyses of two Geraniaceae transcriptomes using next-generation sequencing.

    Science.gov (United States)

    Zhang, Jin; Ruhlman, Tracey A; Mower, Jeffrey P; Jansen, Robert K

    2013-12-29

    Organelle genomes of Geraniaceae exhibit several unusual evolutionary phenomena compared to other angiosperm families including accelerated nucleotide substitution rates, widespread gene loss, reduced RNA editing, and extensive genomic rearrangements. Since most organelle-encoded proteins function in multi-subunit complexes that also contain nuclear-encoded proteins, it is likely that the atypical organellar phenomena affect the evolution of nuclear genes encoding organellar proteins. To begin to unravel the complex co-evolutionary interplay between organellar and nuclear genomes in this family, we sequenced nuclear transcriptomes of two species, Geranium maderense and Pelargonium x hortorum. Normalized cDNA libraries of G. maderense and P. x hortorum were used for transcriptome sequencing. Five assemblers (MIRA, Newbler, SOAPdenovo, SOAPdenovo-trans [SOAPtrans], Trinity) and two next-generation technologies (454 and Illumina) were compared to determine the optimal transcriptome sequencing approach. Trinity provided the highest quality assembly of Illumina data with the deepest transcriptome coverage. An analysis to determine the amount of sequencing needed for de novo assembly revealed diminishing returns of coverage and quality with data sets larger than sixty million Illumina paired end reads for both species. The G. maderense and P. x hortorum transcriptomes contained fewer transcripts encoding the PLS subclass of PPR proteins relative to other angiosperms, consistent with reduced mitochondrial RNA editing activity in Geraniaceae. In addition, transcripts for all six plastid targeted sigma factors were identified in both transcriptomes, suggesting that one of the highly divergent rpoA-like ORFs in the P. x hortorum plastid genome is functional. The findings support the use of the Illumina platform and assemblers optimized for transcriptome assembly, such as Trinity or SOAPtrans, to generate high-quality de novo transcriptomes with broad coverage. In addition

  10. Direct comparisons of Illumina vs. Roche 454 sequencing technologies on the same microbial community DNA sample.

    Science.gov (United States)

    Luo, Chengwei; Tsementzi, Despina; Kyrpides, Nikos; Read, Timothy; Konstantinidis, Konstantinos T

    2012-01-01

    Next-generation sequencing (NGS) is commonly used in metagenomic studies of complex microbial communities but whether or not different NGS platforms recover the same diversity from a sample and their assembled sequences are of comparable quality remain unclear. We compared the two most frequently used platforms, the Roche 454 FLX Titanium and the Illumina Genome Analyzer (GA) II, on the same DNA sample obtained from a complex freshwater planktonic community. Despite the substantial differences in read length and sequencing protocols, the platforms provided a comparable view of the community sampled. For instance, derived assemblies overlapped in ~90% of their total sequences and in situ abundances of genes and genotypes (estimated based on sequence coverage) correlated highly between the two platforms (R(2)>0.9). Evaluation of base-call error, frameshift frequency, and contig length suggested that Illumina offered equivalent, if not better, assemblies than Roche 454. The results from metagenomic samples were further validated against DNA samples of eighteen isolate genomes, which showed a range of genome sizes and G+C% content. We also provide quantitative estimates of the errors in gene and contig sequences assembled from datasets characterized by different levels of complexity and G+C% content. For instance, we noted that homopolymer-associated, single-base errors affected ~1% of the protein sequences recovered in Illumina contigs of 10× coverage and 50% G+C; this frequency increased to ~3% when non-homopolymer errors were also considered. Collectively, our results should serve as a useful practical guide for choosing proper sampling strategies and data possessing protocols for future metagenomic studies.

  11. Combining independent de novo assemblies optimizes the coding transcriptome for nonconventional model eukaryotic organisms.

    Science.gov (United States)

    Cerveau, Nicolas; Jackson, Daniel J

    2016-12-09

    Next-generation sequencing (NGS) technologies are arguably the most revolutionary technical development to join the list of tools available to molecular biologists since PCR. For researchers working with nonconventional model organisms one major problem with the currently dominant NGS platform (Illumina) stems from the obligatory fragmentation of nucleic acid material that occurs prior to sequencing during library preparation. This step creates a significant bioinformatic challenge for accurate de novo assembly of novel transcriptome data. This challenge becomes apparent when a variety of modern assembly tools (of which there is no shortage) are applied to the same raw NGS dataset. With the same assembly parameters these tools can generate markedly different assembly outputs. In this study we present an approach that generates an optimized consensus de novo assembly of eukaryotic coding transcriptomes. This approach does not represent a new assembler, rather it combines the outputs of a variety of established assembly packages, and removes redundancy via a series of clustering steps. We test and validate our approach using Illumina datasets from six phylogenetically diverse eukaryotes (three metazoans, two plants and a yeast) and two simulated datasets derived from metazoan reference genome annotations. All of these datasets were assembled using three currently popular assembly packages (CLC, Trinity and IDBA-tran). In addition, we experimentally demonstrate that transcripts unique to one particular assembly package are likely to be bioinformatic artefacts. For all eight datasets our pipeline generates more concise transcriptomes that in fact possess more unique annotatable protein domains than any of the three individual assemblers we employed. Another measure of assembly completeness (using the purpose built BUSCO databases) also confirmed that our approach yields more information. Our approach yields coding transcriptome assemblies that are more likely to be

  12. RNA-seq analysis and de novo transcriptome assembly of Jerusalem artichoke (Helianthus tuberosus Linne).

    Science.gov (United States)

    Jung, Won Yong; Lee, Sang Sook; Kim, Chul Wook; Kim, Hyun-Soon; Min, Sung Ran; Moon, Jae Sun; Kwon, Suk-Yoon; Jeon, Jae-Heung; Cho, Hye Sun

    2014-01-01

    Jerusalem artichoke (Helianthus tuberosus L.) has long been cultivated as a vegetable and as a source of fructans (inulin) for pharmaceutical applications in diabetes and obesity prevention. However, transcriptomic and genomic data for Jerusalem artichoke remain scarce. In this study, Illumina RNA sequencing (RNA-Seq) was performed on samples from Jerusalem artichoke leaves, roots, stems and two different tuber tissues (early and late tuber development). Data were used for de novo assembly and characterization of the transcriptome. In total 206,215,632 paired-end reads were generated. These were assembled into 66,322 loci with 272,548 transcripts. Loci were annotated by querying against the NCBI non-redundant, Phytozome and UniProt databases, and 40,215 loci were homologous to existing database sequences. Gene Ontology terms were assigned to 19,848 loci, 15,434 loci were matched to 25 Clusters of Eukaryotic Orthologous Groups classifications, and 11,844 loci were classified into 142 Kyoto Encyclopedia of Genes and Genomes pathways. The assembled loci also contained 10,778 potential simple sequence repeats. The newly assembled transcriptome was used to identify loci with tissue-specific differential expression patterns. In total, 670 loci exhibited tissue-specific expression, and a subset of these were confirmed using RT-PCR and qRT-PCR. Gene expression related to inulin biosynthesis in tuber tissue was also investigated. Exsiting genetic and genomic data for H. tuberosus are scarce. The sequence resources developed in this study will enable the analysis of thousands of transcripts and will thus accelerate marker-assisted breeding studies and studies of inulin biosynthesis in Jerusalem artichoke.

  13. The Amaranth Genome: Genome, Transcriptome, and Physical Map Assembly

    Directory of Open Access Journals (Sweden)

    J. W. Clouse

    2016-03-01

    Full Text Available Amaranth ( L. is an emerging pseudocereal native to the New World that has garnered increased attention in recent years because of its nutritional quality, in particular its seed protein and more specifically its high levels of the essential amino acid lysine. It belongs to the Amaranthaceae family, is an ancient paleopolyploid that shows disomic inheritance (2 = 32, and has an estimated genome size of 466 Mb. Here we present a high-quality draft genome sequence of the grain amaranth. The genome assembly consisted of 377 Mb in 3518 scaffolds with an N of 371 kb. Repetitive element analysis predicted that 48% of the genome is comprised of repeat sequences, of which -like elements were the most commonly classified retrotransposon. A de novo transcriptome consisting of 66,370 contigs was assembled from eight different amaranth tissue and abiotic stress libraries. Annotation of the genome identified 23,059 protein-coding genes. Seven grain amaranths (, , and and their putative progenitor ( were resequenced. A single nucleotide polymorphism (SNP phylogeny supported the classification of as the progenitor species of the grain amaranths. Lastly, we generated a de novo physical map for using the BioNano Genomics’ Genome Mapping platform. The physical map spanned 340 Mb and a hybrid assembly using the BioNano physical maps nearly doubled the N of the assembly to 697 kb. Moreover, we analyzed synteny between amaranth and sugar beet ( L. and estimated, using analysis, the age of the most recent polyploidization event in amaranth.

  14. Assembly process of the ITER neutral beam injectors

    Energy Technology Data Exchange (ETDEWEB)

    Graceffa, J., E-mail: joseph.graceffa@iter.org [ITER Organization, Route de Vinon sur Verdon, 13115 Saint Paul lez Durance (France); Boilson, D.; Hemsworth, R.; Petrov, V.; Schunke, B.; Urbani, M. [ITER Organization, Route de Vinon sur Verdon, 13115 Saint Paul lez Durance (France); Pilard, V. [Fusion for Energy, C/ Josep Pla, n°2, Torres Diagonal Litoral, Edificio B3, 08019 Barcelona (Spain)

    2013-10-15

    The ITER neutral beam (NB) injectors are used for heating and diagnostics operations. There are 4 injectors in total, 3 heating neutral beam injectors (HNBs) and one diagnostic neutral beam injector (DNB). Two HNBs and the DNB will start injection into ITER during the hydrogen/helium phase of ITER operations. A third HNB is considered as an upgrade to the ITER heating systems, and the impact of the later installation and use of that injector have to be taken into account when considering the installation and assembly of the whole NB system. It is assumed that if a third HNB is to be installed, it will be installed before the nuclear phase of the ITER project. The total weight of one injector is around 1200 t and it is composed of 18 main components and 36 sets of shielding plates. The overall dimensions are length 20 m, height 10 m and width 5 m. Assembly of the first two HNBs and the DNB will start before the first plasma is produced in ITER, but as the time required to assemble one injector is estimated at around 1.5 year, the assembly will be divided into 2 steps, one prior to first plasma, and the second during the machine second assembly phase. To comply with this challenging schedule the assembly sequence has been defined to allow assembly of three first injectors in parallel. Due to the similar design between the DNB and HNBs it has been decided to use the same tools, which will be designed to accommodate the differences between the two sets of components. This reduces the global cost of the assembly and the overall assembly time for the injector system. The alignment and positioning of the injectors is a major consideration for the injector assembly as the alignment of the beamline components and the beam source are critical if good injector performance is to be achieved. The theoretical axes of the beams are defined relative to the duct liners which are installed in the NB ports. The concept adopted to achieve the required alignment accuracy is to use the

  15. De novo transcriptome assembly of Setatria italica variety Taejin

    Directory of Open Access Journals (Sweden)

    Yeonhwa Jo

    2016-06-01

    Full Text Available Foxtail millet (Setaria italica belonging to the family Poaceae is an important millet that is widely cultivated in East Asia. Of the cultivated millets, the foxtail millet has the longest history and is one of the main food crops in South India and China. Moreover, foxtail millet is a model plant system for biofuel generation utilizing the C4 photosynthetic pathway. In this study, we carried out de novo transcriptome assembly for the foxtail millet variety Taejin collected from Korea using next-generation sequencing. We obtained a total of 8.676 GB raw data by paired-end sequencing. The raw data in this study can be available in NCBI SRA database with accession number of SRR3406552. The Trinity program was used to de novo assemble 145,332 transcripts. Using the TransDecoder program, we predicted 82,925 putative proteins. BLASTP was performed against the Swiss-Prot protein sequence database to annotate the functions of identified proteins, resulting in 20,555 potentially novel proteins. Taken together, this study provides transcriptome data for the foxtail millet variety Taejin by RNA-Seq.

  16. Fuel assembly duct cutting in the FFTF/IEM Cell

    International Nuclear Information System (INIS)

    Gibbons, P.W.

    1985-01-01

    Two mill type slitting cutters are used in the Fast Flux Test Facility (FFTF) Interim Examination and Maintenance (IEM) Cell during the disassembly sequence of a Driver Fuel Assembly. This disassembly is necessary so that selected parts may be examined both in the IEM Cell and elsewhere. The cutters have been in use for two years. During this time eight Driver Fuel assemblies have been taken apart in the IEM Cell. The cutters' operating philosophy and characteristics, as well as lessons learned from a significant equipment failure are presented. 1 ref., 6 figs., 1 tab

  17. An Analysis of Application Generators.

    Science.gov (United States)

    1983-03-01

    query language OUEL in the programming language C, THESEUS [20], which embeds relational operators in the language Euclid. Schmidt [21] reports some...34The Design and Implementation of INGRES," ACM-TODS, Vol. 1. No. 3, 1976,. 33 £ 20. Shopiro,J.E., " THESEUS -A Programming Language for Relational

  18. Assembly, Annotation, and Analysis of Multiple Mycorrhizal Fungal Genomes

    Energy Technology Data Exchange (ETDEWEB)

    Initiative Consortium, Mycorrhizal Genomics; Kuo, Alan; Grigoriev, Igor; Kohler, Annegret; Martin, Francis

    2013-03-08

    Mycorrhizal fungi play critical roles in host plant health, soil community structure and chemistry, and carbon and nutrient cycling, all areas of intense interest to the US Dept. of Energy (DOE) Joint Genome Institute (JGI). To this end we are building on our earlier sequencing of the Laccaria bicolor genome by partnering with INRA-Nancy and the mycorrhizal research community in the MGI to sequence and analyze dozens of mycorrhizal genomes of all Basidiomycota and Ascomycota orders and multiple ecological types (ericoid, orchid, and ectomycorrhizal). JGI has developed and deployed high-throughput sequencing techniques, and Assembly, RNASeq, and Annotation Pipelines. In 2012 alone we sequenced, assembled, and annotated 12 draft or improved genomes of mycorrhizae, and predicted ~;;232831 genes and ~;;15011 multigene families, All of this data is publicly available on JGI MycoCosm (http://jgi.doe.gov/fungi/), which provides access to both the genome data and tools with which to analyze the data. Preliminary comparisons of the current total of 14 public mycorrhizal genomes suggest that 1) short secreted proteins potentially involved in symbiosis are more enriched in some orders than in others amongst the mycorrhizal Agaricomycetes, 2) there are wide ranges of numbers of genes involved in certain functional categories, such as signal transduction and post-translational modification, and 3) novel gene families are specific to some ecological types.

  19. Diverse Supramolecular Nanofiber Networks Assembled by Functional Low-Complexity Domains.

    Science.gov (United States)

    An, Bolin; Wang, Xinyu; Cui, Mengkui; Gui, Xinrui; Mao, Xiuhai; Liu, Yan; Li, Ke; Chu, Cenfeng; Pu, Jiahua; Ren, Susu; Wang, Yanyi; Zhong, Guisheng; Lu, Timothy K; Liu, Cong; Zhong, Chao

    2017-07-25

    Self-assembling supramolecular nanofibers, common in the natural world, are of fundamental interest and technical importance to both nanotechnology and materials science. Despite important advances, synthetic nanofibers still lack the structural and functional diversity of biological molecules, and the controlled assembly of one type of molecule into a variety of fibrous structures with wide-ranging functional attributes remains challenging. Here, we harness the low-complexity (LC) sequence domain of fused in sarcoma (FUS) protein, an essential cellular nuclear protein with slow kinetics of amyloid fiber assembly, to construct random copolymer-like, multiblock, and self-sorted supramolecular fibrous networks with distinct structural features and fluorescent functionalities. We demonstrate the utilities of these networks in the templated, spatially controlled assembly of ligand-decorated gold nanoparticles, quantum dots, nanorods, DNA origami, and hybrid structures. Owing to the distinguishable nanoarchitectures of these nanofibers, this assembly is structure-dependent. By coupling a modular genetic strategy with kinetically controlled complex supramolecular self-assembly, we demonstrate that a single type of protein molecule can be used to engineer diverse one-dimensional supramolecular nanostructures with distinct functionalities.

  20. DNA-mediated self-assembly of tetrahedral plasmonic clusters for metafluids

    Science.gov (United States)

    Schade, Nicholas; Sun, Li; Lee, You-Jin; Fan, Jonathan; Capasso, Federico; Yi, Gi-Ra; Manoharan, Vinothan

    2014-03-01

    We direct the self-assembly of clusters of gold nanospheres with the goal of creating a bulk, isotropic, optical metafluid. We use spherical gold nanoparticles that are exceptionally smooth, monocrystalline, and monodisperse. These particles exhibit highly reproducible scattering spectra compared with commercially available gold colloids. We label them with DNA sequences and mix them together to self-assemble small clusters. By controlling the particle sizes and the interactions between them, we maximize the yield of tetrahedral clusters, the ideal structures for isotropic metamaterials.

