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

De novo assembly of the perennial ryegrass transcriptome using an RNA-Seq strategy.

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Jacqueline D Farrell

Full Text Available Perennial ryegrass is a highly heterozygous outbreeding grass species used for turf and forage production. Heterozygosity can affect de-Bruijn graph assembly making de novo transcriptome assembly of species such as perennial ryegrass challenging. Creating a reference transcriptome from a homozygous perennial ryegrass genotype can circumvent the challenge of heterozygosity. The goals of this study were to perform RNA-sequencing on multiple tissues from a highly inbred genotype to develop a reference transcriptome. This was complemented with RNA-sequencing of a highly heterozygous genotype for SNP calling.De novo transcriptome assembly of the inbred genotype created 185,833 transcripts with an average length of 830 base pairs. Within the inbred reference transcriptome 78,560 predicted open reading frames were found of which 24,434 were predicted as complete. Functional annotation found 50,890 transcripts with a BLASTp hit from the Swiss-Prot non-redundant database, 58,941 transcripts with a Pfam protein domain and 1,151 transcripts encoding putative secreted peptides. To evaluate the reference transcriptome we targeted the high-affinity K+ transporter gene family and found multiple orthologs. Using the longest unique open reading frames as the reference sequence, 64,242 single nucleotide polymorphisms were found. One thousand sixty one open reading frames from the inbred genotype contained heterozygous sites, confirming the high degree of homozygosity.Our study has developed an annotated, comprehensive transcriptome reference for perennial ryegrass that can aid in determining genetic variation, expression analysis, genome annotation, and gene mapping.

De novo Transcriptome Assemblies of Rana (Lithobates catesbeiana and Xenopus laevis Tadpole Livers for Comparative Genomics without Reference Genomes.

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Inanc Birol

Full Text Available In this work we studied the liver transcriptomes of two frog species, the American bullfrog (Rana (Lithobates catesbeiana and the African clawed frog (Xenopus laevis. We used high throughput RNA sequencing (RNA-seq data to assemble and annotate these transcriptomes, and compared how their baseline expression profiles change when tadpoles of the two species are exposed to thyroid hormone. We generated more than 1.5 billion RNA-seq reads in total for the two species under two conditions as treatment/control pairs. We de novo assembled these reads using Trans-ABySS to reconstruct reference transcriptomes, obtaining over 350,000 and 130,000 putative transcripts for R. catesbeiana and X. laevis, respectively. Using available genomics resources for X. laevis, we annotated over 97% of our X. laevis transcriptome contigs, demonstrating the utility and efficacy of our methodology. Leveraging this validated analysis pipeline, we also annotated the assembled R. catesbeiana transcriptome. We used the expression profiles of the annotated genes of the two species to examine the similarities and differences between the tadpole liver transcriptomes. We also compared the gene ontology terms of expressed genes to measure how the animals react to a challenge by thyroid hormone. Our study reports three main conclusions. First, de novo assembly of RNA-seq data is a powerful method for annotating and establishing transcriptomes of non-model organisms. Second, the liver transcriptomes of the two frog species, R. catesbeiana and X. laevis, show many common features, and the distribution of their gene ontology profiles are statistically indistinguishable. Third, although they broadly respond the same way to the presence of thyroid hormone in their environment, their receptor/signal transduction pathways display marked differences.

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

Icarus: visualizer for de novo assembly evaluation.

Science.gov (United States)

Mikheenko, Alla; Valin, Gleb; Prjibelski, Andrey; Saveliev, Vladislav; Gurevich, Alexey

2016-11-01

: Data visualization plays an increasingly important role in NGS data analysis. With advances in both sequencing and computational technologies, it has become a new bottleneck in genomics studies. Indeed, evaluation of de novo genome assemblies is one of the areas that can benefit from the visualization. However, even though multiple quality assessment methods are now available, existing visualization tools are hardly suitable for this purpose. Here, we present Icarus-a novel genome visualizer for accurate assessment and analysis of genomic draft assemblies, which is based on the tool QUAST. Icarus can be used in studies where a related reference genome is available, as well as for non-model organisms. The tool is available online and as a standalone application. http://cab.spbu.ru/software/icarus CONTACT: aleksey.gurevich@spbu.ruSupplementary information: 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.

Spaced Seed Data Structures for De Novo Assembly

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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.

Selecting Superior De Novo Transcriptome Assemblies: Lessons Learned by Leveraging the Best Plant Genome.

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Loren A Honaas

Full Text Available Whereas de novo assemblies of RNA-Seq data are being published for a growing number of species across the tree of life, there are currently no broadly accepted methods for evaluating such assemblies. Here we present a detailed comparison of 99 transcriptome assemblies, generated with 6 de novo assemblers including CLC, Trinity, SOAP, Oases, ABySS and NextGENe. Controlled analyses of de novo assemblies for Arabidopsis thaliana and Oryza sativa transcriptomes provide new insights into the strengths and limitations of transcriptome assembly strategies. We find that the leading assemblers generate reassuringly accurate assemblies for the majority of transcripts. At the same time, we find a propensity for assemblers to fail to fully assemble highly expressed genes. Surprisingly, the instance of true chimeric assemblies is very low for all assemblers. Normalized libraries are reduced in highly abundant transcripts, but they also lack 1000s of low abundance transcripts. We conclude that the quality of de novo transcriptome assemblies is best assessed through consideration of a combination of metrics: 1 proportion of reads mapping to an assembly 2 recovery of conserved, widely expressed genes, 3 N50 length statistics, and 4 the total number of unigenes. We provide benchmark Illumina transcriptome data and introduce SCERNA, a broadly applicable modular protocol for de novo assembly improvement. Finally, our de novo assembly of the Arabidopsis leaf transcriptome revealed ~20 putative Arabidopsis genes lacking in the current annotation.

De novo transcriptome assembly for a non-model species, the blood-sucking bug Triatoma brasiliensis, a vector of Chagas disease.

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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.

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...

Combining independent de novo assemblies optimizes the coding transcriptome for nonconventional model eukaryotic organisms.

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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

Identifying wrong assemblies in de novo short read primary ...

Indian Academy of Sciences (India)

2016-08-05

Aug 5, 2016 ... Most of these assemblies are done using some de novo short read assemblers and other related approaches. .... benchmarking projects like Assemblathon 1, Assemblathon ... from a large insert library (at least 1000 bases).

De novo transcriptome assembly of two different peach cultivars grown in Korea

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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.

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

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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.

Towards accurate de novo assembly for genomes with repeats

NARCIS (Netherlands)

Bucur, Doina

2017-01-01

De novo genome assemblers designed for short k-mer length or using short raw reads are unlikely to recover complex features of the underlying genome, such as repeats hundreds of bases long. We implement a stochastic machine-learning method which obtains accurate assemblies with repeats and

Sequencing and De Novo Transcriptome Assembly of Brachypodium sylvaticum (Poaceae

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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.

Rnnotator: an automated de novo transcriptome assembly pipeline from stranded RNA-Seq reads

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Martin, Jeffrey; Bruno, Vincent M.; Fang, Zhide; Meng, Xiandong; Blow, Matthew; Zhang, Tao; Sherlock, Gavin; Snyder, Michael; Wang, Zhong

2010-11-19

Background: Comprehensive annotation and quantification of transcriptomes are outstanding problems in functional genomics. While high throughput mRNA sequencing (RNA-Seq) has emerged as a powerful tool for addressing these problems, its success is dependent upon the availability and quality of reference genome sequences, thus limiting the organisms to which it can be applied. Results: Here, we describe Rnnotator, an automated software pipeline that generates transcript models by de novo assembly of RNA-Seq data without the need for a reference genome. We have applied the Rnnotator assembly pipeline to two yeast transcriptomes and compared the results to the reference gene catalogs of these organisms. The contigs produced by Rnnotator are highly accurate (95percent) and reconstruct full-length genes for the majority of the existing gene models (54.3percent). Furthermore, our analyses revealed many novel transcribed regions that are absent from well annotated genomes, suggesting Rnnotator serves as a complementary approach to analysis based on a reference genome for comprehensive transcriptomics. Conclusions: These results demonstrate that the Rnnotator pipeline is able to reconstruct full-length transcripts in the absence of a complete reference genome.

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

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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.

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

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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.

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.

A practical guide to build de-novo assemblies for single tissues of non-model organisms: the example of a Neotropical frog

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Santiago Montero-Mendieta

2017-09-01

Full Text Available Whole genome sequencing (WGS is a very valuable resource to understand the evolutionary history of poorly known species. However, in organisms with large genomes, as most amphibians, WGS is still excessively challenging and transcriptome sequencing (RNA-seq represents a cost-effective tool to explore genome-wide variability. Non-model organisms do not usually have a reference genome and the transcriptome must be assembled de-novo. We used RNA-seq to obtain the transcriptomic profile for Oreobates cruralis, a poorly known South American direct-developing frog. In total, 550,871 transcripts were assembled, corresponding to 422,999 putative genes. Of those, we identified 23,500, 37,349, 38,120 and 45,885 genes present in the Pfam, EggNOG, KEGG and GO databases, respectively. Interestingly, our results suggested that genes related to immune system and defense mechanisms are abundant in the transcriptome of O. cruralis. We also present a pipeline to assist with pre-processing, assembling, evaluating and functionally annotating a de-novo transcriptome from RNA-seq data of non-model organisms. Our pipeline guides the inexperienced user in an intuitive way through all the necessary steps to build de-novo transcriptome assemblies using readily available software and is freely available at: https://github.com/biomendi/TRANSCRIPTOME-ASSEMBLY-PIPELINE/wiki.

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.

IDBA-tran: a more robust de novo de Bruijn graph assembler for transcriptomes with uneven expression levels.

Science.gov (United States)

Peng, Yu; Leung, Henry C M; Yiu, Siu-Ming; Lv, Ming-Ju; Zhu, Xin-Guang; Chin, Francis Y L

2013-07-01

RNA sequencing based on next-generation sequencing technology is effective for analyzing transcriptomes. Like de novo genome assembly, de novo transcriptome assembly does not rely on any reference genome or additional annotation information, but is more difficult. In particular, isoforms can have very uneven expression levels (e.g. 1:100), which make it very difficult to identify low-expressed isoforms. One challenge is to remove erroneous vertices/edges with high multiplicity (produced by high-expressed isoforms) in the de Bruijn graph without removing correct ones with not-so-high multiplicity from low-expressed isoforms. Failing to do so will result in the loss of low-expressed isoforms or having complicated subgraphs with transcripts of different genes mixed together due to erroneous vertices/edges. Contributions: Unlike existing tools, which remove erroneous vertices/edges with multiplicities lower than a global threshold, we use a probabilistic progressive approach to iteratively remove them with local thresholds. This enables us to decompose the graph into disconnected components, each containing a few genes, if not a single gene, while retaining many correct vertices/edges of low-expressed isoforms. Combined with existing techniques, IDBA-Tran is able to assemble both high-expressed and low-expressed transcripts and outperform existing assemblers in terms of sensitivity and specificity for both simulated and real data. http://www.cs.hku.hk/~alse/idba_tran. Supplementary data are available at Bioinformatics online.

Defining the maize transcriptome de novo using deep RNA-Seq

Energy Technology Data Exchange (ETDEWEB)

Martin, Jeffrey; Gross, Stephen; Choi, Cindy; Zhang, Tao; Lindquist, Erika; Wei, Chia-Lin; Wang, Zhong

2011-06-01

De novo assembly of the transcriptome is crucial for functional genomics studies in bioenergy research, since many of the organisms lack high quality reference genomes. In a previous study we successfully de novo assembled simple eukaryote transcriptomes exclusively from short Illumina RNA-Seq reads [1]. However, extensive alternative splicing, present in most of the higher eukaryotes, poses a significant challenge for current short read assembly processes. Furthermore, the size of next-generation datasets, often large for plant genomes, presents an informatics challenge. To tackle these challenges we present a combined experimental and informatics strategy for de novo assembly in higher eukaryotes. Using maize as a test case, preliminary results suggest our approach can resolve transcript variants and improve gene annotations.

Defining the maize transcriptome de novo using deep RNA-Seq

Energy Technology Data Exchange (ETDEWEB)

Martin, Jeffrey; Gross, Stephen; Choi, Cindy; Zhang, Tao; Lindquist, Erika; Wei, Chia-Lin; Wang, Zhong

2011-06-02

De novo assembly of the transcriptome is crucial for functional genomics studies in bioenergy research, since many of the organisms lack high quality reference genomes. In a previous study we successfully de novo assembled simple eukaryote transcriptomes exclusively from short Illumina RNA-Seq reads [1]. However, extensive alternative splicing, present in most of the higher eukaryotes, poses a significant challenge for current short read assembly processes. Furthermore, the size of next-generation datasets, often large for plant genomes, presents an informatics challenge. To tackle these challenges we present a combined experimental and informatics strategy for de novo assembly in higher eukaryotes. Using maize as a test case, preliminary results suggest our approach can resolve transcript variants and improve gene annotations.

Optimizing Hybrid de Novo Transcriptome Assembly and Extending Genomic Resources for Giant Freshwater Prawns (Macrobrachium rosenbergii: The Identification of Genes and Markers Associated with Reproduction

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Hyungtaek Jung

2016-05-01

Full Text Available The giant freshwater prawn, Macrobrachium rosenbergii, a sexually dimorphic decapod crustacean is currently the world’s most economically important cultured freshwater crustacean species. Despite its economic importance, there is currently a lack of genomic resources available for this species, and this has limited exploration of the molecular mechanisms that control the M. rosenbergii sex-differentiation system more widely in freshwater prawns. Here, we present the first hybrid transcriptome from M. rosenbergii applying RNA-Seq technologies directed at identifying genes that have potential functional roles in reproductive-related traits. A total of 13,733,210 combined raw reads (1720 Mbp were obtained from Ion-Torrent PGM and 454 FLX. Bioinformatic analyses based on three state-of-the-art assemblers, the CLC Genomic Workbench, Trans-ABySS, and Trinity, that use single and multiple k-mer methods respectively, were used to analyse the data. The influence of multiple k-mers on assembly performance was assessed to gain insight into transcriptome assembly from short reads. After optimisation, de novo assembly resulted in 44,407 contigs with a mean length of 437 bp, and the assembled transcripts were further functionally annotated to detect single nucleotide polymorphisms and simple sequence repeat motifs. Gene expression analysis was also used to compare expression patterns from ovary and testis tissue libraries to identify genes with potential roles in reproduction and sex differentiation. The large transcript set assembled here represents the most comprehensive set of transcriptomic resources ever developed for reproduction traits in M. rosenbergii, and the large number of genetic markers predicted should constitute an invaluable resource for future genetic research studies on M. rosenbergii and can be applied more widely on other freshwater prawn species in the genus Macrobrachium.

FMLRC: Hybrid long read error correction using an FM-index.

Science.gov (United States)

Wang, Jeremy R; Holt, James; McMillan, Leonard; Jones, Corbin D

2018-02-09

Long read sequencing is changing the landscape of genomic research, especially de novo assembly. Despite the high error rate inherent to long read technologies, increased read lengths dramatically improve the continuity and accuracy of genome assemblies. However, the cost and throughput of these technologies limits their application to complex genomes. One solution is to decrease the cost and time to assemble novel genomes by leveraging "hybrid" assemblies that use long reads for scaffolding and short reads for accuracy. We describe a novel method leveraging a multi-string Burrows-Wheeler Transform with auxiliary FM-index to correct errors in long read sequences using a set of complementary short reads. We demonstrate that our method efficiently produces significantly more high quality corrected sequence than existing hybrid error-correction methods. We also show that our method produces more contiguous assemblies, in many cases, than existing state-of-the-art hybrid and long-read only de novo assembly methods. Our method accurately corrects long read sequence data using complementary short reads. We demonstrate higher total throughput of corrected long reads and a corresponding increase in contiguity of the resulting de novo assemblies. Improved throughput and computational efficiency than existing methods will help better economically utilize emerging long read sequencing technologies.

De novo transcriptome assembly of the calanoid copepod Neocalanus flemingeri: A new resource for emergence from diapause.

Science.gov (United States)

Roncalli, Vittoria; Cieslak, Matthew C; Sommer, Stephanie A; Hopcroft, Russell R; Lenz, Petra H

2018-02-01

Copepods, small planktonic crustaceans, are key links between primary producers and upper trophic levels, including many economically important fishes. In the subarctic North Pacific, the life cycle of copepods like Neocalanus flemingeri includes an ontogenetic migration to depth followed by a period of diapause (a type of dormancy) characterized by arrested development and low metabolic activity. The end of diapause is marked by the production of the first brood of eggs. Recent temperature anomalies in the North Pacific have raised concerns about potential negative effects on N. flemingeri. Since diapause is a developmental program, its progress can be tracked using through global gene expression. Thus, a reference transcriptome was developed as a first step towards physiological profiling of diapausing females using high-throughput Illumina sequencing. The de novo transcriptome, the first for this species was designed to investigate the diapause period. RNA-Seq reads were obtained for dormant to reproductive N. flemingeri females. A high quality de novo transcriptome was obtained by first assembling reads from each individual using Trinity software followed by clustering with CAP3 Assembly Program. This assembly consisted of 140,841transcripts (contigs). Bench-marking universal single-copy orthologs analysis identified 85% of core eukaryotic genes, with 79% predicted to be complete. Comparison with other calanoid transcriptomes confirmed its quality and degree of completeness. Trinity assembly of reads originating from multiple individuals led to fragmentation. Thus, the workflow applied here differed from the one recommended by Trinity, but was required to obtain a good assembly. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

De novo transcriptome assembly of Setatria italica variety Taejin

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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.

A consensus approach to vertebrate de novo transcriptome assembly from RNA-seq data: Assembly of the duck (Anas platyrhynchos transcriptome.

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Joanna eMoreton

2014-06-01

Full Text Available For vertebrate organisms where a reference genome is not available, de novo transcriptome assembly enables a cost effective insight into the identification of tissue specific or differentially expressed genes and variation of the coding part of the genome. However, since there are a number of different tools and parameters that can be used to reconstruct transcripts, it is difficult to determine an optimal method. Here we suggest a pipeline based on (1 assessing the performance of three different assembly tools (2 using both single and multiple k-mer approaches (3 examining the influence of the number of reads used in the assembly (4 merging assemblies from different tools. We use an example dataset from the vertebrate Anas platyrhynchos domestica (Pekin duck. We find that taking a subset of data enables a robust assembly to be produced by multiple methods without the need for very high memory capacity. The use of reads mapped back to transcripts (RMBT and CEGMA (Core Eukaryotic Genes Mapping Approach provides useful metrics to determine the completeness of assembly obtained. For this dataset the use of multiple k-mers in the assembly generated a more complete assembly as measured by greater number of RMBT and CEGMA score. Merged single k-mer assemblies are generally smaller but consist of longer transcripts, suggesting an assembly consisting of fewer fragmented transcripts. We suggest that the use of a subset of reads during assembly allows the relatively rapid investigation of assembly characteristics and can guide the user to the most appropriate transcriptome for particular downstream use. Transcriptomes generated by the compared assembly methods and the final merged assembly are freely available for download at http://dx.doi.org/10.6084/m9.figshare.1032613.

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

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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...

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

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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

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.

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

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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.

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

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

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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.

De novo assembly of the perennial ryegrass transcriptome using an RNA-seq strategy

DEFF Research Database (Denmark)

Farrell, Jacqueline Danielle; Byrne, Stephen; Paina, Cristiana

2014-01-01

a homozygous perennial ryegrass genotype can circumvent the challenge of heterozygosity. The goals of this study were to perform RNA-sequencing on multiple tissues from a highly inbred genotype to develop a reference transcriptome. This was complemented with RNA-sequencing of a highly heterozygous genotype...... for SNP calling. Result De novo transcriptome assembly of the inbred genotype created 185,833 transcripts with an average length of 830 base pairs. Within the inbred reference transcriptome 78,560 predicted open reading frames were found of which 24,434 were predicted as complete. Functional annotation...... multiple orthologs. Using the longest unique open reading frames as the reference sequence, 64,242 single nucleotide polymorphisms were found. One thousand sixty one open reading frames from the inbred genotype contained heterozygous sites, confirming the high degree of homozygosity. Conclusion Our study...

De novo transcriptome assembly of shrimp Palaemon serratus

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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.

Improving transcriptome de novo assembly by using a reference genome of a related species: Translational genomics from oil palm to coconut.

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Alix Armero

Full Text Available The palms are a family of tropical origin and one of the main constituents of the ecosystems of these regions around the world. The two main species of palm represent different challenges: coconut (Cocos nucifera L. is a source of multiple goods and services in tropical communities, while oil palm (Elaeis guineensis Jacq is the main protagonist of the oil market. In this study, we present a workflow that exploits the comparative genomics between a target species (coconut and a reference species (oil palm to improve the transcriptomic data, providing a proteome useful to answer functional or evolutionary questions. This workflow reduces redundancy and fragmentation, two inherent problems of transcriptomic data, while preserving the functional representation of the target species. Our approach was validated in Arabidopsis thaliana using Arabidopsis lyrata and Capsella rubella as references species. This analysis showed the high sensitivity and specificity of our strategy, relatively independent of the reference proteome. The workflow increased the length of proteins products in A. thaliana by 13%, allowing, often, to recover 100% of the protein sequence length. In addition redundancy was reduced by a factor greater than 3. In coconut, the approach generated 29,366 proteins, 1,246 of these proteins deriving from new contigs obtained with the BRANCH software. The coconut proteome presented a functional profile similar to that observed in rice and an important number of metabolic pathways related to secondary metabolism. The new sequences found with BRANCH software were enriched in functions related to biotic stress. Our strategy can be used as a complementary step to de novo transcriptome assembly to get a representative proteome of a target species. The results of the current analysis are available on the website PalmComparomics (http://palm-comparomics.southgreen.fr/.

Improving transcriptome de novo assembly by using a reference genome of a related species: Translational genomics from oil palm to coconut.

Science.gov (United States)

Armero, Alix; Baudouin, Luc; Bocs, Stéphanie; This, Dominique

2017-01-01

The palms are a family of tropical origin and one of the main constituents of the ecosystems of these regions around the world. The two main species of palm represent different challenges: coconut (Cocos nucifera L.) is a source of multiple goods and services in tropical communities, while oil palm (Elaeis guineensis Jacq) is the main protagonist of the oil market. In this study, we present a workflow that exploits the comparative genomics between a target species (coconut) and a reference species (oil palm) to improve the transcriptomic data, providing a proteome useful to answer functional or evolutionary questions. This workflow reduces redundancy and fragmentation, two inherent problems of transcriptomic data, while preserving the functional representation of the target species. Our approach was validated in Arabidopsis thaliana using Arabidopsis lyrata and Capsella rubella as references species. This analysis showed the high sensitivity and specificity of our strategy, relatively independent of the reference proteome. The workflow increased the length of proteins products in A. thaliana by 13%, allowing, often, to recover 100% of the protein sequence length. In addition redundancy was reduced by a factor greater than 3. In coconut, the approach generated 29,366 proteins, 1,246 of these proteins deriving from new contigs obtained with the BRANCH software. The coconut proteome presented a functional profile similar to that observed in rice and an important number of metabolic pathways related to secondary metabolism. The new sequences found with BRANCH software were enriched in functions related to biotic stress. Our strategy can be used as a complementary step to de novo transcriptome assembly to get a representative proteome of a target species. The results of the current analysis are available on the website PalmComparomics (http://palm-comparomics.southgreen.fr/).

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...

Identification and assembly of genomes and genetic elements in complex metagenomic samples without using reference genomes.

Science.gov (United States)

Nielsen, H Bjørn; Almeida, Mathieu; Juncker, Agnieszka Sierakowska; Rasmussen, Simon; Li, Junhua; Sunagawa, Shinichi; Plichta, Damian R; Gautier, Laurent; Pedersen, Anders G; Le Chatelier, Emmanuelle; Pelletier, Eric; Bonde, Ida; Nielsen, Trine; Manichanh, Chaysavanh; Arumugam, Manimozhiyan; Batto, Jean-Michel; Quintanilha Dos Santos, Marcelo B; Blom, Nikolaj; Borruel, Natalia; Burgdorf, Kristoffer S; Boumezbeur, Fouad; Casellas, Francesc; Doré, Joël; Dworzynski, Piotr; Guarner, Francisco; Hansen, Torben; Hildebrand, Falk; Kaas, Rolf S; Kennedy, Sean; Kristiansen, Karsten; Kultima, Jens Roat; Léonard, Pierre; Levenez, Florence; Lund, Ole; Moumen, Bouziane; Le Paslier, Denis; Pons, Nicolas; Pedersen, Oluf; Prifti, Edi; Qin, Junjie; Raes, Jeroen; Sørensen, Søren; Tap, Julien; Tims, Sebastian; Ussery, David W; Yamada, Takuji; Renault, Pierre; Sicheritz-Ponten, Thomas; Bork, Peer; Wang, Jun; Brunak, Søren; Ehrlich, S Dusko

2014-08-01

Most current approaches for analyzing metagenomic data rely on comparisons to reference genomes, but the microbial diversity of many environments extends far beyond what is covered by reference databases. De novo segregation of complex metagenomic data into specific biological entities, such as particular bacterial strains or viruses, remains a largely unsolved problem. Here we present a method, based on binning co-abundant genes across a series of metagenomic samples, that enables comprehensive discovery of new microbial organisms, viruses and co-inherited genetic entities and aids assembly of microbial genomes without the need for reference sequences. We demonstrate the method on data from 396 human gut microbiome samples and identify 7,381 co-abundance gene groups (CAGs), including 741 metagenomic species (MGS). We use these to assemble 238 high-quality microbial genomes and identify affiliations between MGS and hundreds of viruses or genetic entities. Our method provides the means for comprehensive profiling of the diversity within complex metagenomic samples.

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

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

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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

Efficient assembly of de novo human artificial chromosomes from large genomic loci

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Stromberg Gregory

2005-07-01

Full Text Available Abstract Background Human Artificial Chromosomes (HACs are potentially useful vectors for gene transfer studies and for functional annotation of the genome because of their suitability for cloning, manipulating and transferring large segments of the genome. However, development of HACs for the transfer of large genomic loci into mammalian cells has been limited by difficulties in manipulating high-molecular weight DNA, as well as by the low overall frequencies of de novo HAC formation. Indeed, to date, only a small number of large (>100 kb genomic loci have been reported to be successfully packaged into de novo HACs. Results We have developed novel methodologies to enable efficient assembly of HAC vectors containing any genomic locus of interest. We report here the creation of a novel, bimolecular system based on bacterial artificial chromosomes (BACs for the construction of HACs incorporating any defined genomic region. We have utilized this vector system to rapidly design, construct and validate multiple de novo HACs containing large (100–200 kb genomic loci including therapeutically significant genes for human growth hormone (HGH, polycystic kidney disease (PKD1 and ß-globin. We report significant differences in the ability of different genomic loci to support de novo HAC formation, suggesting possible effects of cis-acting genomic elements. Finally, as a proof of principle, we have observed sustained ß-globin gene expression from HACs incorporating the entire 200 kb ß-globin genomic locus for over 90 days in the absence of selection. Conclusion Taken together, these results are significant for the development of HAC vector technology, as they enable high-throughput assembly and functional validation of HACs containing any large genomic locus. We have evaluated the impact of different genomic loci on the frequency of HAC formation and identified segments of genomic DNA that appear to facilitate de novo HAC formation. These genomic loci

De novo centriole formation in human cells is error-prone and does not require SAS-6 self-assembly.

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Wang, Won-Jing; Acehan, Devrim; Kao, Chien-Han; Jane, Wann-Neng; Uryu, Kunihiro; Tsou, Meng-Fu Bryan

2015-11-26

Vertebrate centrioles normally propagate through duplication, but in the absence of preexisting centrioles, de novo synthesis can occur. Consistently, centriole formation is thought to strictly rely on self-assembly, involving self-oligomerization of the centriolar protein SAS-6. Here, through reconstitution of de novo synthesis in human cells, we surprisingly found that normal looking centrioles capable of duplication and ciliation can arise in the absence of SAS-6 self-oligomerization. Moreover, whereas canonically duplicated centrioles always form correctly, de novo centrioles are prone to structural errors, even in the presence of SAS-6 self-oligomerization. These results indicate that centriole biogenesis does not strictly depend on SAS-6 self-assembly, and may require preexisting centrioles to ensure structural accuracy, fundamentally deviating from the current paradigm.

A glance at quality score: implication for de novo transcriptome reconstruction of Illumina reads

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Stanley Kimbung Mbandi

2014-02-01

Full Text Available Downstream analyses of short-reads from next-generation sequencing platforms are often preceded by a pre-processing step that removes uncalled and wrongly called bases. Standard approaches rely on their associated base quality scores to retain the read or a portion of it when the score is above a predefined threshold. It is difficult to differentiate sequencing error from biological variation without a reference using quality scores. The effects of quality score based trimming have not been systematically studied in de novo transcriptome assembly. Using RNA-Seq data produced from Illumina, we teased out the effects of quality score base filtering or trimming on de novo transcriptome reconstruction. We showed that assemblies produced from reads subjected to different quality score thresholds contain truncated and missing transfrags when compared to those from untrimmed reads. Our data supports the fact that de novo assembling of untrimmed data is challenging for de Bruijn graph assemblers. However, our results indicates that comparing the assemblies from untrimmed and trimmed read subsets can suggest appropriate filtering parameters and enable selection of the optimum de novo transcriptome assembly in non-model organisms.

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

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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.

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.

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...

Comparing de novo assemblers for 454 transcriptome data

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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

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.

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.

ScanIndel: a hybrid framework for indel detection via gapped alignment, split reads and de novo assembly.

Science.gov (United States)

Yang, Rendong; Nelson, Andrew C; Henzler, Christine; Thyagarajan, Bharat; Silverstein, Kevin A T

2015-12-07

Comprehensive identification of insertions/deletions (indels) across the full size spectrum from second generation sequencing is challenging due to the relatively short read length inherent in the technology. Different indel calling methods exist but are limited in detection to specific sizes with varying accuracy and resolution. We present ScanIndel, an integrated framework for detecting indels with multiple heuristics including gapped alignment, split reads and de novo assembly. Using simulation data, we demonstrate ScanIndel's superior sensitivity and specificity relative to several state-of-the-art indel callers across various coverage levels and indel sizes. ScanIndel yields higher predictive accuracy with lower computational cost compared with existing tools for both targeted resequencing data from tumor specimens and high coverage whole-genome sequencing data from the human NIST standard NA12878. Thus, we anticipate ScanIndel will improve indel analysis in both clinical and research settings. ScanIndel is implemented in Python, and is freely available for academic use at https://github.com/cauyrd/ScanIndel.

De novo transcriptome assembly of Sorghum bicolor variety Taejin

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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.

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

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.

De novo transcriptome assembly of two Vigna angularis varieties collected from Korea

Directory of Open Access Journals (Sweden)

Yeonhwa Jo

2016-06-01

Full Text Available The adzuki bean (Vigna angularis, a member of the family Fabaceae, is widely grown in Asia, from East Asia to the Himalayas. The adzuki bean is known as an ingredient that adds sweetness to diverse desserts made in Eastern Asian countries. Libraries prepared from two V. angularis varieties referred to as Taejin Black and Taejin Red were paired-end sequenced using the Illumina HiSeq 2000 system. The raw data in this study can be available in NCBI SRA database with accession numbers of SRR3406660 and SRR3406553. After de novo transcriptome assembly using Trinity, we obtained 324,219 and 280,056 transcripts from Taejin Black and Taejin Red, respectively. We predicted a total of 238,321 proteins and 179,519 proteins for Taejin Black and Taejin Red, respectively, by the TransDecoder program. We carried out BLASTP on the predicted proteins against the Swiss-Prot protein sequence database to predict the putative functions of identified proteins. Taken together, we provide transcriptomes of two adzuki bean varieties by RNA-Seq, which might be usefully applied to generate molecular markers.

De novo assembly of plant body plan: a step ahead of Deadpool.

Science.gov (United States)

Kareem, Abdul; Radhakrishnan, Dhanya; Sondhi, Yash; Aiyaz, Mohammed; Roy, Merin V; Sugimoto, Kaoru; Prasad, Kalika

2016-08-01

While in the movie Deadpool it is possible for a human to recreate an arm from scratch, in reality plants can even surpass that. Not only can they regenerate lost parts, but also the whole plant body can be reborn from a few existing cells. Despite the decades old realization that plant cells possess the ability to regenerate a complete shoot and root system, it is only now that the underlying mechanisms are being unraveled. De novo plant regeneration involves the initiation of regenerative mass, acquisition of the pluripotent state, reconstitution of stem cells and assembly of regulatory interactions. Recent studies have furthered our understanding on the making of a complete plant system in the absence of embryonic positional cues. We review the recent studies probing the molecular mechanisms of de novo plant regeneration in response to external inductive cues and our current knowledge of direct reprogramming of root to shoot and vice versa. We further discuss how de novo regeneration can be exploited to meet the demands of green culture industries and to serve as a general model to address the fundamental questions of regeneration across the plant kingdom.

De Novo Centromere Formation and Centromeric Sequence Expansion in Wheat and its Wide Hybrids.

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Xiang Guo

2016-04-01

Full Text Available Centromeres typically contain tandem repeat sequences, but centromere function does not necessarily depend on these sequences. We identified functional centromeres with significant quantitative changes in the centromeric retrotransposons of wheat (CRW contents in wheat aneuploids (Triticum aestivum and the offspring of wheat wide hybrids. The CRW signals were strongly reduced or essentially lost in some wheat ditelosomic lines and in the addition lines from the wide hybrids. The total loss of the CRW sequences but the presence of CENH3 in these lines suggests that the centromeres were formed de novo. In wheat and its wide hybrids, which carry large complex genomes or no sequenced genome, we performed CENH3-ChIP-dot-blot methods alone or in combination with CENH3-ChIP-seq and identified the ectopic genomic sequences present at the new centromeres. In adcdition, the transcription of the identified DNA sequences was remarkably increased at the new centromere, suggesting that the transcription of the corresponding sequences may be associated with de novo centromere formation. Stable alien chromosomes with two and three regions containing CRW sequences induced by centromere breakage were observed in the wheat-Th. elongatum hybrid derivatives, but only one was a functional centromere. In wheat-rye (Secale cereale hybrids, the rye centromere-specific sequences spread along the chromosome arms and may have caused centromere expansion. Frequent and significant quantitative alterations in the centromere sequence via chromosomal rearrangement have been systematically described in wheat wide hybridizations, which may affect the retention or loss of the alien chromosomes in the hybrids. Thus, the centromere behavior in wide crosses likely has an important impact on the generation of biodiversity, which ultimately has implications for speciation.

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...

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.

Digital gene expression analysis based on integrated de novo transcriptome assembly of sweet potato [Ipomoea batatas (L. Lam].

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Xiang Tao

Full Text Available BACKGROUND: Sweet potato (Ipomoea batatas L. [Lam.] ranks among the top six most important food crops in the world. It is widely grown throughout the world with high and stable yield, strong adaptability, rich nutrient content, and multiple uses. However, little is known about the molecular biology of this important non-model organism due to lack of genomic resources. Hence, studies based on high-throughput sequencing technologies are needed to get a comprehensive and integrated genomic resource and better understanding of gene expression patterns in different tissues and at various developmental stages. METHODOLOGY/PRINCIPAL FINDINGS: Illumina paired-end (PE RNA-Sequencing was performed, and generated 48.7 million of 75 bp PE reads. These reads were de novo assembled into 128,052 transcripts (≥ 100 bp, which correspond to 41.1 million base pairs, by using a combined assembly strategy. Transcripts were annotated by Blast2GO and 51,763 transcripts got BLASTX hits, in which 39,677 transcripts have GO terms and 14,117 have ECs that are associated with 147 KEGG pathways. Furthermore, transcriptome differences of seven tissues were analyzed by using Illumina digital gene expression (DGE tag profiling and numerous differentially and specifically expressed transcripts were identified. Moreover, the expression characteristics of genes involved in viral genomes, starch metabolism and potential stress tolerance and insect resistance were also identified. CONCLUSIONS/SIGNIFICANCE: The combined de novo transcriptome assembly strategy can be applied to other organisms whose reference genomes are not available. The data provided here represent the most comprehensive and integrated genomic resources for cloning and identifying genes of interest in sweet potato. Characterization of sweet potato transcriptome provides an effective tool for better understanding the molecular mechanisms of cellular processes including development of leaves and storage roots

De Novo Assembly and Characterization of the Transcriptome of Grasshopper Shirakiacris shirakii

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Zhongying Qiu

2016-07-01

Full Text Available Background: The grasshopper Shirakiacris shirakii is an important agricultural pest and feeds mainly on gramineous plants, thereby causing economic damage to a wide range of crops. However, genomic information on this species is extremely limited thus far, and transcriptome data relevant to insecticide resistance and pest control are also not available. Methods: The transcriptome of S. shirakii was sequenced using the Illumina HiSeq platform, and we de novo assembled the transcriptome. Results: Its sequencing produced a total of 105,408,878 clean reads, and the de novo assembly revealed 74,657 unigenes with an average length of 680 bp and N50 of 1057 bp. A total of 28,173 unigenes were annotated for the NCBI non-redundant protein sequences (Nr, NCBI non-redundant nucleotide sequences (Nt, a manually-annotated and reviewed protein sequence database (Swiss-Prot, Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG databases. Based on the Nr annotation results, we manually identified 79 unigenes encoding cytochrome P450 monooxygenases (P450s, 36 unigenes encoding carboxylesterases (CarEs and 36 unigenes encoding glutathione S-transferases (GSTs in S. shirakii. Core RNAi components relevant to miroRNA, siRNA and piRNA pathways, including Pasha, Loquacious, Argonaute-1, Argonaute-2, Argonaute-3, Zucchini, Aubergine, enhanced RNAi-1 and Piwi, were expressed in S. shirakii. We also identified five unigenes that were homologous to the Sid-1 gene. In addition, the analysis of differential gene expressions revealed that a total of 19,764 unigenes were up-regulated and 4185 unigenes were down-regulated in larvae. In total, we predicted 7504 simple sequence repeats (SSRs from 74,657 unigenes. Conclusions: The comprehensive de novo transcriptomic data of S. shirakii will offer a series of valuable molecular resources for better studying insecticide resistance, RNAi and molecular marker discovery in the transcriptome.

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.

Characterization of a de novo duplication of 11p14----p13, using fluorescent in situ hybridization and southern hybridization

NARCIS (Netherlands)

Speleman, F.; Mannens, M.; Redeker, B.; Vercruyssen, M.; van Oostveldt, P.; Leroy, J.; Slater, R.

1991-01-01

A de novo 11p+ chromosome was found in a child with mild mental retardation but no other remarkable dysmorphic characteristics. Banding studies suggested a duplication of regions 11p13 and 11p14 or regions 11p14 and 11p15. Using fluorescent in situ hybridization and digital imaging microscopy, we

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

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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.

Finding Nemo's Genes: A chromosome-scale reference assembly of the genome of the orange clownfish Amphiprion percula

KAUST Repository

Lehmann, Robert; Lightfoot, Damien J; Schunter, Celia Marei; Michell, Craig T; Ohyanagi, Hajime; Mineta, Katsuhiko; Foret, Sylvain; Berumen, Michael L.; Miller, David J; Aranda, Manuel; Gojobori, Takashi; Munday, Philip L; Ravasi, Timothy

2018-01-01

The iconic orange clownfish, Amphiprion percula, is a model organism for studying the ecology and evolution of reef fishes, including patterns of population connectivity, sex change, social organization, habitat selection and adaptation to climate change. Notably, the orange clownfish is the only reef fish for which a complete larval dispersal kernel has been established and was the first fish species for which it was demonstrated that anti-predator responses of reef fishes could be impaired by ocean acidification. Despite its importance, molecular resources for this species remain scarce and until now it lacked a reference genome assembly. Here we present a de novo chromosome-scale assembly of the genome of the orange clownfish Amphiprion percula. We utilized single-molecule real-time sequencing technology from Pacific Biosciences to produce an initial polished assembly comprised of 1,414 contigs, with a contig N50 length of 1.86 Mb. Using Hi-C based chromatin contact maps, 98% of the genome assembly were placed into 24 chromosomes, resulting in a final assembly of 908.8 Mb in length with contig and scaffold N50s of 3.12 and 38.4 Mb, respectively. This makes it one of the most contiguous and complete fish genome assemblies currently available. The genome was annotated with 26,597 protein coding genes and contains 96% of the core set of conserved actinopterygian orthologs. The availability of this reference genome assembly as a community resource will further strengthen the role of the orange clownfish as a model species for research on the ecology and evolution of reef fishes.

Finding Nemo's Genes: A chromosome-scale reference assembly of the genome of the orange clownfish Amphiprion percula

KAUST Repository

Lehmann, Robert

2018-03-08

The iconic orange clownfish, Amphiprion percula, is a model organism for studying the ecology and evolution of reef fishes, including patterns of population connectivity, sex change, social organization, habitat selection and adaptation to climate change. Notably, the orange clownfish is the only reef fish for which a complete larval dispersal kernel has been established and was the first fish species for which it was demonstrated that anti-predator responses of reef fishes could be impaired by ocean acidification. Despite its importance, molecular resources for this species remain scarce and until now it lacked a reference genome assembly. Here we present a de novo chromosome-scale assembly of the genome of the orange clownfish Amphiprion percula. We utilized single-molecule real-time sequencing technology from Pacific Biosciences to produce an initial polished assembly comprised of 1,414 contigs, with a contig N50 length of 1.86 Mb. Using Hi-C based chromatin contact maps, 98% of the genome assembly were placed into 24 chromosomes, resulting in a final assembly of 908.8 Mb in length with contig and scaffold N50s of 3.12 and 38.4 Mb, respectively. This makes it one of the most contiguous and complete fish genome assemblies currently available. The genome was annotated with 26,597 protein coding genes and contains 96% of the core set of conserved actinopterygian orthologs. The availability of this reference genome assembly as a community resource will further strengthen the role of the orange clownfish as a model species for research on the ecology and evolution of reef fishes.

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.

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.

Preliminary neutronics calculation of fusion-fission hybrid reactor breeding spent fuel assembly

International Nuclear Information System (INIS)

Ma Xubo; Chen Yixue; Gao Bin

2013-01-01

The possibility of using the fusion-fission hybrid reactor breeding spent fuel in PWR was preliminarily studied in this paper. According to the fusion-fission hybrid reactor breeding spent fuel characteristics, PWR assembly including fusion-fission hybrid reactor breeding spent fuel was designed. The parameters such as fuel temperature coefficient, moderator temperature coefficient and their variation were investigated. Results show that the neutron properties of uranium-based assembly and hybrid reactor breeding spent fuel assembly are similar. The design of this paper has a smaller uniformity coefficient of power at the same fissile isotope mass percentage. The results will provide technical support for the future fusion-fission hybrid reactor and PWR combined with cycle system. (authors)

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.

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.

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.

Generation of Hybrid Peptide-Silver Nanoparticles for Antibacterial and Antifouling Applications

KAUST Repository

Seferji, Kholoud

2018-01-01

and antifouling agents. Our innovative antibacterial agents are hybrid peptide silver nanoparticles (CH-01-AgNPs) that are created de novo and in situ from a silver nitrate solution (AgNO3) in the presence of ultrashort self-assembling peptides compounds

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.

Fuzzy-Based Adaptive Hybrid Burst Assembly Technique for Optical Burst Switched Networks

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Abubakar Muhammad Umaru

2014-01-01

Full Text Available The optical burst switching (OBS paradigm is perceived as an intermediate switching technology for future all-optical networks. Burst assembly that is the first process in OBS is the focus of this paper. In this paper, an intelligent hybrid burst assembly algorithm that is based on fuzzy logic is proposed. The new algorithm is evaluated against the traditional hybrid burst assembly algorithm and the fuzzy adaptive threshold (FAT burst assembly algorithm via simulation. Simulation results show that the proposed algorithm outperforms the hybrid and the FAT algorithms in terms of burst end-to-end delay, packet end-to-end delay, and packet loss ratio.

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.

De novo assembly and annotation of the Antarctic copepod (Tigriopus kingsejongensis) transcriptome.

Science.gov (United States)

Kim, Hui-Su; Lee, Bo-Young; Han, Jeonghoon; Lee, Young Hwan; Min, Gi-Sik; Kim, Sanghee; Lee, Jae-Seong

2016-08-01

The whole transcriptome of the Antarctic copepod (Tigriopus kingsejongensis) was sequenced using Illumina RNA-seq. De novo assembly was performed with 64,785,098 raw reads using Trinity, which assembled into 81,653 contigs. TransDecoder found 38,250 candidate coding contigs which showed homology to other species by BLAST analysis. Functional gene annotation was performed by Gene Ontology (GO), InterProScan, and KEGG pathway analyses. Finally, we identified a number of expressed gene catalog for T. kingsejongensis that is a useful model animal for gene information-based polar research to uncover molecular mechanisms of environmental adaptation on harsh environments. In particular, we observed highly developing lipid metabolism in T. kingsejongensis directly compared to those of the Far East Pacific coast copepod Tigriopus japonicus at the transcriptome level. Copyright © 2016 Elsevier B.V. All rights reserved.

Structural variation in two human genomes mapped at single-nucleotide resolution by whole genome de novo assembly

DEFF Research Database (Denmark)

Li, Yingrui; Zheng, Hancheng; Luo, Ruibang

2011-01-01

Here we use whole-genome de novo assembly of second-generation sequencing reads to map structural variation (SV) in an Asian genome and an African genome. Our approach identifies small- and intermediate-size homozygous variants (1-50 kb) including insertions, deletions, inversions and their precise...

De novo transcriptome assembly of drought tolerant CAM plants, Agave deserti and Agave tequilana.

Science.gov (United States)

Gross, Stephen M; Martin, Jeffrey A; Simpson, June; Abraham-Juarez, María Jazmín; Wang, Zhong; Visel, Axel

2013-08-19

Agaves are succulent monocotyledonous plants native to xeric environments of North America. Because of their adaptations to their environment, including crassulacean acid metabolism (CAM, a water-efficient form of photosynthesis), and existing technologies for ethanol production, agaves have gained attention both as potential lignocellulosic bioenergy feedstocks and models for exploring plant responses to abiotic stress. However, the lack of comprehensive Agave sequence datasets limits the scope of investigations into the molecular-genetic basis of Agave traits. Here, we present comprehensive, high quality de novo transcriptome assemblies of two Agave species, A. tequilana and A. deserti, built from short-read RNA-seq data. Our analyses support completeness and accuracy of the de novo transcriptome assemblies, with each species having a minimum of approximately 35,000 protein-coding genes. Comparison of agave proteomes to those of additional plant species identifies biological functions of gene families displaying sequence divergence in agave species. Additionally, a focus on the transcriptomics of the A. deserti juvenile leaf confirms evolutionary conservation of monocotyledonous leaf physiology and development along the proximal-distal axis. Our work presents a comprehensive transcriptome resource for two Agave species and provides insight into their biology and physiology. These resources are a foundation for further investigation of agave biology and their improvement for bioenergy development.

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

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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.

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

De novo transcriptome assembly of the mycoheterotrophic plant Monotropa hypopitys

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Alexey V. Beletsky

2017-03-01

Full Text Available Monotropa hypopitys (pinesap is a non-photosynthetic obligately mycoheterotrophic plant of the family Ericaceae. It obtains the carbon and other nutrients from the roots of surrounding autotrophic trees through the associated mycorrhizal fungi. In order to understand the evolutionary changes in the plant genome associated with transition to a heterotrophic lifestyle, we performed de novo transcriptomic analysis of M. hypopitys using next-generation sequencing. We obtained the RNA-Seq data from flowers, flower bracts and roots with haustoria using Illumina HiSeq2500 platform. The raw data obtained in this study can be available in NCBI SRA database with accession number of SRP069226. A total of 10.3 GB raw sequence data were obtained, corresponding to 103,357,809 raw reads. A total of 103,025,683 reads were filtered after removing low-quality reads and trimming the adapter sequences. The Trinity program was used to de novo assemble 98,349 unigens with an N50 of 1342 bp. Using the TransDecoder program, we predicted 43,505 putative proteins. 38,416 unigenes were annotated in the Swiss-Prot protein sequence database using BLASTX. The obtained transcriptomic data will be useful for further studies of the evolution of plant genomes upon transition to a non-photosynthetic lifestyle and the loss of photosynthesis-related functions.

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.

De novo assembly of Phlomis purpurea after challenging with Phytophthora cinnamomi.

Science.gov (United States)

Baldé, Aladje; Neves, Dina; García-Breijo, Francisco J; Pais, Maria Salomé; Cravador, Alfredo

2017-09-06

Phlomis plants are a source of biological active substances with potential applications in the control of phytopathogens. Phlomis purpurea (Lamiaceae) is autochthonous of southern Iberian Peninsula and Morocco and was found to be resistant to Phytophthora cinnamomi. Phlomis purpurea has revealed antagonistic effect in the rhizosphere of Quercus suber and Q. ilex against P. cinnamomi. Phlomis purpurea roots produce bioactive compounds exhibiting antitumor and anti-Phytophthora activities with potential to protect susceptible plants. Although these important capacities of P. purpurea have been demonstrated, there is no transcriptomic or genomic information available in public databases that could bring insights on the genes underlying this anti-oomycete activity. Using Illumina technology we obtained a de novo assembly of P. purpurea transcriptome and differential transcript abundance to identify putative defence related genes in challenged versus non-challenged plants. A total of 1,272,600,000 reads from 18 cDNA libraries were merged and assembled into 215,739 transcript contigs. BLASTX alignment to Nr NCBI database identified 124,386 unique annotated transcripts (57.7%) with significant hits. Functional annotation identified 83,550 out of 124,386 unique transcripts, which were mapped to 141 pathways. 39% of unigenes were assigned GO terms. Their functions cover biological processes, cellular component and molecular functions. Genes associated with response to stimuli, cellular and primary metabolic processes, catalytic and transporter functions were among those identified. Differential transcript abundance analysis using DESeq revealed significant differences among libraries depending on post-challenge times. Comparative cyto-histological studies of P. purpurea roots challenged with P. cinnamomi zoospores and controls revealed specific morphological features (exodermal strips and epi-cuticular layer), that may provide a constitutive efficient barrier against

De novo transcriptome assembly associated with fumonisin production by the rice pathogen Fusarium fujikuroi

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Keerthi S. Guruge

2018-06-01

Full Text Available The present study employed a next-generation sequencing method to assemble a de novo transcriptome database designed to distinguish gene expression changes exhibited by the fumonisin-producing fungus Fusarium fujikuroi when grown under ‘fumonisin-producing’ compared to ‘non-fumonisin-producing’ conditions. The raw data of this study have been deposited at DNA Data Bank of Japan (DDBJ under the accession ID DRA006146. Keywords: Fusarium fujikuroi, Fumonisin, Next-generation sequencing, Transcriptome, Gene-expression

Anisotropic Self-Assembly of Organic–Inorganic Hybrid Microtoroids

KAUST Repository

Al-Rehili, Safa’a

2016-10-24

Toroidal structures based on self-assembly of predesigned building blocks are well-established in the literature, but spontaneous self-organization to prepare such structures has not been reported to date. Here, organic–inorganic hybrid microtoroids synthesized by simultaneous coordination-driven assembly of amphiphilic molecules and hydrophilic polymers are reported. Mixing amphiphilic molecules with iron(III) chloride and hydrophilic polymers in water leads, within minutes, to the formation of starlike nanostructures. A spontaneous self-organization of these nanostructures is then triggered to form stable hybrid microtoroids. Interestingly, the toroids exhibit anisotropic hierarchical growth, giving rise to a layered toroidal framework. These microstructures are mechanically robust and can act as templates to host metallic nanoparticles such as gold and silver. Understanding the nature of spontaneous assembly driven by coordination multiple non-covalent interactions can help explain the well-ordered complexity of many biological organisms in addition to expanding the available tools to mimic such structures at a molecular level.

Assembly of the Boechera retrofracta Genome and Evolutionary Analysis of Apomixis-Associated Genes

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Sergei Kliver

2018-03-01

Full Text Available Closely related to the model plant Arabidopsis thaliana, the genus Boechera is known to contain both sexual and apomictic species or accessions. Boechera retrofracta is a diploid sexually reproducing species and is thought to be an ancestral parent species of apomictic species. Here we report the de novo assembly of the B. retrofracta genome using short Illumina and Roche reads from 1 paired-end and 3 mate pair libraries. The distribution of 23-mers from the paired end library has indicated a low level of heterozygosity and the presence of detectable duplications and triplications. The genome size was estimated to be equal 227 Mb. N50 of the assembled scaffolds was 2.3 Mb. Using a hybrid approach that combines homology-based and de novo methods 27,048 protein-coding genes were predicted. Also repeats, transfer RNA (tRNA and ribosomal RNA (rRNA genes were annotated. Finally, genes of B. retrofracta and 6 other Brassicaceae species were used for phylogenetic tree reconstruction. In addition, we explored the histidine exonuclease APOLLO locus, related to apomixis in Boechera, and proposed model of its evolution through the series of duplications. An assembled genome of B. retrofracta will help in the challenging assembly of the highly heterozygous genomes of hybrid apomictic species.

Comparing genome guided assembly and phased variants based assembly approach to separate the homoeolog transcripts in tetraploid peanut (Arachis hypogaea L.)

Science.gov (United States)

Homoeologous copies of transcripts are abundant in many self-pollinating species including tetraploid peanut, and can impose a challenge to build a transcriptome reference without the merging of homoeologs. De novo transcriptome assembly of tetraploid OLin with single kmer and multiple kmer approach...

De novo transcriptome assembly of ‘Angeleno’ and ‘Lamoon’ Japanese plum cultivars (Prunus salicina

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Máximo González

2016-09-01

De novo transcriptome assembly was performed using CLC Genome Workbench software and a total of 54,584 unique contigs were generated, with an N50 of 1343 base pair (bp and a mean length of 829 bp. This work contributed with a specific Japanese plum skin transcriptome, providing two libraries of contrasting fruit skin color phenotype (yellow and red and increasing substantially the GB of raw data available until now for this specie.

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.

Calibration of the TVO spent BWR reference fuel assembly

International Nuclear Information System (INIS)

Tarvainen, M.; Baecklin, A.; Haakanson, A.

1992-02-01

In 1989 the Support Programmes of Finland (FSP) and Sweden (SSP) initiated a joint task to cross calibrate the burnup of the IAEA spent BWR reference fuel assembly at the TVO AFR storage facility (TVO KPA-STORE) in Finland. The reference assembly, kept separately under the IAEA seal, is used for verification measurements of spent fuel by GBUV method (SG-NDA-38). The cross calibration was performed by establishing a calibration curve, 244 Cm neutron rate versus burnup, using passive neutron assay (PNA) measurements. The declared burnup of the reference assembly was compared with the burnup value deduced from the calibration curve. A calibration line was also established by using the GBUV method with the aid of high resolution gamma ray spectrometry (HRGS). Normalization between the two different facilities was performed using sealed neutron and gamma calibration sources. The results of the passive neutron assay show consistency, better than 1 %, between the declared mean burnup of the reference assembly and the burnup deduced from the calibration curve. The corresponding consistency is within +-2 % for the HRGS measurements

De novo-based transcriptome profiling of male-sterile and fertile watermelon lines.

Science.gov (United States)

Rhee, Sun-Ju; Kwon, Taehyung; Seo, Minseok; Jang, Yoon Jeong; Sim, Tae Yong; Cho, Seoae; Han, Sang-Wook; Lee, Gung Pyo

2017-01-01

The whole-genome sequence of watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai), a valuable horticultural crop worldwide, was released in 2013. Here, we compared a de novo-based approach (DBA) to a reference-based approach (RBA) using RNA-seq data, to aid in efforts to improve the annotation of the watermelon reference genome and to obtain biological insight into male-sterility in watermelon. We applied these techniques to available data from two watermelon lines: the male-sterile line DAH3615-MS and the male-fertile line DAH3615. Using DBA, we newly annotated 855 watermelon transcripts, and found gene functional clusters predicted to be related to stimulus responses, nucleic acid binding, transmembrane transport, homeostasis, and Golgi/vesicles. Among the DBA-annotated transcripts, 138 de novo-exclusive differentially-expressed genes (DEDEGs) related to male sterility were detected. Out of 33 randomly selected newly annotated transcripts and DEDEGs, 32 were validated by RT-qPCR. This study demonstrates the usefulness and reliability of the de novo transcriptome assembly in watermelon, and provides new insights for researchers exploring transcriptional blueprints with regard to the male sterility.

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

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

RNA-seq analysis of Quercus pubescens Leaves: de novo transcriptome assembly, annotation and functional markers development.

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Sara Torre

Full Text Available Quercus pubescens Willd., a species distributed from Spain to southwest Asia, ranks high for drought tolerance among European oaks. Q. pubescens performs a role of outstanding significance in most Mediterranean forest ecosystems, but few mechanistic studies have been conducted to explore its response to environmental constrains, due to the lack of genomic resources. In our study, we performed a deep transcriptomic sequencing in Q. pubescens leaves, including de novo assembly, functional annotation and the identification of new molecular markers. Our results are a pre-requisite for undertaking molecular functional studies, and may give support in population and association genetic studies. 254,265,700 clean reads were generated by the Illumina HiSeq 2000 platform, with an average length of 98 bp. De novo assembly, using CLC Genomics, produced 96,006 contigs, having a mean length of 618 bp. Sequence similarity analyses against seven public databases (Uniprot, NR, RefSeq and KOGs at NCBI, Pfam, InterPro and KEGG resulted in 83,065 transcripts annotated with gene descriptions, conserved protein domains, or gene ontology terms. These annotations and local BLAST allowed identify genes specifically associated with mechanisms of drought avoidance. Finally, 14,202 microsatellite markers and 18,425 single nucleotide polymorphisms (SNPs were, in silico, discovered in assembled and annotated sequences. We completed a successful global analysis of the Q. pubescens leaf transcriptome using RNA-seq. The assembled and annotated sequences together with newly discovered molecular markers provide genomic information for functional genomic studies in Q. pubescens, with special emphasis to response mechanisms to severe constrain of the Mediterranean climate. Our tools enable comparative genomics studies on other Quercus species taking advantage of large intra-specific ecophysiological differences.

Playing hide and seek with repeats in local and global de novo transcriptome assembly of short RNA-seq reads.

Science.gov (United States)

Lima, Leandro; Sinaimeri, Blerina; Sacomoto, Gustavo; Lopez-Maestre, Helene; Marchet, Camille; Miele, Vincent; Sagot, Marie-France; Lacroix, Vincent

2017-01-01

The main challenge in de novo genome assembly of DNA-seq data is certainly to deal with repeats that are longer than the reads. In de novo transcriptome assembly of RNA-seq reads, on the other hand, this problem has been underestimated so far. Even though we have fewer and shorter repeated sequences in transcriptomics, they do create ambiguities and confuse assemblers if not addressed properly. Most transcriptome assemblers of short reads are based on de Bruijn graphs (DBG) and have no clear and explicit model for repeats in RNA-seq data, relying instead on heuristics to deal with them. The results of this work are threefold. First, we introduce a formal model for representing high copy-number and low-divergence repeats in RNA-seq data and exploit its properties to infer a combinatorial characteristic of repeat-associated subgraphs. We show that the problem of identifying such subgraphs in a DBG is NP-complete. Second, we show that in the specific case of local assembly of alternative splicing (AS) events, we can implicitly avoid such subgraphs, and we present an efficient algorithm to enumerate AS events that are not included in repeats. Using simulated data, we show that this strategy is significantly more sensitive and precise than the previous version of KisSplice (Sacomoto et al. in WABI, pp 99-111, 1), Trinity (Grabherr et al. in Nat Biotechnol 29(7):644-652, 2), and Oases (Schulz et al. in Bioinformatics 28(8):1086-1092, 3), for the specific task of calling AS events. Third, we turn our focus to full-length transcriptome assembly, and we show that exploring the topology of DBGs can improve de novo transcriptome evaluation methods. Based on the observation that repeats create complicated regions in a DBG, and when assemblers try to traverse these regions, they can infer erroneous transcripts, we propose a measure to flag transcripts traversing such troublesome regions, thereby giving a confidence level for each transcript. The originality of our work when

Block copolymer-nanoparticle hybrid self-assembly

KAUST Repository

Hoheisel, Tobias N.; Hur, Kahyun; Wiesner, Ulrich B.