  1. The genome sequence of the outbreeding globe artichoke constructed de novo incorporating a phase-aware low-pass sequencing strategy of F1 progeny

    Science.gov (United States)

    Scaglione, Davide; Reyes-Chin-Wo, Sebastian; Acquadro, Alberto; Froenicke, Lutz; Portis, Ezio; Beitel, Christopher; Tirone, Matteo; Mauro, Rosario; Lo Monaco, Antonino; Mauromicale, Giovanni; Faccioli, Primetta; Cattivelli, Luigi; Rieseberg, Loren; Michelmore, Richard; Lanteri, Sergio

    2016-01-01

    Globe artichoke (Cynara cardunculus var. scolymus) is an out-crossing, perennial, multi-use crop species that is grown worldwide and belongs to the Compositae, one of the most successful Angiosperm families. We describe the first genome sequence of globe artichoke. The assembly, comprising of 13,588 scaffolds covering 725 of the 1,084 Mb genome, was generated using ~133-fold Illumina sequencing data and encodes 26,889 predicted genes. Re-sequencing (30×) of globe artichoke and cultivated cardoon (C. cardunculus var. altilis) parental genotypes and low-coverage (0.5 to 1×) genotyping-by-sequencing of 163 F1 individuals resulted in 73% of the assembled genome being anchored in 2,178 genetic bins ordered along 17 chromosomal pseudomolecules. This was achieved using a novel pipeline, SOILoCo (Scaffold Ordering by Imputation with Low Coverage), to detect heterozygous regions and assign parental haplotypes with low sequencing read depth and of unknown phase. SOILoCo provides a powerful tool for de novo genome analysis of outcrossing species. Our data will enable genome-scale analyses of evolutionary processes among crops, weeds, and wild species within and beyond the Compositae, and will facilitate the identification of economically important genes from related species. PMID:26786968

  2. RISC assembly: Coordination between small RNAs and Argonaute proteins.

    Science.gov (United States)

    Kobayashi, Hotaka; Tomari, Yukihide

    2016-01-01

    Non-coding RNAs generally form ribonucleoprotein (RNP) complexes with their partner proteins to exert their functions. Small RNAs, including microRNAs, small interfering RNAs, and PIWI-interacting RNAs, assemble with Argonaute (Ago) family proteins into the effector complex called RNA-induced silencing complex (RISC), which mediates sequence-specific target gene silencing. RISC assembly is not a simple binding between a small RNA and Ago; rather, it follows an ordered multi-step pathway that requires specific accessory factors. Some steps of RISC assembly and RISC-mediated gene silencing are dependent on or facilitated by particular intracellular platforms, suggesting their spatial regulation. In this review, we summarize the currently known mechanisms for RISC assembly of each small RNA class and propose a revised model for the role of the chaperone machinery in the duplex-initiated RISC assembly pathway. This article is part of a Special Issue entitled: Clues to long noncoding RNA taxonomy1, edited by Dr. Tetsuro Hirose and Dr. Shinichi Nakagawa. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. DeepSimulator: a deep simulator for Nanopore sequencing

    KAUST Repository

    Li, Yu; Han, Renmin; Bi, Chongwei; Li, Mo; Wang, Sheng; Gao, Xin

    2017-01-01

    or assembled contigs, we simulate the electrical current signals by a context-dependent deep learning model, followed by a base-calling procedure to yield simulated reads. This workflow mimics the sequencing procedure more naturally. The thorough experiments

  4. Sparc: a sparsity-based consensus algorithm for long erroneous sequencing reads

    Directory of Open Access Journals (Sweden)

    Chengxi Ye

    2016-06-01

    Full Text Available Motivation. The third generation sequencing (3GS technology generates long sequences of thousands of bases. However, its current error rates are estimated in the range of 15–40%, significantly higher than those of the prevalent next generation sequencing (NGS technologies (less than 1%. Fundamental bioinformatics tasks such as de novo genome assembly and variant calling require high-quality sequences that need to be extracted from these long but erroneous 3GS sequences. Results. We describe a versatile and efficient linear complexity consensus algorithm Sparc to facilitate de novo genome assembly. Sparc builds a sparse k-mer graph using a collection of sequences from a targeted genomic region. The heaviest path which approximates the most likely genome sequence is searched through a sparsity-induced reweighted graph as the consensus sequence. Sparc supports using NGS and 3GS data together, which leads to significant improvements in both cost efficiency and computational efficiency. Experiments with Sparc show that our algorithm can efficiently provide high-quality consensus sequences using both PacBio and Oxford Nanopore sequencing technologies. With only 30× PacBio data, Sparc can reach a consensus with error rate <0.5%. With the more challenging Oxford Nanopore data, Sparc can also achieve similar error rate when combined with NGS data. Compared with the existing approaches, Sparc calculates the consensus with higher accuracy, and uses approximately 80% less memory and time. Availability. The source code is available for download at https://github.com/yechengxi/Sparc.

  5. Comparison of the Equine Reference Sequence with Its Sanger Source Data and New Illumina Reads.

    Directory of Open Access Journals (Sweden)

    Jovan Rebolledo-Mendez

    Full Text Available The reference assembly for the domestic horse, EquCab2, published in 2009, was built using approximately 30 million Sanger reads from a Thoroughbred mare named Twilight. Contiguity in the assembly was facilitated using nearly 315 thousand BAC end sequences from Twilight's half brother Bravo. Since then, it has served as the foundation for many genome-wide analyses that include not only the modern horse, but ancient horses and other equid species as well. As data mapped to this reference has accumulated, consistent variation between mapped datasets and the reference, in terms of regions with no read coverage, single nucleotide variants, and small insertions/deletions have become apparent. In many cases, it is not clear whether these differences are the result of true sequence variation between the research subjects' and Twilight's genome or due to errors in the reference. EquCab2 is regarded as "The Twilight Assembly." The objective of this study was to identify inconsistencies between the EquCab2 assembly and the source Twilight Sanger data used to build it. To that end, the original Sanger and BAC end reads have been mapped back to this equine reference and assessed with the addition of approximately 40X coverage of new Illumina Paired-End sequence data. The resulting mapped datasets identify those regions with low Sanger read coverage, as well as variation in genomic content that is not consistent with either the original Twilight Sanger data or the new genomic sequence data generated from Twilight on the Illumina platform. As the haploid EquCab2 reference assembly was created using Sanger reads derived largely from a single individual, the vast majority of variation detected in a mapped dataset comprised of those same Sanger reads should be heterozygous. In contrast, homozygous variations would represent either errors in the reference or contributions from Bravo's BAC end sequences. Our analysis identifies 720,843 homozygous discrepancies

  6. XplorSeq: a software environment for integrated management and phylogenetic analysis of metagenomic sequence data.

    Science.gov (United States)

    Frank, Daniel N

    2008-10-07

    Advances in automated DNA sequencing technology have accelerated the generation of metagenomic DNA sequences, especially environmental ribosomal RNA gene (rDNA) sequences. As the scale of rDNA-based studies of microbial ecology has expanded, need has arisen for software that is capable of managing, annotating, and analyzing the plethora of diverse data accumulated in these projects. XplorSeq is a software package that facilitates the compilation, management and phylogenetic analysis of DNA sequences. XplorSeq was developed for, but is not limited to, high-throughput analysis of environmental rRNA gene sequences. XplorSeq integrates and extends several commonly used UNIX-based analysis tools by use of a Macintosh OS-X-based graphical user interface (GUI). Through this GUI, users may perform basic sequence import and assembly steps (base-calling, vector/primer trimming, contig assembly), perform BLAST (Basic Local Alignment and Search Tool; 123) searches of NCBI and local databases, create multiple sequence alignments, build phylogenetic trees, assemble Operational Taxonomic Units, estimate biodiversity indices, and summarize data in a variety of formats. Furthermore, sequences may be annotated with user-specified meta-data, which then can be used to sort data and organize analyses and reports. A document-based architecture permits parallel analysis of sequence data from multiple clones or amplicons, with sequences and other data stored in a single file. XplorSeq should benefit researchers who are engaged in analyses of environmental sequence data, especially those with little experience using bioinformatics software. Although XplorSeq was developed for management of rDNA sequence data, it can be applied to most any sequencing project. The application is available free of charge for non-commercial use at http://vent.colorado.edu/phyloware.

  7. A first report and complete genome sequence of alfalfa enamovirus from Sudan

    Science.gov (United States)

    A full genome sequence of a viral pathogen, provisionally named alfalfa enamovirus 2 (AEV-2), was reconstructed from short reads obtained by Illumina RNA sequencing of alfalfa sample originating from Sudan. Ambiguous nucleotides in the resultant consensus assembly and identity of the predicted virus...

  8. The Nereus Hybrid Underwater Robotic Vehicle for Global Ocean Science Operations to 11,000m Depth

    Science.gov (United States)

    2008-09-01

    B. Butler. Field trials of the Theseus AUV. Proc. Int. Symp. on Unmanned Untethered Submersible Technology, page 615, 1995. [9] R. M. Eustice, L. L...pages 4257–4264, Apr. 2007. [10] J. Ferguson, A. Pope, B. Butler, and R. Verrall. Theseus AUV-two record breaking missions. Sea Technology, 40(2):65

  9. Programmed assembly of nanoscale structures using peptoids.

    Energy Technology Data Exchange (ETDEWEB)

    Ren, Jianhua (University of the Pacific, Stockton, CA); Russell, Scott (California State University, Stanislaus, Turlock, CA); Morishetti, Kiran (University of the Pacific, Stockton, CA); Robinson, David B.; Zuckermann, Ronald N. (Lawrence Berkeley National Laboratory, Berkeley, CA); Buffleben, George M.; Hjelm, Rex P. (Los Alamos National Laboratory, Los Alamos, NM); Kent, Michael Stuart (Sandia National Laboratories, Albuquerque, NM)

    2011-02-01

    Sequence-specific polymers are the basis of the most promising approaches to bottom-up programmed assembly of nanoscale materials. Examples include artificial peptides and nucleic acids. Another class is oligo(N-functional glycine)s, also known as peptoids, which permit greater sidegroup diversity and conformational control, and can be easier to synthesize and purify. We have developed a set of peptoids that can be used to make inorganic nanoparticles more compatible with biological sequence-specific polymers so that they can be incorporated into nucleic acid or other biologically based nanostructures. Peptoids offer degrees of modularity, versatility, and predictability that equal or exceed other sequence-specific polymers, allowing for rational design of oligomers for a specific purpose. This degree of control will be essential to the development of arbitrarily designed nanoscale structures.

  10. Direct comparisons of Illumina vs. Roche 454 sequencing technologies on the same microbial community DNA sample.

    Directory of Open Access Journals (Sweden)

    Chengwei Luo

    Full Text Available Next-generation sequencing (NGS is commonly used in metagenomic studies of complex microbial communities but whether or not different NGS platforms recover the same diversity from a sample and their assembled sequences are of comparable quality remain unclear. We compared the two most frequently used platforms, the Roche 454 FLX Titanium and the Illumina Genome Analyzer (GA II, on the same DNA sample obtained from a complex freshwater planktonic community. Despite the substantial differences in read length and sequencing protocols, the platforms provided a comparable view of the community sampled. For instance, derived assemblies overlapped in ~90% of their total sequences and in situ abundances of genes and genotypes (estimated based on sequence coverage correlated highly between the two platforms (R(2>0.9. Evaluation of base-call error, frameshift frequency, and contig length suggested that Illumina offered equivalent, if not better, assemblies than Roche 454. The results from metagenomic samples were further validated against DNA samples of eighteen isolate genomes, which showed a range of genome sizes and G+C% content. We also provide quantitative estimates of the errors in gene and contig sequences assembled from datasets characterized by different levels of complexity and G+C% content. For instance, we noted that homopolymer-associated, single-base errors affected ~1% of the protein sequences recovered in Illumina contigs of 10× coverage and 50% G+C; this frequency increased to ~3% when non-homopolymer errors were also considered. Collectively, our results should serve as a useful practical guide for choosing proper sampling strategies and data possessing protocols for future metagenomic studies.

  11. A two-stage stochastic rule-based model to determine pre-assembly buffer content

    Science.gov (United States)

    Gunay, Elif Elcin; Kula, Ufuk

    2018-01-01

    This study considers instant decision-making needs of the automobile manufactures for resequencing vehicles before final assembly (FA). We propose a rule-based two-stage stochastic model to determine the number of spare vehicles that should be kept in the pre-assembly buffer to restore the altered sequence due to paint defects and upstream department constraints. First stage of the model decides the spare vehicle quantities, where the second stage model recovers the scrambled sequence respect to pre-defined rules. The problem is solved by sample average approximation (SAA) algorithm. We conduct a numerical study to compare the solutions of heuristic model with optimal ones and provide following insights: (i) as the mismatch between paint entrance and scheduled sequence decreases, the rule-based heuristic model recovers the scrambled sequence as good as the optimal resequencing model, (ii) the rule-based model is more sensitive to the mismatch between the paint entrance and scheduled sequences for recovering the scrambled sequence, (iii) as the defect rate increases, the difference in recovery effectiveness between rule-based heuristic and optimal solutions increases, (iv) as buffer capacity increases, the recovery effectiveness of the optimization model outperforms heuristic model, (v) as expected the rule-based model holds more inventory than the optimization model.

  12. The Automated Assembly Team contributions to the APRIMED Agile Manufacturing Project

    International Nuclear Information System (INIS)

    Jones, R.E.; Ames, A.L.; Calton, T.L.

    1995-06-01

    The Automated Assembly Team of the APRIMED Project (abbreviated as A') consists of two parts: the Archimedes Project, which is an ongoing project developing automated assembly technology, and the A' Robot Team. Archimedes is a second generation assembly planning system that both provides a general high-level assembly sequencing capability and, for a smaller class of products, facilitates automatic programming of a robotic workcell to assemble them. The A' robot team designed, developed, and implemented a flexible robot workcell which served as the automated factory of the A' project. In this document we briefly describe the role of automated assembly planning in agile manufacturing, and specifically describe the contributions of the Archimedes project and the A' robot team to the A' project. We introduce the concepts of the Archimedes automated assembly planning project, and discuss the enhancements to Archimedes which were developed in response to the needs of the A' project. We also present the work of the A' robot team in designing and developing the A' robot workcell, including all tooling and programming to support assembly of the A' discriminator devices. Finally, we discuss the process changes which these technologies have enabled in the A' project

  13. Discovery of genes related to insecticide resistance in Bactrocera dorsalis by functional genomic analysis of a de novo assembled transcriptome.

    Science.gov (United States)

    Hsu, Ju-Chun; Chien, Ting-Ying; Hu, Chia-Cheng; Chen, Mei-Ju May; Wu, Wen-Jer; Feng, Hai-Tung; Haymer, David S; Chen, Chien-Yu

    2012-01-01

    Insecticide resistance has recently become a critical concern for control of many insect pest species. Genome sequencing and global quantization of gene expression through analysis of the transcriptome can provide useful information relevant to this challenging problem. The oriental fruit fly, Bactrocera dorsalis, is one of the world's most destructive agricultural pests, and recently it has been used as a target for studies of genetic mechanisms related to insecticide resistance. However, prior to this study, the molecular data available for this species was largely limited to genes identified through homology. To provide a broader pool of gene sequences of potential interest with regard to insecticide resistance, this study uses whole transcriptome analysis developed through de novo assembly of short reads generated by next-generation sequencing (NGS). The transcriptome of B. dorsalis was initially constructed using Illumina's Solexa sequencing technology. Qualified reads were assembled into contigs and potential splicing variants (isotigs). A total of 29,067 isotigs have putative homologues in the non-redundant (nr) protein database from NCBI, and 11,073 of these correspond to distinct D. melanogaster proteins in the RefSeq database. Approximately 5,546 isotigs contain coding sequences that are at least 80% complete and appear to represent B. dorsalis genes. We observed a strong correlation between the completeness of the assembled sequences and the expression intensity of the transcripts. The assembled sequences were also used to identify large numbers of genes potentially belonging to families related to insecticide resistance. A total of 90 P450-, 42 GST-and 37 COE-related genes, representing three major enzyme families involved in insecticide metabolism and resistance, were identified. In addition, 36 isotigs were discovered to contain target site sequences related to four classes of resistance genes. Identified sequence motifs were also analyzed to

  14. Molecular Mechanism of Arenavirus Assembly and Budding

    Directory of Open Access Journals (Sweden)

    Shuzo Urata

    2012-10-01

    Full Text Available Arenaviruses have a bisegmented negative-strand RNA genome, which encodes four viral proteins: GP and NP by the S segment and L and Z by the L segment. These four viral proteins possess multiple functions in infection, replication and release of progeny viruses from infected cells. The small RING finger protein, Z protein is a matrix protein that plays a central role in viral assembly and budding. Although all arenaviruses encode Z protein, amino acid sequence alignment showed a huge variety among the species, especially at the C-terminus where the L-domain is located. Recent publications have demonstrated the interactions between viral protein and viral protein, and viral protein and host cellular protein, which facilitate transportation and assembly of viral components to sites of virus egress. This review presents a summary of current knowledge regarding arenavirus assembly and budding, in comparison with other enveloped viruses. We also refer to the restriction of arenavirus production by the antiviral cellular factor, Tetherin/BST-2.

  15. Comparison of bacterial genome assembly software for MinION data and their applicability to medical microbiology.

    Science.gov (United States)

    Judge, Kim; Hunt, Martin; Reuter, Sandra; Tracey, Alan; Quail, Michael A; Parkhill, Julian; Peacock, Sharon J

    2016-09-01

    Translating the Oxford Nanopore MinION sequencing technology into medical microbiology requires on-going analysis that keeps pace with technological improvements to the instrument and release of associated analysis software. Here, we use a multidrug-resistant Enterobacter kobei isolate as a model organism to compare open source software for the assembly of genome data, and relate this to the time taken to generate actionable information. Three software tools (PBcR, Canu and miniasm) were used to assemble MinION data and a fourth (SPAdes) was used to combine MinION and Illumina data to produce a hybrid assembly. All four had a similar number of contigs and were more contiguous than the assembly using Illumina data alone, with SPAdes producing a single chromosomal contig. Evaluation of the four assemblies to represent the genome structure revealed a single large inversion in the SPAdes assembly, which also incorrectly integrated a plasmid into the chromosomal contig. Almost 50 %, 80 % and 90 % of MinION pass reads were generated in the first 6, 9 and 12 h, respectively. Using data from the first 6 h alone led to a less accurate, fragmented assembly, but data from the first 9 or 12 h generated similar assemblies to that from 48 h sequencing. Assemblies were generated in 2 h using Canu, indicating that going from isolate to assembled data is possible in less than 48 h. MinION data identified that genes responsible for resistance were carried by two plasmids encoding resistance to carbapenem and to sulphonamides, rifampicin and aminoglycosides, respectively.

  16. De novo transcriptome assembly of Sorghum bicolor variety Taejin

    Directory of Open Access Journals (Sweden)

    Yeonhwa Jo

    2016-06-01

    Full Text Available Sorghum (Sorghum bicolor, also known as great millet, is one of the most popular cultivated grass species in the world. Sorghum is frequently consumed as food for humans and animals as well as used for ethanol production. In this study, we conducted de novo transcriptome assembly for sorghum variety Taejin by next-generation sequencing, obtaining 8.748 GB of raw data. The raw data in this study can be available in NCBI SRA database with accession number of SRX1715644. Using the Trinity program, we identified 222,161 transcripts from sorghum variety Taejin. We further predicted coding regions within the assembled transcripts by the TransDecoder program, resulting in a total of 148,531 proteins. We carried out BLASTP against the Swiss-Prot protein sequence database to annotate the functions of the identified proteins. To our knowledge, this is the first transcriptome data for a sorghum variety derived from Korea, and it can be usefully applied to the generation of genetic markers.

  17. Fine de novo sequencing of a fungal genome using only SOLiD short read data: verification on Aspergillus oryzae RIB40.

    Directory of Open Access Journals (Sweden)

    Myco Umemura

    Full Text Available The development of next-generation sequencing (NGS technologies has dramatically increased the throughput, speed, and efficiency of genome sequencing. The short read data generated from NGS platforms, such as SOLiD and Illumina, are quite useful for mapping analysis. However, the SOLiD read data with lengths of <60 bp have been considered to be too short for de novo genome sequencing. Here, to investigate whether de novo sequencing of fungal genomes is possible using only SOLiD short read sequence data, we performed de novo assembly of the Aspergillus oryzae RIB40 genome using only SOLiD read data of 50 bp generated from mate-paired libraries with 2.8- or 1.9-kb insert sizes. The assembled scaffolds showed an N50 value of 1.6 Mb, a 22-fold increase than those obtained using only SOLiD short read in other published reports. In addition, almost 99% of the reference genome was accurately aligned by the assembled scaffold fragments in long lengths. The sequences of secondary metabolite biosynthetic genes and clusters, whose products are of considerable interest in fungal studies due to their potential medicinal, agricultural, and cosmetic properties, were also highly reconstructed in the assembled scaffolds. Based on these findings, we concluded that de novo genome sequencing using only SOLiD short reads is feasible and practical for molecular biological study of fungi. We also investigated the effect of filtering low quality data, library insert size, and k-mer size on the assembly performance, and recommend for the assembly use of mild filtered read data where the N50 was not so degraded and the library has an insert size of ∼2.0 kb, and k-mer size 33.

  18. Genome sequence of the oleaginous yeast Rhodotorula toruloides strain CGMCC 2.1609

    Directory of Open Access Journals (Sweden)

    Christine Sambles

    2017-09-01

    Full Text Available Most eukaryotic oleaginous species are yeasts and among them the basidiomycete red yeast, Rhodotorula (Rhodosporidium toruloides (Pucciniomycotina is known to produce high quantities of lipids when grown in nitrogen-limiting media, and has potential for biodiesel production. The genome of the CGMCC 2.1609 strain of this oleaginous red yeast was sequenced using a hybrid of Roche 454 and Illumina technology generating 13× coverage. The de novo assembly was carried out using MIRA and scaffolded using MAQ and BAMBUS. The sequencing and assembly resulted in 365 scaffolds with total genome size of 33.4 Mb. The complete genome sequence of this strain was deposited in GenBank and the accession number is LKER00000000. The annotation is available on Figshare (doi:10.6084/m9.figshare.4754251.

  19. Nanomaterial processing using self-assembly-bottom-up chemical and biological approaches

    International Nuclear Information System (INIS)

    Thiruvengadathan, Rajagopalan; Gangopadhyay, Keshab; Gangopadhyay, Shubhra; Korampally, Venumadhav; Ghosh, Arkasubhra; Chanda, Nripen

    2013-01-01

    Nanotechnology is touted as the next logical sequence in technological evolution. This has led to a substantial surge in research activities pertaining to the development and fundamental understanding of processes and assembly at the nanoscale. Both top-down and bottom-up fabrication approaches may be used to realize a range of well-defined nanostructured materials with desirable physical and chemical attributes. Among these, the bottom-up self-assembly process offers the most realistic solution toward the fabrication of next-generation functional materials and devices. Here, we present a comprehensive review on the physical basis behind self-assembly and the processes reported in recent years to direct the assembly of nanoscale functional blocks into hierarchically ordered structures. This paper emphasizes assembly in the synthetic domain as well in the biological domain, underscoring the importance of biomimetic approaches toward novel materials. In particular, two important classes of directed self-assembly, namely, (i) self-assembly among nanoparticle–polymer systems and (ii) external field-guided assembly are highlighted. The spontaneous self-assembling behavior observed in nature that leads to complex, multifunctional, hierarchical structures within biological systems is also discussed in this review. Recent research undertaken to synthesize hierarchically assembled functional materials have underscored the need as well as the benefits harvested in synergistically combining top-down fabrication methods with bottom-up self-assembly. (review article)

  20. DNA fragments assembly based on nicking enzyme system.

    Directory of Open Access Journals (Sweden)

    Rui-Yan Wang

    Full Text Available A couple of DNA ligation-independent cloning (LIC methods have been reported to meet various requirements in metabolic engineering and synthetic biology. The principle of LIC is the assembly of multiple overlapping DNA fragments by single-stranded (ss DNA overlaps annealing. Here we present a method to generate single-stranded DNA overlaps based on Nicking Endonucleases (NEases for LIC, the method was termed NE-LIC. Factors related to cloning efficiency were optimized in this study. This NE-LIC allows generating 3'-end or 5'-end ss DNA overlaps of various lengths for fragments assembly. We demonstrated that the 10 bp/15 bp overlaps had the highest DNA fragments assembling efficiency, while 5 bp/10 bp overlaps showed the highest efficiency when T4 DNA ligase was added. Its advantage over Sequence and Ligation Independent Cloning (SLIC and Uracil-Specific Excision Reagent (USER was obvious. The mechanism can be applied to many other LIC strategies. Finally, the NEases based LIC (NE-LIC was successfully applied to assemble a pathway of six gene fragments responsible for synthesizing microbial poly-3-hydroxybutyrate (PHB.

  1. The Douglas-fir genome sequence reveals specialization of the photosynthetic apparatus in Pinaceae

    Science.gov (United States)

    David B. Neale; Patrick E. McGuire; Nicholas C. Wheeler; Kristian A. Stevens; Marc W. Crepeau; Charis Cardeno; Aleksey V. Zimin; Daniela Puiu; Geo M. Pertea; U. Uzay Sezen; Claudio Casola; Tomasz E. Koralewski; Robin Paul; Daniel Gonzalez-Ibeas; Sumaira Zaman; Richard Cronn; Mark Yandell; Carson Holt; Charles H. Langley; James A. Yorke; Steven L. Salzberg; Jill L. Wegrzyn

    2017-01-01

    A reference genome sequence for Pseudotsuga menziesii var. menziesii (Mirb.) Franco (Coastal Douglas-fir) is reported, thus providing a reference sequence for a third genus of the family Pinaceae. The contiguity and quality of the genome assembly far exceeds that of other conifer reference genome sequences (contig N50 = 44,136 bp and scaffold N50...

  2. Complete genome sequences of six strains of the genus methylobacterium

    Energy Technology Data Exchange (ETDEWEB)

    Marx, Christopher J [Harvard University; Bringel, Francoise O. [University of Strasbourg; Christoserdova, Ludmila [University of Washington, Seattle; Moulin, Lionel [UMR, France; Farhan Ul Haque, Muhammad [CNRS, Strasbourg, France; Fleischman, Darrell E. [Wright State University, Dayton, OH; Gruffaz, Christelle [CNRS, Strasbourg, France; Jourand, Philippe [UMR, France; Knief, Claudia [ETH Zurich, Switzerland; Lee, Ming-Chun [Harvard University; Muller, Emilie E. L. [CNRS, Strasbourg, France; Nadalig, Thierry [CNRS, Strasbourg, France; Peyraud, Remi [ETH Zurich, Switzerland; Roselli, Sandro [CNRS, Strasbourg, France; Russ, Lina [ETH Zurich, Switzerland; Aguero, Fernan [Universidad Nacional de General San Martin; Goodwin, Lynne A. [Los Alamos National Laboratory (LANL); Ivanova, N [U.S. Department of Energy, Joint Genome Institute; Kyrpides, Nikos C [U.S. Department of Energy, Joint Genome Institute; Lajus, Aurelie [Genoscope/Centre National de la Recherche Scientifique-Unite Mixte de Recherche; Land, Miriam L [ORNL; Medigue, Claudine [Genoscope/Centre National de la Recherche Scientifique-Unite Mixte de Recherche; Mikhailova, Natalia [U.S. Department of Energy, Joint Genome Institute; Nolan, Matt [U.S. Department of Energy, Joint Genome Institute; Woyke, Tanja [U.S. Department of Energy, Joint Genome Institute; Stolyar, Sergey [University of Washington; Vorholt, Julia A. [ETH Zurich, Switzerland; Vuilleumier, Stephane [University of Strasbourg

    2012-01-01

    The complete and assembled genome sequences were determined for six strains of the alphaproteobacterial genus Methylobacterium, chosen for their key adaptations to different plant-associated niches and environmental constraints.

  3. Complete Genome Sequences of Six Strains of the Genus Methylobacterium

    Energy Technology Data Exchange (ETDEWEB)

    Marx, Christopher J [Harvard University; Bringel, Francoise O. [University of Strasbourg; Christoserdova, Ludmila [University of Washington, Seattle; Moulin, Lionel [UMR, France; UI Hague, Muhammad Farhan [University of Strasbourg; Fleischman, Darrell E. [Wright State University, Dayton, OH; Gruffaz, Christelle [CNRS, Strasbourg, France; Jourand, Philippe [UMR, France; Knief, Claudia [ETH Zurich, Switzerland; Lee, Ming-Chun [Harvard University; Muller, Emilie E. L. [CNRS, Strasbourg, France; Nadalig, Thierry [CNRS, Strasbourg, France; Peyraud, Remi [ETH Zurich, Switzerland; Roselli, Sandro [CNRS, Strasbourg, France; Russ, Lina [ETH Zurich, Switzerland; Goodwin, Lynne A. [Los Alamos National Laboratory (LANL); Ivanov, Pavel S. [University of Wyoming, Laramie; Ivanova, N [U.S. Department of Energy, Joint Genome Institute; Kyrpides, Nikos C [U.S. Department of Energy, Joint Genome Institute; Lajus, Aurelie [Genoscope/Centre National de la Recherche Scientifique-Unite Mixte de Recherche; Land, Miriam L [ORNL; Medigue, Claudine [Genoscope/Centre National de la Recherche Scientifique-Unite Mixte de Recherche; Mikhailova, Natalia [U.S. Department of Energy, Joint Genome Institute; Nolan, Matt [U.S. Department of Energy, Joint Genome Institute; Woyke, Tanja [U.S. Department of Energy, Joint Genome Institute; Stolyar, Sergey [University of Washington; Vorholt, Julia A. [ETH Zurich, Switzerland; Vuilleumier, Stephane [University of Strasbourg

    2012-01-01

    The complete and assembled genome sequences were determined for six strains of the alphaproteobacterial genus Methylobacterium, chosen for their key adaptations to different plant-associated niches and environmental constraints.

  4. MOCAT: a metagenomics assembly and gene prediction toolkit.

    Science.gov (United States)

    Kultima, Jens Roat; Sunagawa, Shinichi; Li, Junhua; Chen, Weineng; Chen, Hua; Mende, Daniel R; Arumugam, Manimozhiyan; Pan, Qi; Liu, Binghang; Qin, Junjie; Wang, Jun; Bork, Peer

    2012-01-01

    MOCAT is a highly configurable, modular pipeline for fast, standardized processing of single or paired-end sequencing data generated by the Illumina platform. The pipeline uses state-of-the-art programs to quality control, map, and assemble reads from metagenomic samples sequenced at a depth of several billion base pairs, and predict protein-coding genes on assembled metagenomes. Mapping against reference databases allows for read extraction or removal, as well as abundance calculations. Relevant statistics for each processing step can be summarized into multi-sheet Excel documents and queryable SQL databases. MOCAT runs on UNIX machines and integrates seamlessly with the SGE and PBS queuing systems, commonly used to process large datasets. The open source code and modular architecture allow users to modify or exchange the programs that are utilized in the various processing steps. Individual processing steps and parameters were benchmarked and tested on artificial, real, and simulated metagenomes resulting in an improvement of selected quality metrics. MOCAT can be freely downloaded at http://www.bork.embl.de/mocat/.

  5. MOCAT: a metagenomics assembly and gene prediction toolkit.

    Directory of Open Access Journals (Sweden)

    Jens Roat Kultima

    Full Text Available MOCAT is a highly configurable, modular pipeline for fast, standardized processing of single or paired-end sequencing data generated by the Illumina platform. The pipeline uses state-of-the-art programs to quality control, map, and assemble reads from metagenomic samples sequenced at a depth of several billion base pairs, and predict protein-coding genes on assembled metagenomes. Mapping against reference databases allows for read extraction or removal, as well as abundance calculations. Relevant statistics for each processing step can be summarized into multi-sheet Excel documents and queryable SQL databases. MOCAT runs on UNIX machines and integrates seamlessly with the SGE and PBS queuing systems, commonly used to process large datasets. The open source code and modular architecture allow users to modify or exchange the programs that are utilized in the various processing steps. Individual processing steps and parameters were benchmarked and tested on artificial, real, and simulated metagenomes resulting in an improvement of selected quality metrics. MOCAT can be freely downloaded at http://www.bork.embl.de/mocat/.

  6. Approaches for in silico finishing of microbial genome sequences

    Directory of Open Access Journals (Sweden)

    Frederico Schmitt Kremer

    Full Text Available Abstract The introduction of next-generation sequencing (NGS had a significant effect on the availability of genomic information, leading to an increase in the number of sequenced genomes from a large spectrum of organisms. Unfortunately, due to the limitations implied by the short-read sequencing platforms, most of these newly sequenced genomes remained as “drafts”, incomplete representations of the whole genetic content. The previous genome sequencing studies indicated that finishing a genome sequenced by NGS, even bacteria, may require additional sequencing to fill the gaps, making the entire process very expensive. As such, several in silico approaches have been developed to optimize the genome assemblies and facilitate the finishing process. The present review aims to explore some free (open source, in many cases tools that are available to facilitate genome finishing.

  7. Approaches for in silico finishing of microbial genome sequences.

    Science.gov (United States)

    Kremer, Frederico Schmitt; McBride, Alan John Alexander; Pinto, Luciano da Silva

    The introduction of next-generation sequencing (NGS) had a significant effect on the availability of genomic information, leading to an increase in the number of sequenced genomes from a large spectrum of organisms. Unfortunately, due to the limitations implied by the short-read sequencing platforms, most of these newly sequenced genomes remained as "drafts", incomplete representations of the whole genetic content. The previous genome sequencing studies indicated that finishing a genome sequenced by NGS, even bacteria, may require additional sequencing to fill the gaps, making the entire process very expensive. As such, several in silico approaches have been developed to optimize the genome assemblies and facilitate the finishing process. The present review aims to explore some free (open source, in many cases) tools that are available to facilitate genome finishing.

  8. Combinatorial Models for Assembly and Decomposition of Products

    Directory of Open Access Journals (Sweden)

    A. N. Bojko

    2015-01-01

    Full Text Available The paper discusses the most popular combinatorial models that are used for the synthesis of design solutions at the stage of the assembly process flow preparation. It shows that while assembling the product the relations of parts can be represented as a structure of preferences, which is formed on the basis of objective design restrictions put in at the stage of the product design. This structure is a binary preference relation pre-order. Its symmetrical part is equivalence and describes the entry of parts into the assembly unit. The asymmetric part is a partial order. It specifies part- ordering time in in the course of the assembly process. The structure of preferences is a minimal description of the restrictions and constraints in the assembly process. It can serve as a source for generating multiple assembly sequences of a product and its components, which are allowed by design. This multiplicity increases the likelihood of rational choice under uncertainty, unpredictable changes in the properties of technological or industrial systems.Incomplete dominance relation gives grounds for further examination and better understanding of the project situation. Operation field of the study is limited to a set of disparate elements of the partial order. Different strategies for processing the disparate elements may be offered, e.g. selection of the most informative pairs, comparison of which foremost linearizes the original partial order.

  9. Genome puzzle master (GPM): an integrated pipeline for building and editing pseudomolecules from fragmented sequences.

    Science.gov (United States)

    Zhang, Jianwei; Kudrna, Dave; Mu, Ting; Li, Weiming; Copetti, Dario; Yu, Yeisoo; Goicoechea, Jose Luis; Lei, Yang; Wing, Rod A

    2016-10-15

    Next generation sequencing technologies have revolutionized our ability to rapidly and affordably generate vast quantities of sequence data. Once generated, raw sequences are assembled into contigs or scaffolds. However, these assemblies are mostly fragmented and inaccurate at the whole genome scale, largely due to the inability to integrate additional informative datasets (e.g. physical, optical and genetic maps). To address this problem, we developed a semi-automated software tool-Genome Puzzle Master (GPM)-that enables the integration of additional genomic signposts to edit and build 'new-gen-assemblies' that result in high-quality 'annotation-ready' pseudomolecules. With GPM, loaded datasets can be connected to each other via their logical relationships which accomplishes tasks to 'group,' 'merge,' 'order and orient' sequences in a draft assembly. Manual editing can also be performed with a user-friendly graphical interface. Final pseudomolecules reflect a user's total data package and are available for long-term project management. GPM is a web-based pipeline and an important part of a Laboratory Information Management System (LIMS) which can be easily deployed on local servers for any genome research laboratory. The GPM (with LIMS) package is available at https://github.com/Jianwei-Zhang/LIMS CONTACTS: jzhang@mail.hzau.edu.cn or rwing@mail.arizona.eduSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press.

  10. Immobilization of the enzyme {beta}-lactamase by self-assembly on thin films of a poly(phenyleneethynylene) sequenced with flexible segments containing sulfur atoms

    Energy Technology Data Exchange (ETDEWEB)

    Vazquez, Erika [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Aguilar, Abdieel Esquivel [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Facultad de Ciencias Quimicas, Universidad Autonoma de Coahuila, Blvd. V. Carranza and Ing. J. Cardenas, 25000 Saltillo (Mexico); Moggio, Ivana [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico)]. E-mail: imoggio@ciqa.mx; Arias, Eduardo [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Romero, Jorge [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Barrientos, Hector [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Torres, Jose Roman [Centro de Investigacion en Quimica Aplicada (CIQA), Blvd. Enrique Reyna 140, 25253, Saltillo (Mexico); Luz Reyes Vega, Maria de la [Facultad de Ciencias Quimicas, Universidad Autonoma de Coahuila, Blvd. V. Carranza and Ing. J. Cardenas, 25000 Saltillo (Mexico)

    2007-05-16

    A novel poly(phenyleneethynylene) sequenced in the main conjugated chain with flexible groups containing sulfur atoms has been synthesized by Heck-Sonogashira coupling reaction. Layer-by-layer films of the polymer have been prepared with a linear growth in thickness up to four layers as evidenced by UV-Vis spectroscopy and perfilometry. On the top of these multilayers, the enzyme {beta}-lactamase was deposited by self-assembly. The enzymatic activity was measured by a modified spectrophotometric standard assay method for penicillin G, ampicillin and amoxicillin. A higher and faster activity was obtained for penicillin G and thus preliminary study of the biosensor response by fluorescence was carried out for this antibiotic revealing a decrease in the polymer fluorescence as function of the penicillin G concentration.