2015-01-01

© 2014 Published by Elsevier Ltd. Polymer-inorganic hybrid materials provide exciting opportunities as they may display favorable properties from both constituents that are desired in applications including catalysis and energy conversion and storage. For the preparation of hybrid materials with well-defined morphologies, block copolymer-directed nanoparticle hybrids present a particularly promising approach. As will be described in this review, once the fundamental characteristics for successful nanostructure formation at or close to the thermodynamic equilibrium of these nanocomposites are identified, the approach can be generalized to various materials classes. In addition to the discussion of recent materials developments based on the use of AB diblock copolymers as well as ABC triblock terpolymers, this review will therefore emphasize progress in the fundamental understanding of the underlying formation mechanisms of such hybrid materials. To this end, critical experiments for, as well as theoretical progress in the description of these nanostructured block copolymer-based hybrid materials will be discussed. Rather than providing a comprehensive overview, the review will emphasize work by the Wiesner group at Cornell University, US, on block copolymer-directed nanoparticle assemblies as well as their use in first potential application areas. The results provide powerful design criteria for wet-chemical synthesis methodologies for the generation of functional nanomaterials for applications ranging from microelectronics to catalysis to energy conversion and storage.

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....

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.

De novo assembly of leaf transcriptome in the medicinal plant Andrographis paniculata

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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.

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

De Novo transcriptome assembly of Zingiber officinale cv. Suruchi of Odisha.

Science.gov (United States)

Gaur, Mahendra; Das, Aradhana; Sahoo, Rajesh Kumar; Kar, Basudeba; Nayak, Sanghamitra; Subudhi, Enketeswara

2016-09-01

Zingiber officinale Rosc., known as ginger, is an Asian crop, popularly used in every household kitchen and commercially used in bakery, beverage, food and pharmaceutical industries. The present study deals with de novo transcriptome assembly of an elite ginger cultivar Suruchi by next generation sequencing methodology. From the analysis 10.9 GB raw data was obtained which can be available in NCBI accession number SAMN03761185. We identified 41,969 transcripts using Trinity RNA-Seq from ginger rhizome of Suruchi variety from Odisha. The transcript length varied from 300 bp to 8404 bp with a total length of 3,96,40,526 bp and N50 of 1251 bp. To the best of our knowledge, this is the first transcriptome data of an elite ginger cultivar Suruchi released for Odisha state of India which will help molecular biologists to develop genetic markers for identification of cultivars.

Batch Scheduling for Hybrid Assembly Differentiation Flow Shop to Minimize Total Actual Flow Time

Science.gov (United States)

Maulidya, R.; Suprayogi; Wangsaputra, R.; Halim, A. H.

2018-03-01

A hybrid assembly differentiation flow shop is a three-stage flow shop consisting of Machining, Assembly and Differentiation Stages and producing different types of products. In the machining stage, parts are processed in batches on different (unrelated) machines. In the assembly stage, each part of the different parts is assembled into an assembly product. Finally, the assembled products will further be processed into different types of final products in the differentiation stage. In this paper, we develop a batch scheduling model for a hybrid assembly differentiation flow shop to minimize the total actual flow time defined as the total times part spent in the shop floor from the arrival times until its due date. We also proposed a heuristic algorithm for solving the problems. The proposed algorithm is tested using a set of hypothetic data. The solution shows that the algorithm can solve the problems effectively.

De novo assembly of mitochondrial genomes provides insights into genetic diversity and molecular evolution in wild boars and domestic pigs.

Science.gov (United States)

Ni, Pan; Bhuiyan, Ali Akbar; Chen, Jian-Hai; Li, Jingjin; Zhang, Cheng; Zhao, Shuhong; Du, Xiaoyong; Li, Hua; Yu, Hui; Liu, Xiangdong; Li, Kui

2018-05-10

Up to date, the scarcity of publicly available complete mitochondrial sequences for European wild pigs hampers deeper understanding about the genetic changes following domestication. Here, we have assembled 26 de novo mtDNA sequences of European wild boars from next generation sequencing (NGS) data and downloaded 174 complete mtDNA sequences to assess the genetic relationship, nucleotide diversity, and selection. The Bayesian consensus tree reveals the clear divergence between the European and Asian clade and a very small portion (10 out of 200 samples) of maternal introgression. The overall nucleotides diversities of the mtDNA sequences have been reduced following domestication. Interestingly, the selection efficiencies in both European and Asian domestic pigs are reduced, probably caused by changes in both selection constraints and maternal population size following domestication. This study suggests that de novo assembled mitogenomes can be a great boon to uncover the genetic turnover following domestication. Further investigation is warranted to include more samples from the ever-increasing amounts of NGS data to help us to better understand the process of domestication.

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...

Co-assembly of chitosan and phospholipids into hybrid hydrogels

DEFF Research Database (Denmark)

Mendes, Ana Carina Loureiro; Shekarforoush, Elhamalsadat; Engwer, Christoph

2016-01-01

Novel hybrid hydrogels were formed by adding chitosan (Ch) to phospholipids (P) self-assembled particles in lactic acid. The effect of the phospholipid concentration on the hydrogel properties was investigated and was observed to affect the rate of hydrogel formation and viscoelastic properties...

Identification of lignin genes and regulatory sequences involved in secondary cell wall formation in Acacia auriculiformis and Acacia mangium via de novo transcriptome sequencing

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Cannon Charles H

2011-07-01

Full Text Available Abstract Background Acacia auriculiformis × Acacia mangium hybrids are commercially important trees for the timber and pulp industry in Southeast Asia. Increasing pulp yield while reducing pulping costs are major objectives of tree breeding programs. The general monolignol biosynthesis and secondary cell wall formation pathways are well-characterized but genes in these pathways are poorly characterized in Acacia hybrids. RNA-seq on short-read platforms is a rapid approach for obtaining comprehensive transcriptomic data and to discover informative sequence variants. Results We sequenced transcriptomes of A. auriculiformis and A. mangium from non-normalized cDNA libraries synthesized from pooled young stem and inner bark tissues using paired-end libraries and a single lane of an Illumina GAII machine. De novo assembly produced a total of 42,217 and 35,759 contigs with an average length of 496 bp and 498 bp for A. auriculiformis and A. mangium respectively. The assemblies of A. auriculiformis and A. mangium had a total length of 21,022,649 bp and 17,838,260 bp, respectively, with the largest contig 15,262 bp long. We detected all ten monolignol biosynthetic genes using Blastx and further analysis revealed 18 lignin isoforms for each species. We also identified five contigs homologous to R2R3-MYB proteins in other plant species that are involved in transcriptional regulation of secondary cell wall formation and lignin deposition. We searched the contigs against public microRNA database and predicted the stem-loop structures of six highly conserved microRNA families (miR319, miR396, miR160, miR172, miR162 and miR168 and one legume-specific family (miR2086. Three microRNA target genes were predicted to be involved in wood formation and flavonoid biosynthesis. By using the assemblies as a reference, we discovered 16,648 and 9,335 high quality putative Single Nucleotide Polymorphisms (SNPs in the transcriptomes of A. auriculiformis and A. mangium

De Novo Assembly and Genome Analyses of the Marine-Derived Scopulariopsis brevicaulis Strain LF580 Unravels Life-Style Traits and Anticancerous Scopularide Biosynthetic Gene Cluster.

Science.gov (United States)

Kumar, Abhishek; Henrissat, Bernard; Arvas, Mikko; Syed, Muhammad Fahad; Thieme, Nils; Benz, J Philipp; Sørensen, Jens Laurids; Record, Eric; Pöggeler, Stefanie; Kempken, Frank

2015-01-01

The marine-derived Scopulariopsis brevicaulis strain LF580 produces scopularides A and B, which have anticancerous properties. We carried out genome sequencing using three next-generation DNA sequencing methods. De novo hybrid assembly yielded 621 scaffolds with a total size of 32.2 Mb and 16298 putative gene models. We identified a large non-ribosomal peptide synthetase gene (nrps1) and supporting pks2 gene in the same biosynthetic gene cluster. This cluster and the genes within the cluster are functionally active as confirmed by RNA-Seq. Characterization of carbohydrate-active enzymes and major facilitator superfamily (MFS)-type transporters lead to postulate S. brevicaulis originated from a soil fungus, which came into contact with the marine sponge Tethya aurantium. This marine sponge seems to provide shelter to this fungus and micro-environment suitable for its survival in the ocean. This study also builds the platform for further investigations of the role of life-style and secondary metabolites from S. brevicaulis.

Performance of FFTF reference fuel and control assemblies

International Nuclear Information System (INIS)

Leggett, R.D.; Weber, E.T.

1984-11-01

This paper describes the performance of the reference fuel and control assemblies used in FFTF through the first four cycles of irradiation (446 equivalent full power days, EFPD). These assemblies performed flawlessly through the rigors of the Startup Testing Program, STP, (beginning in late 1979) with its cyclic operation and continued to do so throughout Cycles 1, 2, 3 and 4, the latter ending in April 1984

De novo assembling and primary analysis of genome and transcriptome of gray whale Eschrichtius robustus.

Science.gov (United States)

Moskalev, Alexey А; Kudryavtseva, Anna V; Graphodatsky, Alexander S; Beklemisheva, Violetta R; Serdyukova, Natalya A; Krutovsky, Konstantin V; Sharov, Vadim V; Kulakovskiy, Ivan V; Lando, Andrey S; Kasianov, Artem S; Kuzmin, Dmitry A; Putintseva, Yuliya A; Feranchuk, Sergey I; Shaposhnikov, Mikhail V; Fraifeld, Vadim E; Toren, Dmitri; Snezhkina, Anastasia V; Sitnik, Vasily V

2017-12-28

Gray whale, Eschrichtius robustus (E. robustus), is a single member of the family Eschrichtiidae, which is considered to be the most primitive in the class Cetacea. Gray whale is often described as a "living fossil". It is adapted to extreme marine conditions and has a high life expectancy (77 years). The assembly of a gray whale genome and transcriptome will allow to carry out further studies of whale evolution, longevity, and resistance to extreme environment. In this work, we report the first de novo assembly and primary analysis of the E. robustus genome and transcriptome based on kidney and liver samples. The presented draft genome assembly is complete by 55% in terms of a total genome length, but only by 24% in terms of the BUSCO complete gene groups, although 10,895 genes were identified. Transcriptome annotation and comparison with other whale species revealed robust expression of DNA repair and hypoxia-response genes, which is expected for whales. This preliminary study of the gray whale genome and transcriptome provides new data to better understand the whale evolution and the mechanisms of their adaptation to the hypoxic conditions.

Data of first de-novo transcriptome assembly of a non-model species, hawksbill sea turtle, Eretmochelys imbricate, nesting of the Colombian Caribean.

Science.gov (United States)

Hernández-Fernández, Javier

2017-12-01

The hawksbill sea turtle, Eretmochelys imbricata, is an endangered species of the Caribbean Colombian coast due to anthropic and natural factors that have decreased their population levels. Little is known about the genes that are involved in their immune system, sex determination, aging and others important functions. The data generated represents RNA sequencing and the first de-novo assembly of transcripts expressed in the blood of the hawksbill sea turtle. The raw FASTQ files were deposited in the NCBI SRA database with accession number SRX2653641. A total of 5.7 Gb raw sequence data were obtained, corresponding to 47,555,108 raw reads. Trinity was used to perform a first de-novo assembly, and we were able to identify 47,586 transcripts of the female hawksbill turtle transcriptome with an N50 of 1100 bp. The obtained transcriptome data will be useful for further studies of the physiology, biochemistry and evolution in this species.

De Novo Assembly and Transcriptome Analysis of Wheat with Male Sterility Induced by the Chemical Hybridizing Agent SQ-1.

Directory of Open Access Journals (Sweden)

Qidi Zhu

Full Text Available Wheat (Triticum aestivum L., one of the world's most important food crops, is a strictly autogamous (self-pollinating species with exclusively perfect flowers. Male sterility induced by chemical hybridizing agents has increasingly attracted attention as a tool for hybrid seed production in wheat; however, the molecular mechanisms of male sterility induced by the agent SQ-1 remain poorly understood due to limited whole transcriptome data. Therefore, a comparative analysis of wheat anther transcriptomes for male fertile wheat and SQ-1-induced male sterile wheat was carried out using next-generation sequencing technology. In all, 42,634,123 sequence reads were generated and were assembled into 82,356 high-quality unigenes with an average length of 724 bp. Of these, 1,088 unigenes were significantly differentially expressed in the fertile and sterile wheat anthers, including 643 up-regulated unigenes and 445 down-regulated unigenes. The differentially expressed unigenes with functional annotations were mapped onto 60 pathways using the Kyoto Encyclopedia of Genes and Genomes database. They were mainly involved in coding for the components of ribosomes, photosynthesis, respiration, purine and pyrimidine metabolism, amino acid metabolism, glutathione metabolism, RNA transport and signal transduction, reactive oxygen species metabolism, mRNA surveillance pathways, protein processing in the endoplasmic reticulum, protein export, and ubiquitin-mediated proteolysis. This study is the first to provide a systematic overview comparing wheat anther transcriptomes of male fertile wheat with those of SQ-1-induced male sterile wheat and is a valuable source of data for future research in SQ-1-induced wheat male sterility.

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.

Centrioles: Some Self-Assembly Required

OpenAIRE

Song, Mi Hye; Miliaras, Nicholas B.; Peel, Nina; O'Connell, Kevin F.

2008-01-01

Centrioles play an important role in organizing microtubules and are precisely duplicated once per cell cycle. New (daughter) centrioles typically arise in association with existing (mother) centrioles (canonical assembly), suggesting that mother centrioles direct the formation of daughter centrioles. However, under certain circumstances, centrioles can also self-assemble free of an existing centriole (de novo assembly). Recent work indicates that the canonical and de novo pathways utilize a ...

De novo assembly of maritime pine transcriptome: implications for forest breeding and biotechnology.

Science.gov (United States)

Canales, Javier; Bautista, Rocio; Label, Philippe; Gómez-Maldonado, Josefa; Lesur, Isabelle; Fernández-Pozo, Noe; Rueda-López, Marina; Guerrero-Fernández, Dario; Castro-Rodríguez, Vanessa; Benzekri, Hicham; Cañas, Rafael A; Guevara, María-Angeles; Rodrigues, Andreia; Seoane, Pedro; Teyssier, Caroline; Morel, Alexandre; Ehrenmann, François; Le Provost, Grégoire; Lalanne, Céline; Noirot, Céline; Klopp, Christophe; Reymond, Isabelle; García-Gutiérrez, Angel; Trontin, Jean-François; Lelu-Walter, Marie-Anne; Miguel, Celia; Cervera, María Teresa; Cantón, Francisco R; Plomion, Christophe; Harvengt, Luc; Avila, Concepción; Gonzalo Claros, M; Cánovas, Francisco M

2014-04-01

Maritime pine (Pinus pinasterAit.) is a widely distributed conifer species in Southwestern Europe and one of the most advanced models for conifer research. In the current work, comprehensive characterization of the maritime pine transcriptome was performed using a combination of two different next-generation sequencing platforms, 454 and Illumina. De novo assembly of the transcriptome provided a catalogue of 26 020 unique transcripts in maritime pine trees and a collection of 9641 full-length cDNAs. Quality of the transcriptome assembly was validated by RT-PCR amplification of selected transcripts for structural and regulatory genes. Transcription factors and enzyme-encoding transcripts were annotated. Furthermore, the available sequencing data permitted the identification of polymorphisms and the establishment of robust single nucleotide polymorphism (SNP) and simple-sequence repeat (SSR) databases for genotyping applications and integration of translational genomics in maritime pine breeding programmes. All our data are freely available at SustainpineDB, the P. pinaster expressional database. Results reported here on the maritime pine transcriptome represent a valuable resource for future basic and applied studies on this ecological and economically important pine species. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

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...

A 'LEGO' Hybrid Photon Detector - assembled from standard mass-produced vacuum components

International Nuclear Information System (INIS)

Ferenc, Daniel; Lorenz, Eckart; Mirzoyan, Razmik

2000-01-01

Motivated primarily by the MAGIC atmospheric Cherenkov telescope project, we have developed a 'LEGO' Hybrid Photon Detector (HPD) comprising excellent focusing properties and protection against positive ion feedback. LEGO-HPD is supposed to be assembled from standard high vacuum components, which insures simplicity in the assembly procedure and reliability of operation

De novo assembly of a haplotype-resolved human genome

DEFF Research Database (Denmark)

Cao, Hongzhi; Wu, Honglong; Luo, Ruibang

2015-01-01

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-...

Controlled AFM manipulation of small nanoparticles and assembly of hybrid nanostructures

International Nuclear Information System (INIS)

Kim, Suenne; Shafiei, Farbod; Ratchford, Daniel; Li Xiaoqin

2011-01-01

We demonstrate controlled manipulation of semiconductor and metallic nanoparticles (NPs) with 5-15 nm diameters and assemble these NPs into hybrid structures. The manipulation is accomplished under ambient environment using a commercial atomic force microscope (AFM). There are particular difficulties associated with manipulating NPs this small. In addition to spatial drift, the shape of an asymmetric AFM tip has to be taken into account in order to understand the intended and actual manipulation results. Furthermore, small NPs often attach to the tip via electrostatic interaction and modify the effective tip shape. We suggest a method for detaching the NPs by performing a pseudo-manipulation step. Finally, we show by example the ability to assemble these small NPs into prototypical hybrid nanostructures with well-defined composition and geometry.

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.

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.

De novo transcriptome assembly of a sour cherry cultivar, Schattenmorelle

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Yeonhwa Jo

2015-12-01

Full Text Available Sour cherry (Prunus cerasus in the genus Prunus in the family Rosaceae is one of the most popular stone fruit trees worldwide. Of known sour cherry cultivars, the Schattenmorelle is a famous old sour cherry with a high amount of fruit production. The Schattenmorelle was selected before 1650 and described in the 1800s. This cultivar was named after gardens of the Chateau de Moreille in which the cultivar was initially found. In order to identify new genes and to develop genetic markers for sour cherry, we performed a transcriptome analysis of a sour cherry. We selected the cultivar Schattenmorelle, which is among commercially important cultivars in Europe and North America. We obtained 2.05 GB raw data from the Schattenmorelle (NCBI accession number: SRX1187170. De novo transcriptome assembly using Trinity identified 61,053 transcripts in which N50 was 611 bp. Next, we identified 25,585 protein coding sequences using TransDecoder. The identified proteins were blasted against NCBI's non-redundant database for annotation. Based on blast search, we taxonomically classified the obtained sequences. As a result, we provide the transcriptome of sour cherry cultivar Schattenmorelle using next generation sequencing.

Self-Assembled PbSe Nanowire:Perovskite Hybrids

KAUST Repository

Yang, Zhenyu

2015-12-02

© 2015 American Chemical Society. Inorganic semiconductor nanowires are of interest in nano- and microscale photonic and electronic applications. Here we report the formation of PbSe nanowires based on directional quantum dot alignment and fusion regulated by hybrid organic-inorganic perovskite surface ligands. All material synthesis is carried out at mild temperatures. Passivation of PbSe quantum dots was achieved via a new perovskite ligand exchange. Subsequent in situ ammonium/amine substitution by butylamine enables quantum dots to be capped by butylammonium lead iodide, and this further drives the formation of a PbSe nanowire superlattice in a two-dimensional (2D) perovskite matrix. The average spacing between two adjacent nanowires agrees well with the thickness of single atomic layer of 2D perovskite, consistent with the formation of a new self-assembled semiconductor nanowire:perovskite heterocrystal hybrid.

Self-Assembled PbSe Nanowire:Perovskite Hybrids

KAUST Repository

Yang, Zhenyu; Yassitepe, Emre; Voznyy, Oleksandr; Janmohamed, Alyf; Lan, Xinzheng; Levina, Larissa; Comin, Riccardo; Sargent, Edward H.

2015-01-01

© 2015 American Chemical Society. Inorganic semiconductor nanowires are of interest in nano- and microscale photonic and electronic applications. Here we report the formation of PbSe nanowires based on directional quantum dot alignment and fusion regulated by hybrid organic-inorganic perovskite surface ligands. All material synthesis is carried out at mild temperatures. Passivation of PbSe quantum dots was achieved via a new perovskite ligand exchange. Subsequent in situ ammonium/amine substitution by butylamine enables quantum dots to be capped by butylammonium lead iodide, and this further drives the formation of a PbSe nanowire superlattice in a two-dimensional (2D) perovskite matrix. The average spacing between two adjacent nanowires agrees well with the thickness of single atomic layer of 2D perovskite, consistent with the formation of a new self-assembled semiconductor nanowire:perovskite heterocrystal hybrid.

Faucet: streaming de novo assembly graph construction.

Science.gov (United States)

Rozov, Roye; Goldshlager, Gil; Halperin, Eran; Shamir, Ron

2018-01-01

We present Faucet, a two-pass streaming algorithm for assembly graph construction. Faucet builds an assembly graph incrementally as each read is processed. Thus, reads need not be stored locally, as they can be processed while downloading data and then discarded. We demonstrate this functionality by performing streaming graph assembly of publicly available data, and observe that the ratio of disk use to raw data size decreases as coverage is increased. Faucet pairs the de Bruijn graph obtained from the reads with additional meta-data derived from them. We show these metadata-coverage counts collected at junction k-mers and connections bridging between junction pairs-contain most salient information needed for assembly, and demonstrate they enable cleaning of metagenome assembly graphs, greatly improving contiguity while maintaining accuracy. We compared Fauceted resource use and assembly quality to state of the art metagenome assemblers, as well as leading resource-efficient genome assemblers. Faucet used orders of magnitude less time and disk space than the specialized metagenome assemblers MetaSPAdes and Megahit, while also improving on their memory use; this broadly matched performance of other assemblers optimizing resource efficiency-namely, Minia and LightAssembler. However, on metagenomes tested, Faucet,o outputs had 14-110% higher mean NGA50 lengths compared with Minia, and 2- to 11-fold higher mean NGA50 lengths compared with LightAssembler, the only other streaming assembler available. Faucet is available at https://github.com/Shamir-Lab/Faucet. rshamir@tau.ac.il or eranhalperin@gmail.com. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press.

Centrioles: some self-assembly required.

Science.gov (United States)

Song, Mi Hye; Miliaras, Nicholas B; Peel, Nina; O'Connell, Kevin F

2008-12-01

Centrioles play an important role in organizing microtubules and are precisely duplicated once per cell cycle. New (daughter) centrioles typically arise in association with existing (mother) centrioles (canonical assembly), suggesting that mother centrioles direct the formation of daughter centrioles. However, under certain circumstances, centrioles can also selfassemble free of an existing centriole (de novo assembly). Recent work indicates that the canonical and de novo pathways utilize a common mechanism and that a mother centriole spatially constrains the self-assembly process to occur within its immediate vicinity. Other recently identified mechanisms further regulate canonical assembly so that during each cell cycle, one and only one daughter centriole is assembled per mother centriole.

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.

Surface-enhanced Raman effect in hybrid metal–semiconductor nanoparticle assemblies

International Nuclear Information System (INIS)

Lughi, Vanni; Bonifacio, Alois; Barbone, Matteo; Marsich, Lucia; Sergo, Valter

2013-01-01

Hybrid metal–semiconductor nanoparticles consisting of silver nanoparticle cores (AgNPs) coated with a layer of CdSe quantum dots (QDs) have been studied by Raman spectroscopy. The hybrid nanoparticles were prepared via electrostatic interaction by mixing aqueous suspensions of QDs and AgNPs, where opposite charges on the AgNPs and QDs surfaces were induced by opportunely selected capping agents. Assemblies of such hybrid nanoparticles show an increased intensity of the Raman spectrum of up to 500 times, when compared to that of the sole QDs. This enhancement is attributed to the SERS effect (Surface-enhanced Raman scattering). Such enhancement of the Raman modes suggests several opportunities for further research, both in imaging and sensing applications.

Data of first de-novo transcriptome assembly of a non-model species, hawksbill sea turtle, Eretmochelys imbricate, nesting of the Colombian Caribean

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Javier Hernández-Fernández

2017-12-01

Full Text Available The hawksbill sea turtle, Eretmochelys imbricata, is an endangered species of the Caribbean Colombian coast due to anthropic and natural factors that have decreased their population levels. Little is known about the genes that are involved in their immune system, sex determination, aging and others important functions. The data generated represents RNA sequencing and the first de-novo assembly of transcripts expressed in the blood of the hawksbill sea turtle. The raw FASTQ files were deposited in the NCBI SRA database with accession number SRX2653641. A total of 5.7 Gb raw sequence data were obtained, corresponding to 47,555,108 raw reads. Trinity was used to perform a first de-novo assembly, and we were able to identify 47,586 transcripts of the female hawksbill turtle transcriptome with an N50 of 1100 bp. The obtained transcriptome data will be useful for further studies of the physiology, biochemistry and evolution in this species. Keywords: Hawksbill turtle, Trinity, RNAseq, illumina, N50

Reference quality assembly of the 3.5 Gb genome of Capsicum annuum form a single linked-read library

Science.gov (United States)

Linked-Read sequencing technology has recently been employed successfully for de novo assembly of multiple human genomes, however the utility of this technology for complex plant genomes is unproven. We evaluated the technology for this purpose by sequencing the 3.5 gigabase (Gb) diploid pepper (Cap...

Self-assembly and transformation of hybrid nano-objects and nanostructures under equilibrium and non-equilibrium conditions

Science.gov (United States)

Mann, Stephen

2009-10-01

Understanding how chemically derived processes control the construction and organization of matter across extended and multiple length scales is of growing interest in many areas of materials research. Here we review present equilibrium and non-equilibrium self-assembly approaches to the synthetic construction of discrete hybrid (inorganic-organic) nano-objects and higher-level nanostructured networks. We examine a range of synthetic modalities under equilibrium conditions that give rise to integrative self-assembly (supramolecular wrapping, nanoscale incarceration and nanostructure templating) or higher-order self-assembly (programmed/directed aggregation). We contrast these strategies with processes of transformative self-assembly that use self-organizing media, reaction-diffusion systems and coupled mesophases to produce higher-level hybrid structures under non-equilibrium conditions. Key elements of the constructional codes associated with these processes are identified with regard to existing theoretical knowledge, and presented as a heuristic guideline for the rational design of hybrid nano-objects and nanomaterials.

The Amaranth Genome: Genome, Transcriptome, and Physical Map Assembly

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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.

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.

Environmentally responsive optical microstructured hybrid actuator assemblies and applications thereof

Science.gov (United States)

Aizenberg, Joanna; Aizenberg, Michael; Kim, Philseok

2016-01-05

Microstructured hybrid actuator assemblies in which microactuators carrying designed surface properties to be revealed upon actuation are embedded in a layer of responsive materials. The microactuators in a microactuator array reversibly change their configuration in response to a change in the environment without requiring an external power source to switch their optical properties.

Self-Assembled Core-Shell-Type Lipid-Polymer Hybrid Nanoparticles: Intracellular Trafficking and Relevance for Oral Absorption.

Science.gov (United States)

Li, Qiuxia; Xia, Dengning; Tao, Jinsong; Shen, Aijun; He, Yuan; Gan, Yong; Wang, Chi

2017-10-01

Lipid-polymer hybrid nanoparticles (NPs) are advantageous for drug delivery. However, their intracellular trafficking mechanism and relevance for oral drug absorption are poorly understood. In this study, self-assembled core-shell lipid-polymer hybrid NPs made of poly(lactic-co-glycolic acid) (PLGA) and various lipids were developed to study their differing intracellular trafficking in intestinal epithelial cells and their relevance for oral absorption of a model drug saquinavir (SQV). Our results demonstrated that the endocytosis and exocytosis of hybrid NPs could be changed by varying the kind of lipid. A glyceride mixture (hybrid NPs-1) decreased endocytosis but increased exocytosis in Caco-2 cells, whereas the phospholipid (E200) (hybrid NPs-2) decreased endocytosis but exocytosis was unaffected as compared with PLGA nanoparticles. The transport of hybrid NPs-1 in cells involved various pathways, including caveolae/lipid raft-dependent endocytosis, and clathrin-mediated endocytosis and macropinocytosis, which was different from the other groups of NPs that involved only caveolae/lipid raft-dependent endocytosis. Compared with that of the reference formulation (nanoemulsion), the oral absorption of SQV-loaded hybrid NPs in rats was poor, probably due to the limited drug release and transcytosis of NPs across the intestinal epithelium. In conclusion, the intracellular processing of hybrid NPs in intestinal epithelia can be altered by adding lipids to the NP. However, it appears unfavorable to use PLGA-based NPs to improve oral absorption of SQV compared with nanoemulsion. Our findings will be essential in the development of polymer-based NPs for the oral delivery of drugs with the purpose of improving their oral absorption. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Incomplete sex chromosome dosage compensation in the Indian meal moth, Plodia interpunctella, based on de novo transcriptome assembly.

Science.gov (United States)

Harrison, Peter W; Mank, Judith E; Wedell, Nina

2012-01-01

Males and females experience differences in gene dose for loci in the nonrecombining region of heteromorphic sex chromosomes. If not compensated, this leads to expression imbalances, with the homogametic sex on average exhibiting greater expression due to the doubled gene dose. Many organisms with heteromorphic sex chromosomes display global dosage compensation mechanisms, which equalize gene expression levels between the sexes. However, birds and Schistosoma have been previously shown to lack chromosome-wide dosage compensation mechanisms, and the status in other female heterogametic taxa including Lepidoptera remains unresolved. To further our understanding of dosage compensation in female heterogametic taxa and to resolve its status in the lepidopterans, we assessed the Indian meal moth, Plodia interpunctella. As P. interpunctella lacks a complete reference genome, we conducted de novo transcriptome assembly combined with orthologous genomic location prediction from the related silkworm genome, Bombyx mori, to compare Z-linked and autosomal gene expression levels for each sex. We demonstrate that P. interpunctella lacks complete Z chromosome dosage compensation, female Z-linked genes having just over half the expression level of males and autosomal genes. This finding suggests that the Lepidoptera and possibly all female heterogametic taxa lack global dosage compensation, although more species will need to be sampled to confirm this assertion.