  11. Immobilization of the enzyme β-lactamase by self-assembly on thin films of a poly(phenyleneethynylene) sequenced with flexible segments containing sulfur atoms

    International Nuclear Information System (INIS)

    Vazquez, Erika; Aguilar, Abdieel Esquivel; Moggio, Ivana; Arias, Eduardo; Romero, Jorge; Barrientos, Hector; Torres, Jose Roman; Luz Reyes Vega, Maria de la

    2007-01-01

    A novel poly(phenyleneethynylene) sequenced in the main conjugated chain with flexible groups containing sulfur atoms has been synthesized by Heck-Sonogashira coupling reaction. Layer-by-layer films of the polymer have been prepared with a linear growth in thickness up to four layers as evidenced by UV-Vis spectroscopy and perfilometry. On the top of these multilayers, the enzyme β-lactamase was deposited by self-assembly. The enzymatic activity was measured by a modified spectrophotometric standard assay method for penicillin G, ampicillin and amoxicillin. A higher and faster activity was obtained for penicillin G and thus preliminary study of the biosensor response by fluorescence was carried out for this antibiotic revealing a decrease in the polymer fluorescence as function of the penicillin G concentration

  12. De novo transcriptome sequencing and sequence analysis of the malaria vector Anopheles sinensis (Diptera: Culicidae)

    Science.gov (United States)

    2014-01-01

    Background Anopheles sinensis is the major malaria vector in China and Southeast Asia. Vector control is one of the most effective measures to prevent malaria transmission. However, there is little transcriptome information available for the malaria vector. To better understand the biological basis of malaria transmission and to develop novel and effective means of vector control, there is a need to build a transcriptome dataset for functional genomics analysis by large-scale RNA sequencing (RNA-seq). Methods To provide a more comprehensive and complete transcriptome of An. sinensis, eggs, larvae, pupae, male adults and female adults RNA were pooled together for cDNA preparation, sequenced using the Illumina paired-end sequencing technology and assembled into unigenes. These unigenes were then analyzed in their genome mapping, functional annotation, homology, codon usage bias and simple sequence repeats (SSRs). Results Approximately 51.6 million clean reads were obtained, trimmed, and assembled into 38,504 unigenes with an average length of 571 bp, an N50 of 711 bp, and an average GC content 51.26%. Among them, 98.4% of unigenes could be mapped onto the reference genome, and 69% of unigenes could be annotated with known biological functions. Homology analysis identified certain numbers of An. sinensis unigenes that showed homology or being putative 1:1 orthologues with genomes of other Dipteran species. Codon usage bias was analyzed and 1,904 SSRs were detected, which will provide effective molecular markers for the population genetics of this species. Conclusions Our data and analysis provide the most comprehensive transcriptomic resource and characteristics currently available for An. sinensis, and will facilitate genetic, genomic studies, and further vector control of An. sinensis. PMID:25000941

  13. Whole transcriptome analysis using next-generation sequencing of model species Setaria viridis to support C4 photosynthesis research.

    Science.gov (United States)

    Xu, Jiajia; Li, Yuanyuan; Ma, Xiuling; Ding, Jianfeng; Wang, Kai; Wang, Sisi; Tian, Ye; Zhang, Hui; Zhu, Xin-Guang

    2013-09-01

    Setaria viridis is an emerging model species for genetic studies of C4 photosynthesis. Many basic molecular resources need to be developed to support for this species. In this paper, we performed a comprehensive transcriptome analysis from multiple developmental stages and tissues of S. viridis using next-generation sequencing technologies. Sequencing of the transcriptome from multiple tissues across three developmental stages (seed germination, vegetative growth, and reproduction) yielded a total of 71 million single end 100 bp long reads. Reference-based assembly using Setaria italica genome as a reference generated 42,754 transcripts. De novo assembly generated 60,751 transcripts. In addition, 9,576 and 7,056 potential simple sequence repeats (SSRs) covering S. viridis genome were identified when using the reference based assembled transcripts and the de novo assembled transcripts, respectively. This identified transcripts and SSR provided by this study can be used for both reverse and forward genetic studies based on S. viridis.

  14. De novo transcriptome sequence assembly from coconut leaves and seeds with a focus on factors involved in RNA-directed DNA methylation.

    Science.gov (United States)

    Huang, Ya-Yi; Lee, Chueh-Pai; Fu, Jason L; Chang, Bill Chia-Han; Matzke, Antonius J M; Matzke, Marjori

    2014-09-04

    Coconut palm (Cocos nucifera) is a symbol of the tropics and a source of numerous edible and nonedible products of economic value. Despite its nutritional and industrial significance, coconut remains under-represented in public repositories for genomic and transcriptomic data. We report de novo transcript assembly from RNA-seq data and analysis of gene expression in seed tissues (embryo and endosperm) and leaves of a dwarf coconut variety. Assembly of 10 GB sequencing data for each tissue resulted in 58,211 total unigenes in embryo, 61,152 in endosperm, and 33,446 in leaf. Within each unigene pool, 24,857 could be annotated in embryo, 29,731 could be annotated in endosperm, and 26,064 could be annotated in leaf. A KEGG analysis identified 138, 138, and 139 pathways, respectively, in transcriptomes of embryo, endosperm, and leaf tissues. Given the extraordinarily large size of coconut seeds and the importance of small RNA-mediated epigenetic regulation during seed development in model plants, we used homology searches to identify putative homologs of factors required for RNA-directed DNA methylation in coconut. The findings suggest that RNA-directed DNA methylation is important during coconut seed development, particularly in maturing endosperm. This dataset will expand the genomics resources available for coconut and provide a foundation for more detailed analyses that may assist molecular breeding strategies aimed at improving this major tropical crop. Copyright © 2014 Huang et al.

  15. Sequencing and de novo analysis of a coral larval transcriptome using 454 GSFlx

    Directory of Open Access Journals (Sweden)

    Colbourne John K

    2009-05-01

    Full Text Available Abstract Background New methods are needed for genomic-scale analysis of emerging model organisms that exemplify important biological questions but lack fully sequenced genomes. For example, there is an urgent need to understand the potential for corals to adapt to climate change, but few molecular resources are available for studying these processes in reef-building corals. To facilitate genomics studies in corals and other non-model systems, we describe methods for transcriptome sequencing using 454, as well as strategies for assembling a useful catalog of genes from the output. We have applied these methods to sequence the transcriptome of planulae larvae from the coral Acropora millepora. Results More than 600,000 reads produced in a single 454 sequencing run were assembled into ~40,000 contigs with five-fold average sequencing coverage. Based on sequence similarity with known proteins, these analyses identified ~11,000 different genes expressed in a range of conditions including thermal stress and settlement induction. Assembled sequences were annotated with gene names, conserved domains, and Gene Ontology terms. Targeted searches using these annotations identified the majority of genes associated with essential metabolic pathways and conserved signaling pathways, as well as novel candidate genes for stress-related processes. Comparisons with the genome of the anemone Nematostella vectensis revealed ~8,500 pairs of orthologs and ~100 candidate coral-specific genes. More than 30,000 SNPs were detected in the coral sequences, and a subset of these validated by re-sequencing. Conclusion The methods described here for deep sequencing of the transcriptome should be widely applicable to generate catalogs of genes and genetic markers in emerging model organisms. Our data provide the most comprehensive sequence resource currently available for reef-building corals, and include an extensive collection of potential genetic markers for association and

  16. Genome sequence of the olive tree, Olea europaea.

    Science.gov (United States)

    Cruz, Fernando; Julca, Irene; Gómez-Garrido, Jèssica; Loska, Damian; Marcet-Houben, Marina; Cano, Emilio; Galán, Beatriz; Frias, Leonor; Ribeca, Paolo; Derdak, Sophia; Gut, Marta; Sánchez-Fernández, Manuel; García, Jose Luis; Gut, Ivo G; Vargas, Pablo; Alioto, Tyler S; Gabaldón, Toni

    2016-06-27

    The Mediterranean olive tree (Olea europaea subsp. europaea) was one of the first trees to be domesticated and is currently of major agricultural importance in the Mediterranean region as the source of olive oil. The molecular bases underlying the phenotypic differences among domesticated cultivars, or between domesticated olive trees and their wild relatives, remain poorly understood. Both wild and cultivated olive trees have 46 chromosomes (2n). A total of 543 Gb of raw DNA sequence from whole genome shotgun sequencing, and a fosmid library containing 155,000 clones from a 1,000+ year-old olive tree (cv. Farga) were generated by Illumina sequencing using different combinations of mate-pair and pair-end libraries. Assembly gave a final genome with a scaffold N50 of 443 kb, and a total length of 1.31 Gb, which represents 95 % of the estimated genome length (1.38 Gb). In addition, the associated fungus Aureobasidium pullulans was partially sequenced. Genome annotation, assisted by RNA sequencing from leaf, root, and fruit tissues at various stages, resulted in 56,349 unique protein coding genes, suggesting recent genomic expansion. Genome completeness, as estimated using the CEGMA pipeline, reached 98.79 %. The assembled draft genome of O. europaea will provide a valuable resource for the study of the evolution and domestication processes of this important tree, and allow determination of the genetic bases of key phenotypic traits. Moreover, it will enhance breeding programs and the formation of new varieties.

  17. A hybrid reference-guided de novo assembly approach for generating Cyclospora mitochondrion genomes.

    Science.gov (United States)

    Gopinath, G R; Cinar, H N; Murphy, H R; Durigan, M; Almeria, M; Tall, B D; DaSilva, A J

    2018-01-01

    Cyclospora cayetanensis is a coccidian parasite associated with large and complex foodborne outbreaks worldwide. Linking samples from cyclosporiasis patients during foodborne outbreaks with suspected contaminated food sources, using conventional epidemiological methods, has been a persistent challenge. To address this issue, development of new methods based on potential genomically-derived markers for strain-level identification has been a priority for the food safety research community. The absence of reference genomes to identify nucleotide and structural variants with a high degree of confidence has limited the application of using sequencing data for source tracking during outbreak investigations. In this work, we determined the quality of a high resolution, curated, public mitochondrial genome assembly to be used as a reference genome by applying bioinformatic analyses. Using this reference genome, three new mitochondrial genome assemblies were built starting with metagenomic reads generated by sequencing DNA extracted from oocysts present in stool samples from cyclosporiasis patients. Nucleotide variants were identified in the new and other publicly available genomes in comparison with the mitochondrial reference genome. A consolidated workflow, presented here, to generate new mitochondrion genomes using our reference-guided de novo assembly approach could be useful in facilitating the generation of other mitochondrion sequences, and in their application for subtyping C. cayetanensis strains during foodborne outbreak investigations.

  18. An accurate clone-based haplotyping method by overlapping pool sequencing.

    Science.gov (United States)

    Li, Cheng; Cao, Changchang; Tu, Jing; Sun, Xiao

    2016-07-08

    Chromosome-long haplotyping of human genomes is important to identify genetic variants with differing gene expression, in human evolution studies, clinical diagnosis, and other biological and medical fields. Although several methods have realized haplotyping based on sequencing technologies or population statistics, accuracy and cost are factors that prohibit their wide use. Borrowing ideas from group testing theories, we proposed a clone-based haplotyping method by overlapping pool sequencing. The clones from a single individual were pooled combinatorially and then sequenced. According to the distinct pooling pattern for each clone in the overlapping pool sequencing, alleles for the recovered variants could be assigned to their original clones precisely. Subsequently, the clone sequences could be reconstructed by linking these alleles accordingly and assembling them into haplotypes with high accuracy. To verify the utility of our method, we constructed 130 110 clones in silico for the individual NA12878 and simulated the pooling and sequencing process. Ultimately, 99.9% of variants on chromosome 1 that were covered by clones from both parental chromosomes were recovered correctly, and 112 haplotype contigs were assembled with an N50 length of 3.4 Mb and no switch errors. A comparison with current clone-based haplotyping methods indicated our method was more accurate. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Deep sequencing as a method of typing bluetongue virus isolates.

    Science.gov (United States)

    Rao, Pavuluri Panduranga; Reddy, Yella Narasimha; Ganesh, Kapila; Nair, Shreeja G; Niranjan, Vidya; Hegde, Nagendra R

    2013-11-01

    Bluetongue (BT) is an economically important endemic disease of livestock in tropics and subtropics. In addition, its recent spread to temperate regions like North America and Northern Europe is of serious concern. Rapid serotyping and characterization of BT virus (BTV) is an essential step in the identification of origin of the virus and for controlling the disease. Serotyping of BTV is typically performed by serum neutralization, and of late by nucleotide sequencing. This report describes the near complete genome sequencing and typing of two isolates of BTV using Illumina next generation sequencing platform. Two of the BTV RNAs were multiplexed with ten other unknown samples. Viral RNA was isolated and fragmented, reverse transcribed, the cDNA ends were repaired and ligated with a multiplex oligo. The genome library was amplified using primers complementary to the ligated oligo and subjected to single and paired end sequencing. The raw reads were assembled using a de novo method and reference-based assembly was performed based on the contig data. Near complete sequences of all segments of BTV were obtained with more than 20× coverage, and single read sequencing method was sufficient to identify the genotype and serotype of the virus. The two viruses used in this study were typed as BTV-1 and BTV-9E. Copyright © 2013 Elsevier B.V. All rights reserved.

  20. Comparing and Evaluating Metagenome Assembly Tools from a Microbiologist's Perspective - Not Only Size Matters!

    Directory of Open Access Journals (Sweden)

    John Vollmers

    Full Text Available With the constant improvement in cost-efficiency and quality of Next Generation Sequencing technologies, shotgun-sequencing approaches -such as metagenomics- have nowadays become the methods of choice for studying and classifying microorganisms from various habitats. The production of data has dramatically increased over the past years and processing and analysis steps are becoming more and more of a bottleneck. Limiting factors are partly the availability of computational resources, but mainly the bioinformatics expertise in establishing and applying appropriate processing and analysis pipelines. Fortunately, a large diversity of specialized software tools is nowadays available. Nevertheless, choosing the most appropriate methods for answering specific biological questions can be rather challenging, especially for non-bioinformaticians. In order to provide a comprehensive overview and guide for the microbiological scientific community, we assessed the most common and freely available metagenome assembly tools with respect to their output statistics, their sensitivity for low abundant community members and variability in resulting community profiles as well as their ease-of-use. In contrast to the highly anticipated "Critical Assessment of Metagenomic Interpretation" (CAMI challenge, which uses general mock community-based assembler comparison we here tested assemblers on real Illumina metagenome sequencing data from natural communities of varying complexity sampled from forest soil and algal biofilms. Our observations clearly demonstrate that different assembly tools can prove optimal, depending on the sample type, available computational resources and, most importantly, the specific research goal. In addition, we present detailed descriptions of the underlying principles and pitfalls of publically available assembly tools from a microbiologist's perspective, and provide guidance regarding the user-friendliness, sensitivity and reliability of

  1. Metrology for ITER Assembly

    International Nuclear Information System (INIS)

    Bogusch, E.

    2006-01-01

    The overall dimensions of the ITER Tokamak and the particular assembly sequence preclude the use of conventional optical metrology, mechanical jigs and traditional dimensional control equipment, as used for the assembly of smaller, previous generation, fusion devices. This paper describes the state of the art of the capabilities of available metrology systems, with reference to the previous experience in Fusion engineering and in other industries. Two complementary procedures of transferring datum from the primary datum network on the bioshield to the secondary datum s inside the VV with the desired accuracy of about 0.1 mm is described, one method using the access directly through the ports and the other using transfer techniques, developed during the co-operation with ITER/EFDA. Another important task described is the development of a method for the rapid and easy measurement of the gaps between sectors, required for the production of the customised splice plates between them. The scope of the paper includes the evaluation of the composition and cost of the systems and team of technical staff required to meet the requirements of the assembly procedure. The results from a practical, full-scale demonstration of the methodologies used, using the proposed equipment, is described. This work has demonstrated the feasibility of achieving the necessary accuracies for the successful building of ITER. (author)

  2. Light whole genome sequence for SNP discovery across domestic cat breeds

    Directory of Open Access Journals (Sweden)

    Driscoll Carlos

    2010-06-01

    Full Text Available Abstract Background The domestic cat has offered enormous genomic potential in the veterinary description of over 250 hereditary disease models as well as the occurrence of several deadly feline viruses (feline leukemia virus -- FeLV, feline coronavirus -- FECV, feline immunodeficiency virus - FIV that are homologues to human scourges (cancer, SARS, and AIDS respectively. However, to realize this bio-medical potential, a high density single nucleotide polymorphism (SNP map is required in order to accomplish disease and phenotype association discovery. Description To remedy this, we generated 3,178,297 paired fosmid-end Sanger sequence reads from seven cats, and combined these data with the publicly available 2X cat whole genome sequence. All sequence reads were assembled together to form a 3X whole genome assembly allowing the discovery of over three million SNPs. To reduce potential false positive SNPs due to the low coverage assembly, a low upper-limit was placed on sequence coverage and a high lower-limit on the quality of the discrepant bases at a potential variant site. In all domestic cats of different breeds: female Abyssinian, female American shorthair, male Cornish Rex, female European Burmese, female Persian, female Siamese, a male Ragdoll and a female African wildcat were sequenced lightly. We report a total of 964 k common SNPs suitable for a domestic cat SNP genotyping array and an additional 900 k SNPs detected between African wildcat and domestic cats breeds. An empirical sampling of 94 discovered SNPs were tested in the sequenced cats resulting in a SNP validation rate of 99%. Conclusions These data provide a large collection of mapped feline SNPs across the cat genome that will allow for the development of SNP genotyping platforms for mapping feline diseases.

  3. The War in the North Sea. Volume I. From the Beginning of the War to the First of September, 1914. Part 2. Chapters 5,6; Appendices 14-21; Charts 18-35

    Science.gov (United States)

    1937-01-01

    destroyers of the Iliver type, the assiGirient of the orulsers "Undynion" and ’" Theseus " fron Cruiser ,>uadron XI to ".’, and several other secondary...ton t Hawk« t Royal «rthur ♦ Theseus (X Shortly after outbreak of «ar cruiser squadron .; assigned to Grand fleet and used in Jdockade duty

  4. MATAM: reconstruction of phylogenetic marker genes from short sequencing reads in metagenomes.

    Science.gov (United States)

    Pericard, Pierre; Dufresne, Yoann; Couderc, Loïc; Blanquart, Samuel; Touzet, Hélène

    2018-02-15

    Advances in the sequencing of uncultured environmental samples, dubbed metagenomics, raise a growing need for accurate taxonomic assignment. Accurate identification of organisms present within a community is essential to understanding even the most elementary ecosystems. However, current high-throughput sequencing technologies generate short reads which partially cover full-length marker genes and this poses difficult bioinformatic challenges for taxonomy identification at high resolution. We designed MATAM, a software dedicated to the fast and accurate targeted assembly of short reads sequenced from a genomic marker of interest. The method implements a stepwise process based on construction and analysis of a read overlap graph. It is applied to the assembly of 16S rRNA markers and is validated on simulated, synthetic and genuine metagenomes. We show that MATAM outperforms other available methods in terms of low error rates and recovered fractions and is suitable to provide improved assemblies for precise taxonomic assignments. https://github.com/bonsai-team/matam. pierre.pericard@gmail.com or helene.touzet@univ-lille1.fr. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  5. Whole-Genome de novo Sequencing Of Quail And Grey Partridge

    DEFF Research Database (Denmark)

    Holm, Lars-Erik; Panitz, Frank; Burt, Dave

    2011-01-01

    The development in sequencing methods has made it possible to perform whole genome de novo sequencing of species without large commercial interests. Within the EU-financed QUANTOMICS project (KBBE-2A-222664), we have performed de novo sequencing of quail (Coturnix coturnix) and grey partridge...... (Perdix perdix) on a Genome Analyzer GAII (Illumina) using paired-end sequencing. The amount of generated sequences amounts to 8 to 9 Gb for each species. The analysis and assembly of the generated sequences is ongoing. Access to the whole genome sequence from these two species will enable enhanced...... comparative studies towards the chicken genome and will aid in identifying evolutionarily conserved sequences within the Galliformes. The obtained sequences from quail and partridge represent a beginning of generating the whole genome sequence for these species. The continuation of establishing the genome...

  6. Development of a Feature and Template-Assisted Assembler and Application to the Analysis of a Foot-and-Mouth Disease Virus Genotyping Microarray.

    Directory of Open Access Journals (Sweden)

    Roger W Barrette

    Full Text Available Several RT-PCR and genome sequencing strategies exist for the resolution of Foot-and-Mouth Disease virus (FMDV. While these approaches are relatively straightforward, they can be vulnerable to failure due to the unpredictable nature of FMDV genome sequence variations. Sequence independent single primer amplification (SISPA followed by genotyping microarray offers an attractive unbiased approach to FMDV characterization. Here we describe a custom FMDV microarray and a companion feature and template-assisted assembler software (FAT-assembler capable of resolving virus genome sequence using a moderate number of conserved microarray features. The results demonstrate that this approach may be used to rapidly characterize naturally occurring FMDV as well as an engineered chimeric strain of FMDV. The FAT-assembler, while applied to resolving FMDV genomes, represents a new bioinformatics approach that should be broadly applicable to interpreting microarray genotyping data for other viruses or target organisms.

  7. Meraculous: De Novo Genome Assembly with Short Paired-End Reads

    Energy Technology Data Exchange (ETDEWEB)

    Chapman, Jarrod A.; Ho, Isaac; Sunkara, Sirisha; Luo, Shujun; Schroth, Gary P.; Rokhsar, Daniel S.; Salzberg, Steven L.

    2011-08-18

    We describe a new algorithm, meraculous, for whole genome assembly of deep paired-end short reads, and apply it to the assembly of a dataset of paired 75-bp Illumina reads derived from the 15.4 megabase genome of the haploid yeast Pichia stipitis. More than 95% of the genome is recovered, with no errors; half the assembled sequence is in contigs longer than 101 kilobases and in scaffolds longer than 269 kilobases. Incorporating fosmid ends recovers entire chromosomes. Meraculous relies on an efficient and conservative traversal of the subgraph of the k-mer (deBruijn) graph of oligonucleotides with unique high quality extensions in the dataset, avoiding an explicit error correction step as used in other short-read assemblers. A novel memory-efficient hashing scheme is introduced. The resulting contigs are ordered and oriented using paired reads separated by ~280 bp or ~3.2 kbp, and many gaps between contigs can be closed using paired-end placements. Practical issues with the dataset are described, and prospects for assembling larger genomes are discussed.

  8. Proton-Fueled, Reversible DNA Hybridization Chain Assembly for pH Sensing and Imaging.

    Science.gov (United States)

    Liu, Lan; Liu, Jin-Wen; Huang, Zhi-Mei; Wu, Han; Li, Na; Tang, Li-Juan; Jiang, Jian-Hui

    2017-07-05

    Design of DNA self-assembly with reversible responsiveness to external stimuli is of great interest for diverse applications. We for the first time develop a pH-responsive, fully reversible hybridization chain reaction (HCR) assembly that allows sensitive sensing and imaging of pH in living cells. Our design relies on the triplex forming sequences that form DNA triplex with toehold regions under acidic conditions and then induce a cascade of strand displacement and DNA assembly. The HCR assembly has shown dynamic responses in physiological pH ranges with excellent reversibility and demonstrated the potential for in vitro detection and live-cell imaging of pH. Moreover, this method affords HCR assemblies with highly localized fluorescence responses, offering advantages of improving sensitivity and better selectivity. The proton-fueled, reversible HCR assembly may provide a useful approach for pH-related cell biology study and disease diagnostics.

  9. Draft genome sequence of the silver pomfret fish, Pampus argenteus.

    Science.gov (United States)

    AlMomin, Sabah; Kumar, Vinod; Al-Amad, Sami; Al-Hussaini, Mohsen; Dashti, Talal; Al-Enezi, Khaznah; Akbar, Abrar

    2016-01-01

    Silver pomfret, Pampus argenteus, is a fish species from coastal waters. Despite its high commercial value, this edible fish has not been sequenced. Hence, its genetic and genomic studies have been limited. We report the first draft genome sequence of the silver pomfret obtained using a Next Generation Sequencing (NGS) technology. We assembled 38.7 Gb of nucleotides into scaffolds of 350 Mb with N50 of about 1.5 kb, using high quality paired end reads. These scaffolds represent 63.7% of the estimated silver pomfret genome length. The newly sequenced and assembled genome has 11.06% repetitive DNA regions, and this percentage is comparable to that of the tilapia genome. The genome analysis predicted 16 322 genes. About 91% of these genes showed homology with known proteins. Many gene clusters were annotated to protein and fatty-acid metabolism pathways that may be important in the context of the meat texture and immune system developmental processes. The reference genome can pave the way for the identification of many other genomic features that could improve breeding and population-management strategies, and it can also help characterize the genetic diversity of P. argenteus.

  10. Complete Sequence and Analysis of Coconut Palm (Cocos nucifera) Mitochondrial Genome.

    Science.gov (United States)

    Aljohi, Hasan Awad; Liu, Wanfei; Lin, Qiang; Zhao, Yuhui; Zeng, Jingyao; Alamer, Ali; Alanazi, Ibrahim O; Alawad, Abdullah O; Al-Sadi, Abdullah M; Hu, Songnian; Yu, Jun

    2016-01-01

    Coconut (Cocos nucifera L.), a member of the palm family (Arecaceae), is one of the most economically important crops in tropics, serving as an important source of food, drink, fuel, medicine, and construction material. Here we report an assembly of the coconut (C. nucifera, Oman local Tall cultivar) mitochondrial (mt) genome based on next-generation sequencing data. This genome, 678,653bp in length and 45.5% in GC content, encodes 72 proteins, 9 pseudogenes, 23 tRNAs, and 3 ribosomal RNAs. Within the assembly, we find that the chloroplast (cp) derived regions account for 5.07% of the total assembly length, including 13 proteins, 2 pseudogenes, and 11 tRNAs. The mt genome has a relatively large fraction of repeat content (17.26%), including both forward (tandem) and inverted (palindromic) repeats. Sequence variation analysis shows that the Ti/Tv ratio of the mt genome is lower as compared to that of the nuclear genome and neutral expectation. By combining public RNA-Seq data for coconut, we identify 734 RNA editing sites supported by at least two datasets. In summary, our data provides the second complete mt genome sequence in the family Arecaceae, essential for further investigations on mitochondrial biology of seed plants.

  11. Human Missions to Europa and Titan - Why Not?

    Science.gov (United States)

    Finarelli, Margaret G.

    2004-04-01

    This report describes a long-term development plan to enable human exploration of the outer solar system, with a focus on Europa and Titan. These are two of the most interesting moons of Jupiter and Saturn, respectively, because they are the places in the solar system with the greatest potential for harboring extraterrestrial life. Since human expeditions to these worlds are considered impossible with current capabilities, the proposal of a well-organized sequence of steps towards making this a reality was formulated. The proposed Development Plan, entitled Theseus, is the outcome of a recent multinational study by a group of students in the framework of the Master of Space Studies (MSS) 2004 course at the International Space University (ISU). The Theseus Program includes the necessary development strategies in key scientific and technological areas that are essential for identifying the requirements for the exploration of the outer planetary moons. Some of the topics that are analysed throughout the plan include: scientific observations at Europa and Titan, advanced propulsion and nuclear power systems, in-situ resource utilization, radiation mitigation techniques, closed life support systems, habitation for long-term spaceflight, and artificial gravity. In addition to the scientific and technological aspects of the Theseus Program, it was recognized that before any research and development work may begin, some level of program management must be established. Within this chapter, legal issues, national and international policy, motivation, organization and management, economic considerations, outreach, education, ethics, and social implications are all considered with respect to four possible future scenarios which enable human missions to the outer solar system. The final chapter of the report builds upon the foundations set by Theseus through a case study. This study illustrates how such accomplishments could influence a mission to Europa to search for evidence

  12. Human Missions to Europa and Titan - Why Not?

    Science.gov (United States)

    2004-01-01

    This report describes a long-term development plan to enable human exploration of the outer solar system, with a focus on Europa and Titan. These are two of the most interesting moons of Jupiter and Saturn, respectively, because they are the places in the solar system with the greatest potential for harboring extraterrestrial life. Since human expeditions to these worlds are considered impossible with current capabilities, the proposal of a well-organized sequence of steps towards making this a reality was formulated. The proposed Development Plan, entitled Theseus, is the outcome of a recent multinational study by a group of students in the framework of the Master of Space Studies (MSS) 2004 course at the International Space University (ISU). The Theseus Program includes the necessary development strategies in key scientific and technological areas that are essential for identifying the requirements for the exploration of the outer planetary moons. Some of the topics that are analysed throughout the plan include: scientific observations at Europa and Titan, advanced propulsion and nuclear power systems, in-situ resource utilization, radiation mitigation techniques, closed life support systems, habitation for long-term spaceflight, and artificial gravity. In addition to the scientific and technological aspects of the Theseus Program, it was recognized that before any research and development work may begin, some level of program management must be established. Within this chapter, legal issues, national and international policy, motivation, organization and management, economic considerations, outreach, education, ethics, and social implications are all considered with respect to four possible future scenarios which enable human missions to the outer solar system. The final chapter of the report builds upon the foundations set by Theseus through a case study. This study illustrates how such accomplishments could influence a mission to Europa to search for evidence

  13. Large-scale parallel genome assembler over cloud computing environment.

    Science.gov (United States)

    Das, Arghya Kusum; Koppa, Praveen Kumar; Goswami, Sayan; Platania, Richard; Park, Seung-Jong

    2017-06-01

    The size of high throughput DNA sequencing data has already reached the terabyte scale. To manage this huge volume of data, many downstream sequencing applications started using locality-based computing over different cloud infrastructures to take advantage of elastic (pay as you go) resources at a lower cost. However, the locality-based programming model (e.g. MapReduce) is relatively new. Consequently, developing scalable data-intensive bioinformatics applications using this model and understanding the hardware environment that these applications require for good performance, both require further research. In this paper, we present a de Bruijn graph oriented Parallel Giraph-based Genome Assembler (GiGA), as well as the hardware platform required for its optimal performance. GiGA uses the power of Hadoop (MapReduce) and Giraph (large-scale graph analysis) to achieve high scalability over hundreds of compute nodes by collocating the computation and data. GiGA achieves significantly higher scalability with competitive assembly quality compared to contemporary parallel assemblers (e.g. ABySS and Contrail) over traditional HPC cluster. Moreover, we show that the performance of GiGA is significantly improved by using an SSD-based private cloud infrastructure over traditional HPC cluster. We observe that the performance of GiGA on 256 cores of this SSD-based cloud infrastructure closely matches that of 512 cores of traditional HPC cluster.

  14. The genome sequence of a widespread apex predator, the golden eagle (Aquila chrysaetos.

    Directory of Open Access Journals (Sweden)

    Jacqueline M Doyle

    Full Text Available Biologists routinely use molecular markers to identify conservation units, to quantify genetic connectivity, to estimate population sizes, and to identify targets of selection. Many imperiled eagle populations require such efforts and would benefit from enhanced genomic resources. We sequenced, assembled, and annotated the first eagle genome using DNA from a male golden eagle (Aquila chrysaetos captured in western North America. We constructed genomic libraries that were sequenced using Illumina technology and assembled the high-quality data to a depth of ∼40x coverage. The genome assembly includes 2,552 scaffolds >10 Kb and 415 scaffolds >1.2 Mb. We annotated 16,571 genes that are involved in myriad biological processes, including such disparate traits as beak formation and color vision. We also identified repetitive regions spanning 92 Mb (∼6% of the assembly, including LINES, SINES, LTR-RTs and DNA transposons. The mitochondrial genome encompasses 17,332 bp and is ∼91% identical to the Mountain Hawk-Eagle (Nisaetus nipalensis. Finally, the data reveal that several anonymous microsatellites commonly used for population studies are embedded within protein-coding genes and thus may not have evolved in a neutral fashion. Because the genome sequence includes ∼800,000 novel polymorphisms, markers can now be chosen based on their proximity to functional genes involved in migration, carnivory, and other biological processes.

  15. Getting complete genomes from complex samples using nanopore sequencing

    DEFF Research Database (Denmark)

    Kirkegaard, Rasmus Hansen; Karst, Søren Michael; Albertsen, Mads

    Short read sequencing and metagenomic binning workflows have made it possible to extract bacterial genome bins from environmental microbial samples containing hundreds to thousands of different species. However, these genome bins often do not represent complete genomes, as they are mostly...... fragmented, incomplete and often contaminated with foreign DNA and with no robust strategies to validate the quality. The value of these `draft genomes` have limited, lasting value to the scientific community, as gene synteny is broken and the uncertainty of what is missing. The genetic material most often...... missed is important multi-copy and/or conserved marker genes such as the 16S rRNA gene, as sequence micro-heterogeneity prevents assembly of these genes in the de novo assembly. We demonstrate that using nanopore long reads it is now possible to overcome these issues and make complete genomes from...

  16. DeepSimulator: a deep simulator for Nanopore sequencing

    KAUST Repository

    Li, Yu

    2017-12-23

    Motivation: Oxford Nanopore sequencing is a rapidly developed sequencing technology in recent years. To keep pace with the explosion of the downstream data analytical tools, a versatile Nanopore sequencing simulator is needed to complement the experimental data as well as to benchmark those newly developed tools. However, all the currently available simulators are based on simple statistics of the produced reads, which have difficulty in capturing the complex nature of the Nanopore sequencing procedure, the main task of which is the generation of raw electrical current signals. Results: Here we propose a deep learning based simulator, DeepSimulator, to mimic the entire pipeline of Nanopore sequencing. Starting from a given reference genome or assembled contigs, we simulate the electrical current signals by a context-dependent deep learning model, followed by a base-calling procedure to yield simulated reads. This workflow mimics the sequencing procedure more naturally. The thorough experiments performed across four species show that the signals generated by our context-dependent model are more similar to the experimentally obtained signals than the ones generated by the official context-independent pore model. In terms of the simulated reads, we provide a parameter interface to users so that they can obtain the reads with different accuracies ranging from 83% to 97%. The reads generated by the default parameter have almost the same properties as the real data. Two case studies demonstrate the application of DeepSimulator to benefit the development of tools in de novo assembly and in low coverage SNP detection. Availability: The software can be accessed freely at: https://github.com/lykaust15/DeepSimulator.

  17. Focal plane AIT sequence: evolution from HRG-Spot 5 to Pleiades HR

    Science.gov (United States)

    Le Goff, Roland; Pranyies, Pascal; Toubhans, Isabelle

    2017-11-01

    Optical and geometrical image qualities of Focal Planes, for "push-broom" high resolution remote sensing satellites, require the implementation of specific means and methods for the AIT sequence. Indeed the geometric performances of the focal plane mainly axial focusing and transverse registration, are duly obtained on the basis of adjustment, setting and measurement of optical and CCD components with an accuracy of a few microns. Since the end of the 1970s, EADS-SODERN has developed a series of detection units for earth observation instruments like SPOT and Helios. And EADS-SODERN is now responsible for the development of the Pleiades High Resolution Focal Plane assembly. This paper presents the AIT sequences. We introduce all the efforts, innovative solutions and improvements made on the assembly facilities to match the technical evolutions and breakthrough of the Pleiades HR FP concept in comparison with the previous High Resolution Geometric SPOT 5 Focal Plane. The main evolution drivers are the implementation of strip filters and the realization of 400 mm continuous retinas. For Pleiades HR AIT sequence, three specific integration and measuring benches, corresponding with the different assembly stages, are used: a 3-D non-contact measurement machine for the assembly of detection module, a 3-D measurement machine for mirror integration on the main Focal Plane SiC structure, and a 3-D geometric coordinates control bench to focus detection module lines and to ensure they are well registered together.