Bioinspired synthesis and self-assembly of hybrid organic–inorganic nanomaterials

Energy Technology Data Exchange (ETDEWEB)

Zhang, Honghu [Iowa State Univ., Ames, IA (United States)

2016-12-17

Nature is replete with complex organic–inorganic hierarchical materials of diverse yet specific functions. These materials are intricately designed under physiological conditions through biomineralization and biological self-assembly processes. Tremendous efforts have been devoted to investigating mechanisms of such biomineralization and biological self-assembly processes as well as gaining inspiration to develop biomimetic methods for synthesis and self-assembly of functional nanomaterials. In this work, we focus on the bioinspired synthesis and self-assembly of functional inorganic nanomaterials templated by specialized macromolecules including proteins, DNA and polymers. The in vitro biomineralization process of the magnetite biomineralizing protein Mms6 has been investigated using small-angle X-ray scattering. Templated by Mms6, complex magnetic nanomaterials can be synthesized on surfaces and in the bulk. DNA and synthetic polymers have been exploited to construct macroscopic two- and three-dimensional (2D and 3D) superlattices of gold nanocrystals. Employing X-ray scattering and spectroscopy techniques, the self-assembled structures and the self-assembly mechanisms have been studied, and theoretical models have been developed. Our results show that specialized macromolecules including proteins, DNA and polymers act as effective templates for synthesis and self-assembly of nanomaterials. These bottom-up approaches provide promising routes to fabricate hybrid organic–inorganic nanomaterials with rationally designed hierarchical structures, targeting specific functions.

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

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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.

Exciplex formation and energy transfer in a self-assembled metal-organic hybrid system.

Science.gov (United States)

Haldar, Ritesh; Rao, K Venkata; George, Subi J; Maji, Tapas Kumar

2012-05-07

Exciting assemblies: A metal-organic self-assembly of pyrenebutyric acid (PBA), 1,10-phenanthroline (o-phen), and Mg(II) shows solid-state fluorescence originating from a 1:1 PBA-o-phen exciplex. This exciplex fluorescence is sensitized by another residual PBA chromophore through an excited-state energy-transfer process. The solvent polarity modulates the self-assembly and the corresponding exciplex as well as the energy transfer, resulting in tunable emission of the hybrid (see figure). Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

An efficient enzyme-powered micromotor device fabricated by cyclic alternate hybridization assembly for DNA detection.

Science.gov (United States)

Fu, Shizhe; Zhang, Xueqing; Xie, Yuzhe; Wu, Jie; Ju, Huangxian

2017-07-06

An efficient enzyme-powered micromotor device was fabricated by assembling multiple layers of catalase on the inner surface of a poly(3,4-ethylenedioxythiophene and sodium 4-styrenesulfonate)/Au microtube (PEDOT-PSS/Au). The catalase assembly was achieved by programmed DNA hybridization, which was performed by immobilizing a designed sandwich DNA structure as the sensing unit on the PEDOT-PSS/Au, and then alternately hybridizing with two assisting DNA to bind the enzyme for efficient motor motion. The micromotor device showed unique features of good reproducibility, stability and motion performance. Under optimal conditions, it showed a speed of 420 μm s -1 in 2% H 2 O 2 and even 51 μm s -1 in 0.25% H 2 O 2 . In the presence of target DNA, the sensing unit hybridized with target DNA to release the multi-layer DNA as well as the multi-catalase, resulting in a decrease of the motion speed. By using the speed as a signal, the micromotor device could detect DNA from 10 nM to 1 μM. The proposed micromotor device along with the cyclic alternate DNA hybridization assembly technique provided a new path to fabricate efficient and versatile micromotors, which would be an exceptional tool for rapid and simple detection of biomolecules.

Production of solar chemicals: gaining selectivity with hybrid molecule/semiconductor assemblies.

Science.gov (United States)

Hennessey, Seán; Farràs, Pau

2018-05-29

Research on the production of solar fuels and chemicals has rocketed over the past decade, with a wide variety of systems proposed to harvest solar energy and drive chemical reactions. In this Feature Article we have focused on hybrid molecule/semiconductor assemblies in both powder and supported materials, summarising recent systems and highlighting the enormous possibilities offered by such assemblies to carry out highly demanding chemical reactions with industrial impact. Of relevance is the higher selectivity obtained in visible light-driven organic transformations when using molecular catalysts compared to photocatalytic materials.

De novo design of an RNA tile that self-assembles into a homo-octameric nanoprism

Science.gov (United States)

Yu, Jinwen; Liu, Zhiyu; Jiang, Wen; Wang, Guansong; Mao, Chengde

2015-01-01

Rational, de novo design of RNA nanostructures can potentially integrate a wide array of structural and functional diversities. Such nanostructures have great promises in biomedical applications. Despite impressive progress in this field, all RNA building blocks (or tiles) reported so far are not geometrically well defined. They are generally flexible and can only assemble into a mixture of complexes with different sizes. To achieve defined structures, multiple tiles with different sequences are needed. In this study, we design an RNA tile that can homo-oligomerize into a uniform RNA nanostructure. The designed RNA nanostructure is characterized by gel electrophoresis, atomic force microscopy and cryogenic electron microscopy imaging. We believe that development along this line would help RNA nanotechnology to reach the structural control that is currently associated with DNA nanotechnology.

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.

De novo Genome Assembly and Single Nucleotide Variations for Soybean Mosaic Virus Using Soybean Seed Transcriptome Data

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Yeonhwa Jo

2017-10-01

Full Text Available Soybean is the most important legume crop in the world. Several diseases in soybean lead to serious yield losses in major soybean-producing countries. Moreover, soybean can be infected by diverse viruses. Recently, we carried out a large-scale screening to identify viruses infecting soybean using available soybean transcriptome data. Of the screened transcriptomes, a soybean transcriptome for soybean seed development analysis contains several virus-associated sequences. In this study, we identified five viruses, including soybean mosaic virus (SMV, infecting soybean by de novo transcriptome assembly followed by blast search. We assembled a nearly complete consensus genome sequence of SMV China using transcriptome data. Based on phylogenetic analysis, the consensus genome sequence of SMV China was closely related to SMV isolates from South Korea. We examined single nucleotide variations (SNVs for SMVs in the soybean seed transcriptome revealing 780 SNVs, which were evenly distributed on the SMV genome. Four SNVs, C-U, U-C, A-G, and G-A, were frequently identified. This result demonstrated the quasispecies variation of the SMV genome. Taken together, this study carried out bioinformatics analyses to identify viruses using soybean transcriptome data. In addition, we demonstrated the application of soybean transcriptome data for virus genome assembly and SNV analysis.

Optical assembly of bio-hybrid micro-robots.

Science.gov (United States)

Barroso, Álvaro; Landwerth, Shirin; Woerdemann, Mike; Alpmann, Christina; Buscher, Tim; Becker, Maike; Studer, Armido; Denz, Cornelia

2015-04-01

The combination of micro synthetic structures with bacterial flagella motors represents an actual trend for the construction of self-propelled micro-robots. The development of methods for fabrication of these bacteria-based robots is a first crucial step towards the realization of functional miniature and autonomous moving robots. We present a novel scheme based on optical trapping to fabricate living micro-robots. By using holographic optical tweezers that allow three-dimensional manipulation in real time, we are able to arrange the building blocks that constitute the micro-robot in a defined way. We demonstrate exemplarily that our method enables the controlled assembly of living micro-robots consisting of a rod-shaped prokaryotic bacterium and a single elongated zeolite L crystal, which are used as model of the biological and abiotic components, respectively. We present different proof-of-principle approaches for the site-selective attachment of the bacteria on the particle surface. The propulsion of the optically assembled micro-robot demonstrates the potential of the proposed method as a powerful strategy for the fabrication of bio-hybrid micro-robots.

Biomimetic growth of gallic acid–ZnO hybrid assemblies and their applications

International Nuclear Information System (INIS)

Sarker, Nazmul H.; Barnaby, Stacey N.; Fath, Karl R.; Frayne, Stephen H.; Nakatsuka, Nako; Banerjee, Ipsita A.

2012-01-01

In this study, we probed the biomimetic formation of gallic acid (GA)–ZnO nanoparticle hybrids. It was found that the morphologies formed were dependent upon pH values, resulting in GA–ZnO hybrids of varying shapes such as micro or nanoplates or fibers. The formed supramolecular GA–ZnO hybrids were found to be luminescent as indicated by confocal microscopy and were utilized for the photocatalytic degradation of the organic dye methylene blue. We also explored the bactericidal effects of the hybrids on Staphylococcus aureus (S. aureus) as well as Escherichia Coli (E. Coli). Thus, we have developed a new class of shape-controlled nanohybrid assemblies via mild, green synthetic methods that may be utilized for photocatalytic degradation for environmental remediation as well as for antibacterial applications.

Biomimetic growth of gallic acid-ZnO hybrid assemblies and their applications

Energy Technology Data Exchange (ETDEWEB)

Sarker, Nazmul H.; Barnaby, Stacey N. [Fordham University, Department of Chemistry (United States); Fath, Karl R. [City University of New York and Graduate Center, Department of Biology, Queens College (United States); Frayne, Stephen H.; Nakatsuka, Nako; Banerjee, Ipsita A., E-mail: banerjee@fordham.edu [Fordham University, Department of Chemistry (United States)

2012-03-15

In this study, we probed the biomimetic formation of gallic acid (GA)-ZnO nanoparticle hybrids. It was found that the morphologies formed were dependent upon pH values, resulting in GA-ZnO hybrids of varying shapes such as micro or nanoplates or fibers. The formed supramolecular GA-ZnO hybrids were found to be luminescent as indicated by confocal microscopy and were utilized for the photocatalytic degradation of the organic dye methylene blue. We also explored the bactericidal effects of the hybrids on Staphylococcus aureus (S. aureus) as well as Escherichia Coli (E. Coli). Thus, we have developed a new class of shape-controlled nanohybrid assemblies via mild, green synthetic methods that may be utilized for photocatalytic degradation for environmental remediation as well as for antibacterial applications.

Biomimetic growth of gallic acid-ZnO hybrid assemblies and their applications

Science.gov (United States)

Sarker, Nazmul H.; Barnaby, Stacey N.; Fath, Karl R.; Frayne, Stephen H.; Nakatsuka, Nako; Banerjee, Ipsita A.

2012-03-01

In this study, we probed the biomimetic formation of gallic acid (GA)-ZnO nanoparticle hybrids. It was found that the morphologies formed were dependent upon pH values, resulting in GA-ZnO hybrids of varying shapes such as micro or nanoplates or fibers. The formed supramolecular GA-ZnO hybrids were found to be luminescent as indicated by confocal microscopy and were utilized for the photocatalytic degradation of the organic dye methylene blue. We also explored the bactericidal effects of the hybrids on Staphylococcus aureus ( S. aureus) as well as Escherichia Coli ( E. Coli). Thus, we have developed a new class of shape-controlled nanohybrid assemblies via mild, green synthetic methods that may be utilized for photocatalytic degradation for environmental remediation as well as for antibacterial applications.

De novo assembly of a cotyledon-enriched transcriptome map of Vicia faba (L. for transfer cell research

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Kiruba Shankari eArun Chinnappa

2015-04-01

Full Text Available Vicia faba (L. is an important cool-season grain legume species used widely in agriculture but also in plant physiology research, particularly as an experimental model to study transfer cell (TC development. Adaxial epidermal cells of isolated cotyledons can be induced to form functional TCs, thus providing a valuable experimental system to investigate genetic regulation of TC development. The genome of V. faba is exceedingly large (ca. 13 Gb, however, and limited genomic information is available for this species. To provide a resource for transcript profiling of epidermal TC development, we have undertaken de novo assembly of a cotyledon-enriched transcriptome map for V. faba. Illumina paired-end sequencing of total RNA pooled from different tissues and different stages, including isolated cotyledons induced to form TCs, generated 69.5M reads, of which 65.8M were used for assembly following trimming and quality control. Assembly using a De-Bruijn graph-based approach within CLC Genomics Workbench v6.1 generated 21,297 contigs, of which 80.6% were successfully annotated against GO terms. The assembly was validated against known V. faba cDNAs held in GenBank, including transcripts previously identified as being specifically expressed in epidermal cells across TC trans-differentiation. This cotyledon-enriched transcriptome map therefore provides a valuable tool for future transcript profiling of epidermal TC development, and also enriches the genetic resources available for this important legume crop species.

Hybrid Wing Body Multi-Bay Test Article Analysis and Assembly Final Report

Science.gov (United States)

Velicki, Alexander; Hoffman, Krishna; Linton, Kim A.; Baraja, Jaime; Wu, Hsi-Yung T.; Thrash, Patrick

2017-01-01

This report summarizes work performed by The Boeing Company, through its Boeing Research & Technology organization located in Huntington Beach, California, under the Environmentally Responsible Aviation (ERA) project. The report documents work performed to structurally analyze and assemble a large-scale Multi-bay Box (MBB) Test Article capable of withstanding bending and internal pressure loadings representative of a Hybrid Wing Body (HWB) aircraft. The work included fabrication of tooling elements for use in the fabrication and assembly of the test article.

De Novo Assembly and Phasing of Dikaryotic Genomes from Two Isolates of Puccinia coronata f. sp. avenae, the Causal Agent of Oat Crown Rust

Directory of Open Access Journals (Sweden)

Marisa E. Miller

2018-02-01

Full Text Available Oat crown rust, caused by the fungus Pucinnia coronata f. sp. avenae, is a devastating disease that impacts worldwide oat production. For much of its life cycle, P. coronata f. sp. avenae is dikaryotic, with two separate haploid nuclei that may vary in virulence genotype, highlighting the importance of understanding haplotype diversity in this species. We generated highly contiguous de novo genome assemblies of two P. coronata f. sp. avenae isolates, 12SD80 and 12NC29, from long-read sequences. In total, we assembled 603 primary contigs for 12SD80, for a total assembly length of 99.16 Mbp, and 777 primary contigs for 12NC29, for a total length of 105.25 Mbp; approximately 52% of each genome was assembled into alternate haplotypes. This revealed structural variation between haplotypes in each isolate equivalent to more than 2% of the genome size, in addition to about 260,000 and 380,000 heterozygous single-nucleotide polymorphisms in 12SD80 and 12NC29, respectively. Transcript-based annotation identified 26,796 and 28,801 coding sequences for isolates 12SD80 and 12NC29, respectively, including about 7,000 allele pairs in haplotype-phased regions. Furthermore, expression profiling revealed clusters of coexpressed secreted effector candidates, and the majority of orthologous effectors between isolates showed conservation of expression patterns. However, a small subset of orthologs showed divergence in expression, which may contribute to differences in virulence between 12SD80 and 12NC29. This study provides the first haplotype-phased reference genome for a dikaryotic rust fungus as a foundation for future studies into virulence mechanisms in P. coronata f. sp. avenae.

Three-input gate logic circuits on chemically assembled single-electron transistors with organic and inorganic hybrid passivation layers.

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Majima, Yutaka; Hackenberger, Guillaume; Azuma, Yasuo; Kano, Shinya; Matsuzaki, Kosuke; Susaki, Tomofumi; Sakamoto, Masanori; Teranishi, Toshiharu

2017-01-01

Single-electron transistors (SETs) are sub-10-nm scale electronic devices based on conductive Coulomb islands sandwiched between double-barrier tunneling barriers. Chemically assembled SETs with alkanethiol-protected Au nanoparticles show highly stable Coulomb diamonds and two-input logic operations. The combination of bottom-up and top-down processes used to form the passivation layer is vital for realizing multi-gate chemically assembled SET circuits, as this combination enables us to connect conventional complementary metal oxide semiconductor (CMOS) technologies via planar processes. Here, three-input gate exclusive-OR (XOR) logic operations are demonstrated in passivated chemically assembled SETs. The passivation layer is a hybrid bilayer of self-assembled monolayers (SAMs) and pulsed laser deposited (PLD) aluminum oxide (AlO[Formula: see text]), and top-gate electrodes were prepared on the hybrid passivation layers. Top and two-side-gated SETs showed clear Coulomb oscillation and diamonds for each of the three available gates, and three-input gate XOR logic operation was clearly demonstrated. These results show the potential of chemically assembled SETs to work as logic devices with multi-gate inputs using organic and inorganic hybrid passivation layers.

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

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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.

The wolf reference genome sequence (Canis lupus lupus) and its implications for Canis spp. population genomics

DEFF Research Database (Denmark)

Gopalakrishnan, Shyam; Samaniego Castruita, Jose Alfredo; Sinding, Mikkel Holger Strander

2017-01-01

Background An increasing number of studies are addressing the evolutionary genomics of dog domestication, principally through resequencing dog, wolf and related canid genomes. There is, however, only one de novo assembled canid genome currently available against which to map such data - that of a......Background An increasing number of studies are addressing the evolutionary genomics of dog domestication, principally through resequencing dog, wolf and related canid genomes. There is, however, only one de novo assembled canid genome currently available against which to map such data...... that regardless of the reference genome choice, most evolutionary genomic analyses yield qualitatively similar results, including those exploring the structure between the wolves and dogs using admixture and principal component analysis. However, we do observe differences in the genomic coverage of re-mapped...

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

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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.

Proton-Fueled, Reversible DNA Hybridization Chain Assembly for pH Sensing and Imaging.

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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.

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

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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.

De novo transcriptome assembly, functional annotation and differential gene expression analysis of juvenile and adult E. fetida, a model oligochaete used in ecotoxicological studies

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Michelle Thunders

Full Text Available Abstract Background Earthworms are sensitive to toxic chemicals present in the soil and so are useful indicator organisms for soil health. Eisenia fetida are commonly used in ecotoxicological studies; therefore the assembly of a baseline transcriptome is important for subsequent analyses exploring the impact of toxin exposure on genome wide gene expression. Results This paper reports on the de novo transcriptome assembly of E. fetida using Trinity, a freely available software tool. Trinotate was used to carry out functional annotation of the Trinity generated transcriptome file and the transdecoder generated peptide sequence file along with BLASTX, BLASTP and HMMER searches and were loaded into a Sqlite3 database. To identify differentially expressed transcripts; each of the original sequence files were aligned to the de novo assembled transcriptome using Bowtie and then RSEM was used to estimate expression values based on the alignment. EdgeR was used to calculate differential expression between the two conditions, with an FDR corrected P value cut off of 0.001, this returned six significantly differentially expressed genes. Initial BLASTX hits of these putative genes included hits with annelid ferritin and lysozyme proteins, as well as fungal NADH cytochrome b5 reductase and senescence associated proteins. At a cut off of P = 0.01 there were a further 26 differentially expressed genes. Conclusion These data have been made publicly available, and to our knowledge represent the most comprehensive available transcriptome for E. fetida assembled from RNA sequencing data. This provides important groundwork for subsequent ecotoxicogenomic studies exploring the impact of the environment on global gene expression in E. fetida and other earthworm species.

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

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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.

Integrated fast assembly of free-standing lithium titanate/carbon nanotube/cellulose nanofiber hybrid network film as flexible paper-electrode for lithium-ion batteries.

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Cao, Shaomei; Feng, Xin; Song, Yuanyuan; Xue, Xin; Liu, Hongjiang; Miao, Miao; Fang, Jianhui; Shi, Liyi

2015-05-27

A free-standing lithium titanate (Li4Ti5O12)/carbon nanotube/cellulose nanofiber hybrid network film is successfully assembled by using a pressure-controlled aqueous extrusion process, which is highly efficient and easily to scale up from the perspective of disposable and recyclable device production. This hybrid network film used as a lithium-ion battery (LIB) electrode has a dual-layer structure consisting of Li4Ti5O12/carbon nanotube/cellulose nanofiber composites (hereinafter referred to as LTO/CNT/CNF), and carbon nanotube/cellulose nanofiber composites (hereinafter referred to as CNT/CNF). In the heterogeneous fibrous network of the hybrid film, CNF serves simultaneously as building skeleton and a biosourced binder, which substitutes traditional toxic solvents and synthetic polymer binders. Of importance here is that the CNT/CNF layer is used as a lightweight current collector to replace traditional heavy metal foils, which therefore reduces the total mass of the electrode while keeping the same areal loading of active materials. The free-standing network film with high flexibility is easy to handle, and has extremely good conductivity, up to 15.0 S cm(-1). The flexible paper-electrode for LIBs shows very good high rate cycling performance, and the specific charge/discharge capacity values are up to 142 mAh g(-1) even at a current rate of 10 C. On the basis of the mild condition and fast assembly process, a CNF template fulfills multiple functions in the fabrication of paper-electrode for LIBs, which would offer an ever increasing potential for high energy density, low cost, and environmentally friendly flexible electronics.

De Novo Assembly of the Donkey White Blood Cell Transcriptome and a Comparative Analysis of Phenotype-Associated Genes between Donkeys and Horses.

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Xie, Feng-Yun; Feng, Yu-Long; Wang, Hong-Hui; Ma, Yun-Feng; Yang, Yang; Wang, Yin-Chao; Shen, Wei; Pan, Qing-Jie; Yin, Shen; Sun, Yu-Jiang; Ma, Jun-Yu

2015-01-01

Prior to the mechanization of agriculture and labor-intensive tasks, humans used donkeys (Equus africanus asinus) for farm work and packing. However, as mechanization increased, donkeys have been increasingly raised for meat, milk, and fur in China. To maintain the development of the donkey industry, breeding programs should focus on traits related to these new uses. Compared to conventional marker-assisted breeding plans, genome- and transcriptome-based selection methods are more efficient and effective. To analyze the coding genes of the donkey genome, we assembled the transcriptome of donkey white blood cells de novo. Using transcriptomic deep-sequencing data, we identified 264,714 distinct donkey unigenes and predicted 38,949 protein fragments. We annotated the donkey unigenes by BLAST searches against the non-redundant (NR) protein database. We also compared the donkey protein sequences with those of the horse (E. caballus) and wild horse (E. przewalskii), and linked the donkey protein fragments with mammalian phenotypes. As the outer ear size of donkeys and horses are obviously different, we compared the outer ear size-associated proteins in donkeys and horses. We identified three ear size-associated proteins, HIC1, PRKRA, and KMT2A, with sequence differences among the donkey, horse, and wild horse loci. Since the donkey genome sequence has not been released, the de novo assembled donkey transcriptome is helpful for preliminary investigations of donkey cultivars and for genetic improvement.

RNA-seq de novo Assembly Reveals Differential Gene Expression in Glossina palpalis gambiensis Infected with Trypanosoma brucei gambiense vs. Non-Infected and Self-Cured Flies.

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Hamidou Soumana, Illiassou; Klopp, Christophe; Ravel, Sophie; Nabihoudine, Ibouniyamine; Tchicaya, Bernadette; Parrinello, Hugues; Abate, Luc; Rialle, Stéphanie; Geiger, Anne

2015-01-01

Trypanosoma brucei gambiense (Tbg), causing the sleeping sickness chronic form, completes its developmental cycle within the tsetse fly vector Glossina palpalis gambiensis (Gpg) before its transmission to humans. Within the framework of an anti-vector disease control strategy, a global gene expression profiling of trypanosome infected (susceptible), non-infected, and self-cured (refractory) tsetse flies was performed, on their midguts, to determine differential genes expression resulting from in vivo trypanosomes, tsetse flies (and their microbiome) interactions. An RNAseq de novo assembly was achieved. The assembled transcripts were mapped to reference sequences for functional annotation. Twenty-four percent of the 16,936 contigs could not be annotated, possibly representing untranslated mRNA regions, or Gpg- or Tbg-specific ORFs. The remaining contigs were classified into 65 functional groups. Only a few transposable elements were present in the Gpg midgut transcriptome, which may represent active transpositions and play regulatory roles. One thousand three hundred and seventy three genes differentially expressed (DEGs) between stimulated and non-stimulated flies were identified at day-3 post-feeding; 52 and 1025 between infected and self-cured flies at 10 and 20 days post-feeding, respectively. The possible roles of several DEGs regarding fly susceptibility and refractoriness are discussed. The results provide new means to decipher fly infection mechanisms, crucial to develop anti-vector control strategies.

Large-area self-assembled reduced graphene oxide/electrochemically exfoliated graphene hybrid films for transparent electrothermal heaters

Science.gov (United States)

Sun, Hongyan; Chen, Ding; Ye, Chen; Li, Xinming; Dai, Dan; Yuan, Qilong; Chee, Kuan W. A.; Zhao, Pei; Jiang, Nan; Lin, Cheng-Te

2018-03-01

Graphene shows great promise as a high-efficiency electrothermal film for flexible transparent defoggers/defrosters. However, it remains a great challenge to achieve a good balance between the production cost and the properties of graphene films. Here, we proposed a cost-effective self-assembly method to fabricate high-performance, large-area graphene oxide/electrochemically exfoliated graphene hybrid films for heater applications. The self-assembled graphene hybrid films with the area of 20 × 20 cm2 could be transferred onto arbitrary substrates with nonplanar surfaces and simply patterned with the hard mask. After reduction by hydrogen iodide vapor followed by 800 °C thermal treatment, the hybrid films with the transmittance of 76.2% exhibit good heating characteristics and defogging performance, which reach a saturation temperature of up to 127.5 °C when 40 V was applied for 60 s.

Evaluating de Bruijn graph assemblers on 454 transcriptomic data.

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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.

De novo transcriptome assembly facilitates characterisation of fast-evolving gene families, MHC class I in the bank vole (Myodes glareolus).

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Migalska, M; Sebastian, A; Konczal, M; Kotlík, P; Radwan, J

2017-04-01

The major histocompatibility complex (MHC) plays a central role in the adaptive immune response and is the most polymorphic gene family in vertebrates. Although high-throughput sequencing has increasingly been used for genotyping families of co-amplifying MHC genes, its potential to facilitate early steps in the characterisation of MHC variation in nonmodel organism has not been fully explored. In this study we evaluated the usefulness of de novo transcriptome assembly in characterisation of MHC sequence diversity. We found that although de novo transcriptome assembly of MHC I genes does not reconstruct sequences of individual alleles, it does allow the identification of conserved regions for PCR primer design. Using the newly designed primers, we characterised MHC I sequences in the bank vole. Phylogenetic analysis of the partial MHC I coding sequence (2-4 exons) of the bank vole revealed a lack of orthology to MHC I of other Cricetidae, consistent with the high gene turnover of this region. The diversity of expressed alleles was characterised using ultra-deep sequencing of the third exon that codes for the peptide-binding region of the MHC molecule. High allelic diversity was demonstrated, with 72 alleles found in 29 individuals. Interindividual variation in the number of expressed loci was found, with the number of alleles per individual ranging from 5 to 14. Strong signatures of positive selection were found for 8 amino acid sites, most of which are inferred to bind antigens in human MHC, indicating conservation of structure despite rapid sequence evolution.

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

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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.

De Novo Assembly and Characterization of Sophora japonica Transcriptome Using RNA-seq

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Liucun Zhu

2014-01-01

Full Text Available Sophora japonica Linn (Chinese Scholar Tree is a shrub species belonging to the subfamily Faboideae of the pea family Fabaceae. In this study, RNA sequencing of S. japonica transcriptome was performed to produce large expression datasets for functional genomic analysis. Approximate 86.1 million high-quality clean reads were generated and assembled de novo into 143010 unique transcripts and 57614 unigenes. The average length of unigenes was 901 bps with an N50 of 545 bps. Four public databases, including the NCBI nonredundant protein (NR, Swiss-Prot, Kyoto Encyclopedia of Genes and Genomes (KEGG, and the Cluster of Orthologous Groups (COG, were used to annotate unigenes through NCBI BLAST procedure. A total of 27541 of 57614 unigenes (47.8% were annotated for gene descriptions, conserved protein domains, or gene ontology. Moreover, an interaction network of unigenes in S. japonica was predicted based on known protein-protein interactions of putative orthologs of well-studied plant genomes. The transcriptome data of S. japonica reported here represents first genome-scale investigation of gene expressions in Faboideae plants. We expect that our study will provide a useful resource for further studies on gene expression, genomics, functional genomics, and protein-protein interaction in S. japonica.

RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome

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Dewey Colin N

2011-08-01

Full Text Available Abstract Background RNA-Seq is revolutionizing the way transcript abundances are measured. A key challenge in transcript quantification from RNA-Seq data is the handling of reads that map to multiple genes or isoforms. This issue is particularly important for quantification with de novo transcriptome assemblies in the absence of sequenced genomes, as it is difficult to determine which transcripts are isoforms of the same gene. A second significant issue is the design of RNA-Seq experiments, in terms of the number of reads, read length, and whether reads come from one or both ends of cDNA fragments. Results We present RSEM, an user-friendly software package for quantifying gene and isoform abundances from single-end or paired-end RNA-Seq data. RSEM outputs abundance estimates, 95% credibility intervals, and visualization files and can also simulate RNA-Seq data. In contrast to other existing tools, the software does not require a reference genome. Thus, in combination with a de novo transcriptome assembler, RSEM enables accurate transcript quantification for species without sequenced genomes. On simulated and real data sets, RSEM has superior or comparable performance to quantification methods that rely on a reference genome. Taking advantage of RSEM's ability to effectively use ambiguously-mapping reads, we show that accurate gene-level abundance estimates are best obtained with large numbers of short single-end reads. On the other hand, estimates of the relative frequencies of isoforms within single genes may be improved through the use of paired-end reads, depending on the number of possible splice forms for each gene. Conclusions RSEM is an accurate and user-friendly software tool for quantifying transcript abundances from RNA-Seq data. As it does not rely on the existence of a reference genome, it is particularly useful for quantification with de novo transcriptome assemblies. In addition, RSEM has enabled valuable guidance for cost

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

49 CFR 572.160 - Incorporation by reference.

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2010-10-01

... Six-Year-Old Weighted Child Test Dummy § 572.160 Incorporation by reference. (a) The following... package entitled “Parts List and Drawings, Part 572 Subpart S, Hybrid III Weighted Six-Year Old Child Test... part of a complete dummy assembly; (iii) Drawing No. 167-2020 Revision A, dated December 8, 2005, Spine...

Microwave-Assisted Rapid Synthesis of Self-Assembled T-Nb2 O5 Nanowires for High-Energy Hybrid Supercapacitors.

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Yang, Huiling; Xu, Henghui; Wang, Libin; Zhang, Lei; Huang, Yunhui; Hu, Xianluo

2017-03-23

Recently ion-intercalation hybrid supercapacitors, with high energy density at high power density, have been widely investigated to meet ever-increasing practical demands. Here, a unique hybrid supercapacitor has been designed and fabricated using self-assembled orthorhombic-phase niobium oxide@carbon (T-Nb 2 O 5 @C) nanowires as an anode and commercially available activated carbon as a cathode. The 3D-interconnected T-Nb 2 O 5 @C nanowires have been synthesized through a highly efficient microwave-solvothermal method, combined with subsequent thermal treatment. The experimental parameters (e.g., time and temperature) can be easily programmed, and the synthesis time can be significantly shortened, thus enabling the buildup of abundant recipes for the engineering of scaled-up production. The Li-ion intercalation pseudocapacitance electrode, made from the as-formed self-assembled T-Nb 2 O 5 @C nanowires, shows excellent charge storage and transfer capability. When assembled into a hybrid supercapacitor with a cathode of activated carbon, a high energy density of 60.6 Wh kg -1 and a high power density of 8.5 kW kg -1 with outstanding stability are achieved. In virtue of easy optimization and programmability of the synthetic strategy, and the remarkable electrochemical performance, the self-assembled T-Nb 2 O 5 @C nanowires offer a promising anode for asymmetric hybrid supercapacitors. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

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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.

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

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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/.

New Approaches and Technologies to Sequence de novo Plant reference Genomes (2013 DOE JGI Genomics of Energy and Environment 8th Annual User Meeting)

Energy Technology Data Exchange (ETDEWEB)

Schmutz, Jeremy

2013-03-01

Jeremy Schmutz of the HudsonAlpha Institute for Biotechnology on New approaches and technologies to sequence de novo plant reference genomes at the 8th Annual Genomics of Energy Environment Meeting on March 27, 2013 in Walnut Creek, CA.

De novo transcriptomic analysis of an oleaginous microalga: pathway description and gene discovery for production of next-generation biofuels.

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LingLin Wan

Full Text Available Eustigmatos cf. polyphem is a yellow-green unicellular soil microalga belonging to the eustimatophyte with high biomass and considerable production of triacylglycerols (TAGs for biofuels, which is thus referred to as an oleaginous microalga. The paucity of microalgae genome sequences, however, limits development of gene-based biofuel feedstock optimization studies. Here we describe the sequencing and de novo transcriptome assembly for a non-model microalgae species, E. cf. polyphem, and identify pathways and genes of importance related to biofuel production.We performed the de novo assembly of E. cf. polyphem transcriptome using Illumina paired-end sequencing technology. In a single run, we produced 29,199,432 sequencing reads corresponding to 2.33 Gb total nucleotides. These reads were assembled into 75,632 unigenes with a mean size of 503 bp and an N50 of 663 bp, ranging from 100 bp to >3,000 bp. Assembled unigenes were subjected to BLAST similarity searches and annotated with Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG orthology identifiers. These analyses identified the majority of carbohydrate, fatty acids, TAG and carotenoids biosynthesis and catabolism pathways in E. cf. polyphem.Our data provides the construction of metabolic pathways involved in the biosynthesis and catabolism of carbohydrate, fatty acids, TAG and carotenoids in E. cf. polyphem and provides a foundation for the molecular genetics and functional genomics required to direct metabolic engineering efforts that seek to enhance the quantity and character of microalgae-based biofuel feedstock.

De Novo Assembly of the Donkey White Blood Cell Transcriptome and a Comparative Analysis of Phenotype-Associated Genes between Donkeys and Horses.

Directory of Open Access Journals (Sweden)

Feng-Yun Xie

Full Text Available Prior to the mechanization of agriculture and labor-intensive tasks, humans used donkeys (Equus africanus asinus for farm work and packing. However, as mechanization increased, donkeys have been increasingly raised for meat, milk, and fur in China. To maintain the development of the donkey industry, breeding programs should focus on traits related to these new uses. Compared to conventional marker-assisted breeding plans, genome- and transcriptome-based selection methods are more efficient and effective. To analyze the coding genes of the donkey genome, we assembled the transcriptome of donkey white blood cells de novo. Using transcriptomic deep-sequencing data, we identified 264,714 distinct donkey unigenes and predicted 38,949 protein fragments. We annotated the donkey unigenes by BLAST searches against the non-redundant (NR protein database. We also compared the donkey protein sequences with those of the horse (E. caballus and wild horse (E. przewalskii, and linked the donkey protein fragments with mammalian phenotypes. As the outer ear size of donkeys and horses are obviously different, we compared the outer ear size-associated proteins in donkeys and horses. We identified three ear size-associated proteins, HIC1, PRKRA, and KMT2A, with sequence differences among the donkey, horse, and wild horse loci. Since the donkey genome sequence has not been released, the de novo assembled donkey transcriptome is helpful for preliminary investigations of donkey cultivars and for genetic improvement.

Self-assembling hybrid diamond–biological quantum devices

International Nuclear Information System (INIS)

Albrecht, A; B Plenio, M; Koplovitz, G; Yochelis, S; Paltiel, Y; Retzker, A; Nevo, Y; Shoseyov, O; Jelezko, F; Porath, D

2014-01-01

The realization of scalable arrangements of nitrogen vacancy (NV) centers in diamond remains a key challenge on the way towards efficient quantum information processing, quantum simulation and quantum sensing applications. Although technologies based on implanting NV-centers in bulk diamond crystals or hybrid device approaches have been developed, they are limited by the achievable spatial resolution and by the intricate technological complexities involved in achieving scalability. We propose and demonstrate a novel approach for creating an arrangement of NV-centers, based on the self-assembling capabilities of biological systems and their beneficial nanometer spatial resolution. Here, a self-assembled protein structure serves as a structural scaffold for surface functionalized nanodiamonds, in this way allowing for the controlled creation of NV-structures on the nanoscale and providing a new avenue towards bridging the bio–nano interface. One-, two- as well as three-dimensional structures are within the scope of biological structural assembling techniques. We realized experimentally the formation of regular structures by interconnecting nanodiamonds using biological protein scaffolds. Based on the achievable NV-center distances of 11 nm, we evaluate the expected dipolar coupling interaction with neighboring NV-centers as well as the expected decoherence time. Moreover, by exploiting these couplings, we provide a detailed theoretical analysis on the viability of multiqubit quantum operations, suggest the possibility of individual addressing based on the random distribution of the NV intrinsic symmetry axes and address the challenges posed by decoherence and imperfect couplings. We then demonstrate in the last part that our scheme allows for the high-fidelity creation of entanglement, cluster states and quantum simulation applications. (papers)

Self-assembling hybrid diamond-biological quantum devices

Science.gov (United States)

Albrecht, A.; Koplovitz, G.; Retzker, A.; Jelezko, F.; Yochelis, S.; Porath, D.; Nevo, Y.; Shoseyov, O.; Paltiel, Y.; Plenio, M. B.

2014-09-01

The realization of scalable arrangements of nitrogen vacancy (NV) centers in diamond remains a key challenge on the way towards efficient quantum information processing, quantum simulation and quantum sensing applications. Although technologies based on implanting NV-centers in bulk diamond crystals or hybrid device approaches have been developed, they are limited by the achievable spatial resolution and by the intricate technological complexities involved in achieving scalability. We propose and demonstrate a novel approach for creating an arrangement of NV-centers, based on the self-assembling capabilities of biological systems and their beneficial nanometer spatial resolution. Here, a self-assembled protein structure serves as a structural scaffold for surface functionalized nanodiamonds, in this way allowing for the controlled creation of NV-structures on the nanoscale and providing a new avenue towards bridging the bio-nano interface. One-, two- as well as three-dimensional structures are within the scope of biological structural assembling techniques. We realized experimentally the formation of regular structures by interconnecting nanodiamonds using biological protein scaffolds. Based on the achievable NV-center distances of 11 nm, we evaluate the expected dipolar coupling interaction with neighboring NV-centers as well as the expected decoherence time. Moreover, by exploiting these couplings, we provide a detailed theoretical analysis on the viability of multiqubit quantum operations, suggest the possibility of individual addressing based on the random distribution of the NV intrinsic symmetry axes and address the challenges posed by decoherence and imperfect couplings. We then demonstrate in the last part that our scheme allows for the high-fidelity creation of entanglement, cluster states and quantum simulation applications.

Self-assembly of multiferroic core-shell particulate nanocomposites through DNA-DNA hybridization and magnetic field directed assembly of superstructures

Energy Technology Data Exchange (ETDEWEB)

Sreenivasulu, Gollapudi; Srinivasan, Gopalan, E-mail: srinivas@oakland.edu, E-mail: chavez@oakland.edu [Department of Physics, Oakland University, Rochester, MI 48309-4401 (United States); Lochbiler, Thomas A.; Panda, Manashi; Chavez, Ferman A., E-mail: srinivas@oakland.edu, E-mail: chavez@oakland.edu [Department of Chemistry, Oakland University, Rochester, MI 48309-4401 (United States)

2016-04-15

Multiferroic composites of ferromagnetic and ferroelectric phases are of importance for studies on mechanical strain mediated coupling between the magnetic and electric subsystems. This work is on DNA-assisted self-assembly of superstructures of such composites with nanometer periodicity. The synthesis involved oligomeric DNA-functionalized ferroelectric and ferromagnetic nanoparticles, 600 nm BaTiO{sub 3} (BTO) and 200 nm NiFe{sub 2}O{sub 4} (NFO), respectively. Mixing BTO and NFO particles, possessing complementary DNA sequences, resulted in the formation of ordered core-shell heteronanocomposites held together by DNA hybridization. The composites were imaged by scanning electron microscopy and scanning microwave microscopy. The presence of heteroassemblies along with core-shell architecture is clearly observed. The reversible nature of the DNA hybridization allows for restructuring the composites into mm-long linear chains and 2D-arrays in the presence of a static magnetic field and ring-like structures in a rotating-magnetic field. Strong magneto-electric (ME) coupling in as-assembled composites is evident from static magnetic field H induced polarization and low-frequency magnetoelectric voltage coefficient measurements. Upon annealing the nanocomposites at high temperatures, evidence for the formation of bulk composites with excellent cross-coupling between the electric and magnetic subsystems is obtained by H-induced polarization and low-frequency ME voltage coefficient. The ME coupling strength in the self-assembled composites is measured to be much stronger than in bulk composites with randomly distributed NFO and BTO prepared by direct mixing and sintering.

The UF family of reference hybrid phantoms for computational radiation dosimetry

International Nuclear Information System (INIS)

Lee, Choonsik; Lodwick, Daniel; Hurtado, Jorge; Pafundi, Deanna; Williams, Jonathan L; Bolch, Wesley E

2010-01-01

Computational human phantoms are computer models used to obtain dose distributions within the human body exposed to internal or external radiation sources. In addition, they are increasingly used to develop detector efficiencies for in vivo whole-body counters. Two classes of computational human phantoms have been widely utilized for dosimetry calculation: stylized and voxel phantoms that describe human anatomy through mathematical surface equations and 3D voxel matrices, respectively. Stylized phantoms are flexible in that changes to organ position and shape are possible given avoidance of region overlap, while voxel phantoms are typically fixed to a given patient anatomy, yet can be proportionally scaled to match individuals of larger or smaller stature, but of equivalent organ anatomy. Voxel phantoms provide much better anatomical realism as compared to stylized phantoms which are intrinsically limited by mathematical surface equations. To address the drawbacks of these phantoms, hybrid phantoms based on non-uniform rational B-spline (NURBS) surfaces have been introduced wherein anthropomorphic flexibility and anatomic realism are both preserved. Researchers at the University of Florida have introduced a series of hybrid phantoms representing the ICRP Publication 89 reference newborn, 15 year, and adult male and female. In this study, six additional phantoms are added to the UF family of hybrid phantoms-those of the reference 1 year, 5 year and 10 year child. Head and torso CT images of patients whose ages were close to the targeted ages were obtained under approved protocols. Major organs and tissues were segmented from these images using an image processing software, 3D-DOCTOR(TM). NURBS and polygon mesh surfaces were then used to model individual organs and tissues after importing the segmented organ models to the 3D NURBS modeling software, Rhinoceros(TM). The phantoms were matched to four reference datasets: (1) standard anthropometric data, (2) reference

De novo Transcriptome Assembly of Common Wild Rice (Oryza rufipogon Griff.) and Discovery of Drought-Response Genes in Root Tissue Based on Transcriptomic Data.

Science.gov (United States)

Tian, Xin-Jie; Long, Yan; Wang, Jiao; Zhang, Jing-Wen; Wang, Yan-Yan; Li, Wei-Min; Peng, Yu-Fa; Yuan, Qian-Hua; Pei, Xin-Wu

2015-01-01

The perennial O. rufipogon (common wild rice), which is considered to be the ancestor of Asian cultivated rice species, contains many useful genetic resources, including drought resistance genes. However, few studies have identified the drought resistance and tissue-specific genes in common wild rice. In this study, transcriptome sequencing libraries were constructed, including drought-treated roots (DR) and control leaves (CL) and roots (CR). Using Illumina sequencing technology, we generated 16.75 million bases of high-quality sequence data for common wild rice and conducted de novo assembly and annotation of genes without prior genome information. These reads were assembled into 119,332 unigenes with an average length of 715 bp. A total of 88,813 distinct sequences (74.42% of unigenes) significantly matched known genes in the NCBI NT database. Differentially expressed gene (DEG) analysis showed that 3617 genes were up-regulated and 4171 genes were down-regulated in the CR library compared with the CL library. Among the DEGs, 535 genes were expressed in roots but not in shoots. A similar comparison between the DR and CR libraries showed that 1393 genes were up-regulated and 315 genes were down-regulated in the DR library compared with the CR library. Finally, 37 genes that were specifically expressed in roots were screened after comparing the DEGs identified in the above-described analyses. This study provides a transcriptome sequence resource for common wild rice plants and establishes a digital gene expression profile of wild rice plants under drought conditions using the assembled transcriptome data as a reference. Several tissue-specific and drought-stress-related candidate genes were identified, representing a fully characterized transcriptome and providing a valuable resource for genetic and genomic studies in plants.

Towards micro-assembly of hybrid MOEMS components on a reconfigurable silicon free-space micro-optical bench

International Nuclear Information System (INIS)

Bargiel, S; Gorecki, C; Rabenorosoa, K; Clévy, C; Lutz, P

2010-01-01

The 3D integration of hybrid chips is a viable approach for the micro-optical technologies to reduce the costs of assembly and packaging. In this paper a technology platform for the hybrid integration of MOEMS components on a reconfigurable silicon free-space micro-optical bench (FS-MOB) is presented. In this approach a desired optical component (e.g. micromirror, microlens) is integrated with a removable and adjustable silicon holder which can be manipulated, aligned and fixed in the precisely etched rail of the silicon baseplate by use of a robotic micro-assembly station. An active-based gripping system allows modification of the holder position on the baseplate with nanometre precision. The fabrication processes of the micromachined parts of the micro-optical bench, based on bulk micromachining of standard silicon wafer and SOI wafer, are described. The successful assembly of the holders, equipped with a micromirror and a refractive glass ball microlens, on the baseplate rail is demonstrated.

Improved de novo genomic assembly for the domestic donkey

DEFF Research Database (Denmark)

Renaud, Gabriel; Petersen, Bent; Seguin-Orlando, Andaine

2018-01-01

Donkeys and horses share a common ancestor dating back to about 4 million years ago. Although a high-quality genome assembly at the chromosomal level is available for the horse, current assemblies available for the donkey are limited to moderately sized scaffolds. The absence of a better......-quality assembly for the donkey has hampered studies involving the characterization of patterns of genetic variation at the genome-wide scale. These range from the application of genomic tools to selective breeding and conservation to the more fundamental characterization of the genomic loci underlying speciation...... and domestication. We present a new high-quality donkey genome assembly obtained using the Chicago HiRise assembly technology, providing scaffolds of subchromosomal size. We make use of this new assembly to obtain more accurate measures of heterozygosity for equine species other than the horse, both genome...

A multi-platform draft de novo genome assembly and comparative analysis for the Scarlet Macaw (Ara macao.

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Christopher M Seabury

Full Text Available Data deposition to NCBI Genomes: This Whole Genome Shotgun project has been deposited at DDBJ/EMBL/GenBank under the accession AMXX00000000 (SMACv1.0, unscaffolded genome assembly. The version described in this paper is the first version (AMXX01000000. The scaffolded assembly (SMACv1.1 has been deposited at DDBJ/EMBL/GenBank under the accession AOUJ00000000, and is also the first version (AOUJ01000000. Strong biological interest in traits such as the acquisition and utilization of speech, cognitive abilities, and longevity catalyzed the utilization of two next-generation sequencing platforms to provide the first-draft de novo genome assembly for the large, new world parrot Ara macao (Scarlet Macaw. Despite the challenges associated with genome assembly for an outbred avian species, including 951,507 high-quality putative single nucleotide polymorphisms, the final genome assembly (>1.035 Gb includes more than 997 Mb of unambiguous sequence data (excluding N's. Cytogenetic analyses including ZooFISH revealed complex rearrangements associated with two scarlet macaw macrochromosomes (AMA6, AMA7, which supports the hypothesis that translocations, fusions, and intragenomic rearrangements are key factors associated with karyotype evolution among parrots. In silico annotation of the scarlet macaw genome provided robust evidence for 14,405 nuclear gene annotation models, their predicted transcripts and proteins, and a complete mitochondrial genome. Comparative analyses involving the scarlet macaw, chicken, and zebra finch genomes revealed high levels of nucleotide-based conservation as well as evidence for overall genome stability among the three highly divergent species. Application of a new whole-genome analysis of divergence involving all three species yielded prioritized candidate genes and noncoding regions for parrot traits of interest (i.e., speech, intelligence, longevity which were independently supported by the results of previous human GWAS

Improved de novo genomic assembly for the domestic donkey

Science.gov (United States)

Newton, Richard; Paillot, Romain; Bryant, Neil; Vaudin, Mark

2018-01-01

Donkeys and horses share a common ancestor dating back to about 4 million years ago. Although a high-quality genome assembly at the chromosomal level is available for the horse, current assemblies available for the donkey are limited to moderately sized scaffolds. The absence of a better-quality assembly for the donkey has hampered studies involving the characterization of patterns of genetic variation at the genome-wide scale. These range from the application of genomic tools to selective breeding and conservation to the more fundamental characterization of the genomic loci underlying speciation and domestication. We present a new high-quality donkey genome assembly obtained using the Chicago HiRise assembly technology, providing scaffolds of subchromosomal size. We make use of this new assembly to obtain more accurate measures of heterozygosity for equine species other than the horse, both genome-wide and locally, and to detect runs of homozygosity potentially pertaining to positive selection in domestic donkeys. Finally, this new assembly allowed us to identify fine-scale chromosomal rearrangements between the horse and the donkey that likely played an active role in their divergence and, ultimately, speciation. PMID:29740610

Improved de novo genomic assembly for the domestic donkey.

Science.gov (United States)

Renaud, Gabriel; Petersen, Bent; Seguin-Orlando, Andaine; Bertelsen, Mads Frost; Waller, Andrew; Newton, Richard; Paillot, Romain; Bryant, Neil; Vaudin, Mark; Librado, Pablo; Orlando, Ludovic

2018-04-01

Donkeys and horses share a common ancestor dating back to about 4 million years ago. Although a high-quality genome assembly at the chromosomal level is available for the horse, current assemblies available for the donkey are limited to moderately sized scaffolds. The absence of a better-quality assembly for the donkey has hampered studies involving the characterization of patterns of genetic variation at the genome-wide scale. These range from the application of genomic tools to selective breeding and conservation to the more fundamental characterization of the genomic loci underlying speciation and domestication. We present a new high-quality donkey genome assembly obtained using the Chicago HiRise assembly technology, providing scaffolds of subchromosomal size. We make use of this new assembly to obtain more accurate measures of heterozygosity for equine species other than the horse, both genome-wide and locally, and to detect runs of homozygosity potentially pertaining to positive selection in domestic donkeys. Finally, this new assembly allowed us to identify fine-scale chromosomal rearrangements between the horse and the donkey that likely played an active role in their divergence and, ultimately, speciation.

Fabrication of TiO_2 nanorod assembly grafted rGO (rGO@TiO_2-NR) hybridized flake-like photocatalyst

International Nuclear Information System (INIS)

Lv, Kangle; Fang, Shun; Si, Lingling; Xia, Yang; Ho, Wingkei; Li, Mei

2017-01-01

Highlights: • TiO_2 nanorod assembly grafted with GO hybrid was successfully fabricated. • TiO_2 nanorods can reduce the aggregation of TiO_2 nanoparticles on graphene. • This unique structure facilitates the injection of electron from TiO_2 to graphene. - Abstract: To efficiently separate the photo-generated electron–hole pairs of TiO_2 hybrid, anatase TiO_2 nanorod assembly grafted reduced graphene oxides (rGO@TiO_2-NR) hybrid was successfully fabricated using potassium titanium oxalate (PTO) and graphene oxides (GO) as starting materials and diethylene glycol (DEG) as reductant. The effect of GO content on the structure and photocatalytic activity of rGO@TiO_2-NR composite was systematically studied. Results show that, in the absence of GO, only TiO_2 microsphere assembly is obtained from TiO_2 nanorods. The presence of GO results in the formation of a flake-like TiO_2-nanorod-assembled grafted rGO hybrid. The photocatalytic activity of rGO@TiO_2-NR composite increases first and then decreases with increase in the amount of GO from 0 wt.% to 10 wt.%. The hybridized S4 sample prepared with 4 wt.% GO possesses the highest photocatalytic activity with a constant rate of 0.039 min"−"1 in the photocataytic degradation of Brilliant X-3B dye (X3B); this sample was enhanced more than three times when compared with pure TiO_2 sample (0.012 min"−"1). The enhanced photocatalytic activity of the rGO@TiO_2-NR hybrid was attributed to the strong interaction between TiO_2 nanorods and rGO. The unique hierarchical structure of 1D nanorod assembly TiO_2–rGO flakes facilitates the injection and transfer of photo-generated electrons from TiO_2 to graphene, thus retarding the recombination of electron–hole pairs and enhancing the photocatalytic activity. The enlarged BET surface areas, not only increasing the number of active sites, but also facilitating the adsorption of the dye, and improved light-harvesting ability also contribute to the enhanced photoreactivity

Fabrication of molecular hybrid films of gold nanoparticle and polythiophene by covalent assembly

Energy Technology Data Exchange (ETDEWEB)

Sundaramurthy, Jayaraman, E-mail: jsu2@np.edu.sg [Department of Chemical & Biomolecular Engineering, National University of Singapore, Block E5, 4 Engineering Drive 4, 117576 (Singapore); Environmental & Water Technology Centre of Innovation, Ngee Ann Polytechnic, 599489 (Singapore); Dharmarajan, Rajarathnam [CERAR, University of South Australia, Mawson Lakes, SA 5095 (Australia); Srinivasan, M.P., E-mail: chesmp@nus.edu.sg [Department of Chemical & Biomolecular Engineering, National University of Singapore, Block E5, 4 Engineering Drive 4, 117576 (Singapore)

2015-08-31

This work demonstrates the fabrication of molecular hybrid films comprising gold nanoparticles (AuNPs) incorporated in covalently assembled, substituted polythiophene (poly(3-(2-bromoethoxy)ethoxymethylthiophene-2,5-diyl (PBrEEMT))) films by different surface chemistry routes. AuNPs are incorporated in the immobilized polythiophene matrix due to its affinity for amine and sulfur. The amount of AuNPs present depends on the nature of the incorporation, the extent of film coverage and interaction of thiophene and amine groups. PBrEEMT films functionalized with amine rich polyallylamine immobilize greater numbers of AuNPs due to more extensive gold–amine interactions. Covalent binding between AuNP and PBrEEMT films was accomplished by using pre-functionalised AuNPs (4-aminothiophenol functionalized AuNPs). Atomic force microscopy, field emission scanning electron microscopy and X-ray photoelectron spectroscopy were used to study the morphology and chemical constituents of assembled films. These approaches will pave the way for developing facile methods for nanoparticle incorporation and will also facilitate direct interaction of nanoparticles with the conducting polymer matrix and enhance the electrical properties of the films. - Highlights: • Covalent molecular assembly enabled the fabrication of molecular hybrid films. • Monomeric and polymeric species were employed as intermediate linkers. • Adopted approaches facilitated the direct interaction of gold nanoparticle in films. • The amount of nanoparticle incorporation depended on the extent of film coverage.

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

Stabilization of Pt nanoparticles by single stranded DNA and the binary assembly of Au and Pt nanoparticles without hybridization

International Nuclear Information System (INIS)

Yang, J.; Lee, Jim Yang; Too, Heng-Phon; Chow, Gan-Moog; Gan, Leong M.

2006-01-01

The non-specific interaction between single stranded DNA (ssDNA) and 12 nm Pt nanoparticles is investigated in this work. The data show a strong and non-specific interaction between the two which can be exploited for the stabilization of Pt nanoparticles in aqueous solutions. Based on the experimental findings, a non-hybridization based protocol to assemble 17 nm Au and Pt nanoparticles (12 nm cubic and 3.6 nm spherical) by single-stranded DNA was developed. Transmission electron microscopy (TEM) and UV-visible spectroscopy confirmed that Au and Pt nanoparticles could be assembled by the non-specific interaction in an orderly manner. The experimental results also caution against the potential pitfalls in using DNA melting point analysis to infer metal nanoparticle assembly by DNA hybridization

De Novo Assembly and Comparative Transcriptome Analysis Provide Insight into Lysine Biosynthesis in Toona sinensis Roem.

Science.gov (United States)

Zhang, Xia; Song, Zhenqiao; Liu, Tian; Guo, Linlin; Li, Xingfeng

2016-01-01

Toona sinensis Roem is a popular leafy vegetable in Chinese cuisine and is also used as a traditional Chinese medicine. In this study, leaf samples were collected from the same plant on two development stages and then used for high-throughput Illumina RNA-sequencing (RNA-Seq). 125,884 transcripts and 54,628 unigenes were obtained through de novo assembly. A total of 25,570 could be annotated with known biological functions, which indicated that the T. sinensis leaves and shoots were undergoing multiple developmental processes especially for active metabolic processes. Analysis of differentially expressed unigenes between the two libraries showed that the lysine biosynthesis was an enriched KEGG pathway, and candidate genes involved in the lysine biosynthesis pathway in T. sinensis leaves and shoots were identified. Our results provide a primary analysis of the gene expression files of T. sinensis leaf and shoot on different development stages and afford a valuable resource for genetic and genomic research on plant lysine biosynthesis.

De novo transcriptome assembly and quantification reveal differentially expressed genes between soft-seed and hard-seed pomegranate (Punica granatum L..

Directory of Open Access Journals (Sweden)

Hui Xue

Full Text Available Pomegranate (Punica granatum L. belongs to Punicaceae, and is valued for its social, ecological, economic, and aesthetic values, as well as more recently for its health benefits. The 'Tunisia' variety has softer seeds and big arils that are easily swallowed. It is a widely popular fruit; however, the molecular mechanisms of the formation of hard and soft seeds is not yet clear. We conducted a de novo assembly of the seed transcriptome in P. granatum L. and revealed differential gene expression between the soft-seed and hard-seed pomegranate varieties. A total of 35.1 Gb of data were acquired in this study, including 280,881,106 raw reads. Additionally, de novo transcriptome assembly generated 132,287 transcripts and 105,743 representative unigenes; approximately 13,805 unigenes (37.7% were longer than 1,000 bp. Using bioinformatics annotation libraries, a total of 76,806 unigenes were annotated and, among the high-quality reads, 72.63% had at least one significant match to an existing gene model. Gene expression and differentially expressed genes were analyzed. The seed formation of the two pomegranate cultivars involves lignin biosynthesis and metabolism, including some genes encoding laccase and peroxidase, WRKY, MYB, and NAC transcription factors. In the hard-seed pomegranate, lignin-related genes and cellulose synthesis-related genes were highly expressed; in soft-seed pomegranates, expression of genes related to flavonoids and programmed cell death was slightly higher. We validated selection of the identified genes using qRT-PCR. This is the first transcriptome analysis of P. granatum L. This transcription sequencing greatly enriched the pomegranate molecular database, and the high-quality SSRs generated in this study will aid the gene cloning from pomegranate in the future. It provides important insights into the molecular mechanisms underlying the formation of soft seeds in pomegranate.

De novo transcriptome assembly and quantification reveal differentially expressed genes between soft-seed and hard-seed pomegranate (Punica granatum L.).

Science.gov (United States)

Xue, Hui; Cao, Shangyin; Li, Haoxian; Zhang, Jie; Niu, Juan; Chen, Lina; Zhang, Fuhong; Zhao, Diguang

2017-01-01

Pomegranate (Punica granatum L.) belongs to Punicaceae, and is valued for its social, ecological, economic, and aesthetic values, as well as more recently for its health benefits. The 'Tunisia' variety has softer seeds and big arils that are easily swallowed. It is a widely popular fruit; however, the molecular mechanisms of the formation of hard and soft seeds is not yet clear. We conducted a de novo assembly of the seed transcriptome in P. granatum L. and revealed differential gene expression between the soft-seed and hard-seed pomegranate varieties. A total of 35.1 Gb of data were acquired in this study, including 280,881,106 raw reads. Additionally, de novo transcriptome assembly generated 132,287 transcripts and 105,743 representative unigenes; approximately 13,805 unigenes (37.7%) were longer than 1,000 bp. Using bioinformatics annotation libraries, a total of 76,806 unigenes were annotated and, among the high-quality reads, 72.63% had at least one significant match to an existing gene model. Gene expression and differentially expressed genes were analyzed. The seed formation of the two pomegranate cultivars involves lignin biosynthesis and metabolism, including some genes encoding laccase and peroxidase, WRKY, MYB, and NAC transcription factors. In the hard-seed pomegranate, lignin-related genes and cellulose synthesis-related genes were highly expressed; in soft-seed pomegranates, expression of genes related to flavonoids and programmed cell death was slightly higher. We validated selection of the identified genes using qRT-PCR. This is the first transcriptome analysis of P. granatum L. This transcription sequencing greatly enriched the pomegranate molecular database, and the high-quality SSRs generated in this study will aid the gene cloning from pomegranate in the future. It provides important insights into the molecular mechanisms underlying the formation of soft seeds in pomegranate.