  18. Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants

    Science.gov (United States)

    Machens, Fabian; Coll, Anna; Baebler, Špela; Messerschmidt, Katrin; Gruden, Kristina

    2018-01-01

    Cloning multiple DNA fragments for delivery of several genes of interest into the plant genome is one of the main technological challenges in plant synthetic biology. Despite several modular assembly methods developed in recent years, the plant biotechnology community has not widely adopted them yet, probably due to the lack of appropriate vectors and software tools. Here we present Plant X-tender, an extension of the highly efficient, scar-free and sequence-independent multigene assembly strategy AssemblX, based on overlap-depended cloning methods and rare-cutting restriction enzymes. Plant X-tender consists of a set of plant expression vectors and the protocols for most efficient cloning into the novel vector set needed for plant expression and thus introduces advantages of AssemblX into plant synthetic biology. The novel vector set covers different backbones and selection markers to allow full design flexibility. We have included ccdB counterselection, thereby allowing the transfer of multigene constructs into the novel vector set in a straightforward and highly efficient way. Vectors are available as empty backbones and are fully flexible regarding the orientation of expression cassettes and addition of linkers between them, if required. We optimised the assembly and subcloning protocol by testing different scar-less assembly approaches: the noncommercial SLiCE and TAR methods and the commercial Gibson assembly and NEBuilder HiFi DNA assembly kits. Plant X-tender was applicable even in combination with low efficient homemade chemically competent or electrocompetent Escherichia coli. We have further validated the developed procedure for plant protein expression by cloning two cassettes into the newly developed vectors and subsequently transferred them to Nicotiana benthamiana in a transient expression setup. Thereby we show that multigene constructs can be delivered into plant cells in a streamlined and highly efficient way. Our results will support faster

  19. Plant X-tender: An extension of the AssemblX system for the assembly and expression of multigene constructs in plants.

    Science.gov (United States)

    Lukan, Tjaša; Machens, Fabian; Coll, Anna; Baebler, Špela; Messerschmidt, Katrin; Gruden, Kristina

    2018-01-01

    Cloning multiple DNA fragments for delivery of several genes of interest into the plant genome is one of the main technological challenges in plant synthetic biology. Despite several modular assembly methods developed in recent years, the plant biotechnology community has not widely adopted them yet, probably due to the lack of appropriate vectors and software tools. Here we present Plant X-tender, an extension of the highly efficient, scar-free and sequence-independent multigene assembly strategy AssemblX, based on overlap-depended cloning methods and rare-cutting restriction enzymes. Plant X-tender consists of a set of plant expression vectors and the protocols for most efficient cloning into the novel vector set needed for plant expression and thus introduces advantages of AssemblX into plant synthetic biology. The novel vector set covers different backbones and selection markers to allow full design flexibility. We have included ccdB counterselection, thereby allowing the transfer of multigene constructs into the novel vector set in a straightforward and highly efficient way. Vectors are available as empty backbones and are fully flexible regarding the orientation of expression cassettes and addition of linkers between them, if required. We optimised the assembly and subcloning protocol by testing different scar-less assembly approaches: the noncommercial SLiCE and TAR methods and the commercial Gibson assembly and NEBuilder HiFi DNA assembly kits. Plant X-tender was applicable even in combination with low efficient homemade chemically competent or electrocompetent Escherichia coli. We have further validated the developed procedure for plant protein expression by cloning two cassettes into the newly developed vectors and subsequently transferred them to Nicotiana benthamiana in a transient expression setup. Thereby we show that multigene constructs can be delivered into plant cells in a streamlined and highly efficient way. Our results will support faster

  20. A simple and efficient method for assembling TALE protein based on plasmid library.

    Science.gov (United States)

    Zhang, Zhiqiang; Li, Duo; Xu, Huarong; Xin, Ying; Zhang, Tingting; Ma, Lixia; Wang, Xin; Chen, Zhilong; Zhang, Zhiying

    2013-01-01

    DNA binding domain of the transcription activator-like effectors (TALEs) from Xanthomonas sp. consists of tandem repeats that can be rearranged according to a simple cipher to target new DNA sequences with high DNA-binding specificity. This technology has been successfully applied in varieties of species for genome engineering. However, assembling long TALE tandem repeats remains a big challenge precluding wide use of this technology. Although several new methodologies for efficiently assembling TALE repeats have been recently reported, all of them require either sophisticated facilities or skilled technicians to carry them out. Here, we described a simple and efficient method for generating customized TALE nucleases (TALENs) and TALE transcription factors (TALE-TFs) based on TALE repeat tetramer library. A tetramer library consisting of 256 tetramers covers all possible combinations of 4 base pairs. A set of unique primers was designed for amplification of these tetramers. PCR products were assembled by one step of digestion/ligation reaction. 12 TALE constructs including 4 TALEN pairs targeted to mouse Gt(ROSA)26Sor gene and mouse Mstn gene sequences as well as 4 TALE-TF constructs targeted to mouse Oct4, c-Myc, Klf4 and Sox2 gene promoter sequences were generated by using our method. The construction routines took 3 days and parallel constructions were available. The rate of positive clones during colony PCR verification was 64% on average. Sequencing results suggested that all TALE constructs were performed with high successful rate. This is a rapid and cost-efficient method using the most common enzymes and facilities with a high success rate.

  1. A versatile system for USER cloning-based assembly of expression vectors for mammalian cell engineering.

    Directory of Open Access Journals (Sweden)

    Anne Mathilde Lund

    Full Text Available A new versatile mammalian vector system for protein production, cell biology analyses, and cell factory engineering was developed. The vector system applies the ligation-free uracil-excision based technique--USER cloning--to rapidly construct mammalian expression vectors of multiple DNA fragments and with maximum flexibility, both for choice of vector backbone and cargo. The vector system includes a set of basic vectors and a toolbox containing a multitude of DNA building blocks including promoters, terminators, selectable marker- and reporter genes, and sequences encoding an internal ribosome entry site, cellular localization signals and epitope- and purification tags. Building blocks in the toolbox can be easily combined as they contain defined and tested Flexible Assembly Sequence Tags, FASTs. USER cloning with FASTs allows rapid swaps of gene, promoter or selection marker in existing plasmids and simple construction of vectors encoding proteins, which are fused to fluorescence-, purification-, localization-, or epitope tags. The mammalian expression vector assembly platform currently allows for the assembly of up to seven fragments in a single cloning step with correct directionality and with a cloning efficiency above 90%. The functionality of basic vectors for FAST assembly was tested and validated by transient expression of fluorescent model proteins in CHO, U-2-OS and HEK293 cell lines. In this test, we included many of the most common vector elements for heterologous gene expression in mammalian cells, in addition the system is fully extendable by other users. The vector system is designed to facilitate high-throughput genome-scale studies of mammalian cells, such as the newly sequenced CHO cell lines, through the ability to rapidly generate high-fidelity assembly of customizable gene expression vectors.

  2. Historical Research and Narrative Inquiry: Striking Similarities, Notable Differences

    Science.gov (United States)

    Craig, Cheryl J.

    2005-01-01

    When Theseus sailed from Athens to the island of Crete to slay the Minotaur, a fearsome monster whose food was human flesh and whose home was the labyrinth, Ariadne, the daughter of the Cretan King Minos, gave her new found love, Theseus, a ball of thread to assist him in maneuvering his way through the great maze of winding passages. "Unwind it…

  3. De novo transcriptome assembly of two different peach cultivars grown in Korea

    Directory of Open Access Journals (Sweden)

    Yeonhwa Jo

    2015-12-01

    Full Text Available Peach (Prunus persica is one of the most popular stone fruits worldwide. Next generation sequencing (NGS has facilitated genome and transcriptome analyses of several stone fruit trees. In this study, we conducted de novo transcriptome analyses of two peach cultivars grown in Korea. Leaves of two cultivars, referred to as Jangtaek and Mibaek, were harvested and used for library preparation. The two prepared libraries were paired-end sequenced by the HiSeq2000 system. We obtained 8.14 GB and 9.62 GB sequence data from Jangtaek and Mibaek (NCBI accession numbers: SRS1056585 and SRS1056587, respectively. The Trinity program was used to assemble two transcriptomes de novo, resulting in 110,477 (Jangtaek and 136,196 (Mibaek transcripts. TransDecoder identified possible coding regions in assembled transcripts. The identified proteins were subjected to BLASTP search against NCBI's non-redundant database for functional annotation. This study provides transcriptome data for two peach cultivars, which might be useful for genetic marker development and comparative transcriptome analyses.

  4. Packaging signals in two single-stranded RNA viruses imply a conserved assembly mechanism and geometry of the packaged genome.

    Science.gov (United States)

    Dykeman, Eric C; Stockley, Peter G; Twarock, Reidun

    2013-09-09

    The current paradigm for assembly of single-stranded RNA viruses is based on a mechanism involving non-sequence-specific packaging of genomic RNA driven by electrostatic interactions. Recent experiments, however, provide compelling evidence for sequence specificity in this process both in vitro and in vivo. The existence of multiple RNA packaging signals (PSs) within viral genomes has been proposed, which facilitates assembly by binding coat proteins in such a way that they promote the protein-protein contacts needed to build the capsid. The binding energy from these interactions enables the confinement or compaction of the genomic RNAs. Identifying the nature of such PSs is crucial for a full understanding of assembly, which is an as yet untapped potential drug target for this important class of pathogens. Here, for two related bacterial viruses, we determine the sequences and locations of their PSs using Hamiltonian paths, a concept from graph theory, in combination with bioinformatics and structural studies. Their PSs have a common secondary structure motif but distinct consensus sequences and positions within the respective genomes. Despite these differences, the distributions of PSs in both viruses imply defined conformations for the packaged RNA genomes in contact with the protein shell in the capsid, consistent with a recent asymmetric structure determination of the MS2 virion. The PS distributions identified moreover imply a preferred, evolutionarily conserved assembly pathway with respect to the RNA sequence with potentially profound implications for other single-stranded RNA viruses known to have RNA PSs, including many animal and human pathogens. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. Scoring-and-unfolding trimmed tree assembler: concepts, constructs and comparisons.

    Science.gov (United States)

    Narzisi, Giuseppe; Mishra, Bud

    2011-01-15

    Mired by its connection to a well-known -complete combinatorial optimization problem-namely, the Shortest Common Superstring Problem (SCSP)-historically, the whole-genome sequence assembly (WGSA) problem has been assumed to be amenable only to greedy and heuristic methods. By placing efficiency as their first priority, these methods opted to rely only on local searches, and are thus inherently approximate, ambiguous or error prone, especially, for genomes with complex structures. Furthermore, since choice of the best heuristics depended critically on the properties of (e.g. errors in) the input data and the available long range information, these approaches hindered designing an error free WGSA pipeline. We dispense with the idea of limiting the solutions to just the approximated ones, and instead favor an approach that could potentially lead to an exhaustive (exponential-time) search of all possible layouts. Its computational complexity thus must be tamed through a constrained search (Branch-and-Bound) and quick identification and pruning of implausible overlays. For his purpose, such a method necessarily relies on a set of score functions (oracles) that can combine different structural properties (e.g. transitivity, coverage, physical maps, etc.). We give a detailed description of this novel assembly framework, referred to as Scoring-and-Unfolding Trimmed Tree Assembler (SUTTA), and present experimental results on several bacterial genomes using next-generation sequencing technology data. We also report experimental evidence that the assembly quality strongly depends on the choice of the minimum overlap parameter k. SUTTA's binaries are freely available to non-profit institutions for research and educational purposes at http://www.bioinformatics.nyu.edu.

  6. Why size really matters when sequencing plant genomes

    Czech Academy of Sciences Publication Activity Database

    Kelly, L.J.; Leitch, A.R.; Fay, M. F.; Renny-Byfield, S.; Pellicer, J.; Macas, Jiří; Leitch, I.J.

    2012-01-01

    Roč. 5, č. 4 (2012), s. 415-425 ISSN 1755-0874 Institutional research plan: CEZ:AV0Z50510513 Institutional support: RVO:60077344 Keywords : C-value * genome assembly * genome size evolution * genome sequencing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.924, year: 2012

  7. Ten steps to get started in Genome Assembly and Annotation

    Science.gov (United States)

    Dominguez Del Angel, Victoria; Hjerde, Erik; Sterck, Lieven; Capella-Gutierrez, Salvadors; Notredame, Cederic; Vinnere Pettersson, Olga; Amselem, Joelle; Bouri, Laurent; Bocs, Stephanie; Klopp, Christophe; Gibrat, Jean-Francois; Vlasova, Anna; Leskosek, Brane L.; Soler, Lucile; Binzer-Panchal, Mahesh; Lantz, Henrik

    2018-01-01

    As a part of the ELIXIR-EXCELERATE efforts in capacity building, we present here 10 steps to facilitate researchers getting started in genome assembly and genome annotation. The guidelines given are broadly applicable, intended to be stable over time, and cover all aspects from start to finish of a general assembly and annotation project. Intrinsic properties of genomes are discussed, as is the importance of using high quality DNA. Different sequencing technologies and generally applicable workflows for genome assembly are also detailed. We cover structural and functional annotation and encourage readers to also annotate transposable elements, something that is often omitted from annotation workflows. The importance of data management is stressed, and we give advice on where to submit data and how to make your results Findable, Accessible, Interoperable, and Reusable (FAIR). PMID:29568489

  8. Analysis of Litopenaeus vannamei transcriptome using the next-generation DNA sequencing technique.

    Directory of Open Access Journals (Sweden)

    Chaozheng Li

    Full Text Available BACKGROUND: Pacific white shrimp (Litopenaeus vannamei, the major species of farmed shrimps in the world, has been attracting extensive studies, which require more and more genome background knowledge. The now available transcriptome data of L. vannamei are insufficient for research requirements, and have not been adequately assembled and annotated. METHODOLOGY/PRINCIPAL FINDINGS: This is the first study that used a next-generation high-throughput DNA sequencing technique, the Solexa/Illumina GA II method, to analyze the transcriptome from whole bodies of L. vannamei larvae. More than 2.4 Gb of raw data were generated, and 109,169 unigenes with a mean length of 396 bp were assembled using the SOAP denovo software. 73,505 unigenes (>200 bp with good quality sequences were selected and subjected to annotation analysis, among which 37.80% can be matched in NCBI Nr database, 37.3% matched in Swissprot, and 44.1% matched in TrEMBL. Using BLAST and BLAST2Go softwares, 11,153 unigenes were classified into 25 Clusters of Orthologous Groups of proteins (COG categories, 8171 unigenes were assigned into 51 Gene ontology (GO functional groups, and 18,154 unigenes were divided into 220 Kyoto Encyclopedia of Genes and Genomes (KEGG pathways. To primarily verify part of the results of assembly and annotations, 12 assembled unigenes that are homologous to many embryo development-related genes were chosen and subjected to RT-PCR for electrophoresis and Sanger sequencing analyses, and to real-time PCR for expression profile analyses during embryo development. CONCLUSIONS/SIGNIFICANCE: The L. vannamei transcriptome analyzed using the next-generation sequencing technique enriches the information of L. vannamei genes, which will facilitate our understanding of the genome background of crustaceans, and promote the studies on L. vannamei.

  9. ASSEMBLY TRANSFER SYSTEM DESCRIPTION DOCUMENT

    International Nuclear Information System (INIS)

    Gorpani, B.

    2000-01-01

    The Assembly Transfer System (ATS) receives, cools, and opens rail and truck transportation casks from the Carrier/Cask Handling System (CCHS). The system unloads transportation casks consisting of bare Spent Nuclear Fuel (SNF) assemblies, single element canisters, and Dual Purpose Canisters (DPCs). For casks containing DPCs, the system opens the DPCs and unloads the SNF. The system stages the assemblies, transfer assemblies to and from fuel-blending inventory pools, loads them into Disposal Containers (DCs), temporarily seals and inerts the DC, decontaminates the DC and transfers it to the Disposal Container Handling System. The system also prepares empty casks and DPCs for off-site shipment. Two identical Assembly Transfer System lines are provided in the Waste Handling Building (WHB). Each line operates independently to handle the waste transfer throughput and to support maintenance operations. Each system line primarily consists of wet and dry handling areas. The wet handling area includes a cask transport system, cask and DPC preparation system, and a wet assembly handling system. The basket transport system forms the transition between the wet and dry handling areas. The dry handling area includes the dry assembly handling system, assembly drying system, DC preparation system, and DC transport system. Both the wet and dry handling areas are controlled by the control and tracking system. The system operating sequence begins with moving transportation casks to the cask preparation area. The cask preparation operations consist of cask cavity gas sampling, cask venting, cask cool-down, outer lid removal, and inner shield plug lifting fixture attachment. Casks containing bare SNF (no DPC) are filled with water and placed in the cask unloading pool. The inner shield plugs are removed underwater. For casks containing a DPC, the cask lid(s) is removed, and the DPC is penetrated, sampled, vented, and cooled. A DPC lifting fixture is attached and the cask is placed

  10. Potential for containment leak paths through electrical penetration assemblies under severe accident conditions

    International Nuclear Information System (INIS)

    Sebrell, W.

    1983-07-01

    The leakage behavior of containments beyond design conditions and knowledge of failure modes is required for evaluation of mitigation strategies for severe accidents, risk studies, emergency preparedness planning, and siting. These studies are directed towards assessing the risk and consequences of severe accidents. An accident sequence analysis conducted on a Boiling Water Reactor (BWR), Mark I (MK I), indicated very high temperatures in the dry-well region, which is the location of the majority of electrical penetration assemblies. Because of the high temperatures, it was postulated in the ORNL study that the sealants would fail and all the electrical penetration assemblies would leak before structural failure would occur. Since other containments had similar electrical penetration assemblies, it was concluded that all containments would experience the same type of failure. The results of this study, however, show that this conclusion does not hold for PWRs because in the worst accident sequence, the long time containment gases stabilize to 350 0 F. BWRs, on the other hand, do experience high dry-well temperatures and have a higher potential for leakage

  11. Read length and repeat resolution: Exploring prokaryote genomes using next-generation sequencing technologies

    KAUST Repository

    Cahill, Matt J.

    2010-07-12

    Background: There are a growing number of next-generation sequencing technologies. At present, the most cost-effective options also produce the shortest reads. However, even for prokaryotes, there is uncertainty concerning the utility of these technologies for the de novo assembly of complete genomes. This reflects an expectation that short reads will be unable to resolve small, but presumably abundant, repeats. Methodology/Principal Findings: Using a simple model of repeat assembly, we develop and test a technique that, for any read length, can estimate the occurrence of unresolvable repeats in a genome, and thus predict the number of gaps that would need to be closed to produce a complete sequence. We apply this technique to 818 prokaryote genome sequences. This provides a quantitative assessment of the relative performance of various lengths. Notably, unpaired reads of only 150nt can reconstruct approximately 50% of the analysed genomes with fewer than 96 repeat-induced gaps. Nonetheless, there is considerable variation amongst prokaryotes. Some genomes can be assembled to near contiguity using very short reads while others require much longer reads. Conclusions: Given the diversity of prokaryote genomes, a sequencing strategy should be tailored to the organism under study. Our results will provide researchers with a practical resource to guide the selection of the appropriate read length. 2010 Cahill et al.