De novo transcriptome sequencing of axolotl blastema for identification of differentially expressed genes during limb regeneration

Science.gov (United States)

2013-01-01

Background Salamanders are unique among vertebrates in their ability to completely regenerate amputated limbs through the mediation of blastema cells located at the stump ends. This regeneration is nerve-dependent because blastema formation and regeneration does not occur after limb denervation. To obtain the genomic information of blastema tissues, de novo transcriptomes from both blastema tissues and denervated stump ends of Ambystoma mexicanum (axolotls) 14 days post-amputation were sequenced and compared using Solexa DNA sequencing. Results The sequencing done for this study produced 40,688,892 reads that were assembled into 307,345 transcribed sequences. The N50 of transcribed sequence length was 562 bases. A similarity search with known proteins identified 39,200 different genes to be expressed during limb regeneration with a cut-off E-value exceeding 10-5. We annotated assembled sequences by using gene descriptions, gene ontology, and clusters of orthologous group terms. Targeted searches using these annotations showed that the majority of the genes were in the categories of essential metabolic pathways, transcription factors and conserved signaling pathways, and novel candidate genes for regenerative processes. We discovered and confirmed numerous sequences of the candidate genes by using quantitative polymerase chain reaction and in situ hybridization. Conclusion The results of this study demonstrate that de novo transcriptome sequencing allows gene expression analysis in a species lacking genome information and provides the most comprehensive mRNA sequence resources for axolotls. The characterization of the axolotl transcriptome can help elucidate the molecular mechanisms underlying blastema formation during limb regeneration. PMID:23815514

De novo transcriptome assembly for the tropical grass Panicum maximum Jacq.

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Guilherme Toledo-Silva

Full Text Available Guinea grass (Panicum maximum Jacq. is a tropical African grass often used to feed beef cattle, which is an important economic activity in Brazil. Brazil is the leader in global meat exportation because of its exclusively pasture-raised bovine herds. Guinea grass also has potential uses in bioenergy production due to its elevated biomass generation through the C4 photosynthesis pathway. We generated approximately 13 Gb of data from Illumina sequencing of P. maximum leaves. Four different genotypes were sequenced, and the combined reads were assembled de novo into 38,192 unigenes and annotated; approximately 63% of the unigenes had homology to other proteins in the NCBI non-redundant protein database. Functional classification through COG (Clusters of Orthologous Groups, GO (Gene Ontology and KEGG (Kyoto Encyclopedia of Genes and Genomes analyses showed that the unigenes from Guinea grass leaves are involved in a wide range of biological processes and metabolic pathways, including C4 photosynthesis and lignocellulose generation, which are important for cattle grazing and bioenergy production. The most abundant transcripts were involved in carbon fixation, photosynthesis, RNA translation and heavy metal cellular homeostasis. Finally, we identified a number of potential molecular markers, including 5,035 microsatellites (SSRs and 346,456 single nucleotide polymorphisms (SNPs. To the best of our knowledge, this is the first study to characterize the complete leaf transcriptome of P. maximum using high-throughput sequencing. The biological information provided here will aid in gene expression studies and marker-assisted selection-based breeding research in tropical grasses.

De Novo Transcriptomic Analysis of an Oleaginous Microalga: Pathway Description and Gene Discovery for Production of Next-Generation Biofuels

Science.gov (United States)

Wan, LingLin; Han, Juan; Sang, Min; Li, AiFen; Wu, Hong; Yin, ShunJi; Zhang, ChengWu

2012-01-01

Background Eustigmatos cf. polyphem is a yellow-green unicellular soil microalga belonging to the eustimatophyte with high biomass and considerable production of triacylglycerols (TAGs) for biofuels, which is thus referred to as an oleaginous microalga. The paucity of microalgae genome sequences, however, limits development of gene-based biofuel feedstock optimization studies. Here we describe the sequencing and de novo transcriptome assembly for a non-model microalgae species, E. cf. polyphem, and identify pathways and genes of importance related to biofuel production. Results We performed the de novo assembly of E. cf. polyphem transcriptome using Illumina paired-end sequencing technology. In a single run, we produced 29,199,432 sequencing reads corresponding to 2.33 Gb total nucleotides. These reads were assembled into 75,632 unigenes with a mean size of 503 bp and an N50 of 663 bp, ranging from 100 bp to >3,000 bp. Assembled unigenes were subjected to BLAST similarity searches and annotated with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) orthology identifiers. These analyses identified the majority of carbohydrate, fatty acids, TAG and carotenoids biosynthesis and catabolism pathways in E. cf. polyphem. Conclusions Our data provides the construction of metabolic pathways involved in the biosynthesis and catabolism of carbohydrate, fatty acids, TAG and carotenoids in E. cf. polyphem and provides a foundation for the molecular genetics and functional genomics required to direct metabolic engineering efforts that seek to enhance the quantity and character of microalgae-based biofuel feedstock. PMID:22536352

Illumina-based de novo transcriptome sequencing and analysis

Indian Academy of Sciences (India)

In the present study, we used Illumina HiSeq technology to perform de novo assembly of heart and musk gland transcriptomes from the Chinese forest musk deer. A total of 239,383 transcripts and 176,450 unigenes were obtained, of which 37,329 unigenes were matched to known sequences in the NCBI nonredundant ...

The mitochondrial genomes of Atlas Geckos (Quedenfeldtia): mitogenome assembly from transcriptomes and anchored hybrid enrichment datasets

OpenAIRE

Lyra, Mariana L.; Joger, Ulrich; Schulte, Ulrich; Slimani, Tahar; El Mouden, El Hassan; Bouazza, Abdellah; Künzel, Sven; Lemmon, Alan R.; Moriarty Lemmon, Emily; Vences, Miguel

2017-01-01

The nearly complete mitogenomes of the two species of North African Atlas geckos, Quedenfeldtia moerens and Q. trachyblepharus were assembled from anchored hybrid enrichment data and RNAseq data. Congruent assemblies were obtained for four samples included in both datasets. We recovered the 13 protein-coding genes, 22 tRNA genes, and two rRNA genes for both species, including partial control region. The order of genes agrees with that of other geckos.

Integrated effect of supramolecular self-assembled sandwich-like melamine cyanurate/MoS{sub 2} hybrid sheets on reducing fire hazards of polyamide 6 composites

Energy Technology Data Exchange (ETDEWEB)

Feng, Xiaming [State Key Laboratory of Fire Science, University of Science and Technology of China, Anhui 230026 (China); Suzhou Key Laboratory of Urban Public Safety, Suzhou Institute for Advanced Study, University of Science and Technology of China, Suzhou, Jiangsu 215123 (China); Wang, Xin, E-mail: wxcmx@ustc.edu.cn [State Key Laboratory of Fire Science, University of Science and Technology of China, Anhui 230026 (China); Cai, Wei; Hong, Ningning [State Key Laboratory of Fire Science, University of Science and Technology of China, Anhui 230026 (China); Hu, Yuan, E-mail: yuanhu@ustc.edu.cn [State Key Laboratory of Fire Science, University of Science and Technology of China, Anhui 230026 (China); Suzhou Key Laboratory of Urban Public Safety, Suzhou Institute for Advanced Study, University of Science and Technology of China, Suzhou, Jiangsu 215123 (China); Liew, Kim Meow [Suzhou Key Laboratory of Urban Public Safety, Suzhou Institute for Advanced Study, University of Science and Technology of China, Suzhou, Jiangsu 215123 (China); Department of Architectural and Civil Engineering, City University of Hong Kong, Tat Chee Avenue, Kowloon (Hong Kong)

2016-12-15

A novel strategy of using supramolecular self-assembly for preparing sandwich-like melamine cyanurate/MoS{sub 2} sheets as the hybrid flame retardants for polyamide 6 (PA6) is reported for the first time. The introduction of MoS{sub 2} sheets function not only as a template to induce the formation of two-dimensional melamine cyanurate capping layers but also as a synergist to generate integrated flame-retarding effect of hybrid sheets, as well as a high-performance smoke suppressor to reduce fire hazards of PA6 materials. Once incorporating this well-designed structures (4 wt%) into PA6 matrix, there resulted in a remarkable drop (40%) in the peak heat release rate and a 25% reduction in total heat release. Moreover, the smoke production and pyrolysis gaseous products were efficiently suppressed by the addition of sandwich-like hybrid sheets. The integrated functions consisting of inherent flame retarding effect, physical barrier performance and catalytic activity are believed to the crucial guarantee for the reduced fire hazards of PA6 nanocomposites. Furthermore, this novel strategy with facile and scalable features may provide reference for developing various kinds of MoS{sub 2} based hybrid sheets for diverse applications.

Hybrid pregnant reference phantom series based on adult female ICRP reference phantom

Science.gov (United States)

Rafat-Motavalli, Laleh; Miri-Hakimabad, Hashem; Hoseinian-Azghadi, Elie

2018-03-01

This paper presents boundary representation (BREP) models of pregnant female and her fetus at the end of each trimester. The International Commission on Radiological Protection (ICRP) female reference voxel phantom was used as a base template in development process of the pregnant hybrid phantom series. The differences in shape and location of the displaced maternal organs caused by enlarging uterus were also taken into account. The CT and MR images of fetus specimens and pregnant patients of various ages were used to replace the maternal abdominal pelvic organs of template phantom and insert the fetus inside the gravid uterus. Each fetal model contains 21 different organs and tissues. The skeletal model of the fetus also includes age-dependent cartilaginous and ossified skeletal components. The replaced maternal organ models were converted to NURBS surfaces and then modified to conform to reference values of ICRP Publication 89. The particular feature of current series compared to the previously developed pregnant phantoms is being constructed upon the basis of ICRP reference phantom. The maternal replaced organ models are NURBS surfaces. With this great potential, they might have the feasibility of being converted to high quality polygon mesh phantoms.

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.

Self-Assembly of 1D/2D Hybrid Nanostructures Consisting of a Cd(II Coordination Polymer and NiAl-Layered Double Hydroxides

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Gonzalo Abellán

2015-12-01

Full Text Available The preparation and characterization of a novel hybrid material based on the combination of a 2D-layered double hydroxide (LDH nanosheets and a 1D-coordination polymer (1D-CP has been achieved through a simple mixture of suspensions of both building blocks via an exfoliation/restacking approach. The hybrid material has been thoroughly characterized demonstrating that the 1D-CP moieties are intercalated as well as adsorbed on the surface of the LDH, giving rise to a layered assembly with the coexistence of the functionalities of their initial constituents. This hybrid represents the first example of the assembly of 1D/2D nanomaterials combining LDH with CP and opens the door for a plethora of different functional hybrid systems.

Transcriptomic identification of salt-related genes and de novo assembly in common buckwheat (F. esculentum).

Science.gov (United States)

Lu, Qi-Huan; Wang, Ya-Qi; Song, Jin-Nan; Yang, Hong-Bing

2018-06-01

Common buckwheat (F. esculentum), annually herbaceous crop, is prevalent in people's daily life with the increasing development of economics. Compared with wheat, it is highly praised with high content of rutin and flavonoid. Common buckwheat is recognized as healthy food with good taste, and the product price of which such as noodles, flour, bread and so on are higher than wheat, and the seeds of which are bigger than that of tartary buckwheat, so if common buckwheat are planted more widely, people will spend less money on this healthy and delicious food. However, soil salinity has been a giant problem for agriculture production. The cultivation of salt tolerant crop varieties is an effective way to make full use of saline alkali land, and the highest salinity that the common buckwheat can sow is at 6.0%, so we chose 100 mM as the concentration of NaCl for treatment. Then we conducted transcriptome comparison between control and treatment groups. Potential regulatory genes related salt stress in common buckwheat were identified. A total of 29.36 million clean reads were produced via an illumina sequencing approach. We de novo assembled these reads into a transcriptome dataset containing 43,772 unigenes with N50 length of 1778 bp. A total of 26,672 unigenes could be found matches in public databases. GO, KEGG and Swiss-Prot classification suggested the enrichment of these unigenes in 47 sub-categories, 25 KOG and 129 pathways, respectively. We got 385 differentially expressed genes (DEGs) after comparing the transcriptome data between salt treatment and control groups. There are some genes encoded for responsing to stimulus, cell killing, metabolic process, signaling, multi-organism process, growth and cellular process might be relevant to salt stress in common buckwheat, which will provide a valuable references for the study on mechanism of salt tolerance and will be used as a genetic information for cultivating strong salt tolerant common buckwheat varieties in

Antimicrobial peptide capsids of de novo design.

Science.gov (United States)

De Santis, Emiliana; Alkassem, Hasan; Lamarre, Baptiste; Faruqui, Nilofar; Bella, Angelo; Noble, James E; Micale, Nicola; Ray, Santanu; Burns, Jonathan R; Yon, Alexander R; Hoogenboom, Bart W; Ryadnov, Maxim G

2017-12-22

The spread of bacterial resistance to antibiotics poses the need for antimicrobial discovery. With traditional search paradigms being exhausted, approaches that are altogether different from antibiotics may offer promising and creative solutions. Here, we introduce a de novo peptide topology that-by emulating the virus architecture-assembles into discrete antimicrobial capsids. Using the combination of high-resolution and real-time imaging, we demonstrate that these artificial capsids assemble as 20-nm hollow shells that attack bacterial membranes and upon landing on phospholipid bilayers instantaneously (seconds) convert into rapidly expanding pores causing membrane lysis (minutes). The designed capsids show broad antimicrobial activities, thus executing one primary function-they destroy bacteria on contact.

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.

Quantitative characterization of colloidal assembly of graphene oxide-silver nanoparticle hybrids using aerosol differential mobility-coupled mass analyses.

Science.gov (United States)

Nguyen, Thai Phuong; Chang, Wei-Chang; Lai, Yen-Chih; Hsiao, Ta-Chih; Tsai, De-Hao

2017-10-01

In this work, we develop an aerosol-based, time-resolved ion mobility-coupled mass characterization method to investigate colloidal assembly of graphene oxide (GO)-silver nanoparticle (AgNP) hybrid nanostructure on a quantitative basis. Transmission electron microscopy (TEM) and zeta potential (ZP) analysis were used to provide visual information and elemental-based particle size distributions, respectively. Results clearly show a successful controlled assembly of GO-AgNP by electrostatic-directed heterogeneous aggregation between GO and bovine serum albumin (BSA)-functionalized AgNP under an acidic environment. Additionally, physical size, mass, and conformation (i.e., number of AgNP per nanohybrid) of GO-AgNP were shown to be proportional to the number concentration ratio of AgNP to GO (R) and the selected electrical mobility diameter. An analysis of colloidal stability of GO-AgNP indicates that the stability increased with its absolute ZP, which was dependent on R and environmental pH. The work presented here provides a proof of concept for systematically synthesizing hybrid colloidal nanomaterials through the tuning of surface chemistry in aqueous phase with the ability in quantitative characterization. Graphical Abstract Colloidal assembly of graphene oxide-silver nanoparticle hybrids characterized by aerosol differential mobility-coupled mass analyses.

De novo identification of viral pathogens from cell culture hologenomes

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Patowary Ashok

2012-01-01

Full Text Available Abstract Background Fast, specific identification and surveillance of pathogens is the cornerstone of any outbreak response system, especially in the case of emerging infectious diseases and viral epidemics. This process is generally tedious and time-consuming thus making it ineffective in traditional settings. The added complexity in these situations is the non-availability of pure isolates of pathogens as they are present as mixed genomes or hologenomes. Next-generation sequencing approaches offer an attractive solution in this scenario as it provides adequate depth of sequencing at fast and affordable costs, apart from making it possible to decipher complex interactions between genomes at a scale that was not possible before. The widespread application of next-generation sequencing in this field has been limited by the non-availability of an efficient computational pipeline to systematically analyze data to delineate pathogen genomes from mixed population of genomes or hologenomes. Findings We applied next-generation sequencing on a sample containing mixed population of genomes from an epidemic with appropriate processing and enrichment. The data was analyzed using an extensive computational pipeline involving mapping to reference genome sets and de-novo assembly. In depth analysis of the data generated revealed the presence of sequences corresponding to Japanese encephalitis virus. The genome of the virus was also independently de-novo assembled. The presence of the virus was in addition, verified using standard molecular biology techniques. Conclusions Our approach can accurately identify causative pathogens from cell culture hologenome samples containing mixed population of genomes and in principle can be applied to patient hologenome samples without any background information. This methodology could be widely applied to identify and isolate pathogen genomes and understand their genomic variability during outbreaks.

Fabrication of TiO2 nanorod assembly grafted rGO (rGO@TiO2-NR) hybridized flake-like photocatalyst

Science.gov (United States)

Lv, Kangle; Fang, Shun; Si, Lingling; Xia, Yang; Ho, Wingkei; Li, Mei

2017-01-01

To efficiently separate the photo-generated electron-hole pairs of TiO2 hybrid, anatase TiO2 nanorod assembly grafted reduced graphene oxides (rGO@TiO2-NR) hybrid was successfully fabricated using potassium titanium oxalate (PTO) and graphene oxides (GO) as starting materials and diethylene glycol (DEG) as reductant. The effect of GO content on the structure and photocatalytic activity of rGO@TiO2-NR composite was systematically studied. Results show that, in the absence of GO, only TiO2 microsphere assembly is obtained from TiO2 nanorods. The presence of GO results in the formation of a flake-like TiO2-nanorod-assembled grafted rGO hybrid. The photocatalytic activity of rGO@TiO2-NR composite increases first and then decreases with increase in the amount of GO from 0 wt.% to 10 wt.%. The hybridized S4 sample prepared with 4 wt.% GO possesses the highest photocatalytic activity with a constant rate of 0.039 min-1 in the photocataytic degradation of Brilliant X-3B dye (X3B); this sample was enhanced more than three times when compared with pure TiO2 sample (0.012 min-1). The enhanced photocatalytic activity of the rGO@TiO2-NR hybrid was attributed to the strong interaction between TiO2 nanorods and rGO. The unique hierarchical structure of 1D nanorod assembly TiO2-rGO flakes facilitates the injection and transfer of photo-generated electrons from TiO2 to graphene, thus retarding the recombination of electron-hole pairs and enhancing the photocatalytic activity. The enlarged BET surface areas, not only increasing the number of active sites, but also facilitating the adsorption of the dye, and improved light-harvesting ability also contribute to the enhanced photoreactivity of rGO@TiO2-NR hybrid.

De novo clustering methods outperform reference-based methods for assigning 16S rRNA gene sequences to operational taxonomic units

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Sarah L. Westcott

2015-12-01

Full Text Available Background. 16S rRNA gene sequences are routinely assigned to operational taxonomic units (OTUs that are then used to analyze complex microbial communities. A number of methods have been employed to carry out the assignment of 16S rRNA gene sequences to OTUs leading to confusion over which method is optimal. A recent study suggested that a clustering method should be selected based on its ability to generate stable OTU assignments that do not change as additional sequences are added to the dataset. In contrast, we contend that the quality of the OTU assignments, the ability of the method to properly represent the distances between the sequences, is more important.Methods. Our analysis implemented six de novo clustering algorithms including the single linkage, complete linkage, average linkage, abundance-based greedy clustering, distance-based greedy clustering, and Swarm and the open and closed-reference methods. Using two previously published datasets we used the Matthew’s Correlation Coefficient (MCC to assess the stability and quality of OTU assignments.Results. The stability of OTU assignments did not reflect the quality of the assignments. Depending on the dataset being analyzed, the average linkage and the distance and abundance-based greedy clustering methods generated OTUs that were more likely to represent the actual distances between sequences than the open and closed-reference methods. We also demonstrated that for the greedy algorithms VSEARCH produced assignments that were comparable to those produced by USEARCH making VSEARCH a viable free and open source alternative to USEARCH. Further interrogation of the reference-based methods indicated that when USEARCH or VSEARCH were used to identify the closest reference, the OTU assignments were sensitive to the order of the reference sequences because the reference sequences can be identical over the region being considered. More troubling was the observation that while both USEARCH and

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.

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

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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.

De novo assembly and comparison of the ovarian transcriptomes of the common Chinese cuttlefish (Sepiella japonica with different gonadal development

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Zhenming Lü

2016-03-01

Full Text Available The common Chinese cuttlefish (Sepiella japonica has been considered one of the most economically important marine Cephalopod species in East Asia and seed breeding technology has been established for massive aquaculture and stock enhancement. In the present study, we used Illumina HiSeq2000 to sequence, assemble and annotate the transcriptome of the ovary tissues of S. japonica for the first time. A total of 53,116,650 and 53,446,640 reads were obtained from the immature and matured ovaries, respectively (NCBI SRA database SRX1409472 and SRX1409473, and 70,039 contigs (N50 = 1443 bp were obtained after de novo assembling with Trinity software. Digital gene expression analysis reveals 47,288 contigs show differential expression profile and 793 contigs are highly expressed in the immature ovary, while 38 contigs are highly expressed in the mature ovary with FPKM >100. We hope that the ovarian transcriptome and those stage-enriched transcripts of S. japonica can provide some insight into the understanding of genome-wide transcriptome profile of cuttlefish gonad tissue and give useful information in cuttlefish gonad development. Keywords: Cuttlefish, Gonad development, Transcriptome

Identifying wrong assemblies in de novo short read primary

Indian Academy of Sciences (India)

Finally, some mis-assembly detecting tools have been evaluated for their ability to detect the wrongly assembledprimary contigs, suggesting a lot of scope for improvement in this area. The present work also proposes a simpleunsupervised learning-based novel approach to identify mis-assemblies in the contigs which was ...

De novo assembly and transcriptome analysis of five major tissues of Jatropha curcas L. using GS FLX titanium platform of 454 pyrosequencing

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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

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

Modeling the Hybrid Flow Shop Scheduling Problem Followed by an Assembly Stage Considering Aging Effects and Preventive Maintenance Activities

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Seyyed Mohammad Hassan Hosseini

2016-05-01

Full Text Available Scheduling problem for the hybrid flow shop scheduling problem (HFSP followed by an assembly stage considering aging effects additional preventive and maintenance activities is studied in this paper. In this production system, a number of products of different kinds are produced. Each product is assembled with a set of several parts. The first stage is a hybrid flow shop to produce parts. All machines can process all kinds of parts in this stage but each machine can process only one part at the same time. The second stage is a single assembly machine or a single assembly team of workers. The aim is to schedule the parts on the machines and assembly sequence and also determine when the preventive maintenance activities get done in order to minimize the completion time of all products (makespan. A mathematical modeling is presented and its validation is shown by solving an example in small scale. Since this problem has been proved strongly NP-hard, in order to solve the problem in medium and large scale, four heuristic algorithms is proposed based on the Johnson’s algorithm. The numerical experiments are used to run the mathematical model and evaluate the performance of the proposed algorithms.

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.

De novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome to identify putative genes involved in the aquatic adaptation and immune response.

Science.gov (United States)

Gui, Duan; Jia, Kuntong; Xia, Jia; Yang, Lili; Chen, Jialin; Wu, Yuping; Yi, Meisheng

2013-01-01

The Indo-Pacific humpback dolphin (Sousa chinensis), a marine mammal species inhabited in the waters of Southeast Asia, South Africa and Australia, has attracted much attention because of the dramatic decline in population size in the past decades, which raises the concern of extinction. So far, this species is poorly characterized at molecular level due to little sequence information available in public databases. Recent advances in large-scale RNA sequencing provide an efficient approach to generate abundant sequences for functional genomic analyses in the species with un-sequenced genomes. We performed a de novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome by Illumina sequencing. 108,751 high quality sequences from 47,840,388 paired-end reads were generated, and 48,868 and 46,587 unigenes were functionally annotated by BLAST search against the NCBI non-redundant and Swiss-Prot protein databases (E-valueIndo-Pacific humpback dolphin, an endangered species. The de novo transcriptome analysis of the unique transcripts will provide valuable sequence information for discovery of new genes, characterization of gene expression, investigation of various pathways and adaptive evolution, as well as identification of genetic markers.

De novo transcriptome assembly and its annotation for the aposematic wood tiger moth (Parasemia plantaginis

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Juan A. Galarza

2017-06-01

Full Text Available In this paper we report the public availability of transcriptome resources for the aposematic wood tiger moth (Parasemia plantaginis. A comprehensive assembly methods, quality statistics, and annotation are provided. This reference transcriptome may serve as a useful resource for investigating functional gene activity in aposematic Lepidopteran species. All data is freely available at the European Nucleotide Archive (http://www.ebi.ac.uk/ena under study accession number: PRJEB14172.

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

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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

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

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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.

Fabrication of TiO{sub 2} nanorod assembly grafted rGO (rGO@TiO{sub 2}-NR) hybridized flake-like photocatalyst

Energy Technology Data Exchange (ETDEWEB)

Lv, Kangle, E-mail: lvkangle@mail.scuec.edu.cn [Department of Science and Environmental Studies, The Hong Kong Institute of Education, Taipo, N.T., Hong Kong (China); Key Laboratory of Catalysis and Materials Science of the State Ethnic Affairs Commission and Ministry of Education, College of Resources and Environmental Science, South-Central University for Nationalities, Wuhan 430074 (China); Fang, Shun; Si, Lingling; Xia, Yang [Key Laboratory of Catalysis and Materials Science of the State Ethnic Affairs Commission and Ministry of Education, College of Resources and Environmental Science, South-Central University for Nationalities, Wuhan 430074 (China); Ho, Wingkei, E-mail: keithho@ied.edu.hk [Department of Science and Environmental Studies, The Hong Kong Institute of Education, Taipo, N.T., Hong Kong (China); Li, Mei [Key Laboratory of Catalysis and Materials Science of the State Ethnic Affairs Commission and Ministry of Education, College of Resources and Environmental Science, South-Central University for Nationalities, Wuhan 430074 (China)

2017-01-01

Highlights: • TiO{sub 2} nanorod assembly grafted with GO hybrid was successfully fabricated. • TiO{sub 2} nanorods can reduce the aggregation of TiO{sub 2} nanoparticles on graphene. • This unique structure facilitates the injection of electron from TiO{sub 2} to graphene. - Abstract: To efficiently separate the photo-generated electron–hole pairs of TiO{sub 2} hybrid, anatase TiO{sub 2} nanorod assembly grafted reduced graphene oxides (rGO@TiO{sub 2}-NR) hybrid was successfully fabricated using potassium titanium oxalate (PTO) and graphene oxides (GO) as starting materials and diethylene glycol (DEG) as reductant. The effect of GO content on the structure and photocatalytic activity of rGO@TiO{sub 2}-NR composite was systematically studied. Results show that, in the absence of GO, only TiO{sub 2} microsphere assembly is obtained from TiO{sub 2} nanorods. The presence of GO results in the formation of a flake-like TiO{sub 2}-nanorod-assembled grafted rGO hybrid. The photocatalytic activity of rGO@TiO{sub 2}-NR composite increases first and then decreases with increase in the amount of GO from 0 wt.% to 10 wt.%. The hybridized S4 sample prepared with 4 wt.% GO possesses the highest photocatalytic activity with a constant rate of 0.039 min{sup −1} in the photocataytic degradation of Brilliant X-3B dye (X3B); this sample was enhanced more than three times when compared with pure TiO{sub 2} sample (0.012 min{sup −1}). The enhanced photocatalytic activity of the rGO@TiO{sub 2}-NR hybrid was attributed to the strong interaction between TiO{sub 2} nanorods and rGO. The unique hierarchical structure of 1D nanorod assembly TiO{sub 2}–rGO flakes facilitates the injection and transfer of photo-generated electrons from TiO{sub 2} to graphene, thus retarding the recombination of electron–hole pairs and enhancing the photocatalytic activity. The enlarged BET surface areas, not only increasing the number of active sites, but also facilitating the adsorption of

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.

UniNovo: a universal tool for de novo peptide sequencing.

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Jeong, Kyowon; Kim, Sangtae; Pevzner, Pavel A

2013-08-15

Mass spectrometry (MS) instruments and experimental protocols are rapidly advancing, but de novo peptide sequencing algorithms to analyze tandem mass (MS/MS) spectra are lagging behind. Although existing de novo sequencing tools perform well on certain types of spectra [e.g. Collision Induced Dissociation (CID) spectra of tryptic peptides], their performance often deteriorates on other types of spectra, such as Electron Transfer Dissociation (ETD), Higher-energy Collisional Dissociation (HCD) spectra or spectra of non-tryptic digests. Thus, rather than developing a new algorithm for each type of spectra, we develop a universal de novo sequencing algorithm called UniNovo that works well for all types of spectra or even for spectral pairs (e.g. CID/ETD spectral pairs). UniNovo uses an improved scoring function that captures the dependences between different ion types, where such dependencies are learned automatically using a modified offset frequency function. The performance of UniNovo is compared with PepNovo+, PEAKS and pNovo using various types of spectra. The results show that the performance of UniNovo is superior to other tools for ETD spectra and superior or comparable with others for CID and HCD spectra. UniNovo also estimates the probability that each reported reconstruction is correct, using simple statistics that are readily obtained from a small training dataset. We demonstrate that the estimation is accurate for all tested types of spectra (including CID, HCD, ETD, CID/ETD and HCD/ETD spectra of trypsin, LysC or AspN digested peptides). UniNovo is implemented in JAVA and tested on Windows, Ubuntu and OS X machines. UniNovo is available at http://proteomics.ucsd.edu/Software/UniNovo.html along with the manual.

Genomes correction and assembling: present methods and tools

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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.

Assembly of viral genomes from metagenomes

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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.

Hybrid CMS methods with model reduction for assembly of structures

Science.gov (United States)

Farhat, Charbel

1991-01-01

Future on-orbit structures will be designed and built in several stages, each with specific control requirements. Therefore there must be a methodology which can predict the dynamic characteristics of the assembled structure, based on the dynamic characteristics of the subassemblies and their interfaces. The methodology developed by CSC to address this issue is Hybrid Component Mode Synthesis (HCMS). HCMS distinguishes itself from standard component mode synthesis algorithms in the following features: (1) it does not require the subcomponents to have displacement compatible models, which makes it ideal for analyzing the deployment of heterogeneous flexible multibody systems, (2) it incorporates a second-level model reduction scheme at the interface, which makes it much faster than other algorithms and therefore suitable for control purposes, and (3) it does answer specific questions such as 'how does the global fundamental frequency vary if I change the physical parameters of substructure k by a specified amount?'. Because it is based on an energy principle rather than displacement compatibility, this methodology can also help the designer to define an assembly process. Current and future efforts are devoted to applying the HCMS method to design and analyze docking and berthing procedures in orbital construction.

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

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.

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.

Self-Assembly of Fluorescent Hybrid Core-Shell Nanoparticles and Their Application.

Science.gov (United States)

Wang, Chun; Tang, Fu; Wang, Xiaoyu; Li, Lidong

2015-06-24

In this work, a fluorescent hybrid core-shell nanoparticle was prepared by coating a functional polymer shell onto silver nanoparticles via a facile one-pot method. The biomolecule poly-L-lysine (PLL) was chosen as the polymer shell and assembled onto the silver core via the amine-reactive cross-linker, 3,3'-dithiobis(sulfosuccinimidylpropionate). The fluorescent anticancer drug, doxorubicin, was incorporated into the PLL shell through the same linkage. As the cross-linker possesses a thiol-cleavable disulfide bond, disassembly of the PLL shell was observed in the presence of glutathione, leading to controllable doxorubicin release. The silver core there provided an easily modified surface to facilitate the shell coating and ensures the efficient separation of as-prepared nanoparticles from their reaction mixture through centrifugation. Cell assays show that the prepared hybrid fluorescent nanoparticles can internalize into cells possessing excellent biocompatibility prior to the release of doxorubicin, terminating cancer cells efficiently as the doxorubicin is released at the intracellular glutathione level. Such properties are important for designing smart containers for target drug delivery and cellular imaging.

Analysis of insecticide resistance-related genes of the Carmine spider mite Tetranychus cinnabarinus based on a de novo assembled transcriptome.

Science.gov (United States)

Xu, Zhifeng; Zhu, Wenyi; Liu, Yanchao; Liu, Xing; Chen, Qiushuang; Peng, Miao; Wang, Xiangzun; Shen, Guangmao; He, Lin

2014-01-01

The carmine spider mite (CSM), Tetranychus cinnabarinus, is an important pest mite in agriculture, because it can develop insecticide resistance easily. To gain valuable gene information and molecular basis for the future insecticide resistance study of CSM, the first transcriptome analysis of CSM was conducted. A total of 45,016 contigs and 25,519 unigenes were generated from the de novo transcriptome assembly, and 15,167 unigenes were annotated via BLAST querying against current databases, including nr, SwissProt, the Clusters of Orthologous Groups (COGs), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO). Aligning the transcript to Tetranychus urticae genome, the 19255 (75.45%) of the transcripts had significant (e-value insecticide resistance in arthropod were generated from CSM transcriptome, including 53 P450-, 22 GSTs-, 23 CarEs-, 1 AChE-, 7 GluCls-, 9 nAChRs-, 8 GABA receptor-, 1 sodium channel-, 6 ATPase- and 12 Cyt b genes. We developed significant molecular resources for T. cinnabarinus putatively involved in insecticide resistance. The transcriptome assembly analysis will significantly facilitate our study on the mechanism of adapting environmental stress (including insecticide) in CSM at the molecular level, and will be very important for developing new control strategies against this pest mite.

De Novo Assembly and Phasing of Dikaryotic Genomes from Two Isolates of Puccinia coronata f. sp. avenae, the Causal Agent of Oat Crown Rust.

Science.gov (United States)

Miller, Marisa E; Zhang, Ying; Omidvar, Vahid; Sperschneider, Jana; Schwessinger, Benjamin; Raley, Castle; Palmer, Jonathan M; Garnica, Diana; Upadhyaya, Narayana; Rathjen, John; Taylor, Jennifer M; Park, Robert F; Dodds, Peter N; Hirsch, Cory D; Kianian, Shahryar F; Figueroa, Melania

2018-02-20

Oat crown rust, caused by the fungus Pucinnia coronata f. sp. avenae , is a devastating disease that impacts worldwide oat production. For much of its life cycle, P. coronata f. sp. avenae is dikaryotic, with two separate haploid nuclei that may vary in virulence genotype, highlighting the importance of understanding haplotype diversity in this species. We generated highly contiguous de novo genome assemblies of two P. coronata f. sp. avenae isolates, 12SD80 and 12NC29, from long-read sequences. In total, we assembled 603 primary contigs for 12SD80, for a total assembly length of 99.16 Mbp, and 777 primary contigs for 12NC29, for a total length of 105.25 Mbp; approximately 52% of each genome was assembled into alternate haplotypes. This revealed structural variation between haplotypes in each isolate equivalent to more than 2% of the genome size, in addition to about 260,000 and 380,000 heterozygous single-nucleotide polymorphisms in 12SD80 and 12NC29, respectively. Transcript-based annotation identified 26,796 and 28,801 coding sequences for isolates 12SD80 and 12NC29, respectively, including about 7,000 allele pairs in haplotype-phased regions. Furthermore, expression profiling revealed clusters of coexpressed secreted effector candidates, and the majority of orthologous effectors between isolates showed conservation of expression patterns. However, a small subset of orthologs showed divergence in expression, which may contribute to differences in virulence between 12SD80 and 12NC29. This study provides the first haplotype-phased reference genome for a dikaryotic rust fungus as a foundation for future studies into virulence mechanisms in P. coronata f. sp. avenae IMPORTANCE Disease management strategies for oat crown rust are challenged by the rapid evolution of Puccinia coronata f. sp. avenae , which renders resistance genes in oat varieties ineffective. Despite the economic importance of understanding P. coronata f. sp. avenae , resources to study the

Improvement of genome assembly completeness and identification of novel full-length protein-coding genes by RNA-seq in the giant panda genome.

Science.gov (United States)

Chen, Meili; Hu, Yibo; Liu, Jingxing; Wu, Qi; Zhang, Chenglin; Yu, Jun; Xiao, Jingfa; Wei, Fuwen; Wu, Jiayan

2015-12-11

High-quality and complete gene models are the basis of whole genome analyses. The giant panda (Ailuropoda melanoleuca) genome was the first genome sequenced on the basis of solely short reads, but the genome annotation had lacked the support of transcriptomic evidence. In this study, we applied RNA-seq to globally improve the genome assembly completeness and to detect novel expressed transcripts in 12 tissues from giant pandas, by using a transcriptome reconstruction strategy that combined reference-based and de novo methods. Several aspects of genome assembly completeness in the transcribed regions were effectively improved by the de novo assembled transcripts, including genome scaffolding, the detection of small-size assembly errors, the extension of scaffold/contig boundaries, and gap closure. Through expression and homology validation, we detected three groups of novel full-length protein-coding genes. A total of 12.62% of the novel protein-coding genes were validated by proteomic data. GO annotation analysis showed that some of the novel protein-coding genes were involved in pigmentation, anatomical structure formation and reproduction, which might be related to the development and evolution of the black-white pelage, pseudo-thumb and delayed embryonic implantation of giant pandas. The updated genome annotation will help further giant panda studies from both structural and functional perspectives.

Description of a reference design tokamak for the Technology Test Assembly

International Nuclear Information System (INIS)

Haubenreich, P.N.

1975-10-01

Early conceptual studies for the Technology Test Assembly involved a reference conceptual design for a tokamak with superconducting toroidal field magnets. The TF magnet conductors are NbTi filaments in a copper matrix. The 24 TF coils and associated structure operate at 4--5 K and a maximum field (at the windings) of 75 kG. Principal dimensions of the machine are: TF coil bore, 1.8 x 2.4 m (oval); major radius, 2.25 m; plasma minor radius, 0.6 m. A preliminary but detailed cost estimate for the reference machine was prepared to serve as an anchor point for cost scaling for larger machines in subsequent TTA parameter studies

De novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome to identify putative genes involved in the aquatic adaptation and immune response.

Directory of Open Access Journals (Sweden)

Duan Gui

Full Text Available BACKGROUND: The Indo-Pacific humpback dolphin (Sousa chinensis, a marine mammal species inhabited in the waters of Southeast Asia, South Africa and Australia, has attracted much attention because of the dramatic decline in population size in the past decades, which raises the concern of extinction. So far, this species is poorly characterized at molecular level due to little sequence information available in public databases. Recent advances in large-scale RNA sequencing provide an efficient approach to generate abundant sequences for functional genomic analyses in the species with un-sequenced genomes. PRINCIPAL FINDINGS: We performed a de novo assembly of the Indo-Pacific humpback dolphin leucocyte transcriptome by Illumina sequencing. 108,751 high quality sequences from 47,840,388 paired-end reads were generated, and 48,868 and 46,587 unigenes were functionally annotated by BLAST search against the NCBI non-redundant and Swiss-Prot protein databases (E-value<10(-5, respectively. In total, 16,467 unigenes were clustered into 25 functional categories by searching against the COG database, and BLAST2GO search assigned 37,976 unigenes to 61 GO terms. In addition, 36,345 unigenes were grouped into 258 KEGG pathways. We also identified 9,906 simple sequence repeats and 3,681 putative single nucleotide polymorphisms as potential molecular markers in our assembled sequences. A large number of unigenes were predicted to be involved in immune response, and many genes were predicted to be relevant to adaptive evolution and cetacean-specific traits. CONCLUSION: This study represented the first transcriptome analysis of the Indo-Pacific humpback dolphin, an endangered species. The de novo transcriptome analysis of the unique transcripts will provide valuable sequence information for discovery of new genes, characterization of gene expression, investigation of various pathways and adaptive evolution, as well as identification of genetic markers.

Self-Assembled CNT-Polymer Hybrids in Single-Walled Carbon Nanotubes Dispersed Aqueous Triblock Copolymer Solutions

Science.gov (United States)

Vijayaraghavan, D.; Manjunatha, A. S.; Poojitha, C. G.

2018-04-01

We have carried out scanning electron microscopy (SEM), differential scanning calorimetry (DSC), small angle X-ray scattering (SAXS), electrical conductivity, and 1H NMR studies as a function of temperature on single-walled carbon nanotubes (SWCNTs) dispersed aqueous triblock copolymer (P123) solutions. The single-walled carbon nanotubes in this system aggregate to form bundles, and the bundles aggregate to form net-like structures. Depending on the temperature and phases of the polymer, this system exhibits three different self-assembled CNT-polymer hybrids. We find CNT-unimer hybrid at low temperatures, CNT-micelle hybrid at intermediate temperatures wherein the polymer micelles are adsorbed in the pores of the CNT nets, and another type of CNT-micelle hybrid at high temperatures wherein the polymer micelles are adsorbed on the surface of the CNT bundles. Our DSC thermogram showed two peaks related to these structural changes in the CNT-polymer hybrids. Temperature dependence of the 1H NMR chemical shifts of the molecular groups of the polymer and the AC electrical conductivity of the composite also showed discontinuous changes at the temperatures at which the CNT-polymer hybrid's structural changes are seen. Interestingly, for a higher CNT concentration (0.5 wt.%) in the system, the aggregated polymer micelles adsorbed on the CNTs exhibit cone-like and cube-like morphologies at the intermediate and at high temperatures respectively.

Molecular analysis of "de novo" purine biosynthesis in solanaceous species and in Arabidopsis thaliana

DEFF Research Database (Denmark)

van der Graaff, Eric; Hooykaas, Paul; Lein, Wolfgang

2004-01-01

Purine nucleotides are essential components to sustain plant growth and development. In plants they are either synthesized "de novo" during the process of purine biosynthesis or are recycled from purine bases and purine nucleosides throughout the salvage pathway. Comparison between animals...... biosynthesis pathway in plants, and the in planta functional analysis of PRPP (5-phosphoribosyl-1-pyrophoshate) amidotransferase (ATase), catalyzing the first committed step of the "de novo" purine biosynthesis. The cloning of the genes involved in the purine biosynthesis pathway was attained by a screening...... strategy with heterologous cDNA probes and by using S. cerevisiae mutants for complementation. Southern hybridization showed a complex genomic organization for these genes in solanaceous species and their organ- and developmental specific expression was analyzed by Northern hybridization. The specific role...

Customization and design of directed self-assembly using hybrid prepatterns

Science.gov (United States)

Cheng, Joy; Doerk, Gregory S.; Rettner, Charles T.; Singh, Gurpreet; Tjio, Melia; Truong, Hoa; Arellano, Noel; Balakrishnan, Srinivasan; Brink, Markus; Tsai, Hsinyu; Liu, Chi-Chun; Guillorn, Michael; Sanders, Daniel P.

2015-03-01

Diminishing error tolerance renders the customization of patterns created through directed self-assembly (DSA) extremely challenging at tighter pitch. A self-aligned customization scheme can be achieved using a hybrid prepattern comprising both organic and inorganic regions that serves as a guiding prepattern to direct the self-assembly of the block copolymers as well as a cut mask pattern for the DSA arrays aligned to it. In this paper, chemoepitaxy-based self-aligned customization is demonstrated using two types of organic-inorganic prepatterns. CHEETAH prepattern for "CHemoepitaxy Etch Trim using a self-Aligned Hardmask" of preferential hydrogen silsesquioxane (HSQ, inorganic resist), non-preferential organic underlayer is fabricated using electron beam lithography. Customized trench or hole arrays can be achieved through co-transfer of DSA-formed arrays and CHEETAH prepattern. Herein, we also introduce a tone-reversed version called reverse-CHEETAH (or rCHEETAH) in which customized line segments can be achieved through co-transfer of DSA-formed arrays formed on a prepattern wherein the inorganic HSQ regions are nonpreferential and the organic regions are PMMA preferential. Examples of two-dimensional self-aligned customization including 25nm pitch fin structures and an 8-bar "IBM" illustrate the versatility of this customization scheme using rCHEETAH.

Hybrid materials for optics and photonics.

Science.gov (United States)

Lebeau, Benedicte; Innocenzi, Plinio

2011-02-01

The interest in organic-inorganic hybrids as materials for optics and photonics started more than 25 years ago and since then has known a continuous and strong growth. The high versatility of sol-gel processing offers a wide range of possibilities to design tailor-made materials in terms of structure, texture, functionality, properties and shape modelling. From the first hybrid material with optical functional properties that has been obtained by incorporation of an organic dye in a silica matrix, the research in the field has quickly evolved towards more sophisticated systems, such as multifunctional and/or multicomponent materials, nanoscale and self-assembled hybrids and devices for integrated optics. In the present critical review, we have focused our attention on three main research areas: passive and active optical hybrid sol-gel materials, and integrated optics. This is far from exhaustive but enough to give an overview of the huge potential of these materials in photonics and optics (254 references).

Induced assembly and photoluminescence of lanthanum (Tb, Eu, Dy) complexes/ZnO/polyethylene glycol hybrid phosphors

Energy Technology Data Exchange (ETDEWEB)

Yan Bing [Department of Chemistry, Tongji University, Siping Road 1239, Shanghai 200092 (China)]. E-mail: byan@tongji.edu.cn; Chen Xi [Department of Chemistry, Tongji University, Siping Road 1239, Shanghai 200092 (China); Wu Jianhua [Department of Chemistry, Tongji University, Siping Road 1239, Shanghai 200092 (China)

2007-08-31

Some novel kinds of hybrid phosphors were assembled with lanthanum (Tb, Eu, Dy) complexes (with four kinds of terbium complexes is 2,4-dihydroxybenzonic acid (DHBA), 1,10-phenanthroline (phen), acetylacetone (AA) and nicotinic acid (Nic), respectively) doped ZnO/PEG particles by co-precipitation approach derived from Zn(CH{sub 3}COO){sub 2} (Zn(AC){sub 2}), NaOH, PEG as precursors at room temperature. The characteristic luminescence spectra for f-f transitions of Tb{sup 3+}, Eu{sup 3+}, Dy{sup 3+} were observed. It is worthy to point out that ZnO is the excellent host for lanthanum ions by the assembly of PEG matrices.

De Novo Genome and Transcriptome Assembly of the Canadian Beaver (Castor canadensis

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Si Lok

2017-02-01

Full Text Available The Canadian beaver (Castor canadensis is the largest indigenous rodent in North America. We report a draft annotated assembly of the beaver genome, the first for a large rodent and the first mammalian genome assembled directly from uncorrected and moderate coverage (< 30 × long reads generated by single-molecule sequencing. The genome size is 2.7 Gb estimated by k-mer analysis. We assembled the beaver genome using the new Canu assembler optimized for noisy reads. The resulting assembly was refined using Pilon supported by short reads (80 × and checked for accuracy by congruency against an independent short read assembly. We scaffolded the assembly using the exon–gene models derived from 9805 full-length open reading frames (FL-ORFs constructed from the beaver leukocyte and muscle transcriptomes. The final assembly comprised 22,515 contigs with an N50 of 278,680 bp and an N50-scaffold of 317,558 bp. Maximum contig and scaffold lengths were 3.3 and 4.2 Mb, respectively, with a combined scaffold length representing 92% of the estimated genome size. The completeness and accuracy of the scaffold assembly was demonstrated by the precise exon placement for 91.1% of the 9805 assembled FL-ORFs and 83.1% of the BUSCO (Benchmarking Universal Single-Copy Orthologs gene set used to assess the quality of genome assemblies. Well-represented were genes involved in dentition and enamel deposition, defining characteristics of rodents with which the beaver is well-endowed. The study provides insights for genome assembly and an important genomics resource for Castoridae and rodent evolutionary biology.

Hybrid finite volume/ finite element method for radiative heat transfer in graded index media

Science.gov (United States)

Zhang, L.; Zhao, J. M.; Liu, L. H.; Wang, S. Y.

2012-09-01

The rays propagate along curved path determined by the Fermat principle in the graded index medium. The radiative transfer equation in graded index medium (GRTE) contains two specific redistribution terms (with partial derivatives to the angular coordinates) accounting for the effect of the curved ray path. In this paper, the hybrid finite volume with finite element method (hybrid FVM/FEM) (P.J. Coelho, J. Quant. Spectrosc. Radiat. Transf., vol. 93, pp. 89-101, 2005) is extended to solve the radiative heat transfer in two-dimensional absorbing-emitting-scattering graded index media, in which the spatial discretization is carried out using a FVM, while the angular discretization is by a FEM. The FEM angular discretization is demonstrated to be preferable in dealing with the redistribution terms in the GRTE. Two stiff matrix assembly schemes of the angular FEM discretization, namely, the traditional assembly approach and a new spherical assembly approach (assembly on the unit sphere of the solid angular space), are discussed. The spherical assembly scheme is demonstrated to give better results than the traditional assembly approach. The predicted heat flux distributions and temperature distributions in radiative equilibrium are determined by the proposed method and compared with the results available in other references. The proposed hybrid FVM/FEM method can predict the radiative heat transfer in absorbing-emitting-scattering graded index medium with good accuracy.

Hybrid finite volume/ finite element method for radiative heat transfer in graded index media

International Nuclear Information System (INIS)

Zhang, L.; Zhao, J.M.; Liu, L.H.; Wang, S.Y.

2012-01-01

The rays propagate along curved path determined by the Fermat principle in the graded index medium. The radiative transfer equation in graded index medium (GRTE) contains two specific redistribution terms (with partial derivatives to the angular coordinates) accounting for the effect of the curved ray path. In this paper, the hybrid finite volume with finite element method (hybrid FVM/FEM) (P.J. Coelho, J. Quant. Spectrosc. Radiat. Transf., vol. 93, pp. 89-101, 2005) is extended to solve the radiative heat transfer in two-dimensional absorbing-emitting-scattering graded index media, in which the spatial discretization is carried out using a FVM, while the angular discretization is by a FEM. The FEM angular discretization is demonstrated to be preferable in dealing with the redistribution terms in the GRTE. Two stiff matrix assembly schemes of the angular FEM discretization, namely, the traditional assembly approach and a new spherical assembly approach (assembly on the unit sphere of the solid angular space), are discussed. The spherical assembly scheme is demonstrated to give better results than the traditional assembly approach. The predicted heat flux distributions and temperature distributions in radiative equilibrium are determined by the proposed method and compared with the results available in other references. The proposed hybrid FVM/FEM method can predict the radiative heat transfer in absorbing-emitting-scattering graded index medium with good accuracy.

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.

Three-dimensional graphene-polyaniline hybrid hollow spheres by layer-by-layer assembly for application in supercapacitor

International Nuclear Information System (INIS)

Luo, Jing; Ma, Qiang; Gu, Huahao; Zheng, Yuan; Liu, Xiaoya

2015-01-01

Highlights: •A graphene-polyaniline (GR-PANI) hybrid hollow sphere is fabricated by layer-by-layer (LBL) assembly technique. •The GR-PANI hollow sphere has higher specific capacitance than stacked GR-PANI LBL film. •64% of its initial capacitance is maintained with the current density increased from 0.5 to 20 A g −1 . •A high capacity retention rate of 83% after 1000 cycles can be achieved. -- Abstract: A novel kind of three-dimensional graphene-polyaniline hybrid hollow sphere (RGO-PANI HS) has been prepared via layer-by-layer (LBL) assembly of negatively-charged reduced graphene oxide (RGO) and positively charged polyaniline (PANI) on polystyrene (PS) microsphere, followed by the removal of the PS template. The hollow structure of the obtained RGO-PANI HS is confirmed by transmission electron microscopy (TEM). When used as the electrode materials for supercapacitor, the specific capacitance of the RGO-PANI HS reaches 381 F/g at a current density of 4.0 A/g, which is much higher than 251 F/g of the stacked RGO-PANI LBL film. The higher specific capacitance of RGO-PANI HS should be attributed to its unique hollow structure which provides a larger accessible surface area and facilitate the charge and ion transport. In addition, its specific capacitance can be facilely tailored by changing the assembly cycle number. Furthermore, good cycling stability is also demonstrated with 83% of the original capacitance value maintained after 1000 charging/discharging cycles

Flip chip assembly of thinned chips for hybrid pixel detector applications

International Nuclear Information System (INIS)

Fritzsch, T; Zoschke, K; Rothermund, M; Oppermann, H; Woehrmann, M; Ehrmann, O; Lang, K D; Huegging, F

2014-01-01

There is a steady trend to ultra-thin microelectronic devices. Especially for future particle detector systems a reduced readout chip thickness is required to limit the loss of tracking precision due to scattering. The reduction of silicon thickness is performed at wafer level in a two-step thinning process. To minimize the risk of wafer breakage the thinned wafer needs to be handled by a carrier during the whole process chain of wafer bumping. Another key process is the flip chip assembly of thinned readout chips onto thin sensor tiles. Besides the prevention of silicon breakage the minimization of chip warpage is one additional task for a high yield and reliable flip chip process. A new technology using glass carrier wafer will be described in detail. The main advantage of this technology is the combination of a carrier support during wafer processing and the chip support during flip chip assembly. For that a glass wafer is glue-bonded onto the backside of the thinned readout chip wafer. After the bump deposition process the glass-readout chip stack is diced in one step. Finally the glass carrier chip is released by laser illumination after flip chip assembly of the readout chip onto sensor tile. The results of the flip chip assembly process development for the ATLAS IBL upgrade are described more in detail. The new ATLAS FEI4B chip with a size of 20 × 19 mm 2 is flip chip bonded with a thickness of only 150 μm, but the capability of this technology has been demonstrated on hybrid modules with a reduced readout chip thickness of down to 50 μm which is a major step for ultra-thin electronic systems

A high resolution radiation hybrid map of bovine chromosome 14 identifies scaffold rearrangement in the latest bovine assembly

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Wang Zhiquan

2007-07-01

Full Text Available Abstract Background Radiation hybrid (RH maps are considered to be a tool of choice for fine mapping closely linked loci, considering that the resolution of linkage maps is determined by the number of informative meiosis and recombination events which may require very large mapping populations. Accurately defining the marker order on chromosomes is crucial for correct identification of quantitative trait loci (QTL, haplotype map construction and refinement of candidate gene searches. Results A 12 k Radiation hybrid map of bovine chromosome 14 was constructed using 843 single nucleotide polymorphism markers. The resulting map was aligned with the latest version of the bovine assembly (Btau_3.1 as well as other previously published RH maps. The resulting map identified distinct regions on Bovine chromosome 14 where discrepancies between this RH map and the bovine assembly occur. A major region of discrepancy was found near the centromere involving the arrangement and order of the scaffolds from the assembly. The map further confirms previously published conserved synteny blocks with human chromosome 8. As well, it identifies an extra breakpoint and conserved synteny block previously undetected due to lower marker density. This conserved synteny block is in a region where markers between the RH map presented here and the latest sequence assembly are in very good agreement. Conclusion The increase of publicly available markers shifts the rate limiting step from marker discovery to the correct identification of their order for further use by the research community. This high resolution map of bovine chromosome 14 will facilitate identification of regions in the sequence assembly where additional information is required to resolve marker ordering.

Self-assembled hierarchical graphene/polyaniline hybrid aerogels for electrochemical capacitive energy storage

International Nuclear Information System (INIS)

Yang, Fan; Xu, Maowen; Bao, Shu-Juan; Wei, Hua; Chai, Hui

2014-01-01

In this work, polyaniline nanowires (PANI-NWs) act as spacers, incorporated with graphene oxide and self-assembled into graphene/PANI hybrid aerogels through a facile hydrothermal route. The as-synthesized samples have been characterized by X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectra, X-ray photoelectron spectroscopy (XPS), contact angle measurement, field emission scanning electron microscopy (FESEM) and transmission electron microscopy (TEM) for their microstructure, morphology and relative affinities toward water. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) measurements have been used to study the effects of composition, microstructure and morphology of the samples on their capacitive performance. The experimental results indicate that the PANI can effectively tailor the microstructures and electrochemical performances of the products. The as-prepared materials with an appropriate proportion of PANI nanowires can efficiently prevent the adjacent graphene sheets from aggregation and provide fast ionic channels for electrochemical energy storage. A specific capacitance of 520.3 F g −1 has been achieved from graphene/PANI hybrid aerogel, which also exhibits excellent cycling stability

Hybrid state‐space time integration in a rotating frame of reference

DEFF Research Database (Denmark)

Krenk, Steen; Nielsen, Martin Bjerre

2011-01-01

displacements and the global velocities are represented by the same shape functions. This leads to a simple generalization of the corresponding equations of motion in a stationary frame in which all inertial effects are represented via the classic global mass matrix. The formulation introduces two gyroscopic......A time integration algorithm is developed for the equations of motion of a flexible body in a rotating frame of reference. The equations are formulated in a hybrid state‐space, formed by the local displacement components and the global velocity components. In the spatial discretization the local...... terms, while the centrifugal forces are represented implicitly via the hybrid state‐space format. An angular momentum and energy conserving algorithm is developed, in which the angular velocity of the frame is represented by its mean value. A consistent algorithmic damping scheme is identified...

Developmental gene discovery in a hemimetabolous insect: de novo assembly and annotation of a transcriptome for the cricket Gryllus bimaculatus.

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Victor Zeng

Full Text Available Most genomic resources available for insects represent the Holometabola, which are insects that undergo complete metamorphosis like beetles and flies. In contrast, the Hemimetabola (direct developing insects, representing the basal branches of the insect tree, have very few genomic resources. We have therefore created a large and publicly available transcriptome for the hemimetabolous insect Gryllus bimaculatus (cricket, a well-developed laboratory model organism whose potential for functional genetic experiments is currently limited by the absence of genomic resources. cDNA was prepared using mRNA obtained from adult ovaries containing all stages of oogenesis, and from embryo samples on each day of embryogenesis. Using 454 Titanium pyrosequencing, we sequenced over four million raw reads, and assembled them into 21,512 isotigs (predicted transcripts and 120,805 singletons with an average coverage per base pair of 51.3. We annotated the transcriptome manually for over 400 conserved genes involved in embryonic patterning, gametogenesis, and signaling pathways. BLAST comparison of the transcriptome against the NCBI non-redundant protein database (nr identified significant similarity to nr sequences for 55.5% of transcriptome sequences, and suggested that the transcriptome may contain 19,874 unique transcripts. For predicted transcripts without significant similarity to known sequences, we assessed their similarity to other orthopteran sequences, and determined that these transcripts contain recognizable protein domains, largely of unknown function. We created a searchable, web-based database to allow public access to all raw, assembled and annotated data. This database is to our knowledge the largest de novo assembled and annotated transcriptome resource available for any hemimetabolous insect. We therefore anticipate that these data will contribute significantly to more effective and higher-throughput deployment of molecular analysis tools in

Integrative self-assembly of functional hybrid nanoconstructs by inorganic wrapping of single biomolecules, biomolecule arrays and organic supramolecular assemblies.

Science.gov (United States)

Patil, Avinash J; Li, Mei; Mann, Stephen

2013-08-21

Synthesis of functional hybrid nanoscale objects has been a core focus of the rapidly progressing field of nanomaterials science. In particular, there has been significant interest in the integration of evolutionally optimized biological systems such as proteins, DNA, virus particles and cells with functional inorganic building blocks to construct mesoscopic architectures and nanostructured materials. However, in many cases the fragile nature of the biomolecules seriously constrains their potential applications. As a consequence, there is an on-going quest for the development of novel strategies to modulate the thermal and chemical stabilities, and performance of biomolecules under adverse conditions. This feature article highlights new methods of "inorganic molecular wrapping" of single or multiple protein molecules, individual double-stranded DNA helices, lipid bilayer vesicles and self-assembled organic dye superstructures using inorganic building blocks to produce bio-inorganic nanoconstructs with core-shell type structures. We show that spatial isolation of the functional biological nanostructures as "armour-plated" enzyme molecules or polynucleotide strands not only maintains their intact structure and biochemical properties, but also enables the fabrication of novel hybrid nanomaterials for potential applications in diverse areas of bionanotechnology.