  12. Read length and repeat resolution: exploring prokaryote genomes using next-generation sequencing technologies.

    Directory of Open Access Journals (Sweden)

    Matt J Cahill

    Full Text Available BACKGROUND: There are a growing number of next-generation sequencing technologies. At present, the most cost-effective options also produce the shortest reads. However, even for prokaryotes, there is uncertainty concerning the utility of these technologies for the de novo assembly of complete genomes. This reflects an expectation that short reads will be unable to resolve small, but presumably abundant, repeats. METHODOLOGY/PRINCIPAL FINDINGS: Using a simple model of repeat assembly, we develop and test a technique that, for any read length, can estimate the occurrence of unresolvable repeats in a genome, and thus predict the number of gaps that would need to be closed to produce a complete sequence. We apply this technique to 818 prokaryote genome sequences. This provides a quantitative assessment of the relative performance of various lengths. Notably, unpaired reads of only 150nt can reconstruct approximately 50% of the analysed genomes with fewer than 96 repeat-induced gaps. Nonetheless, there is considerable variation amongst prokaryotes. Some genomes can be assembled to near contiguity using very short reads while others require much longer reads. CONCLUSIONS: Given the diversity of prokaryote genomes, a sequencing strategy should be tailored to the organism under study. Our results will provide researchers with a practical resource to guide the selection of the appropriate read length.

  13. Read length and repeat resolution: Exploring prokaryote genomes using next-generation sequencing technologies

    KAUST Repository

    Cahill, Matt J.; Kö ser, Claudio U.; Ross, Nicholas E.; Archer, John A.C.

    2010-01-01

    Background: There are a growing number of next-generation sequencing technologies. At present, the most cost-effective options also produce the shortest reads. However, even for prokaryotes, there is uncertainty concerning the utility of these technologies for the de novo assembly of complete genomes. This reflects an expectation that short reads will be unable to resolve small, but presumably abundant, repeats. Methodology/Principal Findings: Using a simple model of repeat assembly, we develop and test a technique that, for any read length, can estimate the occurrence of unresolvable repeats in a genome, and thus predict the number of gaps that would need to be closed to produce a complete sequence. We apply this technique to 818 prokaryote genome sequences. This provides a quantitative assessment of the relative performance of various lengths. Notably, unpaired reads of only 150nt can reconstruct approximately 50% of the analysed genomes with fewer than 96 repeat-induced gaps. Nonetheless, there is considerable variation amongst prokaryotes. Some genomes can be assembled to near contiguity using very short reads while others require much longer reads. Conclusions: Given the diversity of prokaryote genomes, a sequencing strategy should be tailored to the organism under study. Our results will provide researchers with a practical resource to guide the selection of the appropriate read length. 2010 Cahill et al.

  14. DNA Self-Assembly: From Chirality to Evolution

    Directory of Open Access Journals (Sweden)

    Youri Timsit

    2013-04-01

    Full Text Available Transient or long-term DNA self-assembly participates in essential genetic functions. The present review focuses on tight DNA-DNA interactions that have recently been found to play important roles in both controlling DNA higher-order structures and their topology. Due to their chirality, double helices are tightly packed into stable right-handed crossovers. Simple packing rules that are imposed by DNA geometry and sequence dictate the overall architecture of higher order DNA structures. Close DNA-DNA interactions also provide the missing link between local interactions and DNA topology, thus explaining how type II DNA topoisomerases may sense locally the global topology. Finally this paper proposes that through its influence on DNA self-assembled structures, DNA chirality played a critical role during the early steps of evolution.

  15. Draft genome sequences of seven isolates of Phytophthora ramorum EU2 from Northern Ireland

    Directory of Open Access Journals (Sweden)

    Lourdes de la Mata Saez

    2015-12-01

    Full Text Available Here we present draft-quality genome sequence assemblies for the oomycete Phytophthora ramorum genetic lineage EU2. We sequenced genomes of seven isolates collected in Northern Ireland between 2010 and 2012. Multiple genome sequences from P. ramorum EU2 will be valuable for identifying genetic variation within the clonal lineage that can be useful for tracking its spread.

  16. The genomic sequence of the Chinese hamster ovary (CHO)-K1 cell line

    DEFF Research Database (Denmark)

    Xu, Xun; Pan, Shengkai; Liu, Xin

    2011-01-01

    Chinese hamster ovary (CHO)-derived cell lines are the preferred host cells for the production of therapeutic proteins. Here we present a draft genomic sequence of the CHO-K1 ancestral cell line. The assembly comprises 2.45 Gb of genomic sequence, with 24,383 predicted genes. We associate most of...

  17. WebPrInSeS: automated full-length clone sequence identification and verification using high-throughput sequencing data.

    Science.gov (United States)

    Massouras, Andreas; Decouttere, Frederik; Hens, Korneel; Deplancke, Bart

    2010-07-01

    High-throughput sequencing (HTS) is revolutionizing our ability to obtain cheap, fast and reliable sequence information. Many experimental approaches are expected to benefit from the incorporation of such sequencing features in their pipeline. Consequently, software tools that facilitate such an incorporation should be of great interest. In this context, we developed WebPrInSeS, a web server tool allowing automated full-length clone sequence identification and verification using HTS data. WebPrInSeS encompasses two separate software applications. The first is WebPrInSeS-C which performs automated sequence verification of user-defined open-reading frame (ORF) clone libraries. The second is WebPrInSeS-E, which identifies positive hits in cDNA or ORF-based library screening experiments such as yeast one- or two-hybrid assays. Both tools perform de novo assembly using HTS data from any of the three major sequencing platforms. Thus, WebPrInSeS provides a highly integrated, cost-effective and efficient way to sequence-verify or identify clones of interest. WebPrInSeS is available at http://webprinses.epfl.ch/ and is open to all users.

  18. Drosophila homologues of adenomatous polyposis coli (APC) and the formin diaphanous collaborate by a conserved mechanism to stimulate actin filament assembly.

    Science.gov (United States)

    Jaiswal, Richa; Stepanik, Vince; Rankova, Aneliya; Molinar, Olivia; Goode, Bruce L; McCartney, Brooke M

    2013-05-10

    Vertebrate APC collaborates with Dia through its Basic domain to assemble actin filaments. Despite limited sequence homology between the vertebrate and Drosophila APC Basic domains, Drosophila APC1 collaborates with Dia to stimulate actin assembly in vitro. The mechanism of actin assembly is highly conserved over evolution. APC-Dia collaborations may be crucial in a wide range of animal cells. Adenomatous polyposis coli (APC) is a large multidomain protein that regulates the cytoskeleton. Recently, it was shown that vertebrate APC through its Basic domain directly collaborates with the formin mDia1 to stimulate actin filament assembly in the presence of nucleation barriers. However, it has been unclear whether these activities extend to homologues of APC and Dia in other organisms. Drosophila APC and Dia are each required to promote actin furrow formation in the syncytial embryo, suggesting a potential collaboration in actin assembly, but low sequence homology between the Basic domains of Drosophila and vertebrate APC has left their functional and mechanistic parallels uncertain. To address this question, we purified Drosophila APC1 and Dia and determined their individual and combined effects on actin assembly using both bulk fluorescence assays and total internal reflection fluorescence microscopy. Our data show that APC1, similar to its vertebrate homologue, bound to actin monomers and nucleated and bundled filaments. Further, Drosophila Dia nucleated actin assembly and protected growing filament barbed ends from capping protein. Drosophila APC1 and Dia directly interacted and collaborated to promote actin assembly in the combined presence of profilin and capping protein. Thus, despite limited sequence homology, Drosophila and vertebrate APCs exhibit highly related activities and mechanisms and directly collaborate with formins. These results suggest that APC-Dia interactions in actin assembly are conserved and may underlie important in vivo functions in a broad

  19. Population genetic analysis of shotgun assemblies of genomic sequences from multiple individuals

    DEFF Research Database (Denmark)

    Hellmann, Ines; Mang, Yuan; Gu, Zhiping

    2008-01-01

    We introduce a simple, broadly applicable method for obtaining estimates of nucleotide diversity from genomic shotgun sequencing data. The method takes into account the special nature of these data: random sampling of genomic segments from one or more individuals and a relatively high error rate...... for individual reads. Applying this method to data from the Celera human genome sequencing and SNP discovery project, we obtain estimates of nucleotide diversity in windows spanning the human genome and show that the diversity to divergence ratio is reduced in regions of low recombination. Furthermore, we show...

  20. Assembly & Metrology of First Wall Components of SST-1

    Science.gov (United States)

    Parekh, Tejas; Santra, Prosenjit; Biswas, Prabal; Patel, Hiteshkumar; Paravastu, Yuvakiran; Jaiswal, Snehal; Chauhan, Pradeep; Babu, Gattu Ramesh; A, Arun Prakash; Bhavsar, Dhaval; Raval, Dilip C.; Khan, Ziauddin; Pradhan, Subrata

    2017-04-01

    First Wall Components (FWC) of SST-1 tokamak, which are in the immediate vicinity of plasma comprises of limiters, divertors, baffles, passive stabilizers are designed to operate long duration (1000 s) discharges of elongated plasma. All FWC consists of a copper alloy heat sink modules with SS cooling tubes brazed onto it, graphite tiles acting as armour material facing the plasma, and are mounted to the vacuum vessels with suitable Inconel support structures at ring & port locations. The FWC are very recently assembled and commissioned successfully inside the vacuum vessel of SST-1 undergoing a meticulous planning of assembly sequence, quality checks at every stage of the assembly process. This paper will present the metrology aspects & procedure of each FWC, both outside the vacuum vessel, and inside the vessel, assembly tolerances, tools, equipment and jig/fixtures, used at each stage of assembly, starting from location of support bases on vessel rings, fixing of copper modules on support structures, around 3800 graphite tile mounting on 136 copper modules with proper tightening torques, till final toroidal and poloidal geometry of the in-vessel components are obtained within acceptable limits, also ensuring electrical continuity of passive stabilizers to form a closed saddle loop, electrical isolation of passive stabilizers from vacuum vessel.

  1. De novo assembly of the Aedes aegypti genome using Hi-C yields chromosome-length scaffolds.

    Science.gov (United States)

    Dudchenko, Olga; Batra, Sanjit S; Omer, Arina D; Nyquist, Sarah K; Hoeger, Marie; Durand, Neva C; Shamim, Muhammad S; Machol, Ido; Lander, Eric S; Aiden, Aviva Presser; Aiden, Erez Lieberman

    2017-04-07

    The Zika outbreak, spread by the Aedes aegypti mosquito, highlights the need to create high-quality assemblies of large genomes in a rapid and cost-effective way. Here we combine Hi-C data with existing draft assemblies to generate chromosome-length scaffolds. We validate this method by assembling a human genome, de novo, from short reads alone (67× coverage). We then combine our method with draft sequences to create genome assemblies of the mosquito disease vectors Ae aegypti and Culex quinquefasciatus , each consisting of three scaffolds corresponding to the three chromosomes in each species. These assemblies indicate that almost all genomic rearrangements among these species occur within, rather than between, chromosome arms. The genome assembly procedure we describe is fast, inexpensive, and accurate, and can be applied to many species. Copyright © 2017, American Association for the Advancement of Science.

  2. Characterizing novel endogenous retroviruses from genetic variation inferred from short sequence reads

    DEFF Research Database (Denmark)

    Mourier, Tobias; Mollerup, Sarah; Vinner, Lasse

    2015-01-01

    From Illumina sequencing of DNA from brain and liver tissue from the lion, Panthera leo, and tumor samples from the pike-perch, Sander lucioperca, we obtained two assembled sequence contigs with similarity to known retroviruses. Phylogenetic analyses suggest that the pike-perch retrovirus belongs...... to the epsilonretroviruses, and the lion retrovirus to the gammaretroviruses. To determine if these novel retroviral sequences originate from an endogenous retrovirus or from a recently integrated exogenous retrovirus, we assessed the genetic diversity of the parental sequences from which the short Illumina reads...

  3. Data for ion and seed dependent fibril assembly of a spidroin core domain

    Directory of Open Access Journals (Sweden)

    Martin Humenik

    2015-09-01

    Full Text Available This data article includes size exclusion chromatography data of soluble eADF4(C16, an engineered spider silk variant based on the core domain sequence of the natural dragline silk protein ADF4 of Araneus diadematus, in combination with light scattering; the protein is monomeric before assembly. The assembled mature fibrils were visualized by transmission electron microscopy (TEM and atomic force microscopy (AFM. Sonicated fibrils were used as seeds to by-pass the nucleation lag phase in eADF4(C16 assembly. We also provide data on the sedimentation kinetics of spider silk in the presence of different NaCl concentrations revealing very slow protein aggregation in comparison to the fast assembly triggered by phosphate ions published previously [1]. Experiments in the Data article represent supporting material for our work published recently [1], which described the assembly mechanism of recombinant eADF4(C16 fibrils.

  4. Genome Sequence of the Necrotrophic Plant Pathogen Alternaria brassicicola Abra43

    Science.gov (United States)

    Belmas, Elodie; Briand, Martial; Kwasiborski, Anthony; Colou, Justine; N’Guyen, Guillaume; Iacomi, Béatrice; Grappin, Philippe; Campion, Claire; Simoneau, Philippe; Barret, Matthieu

    2018-01-01

    ABSTRACT Alternaria brassicicola causes dark spot (or black spot) disease, which is one of the most common and destructive fungal diseases of Brassicaceae spp. worldwide. Here, we report the draft genome sequence of strain Abra43. The assembly comprises 29 scaffolds, with an N50 value of 2.1 Mb. The assembled genome was 31,036,461 bp in length, with a G+C content of 50.85%. PMID:29439047

  5. De novo assembly and transcriptome analysis of five major tissues of Jatropha curcas L. using GS FLX titanium platform of 454 pyrosequencing

    Directory of Open Access Journals (Sweden)

    Parani Madasamy

    2011-04-01

    Full Text Available Abstract Background Jatropha curcas L. is an important non-edible oilseed crop with promising future in biodiesel production. However, factors like oil yield, oil composition, toxic compounds in oil cake, pests and diseases limit its commercial potential. Well established genetic engineering methods using cloned genes could be used to address these limitations. Earlier, 10,983 unigenes from Sanger sequencing of ESTs, and 3,484 unique assembled transcripts from 454 pyrosequencing of uncloned cDNAs were reported. In order to expedite the process of gene discovery, we have undertaken 454 pyrosequencing of normalized cDNAs prepared from roots, mature leaves, flowers, developing seeds, and embryos of J. curcas. Results From 383,918 raw reads, we obtained 381,957 quality-filtered and trimmed reads that are suitable for the assembly of transcript sequences. De novo contig assembly of these reads generated 17,457 assembled transcripts (contigs and 54,002 singletons. Average length of the assembled transcripts was 916 bp. About 30% of the transcripts were longer than 1000 bases, and the size of the longest transcript was 7,173 bases. BLASTX analysis revealed that 2,589 of these transcripts are full-length. The assembled transcripts were validated by RT-PCR analysis of 28 transcripts. The results showed that the transcripts were correctly assembled and represent actively expressed genes. KEGG pathway mapping showed that 2,320 transcripts are related to major biochemical pathways including the oil biosynthesis pathway. Overall, the current study reports 14,327 new assembled transcripts which included 2589 full-length transcripts and 27 transcripts that are directly involved in oil biosynthesis. Conclusion The large number of transcripts reported in the current study together with existing ESTs and transcript sequences will serve as an invaluable genetic resource for crop improvement in jatropha. Sequence information of those genes that are involved in oil

  6. De novo assembly and transcriptome analysis of five major tissues of Jatropha curcas L. using GS FLX titanium platform of 454 pyrosequencing.

    Science.gov (United States)

    Natarajan, Purushothaman; Parani, Madasamy

    2011-04-15

    Jatropha curcas L. is an important non-edible oilseed crop with promising future in biodiesel production. However, factors like oil yield, oil composition, toxic compounds in oil cake, pests and diseases limit its commercial potential. Well established genetic engineering methods using cloned genes could be used to address these limitations. Earlier, 10,983 unigenes from Sanger sequencing of ESTs, and 3,484 unique assembled transcripts from 454 pyrosequencing of uncloned cDNAs were reported. In order to expedite the process of gene discovery, we have undertaken 454 pyrosequencing of normalized cDNAs prepared from roots, mature leaves, flowers, developing seeds, and embryos of J. curcas. From 383,918 raw reads, we obtained 381,957 quality-filtered and trimmed reads that are suitable for the assembly of transcript sequences. De novo contig assembly of these reads generated 17,457 assembled transcripts (contigs) and 54,002 singletons. Average length of the assembled transcripts was 916 bp. About 30% of the transcripts were longer than 1000 bases, and the size of the longest transcript was 7,173 bases. BLASTX analysis revealed that 2,589 of these transcripts are full-length. The assembled transcripts were validated by RT-PCR analysis of 28 transcripts. The results showed that the transcripts were correctly assembled and represent actively expressed genes. KEGG pathway mapping showed that 2,320 transcripts are related to major biochemical pathways including the oil biosynthesis pathway. Overall, the current study reports 14,327 new assembled transcripts which included 2589 full-length transcripts and 27 transcripts that are directly involved in oil biosynthesis. The large number of transcripts reported in the current study together with existing ESTs and transcript sequences will serve as an invaluable genetic resource for crop improvement in jatropha. Sequence information of those genes that are involved in oil biosynthesis could be used for metabolic engineering of

  7. Sequence-Based Analysis of Structural Organization and Composition of the Cultivated Sunflower (Helianthus annuus L.) Genome

    Science.gov (United States)

    Gill, Navdeep; Buti, Matteo; Kane, Nolan; Bellec, Arnaud; Helmstetter, Nicolas; Berges, Hélène; Rieseberg, Loren H.

    2014-01-01

    Sunflower is an important oilseed crop, as well as a model system for evolutionary studies, but its 3.6 gigabase genome has proven difficult to assemble, in part because of the high repeat content of its genome. Here we report on the sequencing, assembly, and analyses of 96 randomly chosen BACs from sunflower to provide additional information on the repeat content of the sunflower genome, assess how repetitive elements in the sunflower genome are organized relative to genes, and compare the genomic distribution of these repeats to that found in other food crops and model species. We also examine the expression of transposable element-related transcripts in EST databases for sunflower to determine the representation of repeats in the transcriptome and to measure their transcriptional activity. Our data confirm previous reports in suggesting that the sunflower genome is >78% repetitive. Sunflower repeats share very little similarity to other plant repeats such as those of Arabidopsis, rice, maize and wheat; overall 28% of repeats are “novel” to sunflower. The repetitive sequences appear to be randomly distributed within the sequenced BACs. Assuming the 96 BACs are representative of the genome as a whole, then approximately 5.2% of the sunflower genome comprises non TE-related genic sequence, with an average gene density of 18kbp/gene. Expression levels of these transposable elements indicate tissue specificity and differential expression in vegetative and reproductive tissues, suggesting that expressed TEs might contribute to sunflower development. The assembled BACs will also be useful for assessing the quality of several different draft assemblies of the sunflower genome and for annotating the reference sequence. PMID:24833511

  8. Sequence-Based Analysis of Structural Organization and Composition of the Cultivated Sunflower (Helianthus annuus L. Genome

    Directory of Open Access Journals (Sweden)

    Navdeep Gill

    2014-04-01

    Full Text Available Sunflower is an important oilseed crop, as well as a model system for evolutionary studies, but its 3.6 gigabase genome has proven difficult to assemble, in part because of the high repeat content of its genome. Here we report on the sequencing, assembly, and analyses of 96 randomly chosen BACs from sunflower to provide additional information on the repeat content of the sunflower genome, assess how repetitive elements in the sunflower genome are organized relative to genes, and compare the genomic distribution of these repeats to that found in other food crops and model species. We also examine the expression of transposable element-related transcripts in EST databases for sunflower to determine the representation of repeats in the transcriptome and to measure their transcriptional activity. Our data confirm previous reports in suggesting that the sunflower genome is >78% repetitive. Sunflower repeats share very little similarity to other plant repeats such as those of Arabidopsis, rice, maize and wheat; overall 28% of repeats are “novel” to sunflower. The repetitive sequences appear to be randomly distributed within the sequenced BACs. Assuming the 96 BACs are representative of the genome as a whole, then approximately 5.2% of the sunflower genome comprises non TE-related genic sequence, with an average gene density of 18kbp/gene. Expression levels of these transposable elements indicate tissue specificity and differential expression in vegetative and reproductive tissues, suggesting that expressed TEs might contribute to sunflower development. The assembled BACs will also be useful for assessing the quality of several different draft assemblies of the sunflower genome and for annotating the reference sequence.

  9. Ultra-Precise Assembly of Micro-Electromechanical Systems (MEMS) Components

    Energy Technology Data Exchange (ETDEWEB)

    Feddema, J.T.; Simon, R.; Polosky, M.; Christenson, T.

    1999-04-01

    This report summarizes a three year effort to develop an automated microassembly workcell for the assembly of LIGA (Lithography Galvonoforming Abforming) parts. Over the last several years, Sandia has developed processes for producing surface machined silicon and LIGA parts for use in weapons surety devices. Some of these parts have outside dimensions as small as 100 micron, and most all have submicron tolerances. Parts this small and precise are extremely difficult to assembly by hand. Therefore, in this project, we investigated the technologies required to develop a robotic workcell to assembly these parts. In particular, we concentrated on micro-grippers, visual servoing, micro-assembly planning, and parallel assembly. Three different micro-grippers were tested: a pneumatic probe, a thermally actuated polysilicon tweezer, and a LIGA fabricated tweezer. Visual servoing was used to accuracy position two parts relative to one another. Fourier optics methods were used to generate synthetic microscope images from CAD drawings. These synthetic images are used off-line to test image processing routines under varying magnifications and depths of field. They also provide reference image features which are used to visually servo the part to the desired position. We also investigated a new aspect of fine motion planning for the micro-domain. As parts approach 1-10 {micro}m or less in outside dimensions, interactive forces such as van der Waals and electrostatic forces become major factors which greatly change the assembly sequence and path plans. We developed the mathematics required to determine the goal regions for pick up, holding, and release of a micro-sphere being handled by a rectangular tool. Finally, we implemented and tested the ability to assemble an array of LIGA parts attached to two 3 inch diameter wafers. In this way, hundreds of parts can be assembled in parallel rather than assembling each part individually.

  10. The Douglas-Fir Genome Sequence Reveals Specialization of the Photosynthetic Apparatus in Pinaceae

    Directory of Open Access Journals (Sweden)

    David B. Neale

    2017-09-01

    Full Text Available A reference genome sequence for Pseudotsuga menziesii var. menziesii (Mirb. Franco (Coastal Douglas-fir is reported, thus providing a reference sequence for a third genus of the family Pinaceae. The contiguity and quality of the genome assembly far exceeds that of other conifer reference genome sequences (contig N50 = 44,136 bp and scaffold N50 = 340,704 bp. Incremental improvements in sequencing and assembly technologies are in part responsible for the higher quality reference genome, but it may also be due to a slightly lower exact repeat content in Douglas-fir vs. pine and spruce. Comparative genome annotation with angiosperm species reveals gene-family expansion and contraction in Douglas-fir and other conifers which may account for some of the major morphological and physiological differences between the two major plant groups. Notable differences in the size of the NDH-complex gene family and genes underlying the functional basis of shade tolerance/intolerance were observed. This reference genome sequence not only provides an important resource for Douglas-fir breeders and geneticists but also sheds additional light on the evolutionary processes that have led to the divergence of modern angiosperms from the more ancient gymnosperms.

  11. [Complete genome sequencing of polymalic acid-producing strain Aureobasidium pullulans CCTCC M2012223].

    Science.gov (United States)

    Wang, Yongkang; Song, Xiaodan; Li, Xiaorong; Yang, Sang-tian; Zou, Xiang

    2017-01-04

    To explore the genome sequence of Aureobasidium pullulans CCTCC M2012223, analyze the key genes related to the biosynthesis of important metabolites, and provide genetic background for metabolic engineering. Complete genome of A. pullulans CCTCC M2012223 was sequenced by Illumina HiSeq high throughput sequencing platform. Then, fragment assembly, gene prediction, functional annotation, and GO/COG cluster were analyzed in comparison with those of other five A. pullulans varieties. The complete genome sequence of A. pullulans CCTCC M2012223 was 30756831 bp with an average GC content of 47.49%, and 9452 genes were successfully predicted. Genome-wide analysis showed that A. pullulans CCTCC M2012223 had the biggest genome assembly size. Protein sequences involved in the pullulan and polymalic acid pathway were highly conservative in all of six A. pullulans varieties. Although both A. pullulans CCTCC M2012223 and A. pullulans var. melanogenum have a close affinity, some point mutation and inserts were occurred in protein sequences involved in melanin biosynthesis. Genome information of A. pullulans CCTCC M2012223 was annotated and genes involved in melanin, pullulan and polymalic acid pathway were compared, which would provide a theoretical basis for genetic modification of metabolic pathway in A. pullulans.

  12. Genome sequences of three strains of Aspergillus flavus for the biological control of Aflatoxin

    Science.gov (United States)

    The genomes of three strains of Aspergillus flavus with demonstrated utility for the biological control of aflatoxin were sequenced. These sequences were assembled with MIRA and annotated with Augustus using A. flavus strain 3357 (NCBI EQ963472) as a reference. Each strain had a genome of 36.3 to ...

  13. USER-CENTERED DESIGN SEBAGAI METODE PENINGKATAN KONTROL KOGNITIF PADA LINI ASSEMBLY (STUDI KASUS PT. KUBOTA INDONESIA

    Directory of Open Access Journals (Sweden)

    Novie Susanto

    2012-02-01

    Full Text Available In developing countries like Indonesia, the rationalization of production processes and systems will increase.Challenge that emerged significantly is the increase in planning efficiency. A prospective approach to reduce theplanning effort is the development of a production system capable of optimizing itself during the process. On theassembly line PT. Kubota Indonesia found that the workers perform assembly process with a sequence that is notplanned as a result of the lack of a standardized sequence of process companies and lead to cognitive controlthat are not stable and continuous learning process. Because this issue contrary to the concept of efficiency inplanning, it takes an automated system with a complete and fully optimized (self-optimizing production system.The purpose of this study is to apply the planning efficiency through implementation of optimized productionsystem (self-optimizing production systems in assembly-line PT. Kubota Indonesia. Data obtaining done by thedetermination of primary and secondary variables include the perception of cognitive control, a solution basedon the memory (memory, and action. Repairs are carried out is by applying self-optimizing production systemconsisting of 4 steps: the development of cognitive architecture is to determine the sequence of work andapplying it to extend the software, implementation of individual modules is through the use of hand screw, thedevelopment of human machine interfaces in hand tool screw is through the explanation task analysis andimplementation and evaluation of improvement is the use of robotic assembly in the assembly processenvironment.

  14. The Physalis peruviana leaf transcriptome: assembly, annotation and gene model prediction

    Directory of Open Access Journals (Sweden)

    Garzón-Martínez Gina A

    2012-04-01

    Full Text Available Abstract Background Physalis peruviana commonly known as Cape gooseberry is a member of the Solanaceae family that has an increasing popularity due to its nutritional and medicinal values. A broad range of genomic tools is available for other Solanaceae, including tomato and potato. However, limited genomic resources are currently available for Cape gooseberry. Results We report the generation of a total of 652,614 P. peruviana Expressed Sequence Tags (ESTs, using 454 GS FLX Titanium technology. ESTs, with an average length of 371 bp, were obtained from a normalized leaf cDNA library prepared using a Colombian commercial variety. De novo assembling was performed to generate a collection of 24,014 isotigs and 110,921 singletons, with an average length of 1,638 bp and 354 bp, respectively. Functional annotation was performed using NCBI’s BLAST tools and Blast2GO, which identified putative functions for 21,191 assembled sequences, including gene families involved in all the major biological processes and molecular functions as well as defense response and amino acid metabolism pathways. Gene model predictions in P. peruviana were obtained by using the genomes of Solanum lycopersicum (tomato and Solanum tuberosum (potato. We predict 9,436 P. peruviana sequences with multiple-exon models and conserved intron positions with respect to the potato and tomato genomes. Additionally, to study species diversity we developed 5,971 SSR markers from assembled ESTs. Conclusions We present the first comprehensive analysis of the Physalis peruviana leaf transcriptome, which will provide valuable resources for development of genetic tools in the species. Assembled transcripts with gene models could serve as potential candidates for marker discovery with a variety of applications including: functional diversity, conservation and improvement to increase productivity and fruit quality. P. peruviana was estimated to be phylogenetically branched out before the

  15. The Physalis peruviana leaf transcriptome: assembly, annotation and gene model prediction.

    Science.gov (United States)

    Garzón-Martínez, Gina A; Zhu, Z Iris; Landsman, David; Barrero, Luz S; Mariño-Ramírez, Leonardo

    2012-04-25

    Physalis peruviana commonly known as Cape gooseberry is a member of the Solanaceae family that has an increasing popularity due to its nutritional and medicinal values. A broad range of genomic tools is available for other Solanaceae, including tomato and potato. However, limited genomic resources are currently available for Cape gooseberry. We report the generation of a total of 652,614 P. peruviana Expressed Sequence Tags (ESTs), using 454 GS FLX Titanium technology. ESTs, with an average length of 371 bp, were obtained from a normalized leaf cDNA library prepared using a Colombian commercial variety. De novo assembling was performed to generate a collection of 24,014 isotigs and 110,921 singletons, with an average length of 1,638 bp and 354 bp, respectively. Functional annotation was performed using NCBI's BLAST tools and Blast2GO, which identified putative functions for 21,191 assembled sequences, including gene families involved in all the major biological processes and molecular functions as well as defense response and amino acid metabolism pathways. Gene model predictions in P. peruviana were obtained by using the genomes of Solanum lycopersicum (tomato) and Solanum tuberosum (potato). We predict 9,436 P. peruviana sequences with multiple-exon models and conserved intron positions with respect to the potato and tomato genomes. Additionally, to study species diversity we developed 5,971 SSR markers from assembled ESTs. We present the first comprehensive analysis of the Physalis peruviana leaf transcriptome, which will provide valuable resources for development of genetic tools in the species. Assembled transcripts with gene models could serve as potential candidates for marker discovery with a variety of applications including: functional diversity, conservation and improvement to increase productivity and fruit quality. P. peruviana was estimated to be phylogenetically branched out before the divergence of five other Solanaceae family members, S

  16. De novo genome assembly and annotation of Australia's largest freshwater fish, the Murray cod (Maccullochella peelii), from Illumina and Nanopore sequencing read.

    Science.gov (United States)

    Austin, Christopher M; Tan, Mun Hua; Harrisson, Katherine A; Lee, Yin Peng; Croft, Laurence J; Sunnucks, Paul; Pavlova, Alexandra; Gan, Han Ming

    2017-08-01

    One of the most iconic Australian fish is the Murray cod, Maccullochella peelii (Mitchell 1838), a freshwater species that can grow to ∼1.8 metres in length and live to age ≥48 years. The Murray cod is of a conservation concern as a result of strong population contractions, but it is also popular for recreational fishing and is of growing aquaculture interest. In this study, we report the whole genome sequence of the Murray cod to support ongoing population genetics, conservation, and management research, as well as to better understand the evolutionary ecology and history of the species. A draft Murray cod genome of 633 Mbp (N50 = 109 974bp; BUSCO and CEGMA completeness of 94.2% and 91.9%, respectively) with an estimated 148 Mbp of putative repetitive sequences was assembled from the combined sequencing data of 2 fish individuals with an identical maternal lineage; 47.2 Gb of Illumina HiSeq data and 804 Mb of Nanopore data were generated from the first individual while 23.2 Gb of Illumina MiSeq data were generated from the second individual. The inclusion of Nanopore reads for scaffolding followed by subsequent gap-closing using Illumina data led to a 29% reduction in the number of scaffolds and a 55% and 54% increase in the scaffold and contig N50, respectively. We also report the first transcriptome of Murray cod that was subsequently used to annotate the Murray cod genome, leading to the identification of 26 539 protein-coding genes. We present the whole genome of the Murray cod and anticipate this will be a catalyst for a range of genetic, genomic, and phylogenetic studies of the Murray cod and more generally other fish species of the Percichthydae family. © The Authors 2017. Published by Oxford University Press.

  17. Rapid development of microsatellite markers for Callosobruchus chinensis using Illumina paired-end sequencing.

    Directory of Open Access Journals (Sweden)

    Can-Xing Duan

    Full Text Available BACKGROUND: The adzuki bean weevil, Callosobruchus chinensis L., is one of the most destructive pests of stored legume seeds such as mungbean, cowpea, and adzuki bean, which usually cause considerable loss in the quantity and quality of stored seeds during transportation and storage. However, a lack of genetic information of this pest results in a series of genetic questions remain largely unknown, including population genetic structure, kinship, biotype abundance, and so on. Co-dominant microsatellite markers offer a great resolving power to determine these events. Here, we report rapid microsatellite isolation from C. chinensis via high-throughput sequencing. PRINCIPAL FINDINGS: In this study, 94,560,852 quality-filtered and trimmed reads were obtained for the assembly of genome using Illumina paired-end sequencing technology. In total, the genome with total length of 497,124,785 bp, comprising 403,113 high quality contigs was generated with de novo assembly. More than 6800 SSR loci were detected and a suit of 6303 primer pair sequences were designed and 500 of them were randomly selected for validation. Of these, 196 pair of primers, i.e. 39.2%, produced reproducible amplicons that were polymorphic among 8 C. chinensis genotypes collected from different geographical regions. Twenty out of 196 polymorphic SSR markers were used to analyze the genetic diversity of 18 C. chinensis populations. The results showed the twenty SSR loci were highly polymorphic among these populations. CONCLUSIONS: This study presents a first report of genome sequencing and de novo assembly for C. chinensis and demonstrates the feasibility of generating a large scale of sequence information and SSR loci isolation by Illumina paired-end sequencing. Our results provide a valuable resource for C. chinensis research. These novel markers are valuable for future genetic mapping, trait association, genetic structure and kinship among C. chinensis.

  18. A gene-based high-resolution comparative radiation hybrid map as a framework for genome sequence assembly of a bovine chromosome 6 region associated with QTL for growth, body composition, and milk performance traits

    Directory of Open Access Journals (Sweden)

    Laurent Pascal

    2006-03-01

    Full Text Available Abstract Background A number of different quantitative trait loci (QTL for various phenotypic traits, including milk production, functional, and conformation traits in dairy cattle as well as growth and body composition traits in meat cattle, have been mapped consistently in the middle region of bovine chromosome 6 (BTA6. Dense genetic and physical maps and, ultimately, a fully annotated genome sequence as well as their mutual connections are required to efficiently identify genes and gene variants responsible for genetic variation of phenotypic traits. A comprehensive high-resolution gene-rich map linking densely spaced bovine markers and genes to the annotated human genome sequence is required as a framework to facilitate this approach for the region on BTA6 carrying the QTL. Results Therefore, we constructed a high-resolution radiation hybrid (RH map for the QTL containing chromosomal region of BTA6. This new RH map with a total of 234 loci including 115 genes and ESTs displays a substantial increase in loci density compared to existing physical BTA6 maps. Screening the available bovine genome sequence resources, a total of 73 loci could be assigned to sequence contigs, which were already identified as specific for BTA6. For 43 loci, corresponding sequence contigs, which were not yet placed on the bovine genome assembly, were identified. In addition, the improved potential of this high-resolution RH map for BTA6 with respect to comparative mapping was demonstrated. Mapping a large number of genes on BTA6 and cross-referencing them with map locations in corresponding syntenic multi-species chromosome segments (human, mouse, rat, dog, chicken achieved a refined accurate alignment of conserved segments and evolutionary breakpoints across the species included. Conclusion The gene-anchored high-resolution RH map (1 locus/300 kb for the targeted region of BTA6 presented here will provide a valuable platform to guide high-quality assembling and

  19. Facile Site-Directed Mutagenesis of Large Constructs Using Gibson Isothermal DNA Assembly.

    Science.gov (United States)

    Yonemoto, Isaac T; Weyman, Philip D

    2017-01-01

    Site-directed mutagenesis is a commonly used molecular biology technique to manipulate biological sequences, and is especially useful for studying sequence determinants of enzyme function or designing proteins with improved activity. We describe a strategy using Gibson Isothermal DNA Assembly to perform site-directed mutagenesis on large (>~20 kbp) constructs that are outside the effective range of standard techniques such as QuikChange II (Agilent Technologies), but more reliable than traditional cloning using restriction enzymes and ligation.

  20. Rhipicephalus microplus strain Deutsch, 10 BAC clone sequences

    Science.gov (United States)

    The cattle tick, Rhipicephalus (Boophilus) microplus, has a genome over 2.4 times the size of the human genome, and with over 70% of repetitive DNA, this genome would prove very costly to sequence at today's prices and difficult to assemble and analyze. We used labeled DNA probes from the coding reg...