Integrative self-assembly of functional hybrid nanoconstructs by inorganic wrapping of single biomolecules, biomolecule arrays and organic supramolecular assemblies

Science.gov (United States)

Patil, Avinash J.; Li, Mei; Mann, Stephen

2013-07-01

Synthesis of functional hybrid nanoscale objects has been a core focus of the rapidly progressing field of nanomaterials science. In particular, there has been significant interest in the integration of evolutionally optimized biological systems such as proteins, DNA, virus particles and cells with functional inorganic building blocks to construct mesoscopic architectures and nanostructured materials. However, in many cases the fragile nature of the biomolecules seriously constrains their potential applications. As a consequence, there is an on-going quest for the development of novel strategies to modulate the thermal and chemical stabilities, and performance of biomolecules under adverse conditions. This feature article highlights new methods of ``inorganic molecular wrapping'' of single or multiple protein molecules, individual double-stranded DNA helices, lipid bilayer vesicles and self-assembled organic dye superstructures using inorganic building blocks to produce bio-inorganic nanoconstructs with core-shell type structures. We show that spatial isolation of the functional biological nanostructures as ``armour-plated'' enzyme molecules or polynucleotide strands not only maintains their intact structure and biochemical properties, but also enables the fabrication of novel hybrid nanomaterials for potential applications in diverse areas of bionanotechnology.

Intelligent controller of a flexible hybrid robot machine for ITER assembly and maintenance

International Nuclear Information System (INIS)

Al-saedi, Mazin I.; Wu, Huapeng; Handroos, Heikki

2014-01-01

Highlights: • Studying flexible multibody dynamic of hybrid parallel robot. • Investigating fuzzy-PD controller to control a hybrid flexible hydraulically driven robot. • Investigating ANFIS-PD controller to control a hybrid flexible robot. Compare to traditional PID this method gives better performance. • Using the equilibrium of reaction forces between the parallel and serial parts of hybrid robot to control the serial part hydraulically driven. - Abstract: The assembly and maintenance of International Thermonuclear Experimental Reactor (ITER) vacuum vessel (VV) is highly challenging since the tasks performed by the robot involve welding, material handling, and machine cutting from inside the VV. To fulfill the tasks in ITER application, this paper presents a hybrid redundant manipulator with four DOFs provided by serial kinematic axes and six DOFs by parallel mechanism. Thus, in machining, to achieve greater end-effector trajectory tracking accuracy for surface quality, a robust control of the actuators for the flexible link has to be deduced. In this paper, the intelligent control of a hydraulically driven parallel robot part based on the dynamic model and two control schemes have been investigated: (1) fuzzy-PID self tuning controller composed of the conventional PID control and with fuzzy logic; (2) adaptive neuro-fuzzy inference system-PID (ANFIS-PID) self tuning of the gains of the PID controller, which are implemented independently to control each hydraulic cylinder of the parallel robot based on rod position predictions. The obtained results of the fuzzy-PID and ANFIS-PID self tuning controller can reduce more tracking errors than the conventional PID controller. Subsequently, the serial component of the hybrid robot can be analyzed using the equilibrium of reaction forces at the universal joint connections of the hexa-element. To achieve precise positional control of the end effector for maximum precision machining, the hydraulic cylinder should

Intelligent controller of a flexible hybrid robot machine for ITER assembly and maintenance

Energy Technology Data Exchange (ETDEWEB)

Al-saedi, Mazin I., E-mail: mazin.al-saedi@lut.fi; Wu, Huapeng; Handroos, Heikki

2014-10-15

Highlights: • Studying flexible multibody dynamic of hybrid parallel robot. • Investigating fuzzy-PD controller to control a hybrid flexible hydraulically driven robot. • Investigating ANFIS-PD controller to control a hybrid flexible robot. Compare to traditional PID this method gives better performance. • Using the equilibrium of reaction forces between the parallel and serial parts of hybrid robot to control the serial part hydraulically driven. - Abstract: The assembly and maintenance of International Thermonuclear Experimental Reactor (ITER) vacuum vessel (VV) is highly challenging since the tasks performed by the robot involve welding, material handling, and machine cutting from inside the VV. To fulfill the tasks in ITER application, this paper presents a hybrid redundant manipulator with four DOFs provided by serial kinematic axes and six DOFs by parallel mechanism. Thus, in machining, to achieve greater end-effector trajectory tracking accuracy for surface quality, a robust control of the actuators for the flexible link has to be deduced. In this paper, the intelligent control of a hydraulically driven parallel robot part based on the dynamic model and two control schemes have been investigated: (1) fuzzy-PID self tuning controller composed of the conventional PID control and with fuzzy logic; (2) adaptive neuro-fuzzy inference system-PID (ANFIS-PID) self tuning of the gains of the PID controller, which are implemented independently to control each hydraulic cylinder of the parallel robot based on rod position predictions. The obtained results of the fuzzy-PID and ANFIS-PID self tuning controller can reduce more tracking errors than the conventional PID controller. Subsequently, the serial component of the hybrid robot can be analyzed using the equilibrium of reaction forces at the universal joint connections of the hexa-element. To achieve precise positional control of the end effector for maximum precision machining, the hydraulic cylinder should

De novo transcriptome sequencing of the Octopus vulgaris hemocytes using Illumina RNA-Seq technology: response to the infection by the gastrointestinal parasite Aggregata octopiana.

Science.gov (United States)

Castellanos-Martínez, Sheila; Arteta, David; Catarino, Susana; Gestal, Camino

2014-01-01

Octopus vulgaris is a highly valuable species of great commercial interest and excellent candidate for aquaculture diversification; however, the octopus' well-being is impaired by pathogens, of which the gastrointestinal coccidian parasite Aggregata octopiana is one of the most important. The knowledge of the molecular mechanisms of the immune response in cephalopods, especially in octopus is scarce. The transcriptome of the hemocytes of O. vulgaris was de novo sequenced using the high-throughput paired-end Illumina technology to identify genes involved in immune defense and to understand the molecular basis of octopus tolerance/resistance to coccidiosis. A bi-directional mRNA library was constructed from hemocytes of two groups of octopus according to the infection by A. octopiana, sick octopus, suffering coccidiosis, and healthy octopus, and reads were de novo assembled together. The differential expression of transcripts was analysed using the general assembly as a reference for mapping the reads from each condition. After sequencing, a total of 75,571,280 high quality reads were obtained from the sick octopus group and 74,731,646 from the healthy group. The general transcriptome of the O. vulgaris hemocytes was assembled in 254,506 contigs. A total of 48,225 contigs were successfully identified, and 538 transcripts exhibited differential expression between groups of infection. The general transcriptome revealed genes involved in pathways like NF-kB, TLR and Complement. Differential expression of TLR-2, PGRP, C1q and PRDX genes due to infection was validated using RT-qPCR. In sick octopuses, only TLR-2 was up-regulated in hemocytes, but all of them were up-regulated in caecum and gills. The transcriptome reported here de novo establishes the first molecular clues to understand how the octopus immune system works and interacts with a highly pathogenic coccidian. The data provided here will contribute to identification of biomarkers for octopus resistance against

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.

De novo assembly and characterization of the transcriptome of the parasitic weed dodder identifies genes associated with plant parasitism.

Science.gov (United States)

Ranjan, Aashish; Ichihashi, Yasunori; Farhi, Moran; Zumstein, Kristina; Townsley, Brad; David-Schwartz, Rakefet; Sinha, Neelima R

2014-11-01

Parasitic flowering plants are one of the most destructive agricultural pests and have major impact on crop yields throughout the world. Being dependent on finding a host plant for growth, parasitic plants penetrate their host using specialized organs called haustoria. Haustoria establish vascular connections with the host, which enable the parasite to steal nutrients and water. The underlying molecular and developmental basis of parasitism by plants is largely unknown. In order to investigate the process of parasitism, RNAs from different stages (i.e. seed, seedling, vegetative strand, prehaustoria, haustoria, and flower) were used to de novo assemble and annotate the transcriptome of the obligate plant stem parasite dodder (Cuscuta pentagona). The assembled transcriptome was used to dissect transcriptional dynamics during dodder development and parasitism and identified key gene categories involved in the process of plant parasitism. Host plant infection is accompanied by increased expression of parasite genes underlying transport and transporter categories, response to stress and stimuli, as well as genes encoding enzymes involved in cell wall modifications. By contrast, expression of photosynthetic genes is decreased in the dodder infective stages compared with normal stem. In addition, genes relating to biosynthesis, transport, and response of phytohormones, such as auxin, gibberellins, and strigolactone, were differentially expressed in the dodder infective stages compared with stems and seedlings. This analysis sheds light on the transcriptional changes that accompany plant parasitism and will aid in identifying potential gene targets for use in controlling the infestation of crops by parasitic weeds. © 2014 American Society of Plant Biologists. All Rights Reserved.

Novos paradigmas literários

Directory of Open Access Journals (Sweden)

Denise Azevedo Duarte Guimarães

2005-12-01

Full Text Available O artigo estuda a emergência de novos paradigmas literários, procurando refletir acerca das textualidades contemporâneas. Focaliza os hipertextos informatizados e a poesia multimídia, com o intuito de desvendar como estão sendo criados novos procedimentos expressivos e em que medida eles podem ser identificados com reflexões teóricas anteriores acerca do texto literário impresso. Remete a questões ligadas à leitura dos diferentes tipos de signos e aos modos como eles se integram para a constituição dessas novíssimas linguagens híbridas em novos suportes.El artículo estudia la emergencia de nuevos paradigmas literarios, procurando reflejar acerca de las textualidades contemporáneas. Enfoca los hipertextos informatizados y la poesía multimedia, intentando desvendar cómo están siendo creados nuevos procedimientos expresivos y en qué medida ellos pueden ser identificados a reflexiones teóricas anteriores acerca del texto literario impreso. Remite a cuestiones ligadas a la lectura de los diferentes tipos de signos y a los modos cómo ellos se interaccionan para la constitución de los novísimos lenguajes híbridos en nuevos supuestos.This article investigates the emergence of new literary paradigms as it tries to understand new contemporary textualities. It analyses some hypertexts and multimedia poetry trying to trace how new expressive procedures are being created. How can these new languages be identified and what are their relations to previous theories which dealt with the literary printed text? This study approaches questions linked to the reading of different types of signs and the modes they function towards the fabrication of these new hybrid languages.

Photonic hybrid assembly through flexible waveguides

NARCIS (Netherlands)

Wörhoff, Kerstin; Prak, Albert; postma, F; Leinse, A; Wu, K.; Peters, T.J.; Tichem, M.; Amaning-Appiah, B.; Renukappa, V.; Vollrath, G.; Balcells-Ventura, J.; Uhlig, P.; Seyfried, M.; Rose, D.; Santos, Raquel; Leijtens, XJM; Flintham, B.; Wale, M.; Robbins, D.; Vivien, Laurent; Pavesi, Lorenzo; Pelli, Stefano

2016-01-01

Fully automated, high precision, cost-effective assembly technology for photonic packages remains one of the main challenges in photonic component manufacturing. Next to the cost aspect the most demanding assembly task for multiport photonic integrated circuits (PICs) is the high-precision (±0.1

Role of de novo biosynthesis in ecosystem scale monoterpene emissions from a boreal Scots pine forest

Directory of Open Access Journals (Sweden)

R. Taipale

2011-08-01

Full Text Available Monoterpene emissions from Scots pine have traditionally been assumed to originate as evaporation from specialized storage pools. More recently, the significance of de novo emissions, originating directly from monoterpene biosynthesis, has been recognized. To study the role of biosynthesis at the ecosystem scale, we measured monoterpene emissions from a Scots pine dominated forest in southern Finland using the disjunct eddy covariance method combined with proton transfer reaction mass spectrometry. The interpretation of the measurements was based on a correlation analysis and a hybrid emission algorithm describing both de novo and pool emissions. During the measurement period May–August 2007, the monthly medians of daytime emissions were 200, 290, 180, and 200 μg m⁻² h⁻¹. The emissions were partly light dependent, probably due to de novo biosynthesis. The emission potential for both de novo and pool emissions exhibited a decreasing summertime trend. The ratio of the de novo emission potential to the total emission potential varied between 30 % and 46 %. Although the monthly changes were not significant, the ratio always differed statistically from zero, suggesting that the role of de novo biosynthesis was observable. Given the uncertainties in this study, we conclude that more accurate estimates of the contribution of de novo emissions are required for improving monoterpene emission algorithms for Scots pine dominated forests.

De novo transcriptome assembly and analysis of differential gene expression in response to drought in European beech.

Directory of Open Access Journals (Sweden)

Markus Müller

Full Text Available Despite the ecological and economic importance of European beech (Fagus sylvatica L. genomic resources of this species are still limited. This hampers an understanding of the molecular basis of adaptation to stress. Since beech will most likely be threatened by the consequences of climate change, an understanding of adaptive processes to climate change-related drought stress is of major importance. Here, we used RNA-seq to provide the first drought stress-related transcriptome of beech. In a drought stress trial with beech saplings, 50 samples were taken for RNA extraction at five points in time during a soil desiccation experiment. De novo transcriptome assembly and analysis of differential gene expression revealed 44,335 contigs, and 662 differentially expressed genes between the stress and normally watered control group. Gene expression was specific to the different time points, and only five genes were significantly differentially expressed between the stress and control group on all five sampling days. GO term enrichment showed that mostly genes involved in lipid- and homeostasis-related processes were upregulated, whereas genes involved in oxidative stress response were downregulated in the stressed seedlings. This study gives first insights into the genomic drought stress response of European beech, and provides new genetic resources for adaptation research in this species.

Capacitively coupled hybrid pixel assemblies for the CLIC vertex detector

CERN Document Server

AUTHOR|(SzGeCERN)734627; Benoit, Mathieu; Dannheim, Dominik; Dette, Karola; Hynds, Daniel; Kulis, Szymon; Peric, Ivan; Petric, Marko; Redford, Sophie; Sicking, Eva; Valerio, Pierpaolo

2016-01-01

The vertex detector at the proposed CLIC multi-TeV linear e+e- collider must have minimal material content and high spatial resolution, combined with accurate time-stamping to cope with the expected high rate of beam-induced backgrounds. One of the options being considered is the use of active sensors implemented in a commercial high-voltage CMOS process, capacitively coupled to hybrid pixel ASICs. A prototype of such an assembly, using two custom designed chips (CCPDv3 as active sensor glued to a CLICpix readout chip), has been characterised both in the lab and in beam tests at the CERN SPS using 120 GeV/c positively charged hadrons. Results of these characterisation studies are presented both for single and dual amplification stages in the active sensor. Pixel cross-coupling results are also presented, showing the sensitivity to placement precision and planarity of the glue layer.

Photonic hybrid assembly through flexible waveguides

NARCIS (Netherlands)

Wörhoff, K.; Prak, A.; Postma, F.; Leinse, A.; Wu, K.; Peters, T. J.; Tichem, M.; Amaning-Appiah, B.; Renukappa, V.; Vollrath, G.; Balcells-Ventura, J.; Uhlig, P.; Seyfried, M.; Rose, D.; Santos, R.; Leijtens, X.J.M.; Flintham, B.; Wale, M.; Robbins, D.

2016-01-01

Fully automated, high precision, cost-effective assembly technology for photonic packages remains one of the main challenges in photonic component manufacturing. Next to the cost aspect the most demanding assembly task for multiport photonic integrated circuits (PICs) is the high-precision (±0.1 μm)

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...

Assembling high activity phosphotriesterase composites using hybrid nanoparticle peptide-DNA scaffolded architectures

Science.gov (United States)

Breger, Joyce C.; Buckhout-White, Susan; Walper, Scott A.; Oh, Eunkeu; Susumu, Kimihiro; Ancona, Mario G.; Medintz, Igor L.

2017-06-01

Nanoparticle (NP) display potentially offers a new way to both stabilize and, in many cases, enhance enzyme activity over that seen for native protein in solution. However, the large, globular and sometimes multimeric nature of many enzymes limits their ability to attach directly to the surface of NPs, especially when the latter are colloidally stabilized with bulky PEGylated ligands. Engineering extended protein linkers into the enzymes to achieve direct attachment through the PEG surface often detrimentally alters the enzymes catalytic ability. Here, we demonstrate an alternate, hybrid biomaterials-based approach to achieving directed enzyme assembly on PEGylated NPs. We self-assemble a unique architecture consisting of a central semiconductor quantum dot (QD) scaffold displaying controlled ratios of extended peptide-DNA linkers which penetrate through the PEG surface to directly couple enzymes to the QD surface. As a test case, we utilize phosphotriesterase (PTE), an enzyme of bio-defense interest due to its ability to hydrolyze organophosphate nerve agents. Moreover, this unique approach still allows PTE to maintain enhanced activity while also suggesting the ability of DNA to enhance enzyme activity in and of itself.

Genetically encoded lipid-polypeptide hybrid biomaterials that exhibit temperature-triggered hierarchical self-assembly

Science.gov (United States)

Mozhdehi, Davoud; Luginbuhl, Kelli M.; Simon, Joseph R.; Dzuricky, Michael; Berger, Rüdiger; Varol, H. Samet; Huang, Fred C.; Buehne, Kristen L.; Mayne, Nicholas R.; Weitzhandler, Isaac; Bonn, Mischa; Parekh, Sapun H.; Chilkoti, Ashutosh

2018-05-01

Post-translational modification of proteins is a strategy widely used in biological systems. It expands the diversity of the proteome and allows for tailoring of both the function and localization of proteins within cells as well as the material properties of structural proteins and matrices. Despite their ubiquity in biology, with a few exceptions, the potential of post-translational modifications in biomaterials synthesis has remained largely untapped. As a proof of concept to demonstrate the feasibility of creating a genetically encoded biohybrid material through post-translational modification, we report here the generation of a family of three stimulus-responsive hybrid materials—fatty-acid-modified elastin-like polypeptides—using a one-pot recombinant expression and post-translational lipidation methodology. These hybrid biomaterials contain an amphiphilic domain, composed of a β-sheet-forming peptide that is post-translationally functionalized with a C14 alkyl chain, fused to a thermally responsive elastin-like polypeptide. They exhibit temperature-triggered hierarchical self-assembly across multiple length scales with varied structure and material properties that can be controlled at the sequence level.

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

De novo assembly of a transcriptome for Calanus finmarchicus (Crustacea, Copepoda--the dominant zooplankter of the North Atlantic Ocean.

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Petra H Lenz

Full Text Available Assessing the impact of global warming on the food web of the North Atlantic will require difficult-to-obtain physiological data on a key copepod crustacean, Calanus finmarchicus. The de novo transcriptome presented here represents a new resource for acquiring such data. It was produced from multiplexed gene libraries using RNA collected from six developmental stages: embryo, early nauplius (NI-II, late nauplius (NV-VI, early copepodite (CI-II, late copepodite (CV and adult (CVI female. Over 400,000,000 paired-end reads (100 base-pairs long were sequenced on an Illumina instrument, and assembled into 206,041 contigs using Trinity software. Coverage was estimated to be at least 65%. A reference transcriptome comprising 96,090 unique components ("comps" was annotated using Blast2GO. 40% of the comps had significant blast hits. 11% of the comps were successfully annotated with gene ontology (GO terms. Expression of many comps was found to be near zero in one or more developmental stages suggesting that 35 to 48% of the transcriptome is "silent" at any given life stage. Transcripts involved in lipid biosynthesis pathways, critical for the C. finmarchicus life cycle, were identified and their expression pattern during development was examined. Relative expression of three transcripts suggests wax ester biosynthesis in late copepodites, but triacylglyceride biosynthesis in adult females. Two of these transcripts may be involved in the preparatory phase of diapause. A key environmental challenge for C. finmarchicus is the seasonal exposure to the dinoflagellate Alexandrium fundyense with high concentrations of saxitoxins, neurotoxins that block voltage-gated sodium channels. Multiple contigs encoding putative voltage-gated sodium channels were identified. They appeared to be the result of both alternate splicing and gene duplication. This is the first report of multiple NaV1 genes in a protostome. These data provide new insights into the transcriptome

The wolf reference genome sequence (Canis lupus lupus) and its implications for Canis spp. population genomics

DEFF Research Database (Denmark)

Gopalakrishnan, Shyam; Samaniego Castruita, Jose Alfredo; Sinding, Mikkel Holger Strander

2017-01-01

Background An increasing number of studies are addressing the evolutionary genomics of dog domestication, principally through resequencing dog, wolf and related canid genomes. There is, however, only one de novo assembled canid genome currently available against which to map such data - that of a......Background An increasing number of studies are addressing the evolutionary genomics of dog domestication, principally through resequencing dog, wolf and related canid genomes. There is, however, only one de novo assembled canid genome currently available against which to map such data...

MRUniNovo: an efficient tool for de novo peptide sequencing utilizing the hadoop distributed computing framework.

Science.gov (United States)

Li, Chuang; Chen, Tao; He, Qiang; Zhu, Yunping; Li, Kenli

2017-03-15

Tandem mass spectrometry-based de novo peptide sequencing is a complex and time-consuming process. The current algorithms for de novo peptide sequencing cannot rapidly and thoroughly process large mass spectrometry datasets. In this paper, we propose MRUniNovo, a novel tool for parallel de novo peptide sequencing. MRUniNovo parallelizes UniNovo based on the Hadoop compute platform. Our experimental results demonstrate that MRUniNovo significantly reduces the computation time of de novo peptide sequencing without sacrificing the correctness and accuracy of the results, and thus can process very large datasets that UniNovo cannot. MRUniNovo is an open source software tool implemented in java. The source code and the parameter settings are available at http://bioinfo.hupo.org.cn/MRUniNovo/index.php. s131020002@hnu.edu.cn ; taochen1019@163.com. 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

Hybrid Silicon-Based Organic/Inorganic Block Copolymers with Sol-Gel Active Moieties: Synthetic Advances, Self-Assembly and Applications in Biomedicine and Materials Science.

Science.gov (United States)

Czarnecki, Sebastian; Bertin, Annabelle

2018-03-07

Hybrid silicon-based organic/inorganic (multi)block copolymers are promising polymeric precursors to create robust nano-objects and nanomaterials due to their sol-gel active moieties via self-assembly in solution or in bulk. Such nano-objects and nanomaterials have great potential in biomedicine as nanocarriers or scaffolds for bone regeneration as well as in materials science as Pickering emulsifiers, photonic crystals or coatings/films with antibiofouling, antibacterial or water- and oil-repellent properties. Thus, this Review outlines recent synthetic efforts in the preparation of these hybrid inorganic/organic block copolymers, gives an overview of their self-assembled structures and finally presents recent examples of their use in the biomedical field and material science. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Analysis of 62 hybrid assembled human Y chromosomes exposes rapid structural changes and high rates of gene conversion.

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Laurits Skov

2017-08-01

Full Text Available The human Y-chromosome does not recombine across its male-specific part and is therefore an excellent marker of human migrations. It also plays an important role in male fertility. However, its evolution is difficult to fully understand because of repetitive sequences, inverted repeats and the potentially large role of gene conversion. Here we perform an evolutionary analysis of 62 Y-chromosomes of Danish descent sequenced using a wide range of library insert sizes and high coverage, thus allowing large regions of these chromosomes to be well assembled. These include 17 father-son pairs, which we use to validate variation calling. Using a recent method that can integrate variants based on both mapping and de novo assembly, we genotype 10898 SNVs and 2903 indels (max length of 27241 bp in our sample and show by father-son concordance and experimental validation that the non-recurrent SNP and indel variation on the Y chromosome tree is called very accurately. This includes variation called in a 0.9 Mb centromeric heterochromatic region, which is by far the most variable in the Y chromosome. Among the variation is also longer sequence-stretches not present in the reference genome but shared with the chimpanzee Y chromosome. We analyzed 2.7 Mb of large inverted repeats (palindromes for variation patterns among the two palindrome arms and identified 603 mutation and 416 gene conversions events. We find clear evidence for GC-biased gene conversion in the palindromes (and a balancing AT mutation bias, but irrespective of this, also a strong bias towards gene conversion towards the ancestral state, suggesting that palindromic gene conversion may alleviate Muller's ratchet. Finally, we also find a large number of large-scale gene duplications and deletions in the palindromic regions (at least 24 and find that such events can consist of complex combinations of simultaneous insertions and deletions of long stretches of the Y chromosome.

Annotation of a hybrid partial genome of the Coffee Rust (Hemileia vastatrix contributes to the gene repertoire catalogue of the Pucciniales

Directory of Open Access Journals (Sweden)

Marco Aurelio Cristancho

2014-10-01

Full Text Available Coffee leaf rust caused by the fungus Hemileia vastatrix is the most damaging disease to coffee worldwide. The pathogen has recently appeared in multiple outbreaks in coffee producing countries resulting in significant yield losses and increases in costs related to its control. New races/isolates are constantly emerging as evidenced by the presence of the fungus in plants that were previously resistant. Genomic studies are opening new avenues for the study of the evolution of pathogens, the detailed description of plant-pathogen interactions and the development of molecular techniques for the identification of individual isolates. For this purpose we sequenced 8 different H. vastatrix isolates using NGS technologies and gathered partial genome assemblies due to the large repetitive content in the coffee rust hybrid genome; 74.4% of the assembled contigs harbor repetitive sequences. A hybrid assembly of 333Mb was built based on the 8 isolates; this assembly was used for subsequent analyses.Analysis of the conserved gene space showed that the hybrid H. vastatrix genome, though highly fragmented, had a satisfactory level of completion with 91.94% of core protein-coding orthologous genes present. RNA-Seq from urediniospores was used to guide the de novo annotation of the H. vastatrix gene complement. In total, 14,445 genes organized in 3,921 families were uncovered; a considerable proportion of the predicted proteins (73.8% were homologous to other Pucciniales species genomes. Several gene families related to the fungal lifestyle were identified, particularly 483 predicted secreted proteins that represent candidate effector genes and will provide interesting hints to decipher virulence in the coffee rust fungus. The genome sequence of Hva will serve as a template to understand the molecular mechanisms used by this fungus to attack the coffee plant, to study the diversity of this species and for the development of molecular markers to distinguish

Software design of the hybrid robot machine for ITER vacuum vessel assembly and maintenance

Energy Technology Data Exchange (ETDEWEB)

Li, Ming, E-mail: Ming.Li@lut.fi [Laboratory of Intelligent Machines, Lappeenranta University of Technology (Finland); Wu, Huapeng; Handroos, Heikki [Laboratory of Intelligent Machines, Lappeenranta University of Technology (Finland); Yang, Guangyou [School of Mechanical Engineering, Hubei University of Technology, Wuhan (China)

2013-10-15

A specific software design is elaborated in this paper for the hybrid robot machine used for the ITER vacuum vessel (VV) assembly and maintenance. In order to provide the multi-machining-function as well as the complicated, flexible and customizable GUI designing satisfying the non-standardized VV assembly process in one hand, and in another hand guarantee the stringent machining precision in the real-time motion control of robot machine, a client–server-control software architecture is proposed, which separates the user interaction, data communication and robot control implementation into different software layers. Correspondingly, three particular application protocols upon the TCP/IP are designed to transmit the data, command and status between the client and the server so as to deal with the abundant data streaming in the software. In order not to be affected by the graphic user interface (GUI) modification process in the future experiment in VV assembly working field, the real-time control system is realized as a stand-alone module in the architecture to guarantee the controlling performance of the robot machine. After completing the software development, a milling operation is tested on the robot machine, and the result demonstrates that both the specific GUI operability and the real-time motion control performance could be guaranteed adequately in the software design.

Software design of the hybrid robot machine for ITER vacuum vessel assembly and maintenance

International Nuclear Information System (INIS)

Li, Ming; Wu, Huapeng; Handroos, Heikki; Yang, Guangyou

2013-01-01

A specific software design is elaborated in this paper for the hybrid robot machine used for the ITER vacuum vessel (VV) assembly and maintenance. In order to provide the multi-machining-function as well as the complicated, flexible and customizable GUI designing satisfying the non-standardized VV assembly process in one hand, and in another hand guarantee the stringent machining precision in the real-time motion control of robot machine, a client–server-control software architecture is proposed, which separates the user interaction, data communication and robot control implementation into different software layers. Correspondingly, three particular application protocols upon the TCP/IP are designed to transmit the data, command and status between the client and the server so as to deal with the abundant data streaming in the software. In order not to be affected by the graphic user interface (GUI) modification process in the future experiment in VV assembly working field, the real-time control system is realized as a stand-alone module in the architecture to guarantee the controlling performance of the robot machine. After completing the software development, a milling operation is tested on the robot machine, and the result demonstrates that both the specific GUI operability and the real-time motion control performance could be guaranteed adequately in the software design

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.

rDNA genetic imbalance and nucleolar chromatin restructuring is induced by distant hybridization between Raphanus sativus and Brassica alboglabra.

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Hong Long

Full Text Available The expression of rDNA in hybrids inherited from only one progenitor refers to nucleolar dominance. The molecular basis for choosing which genes to silence remains unclear. We report genetic imbalance induced by distant hybridization correlates with formation of rDNA genes (NORs in the hybrids between Raphanus sativus L. and Brassica alboglabra Bailey. Moreover, increased CCGG methylation of rDNA in F1 hybrids is concomitant with Raphanus-derived rDNA gene silencing and rDNA transcriptional inactivity revealed by nucleolar configuration restriction. Newly formed rDNA gene locus occurred through chromosomal in F1 hybrids via chromosomal imbalance. NORs are gained de novo, lost, and/or transposed in the new genome. Inhibition of methyltransferases leads to changes in nucleolar architecture, implicating a key role of methylation in control of nucleolar dominance and vital nucleolar configuration transition. Our findings suggest that gene imbalance and methylation-related chromatin restructuring is important for rDNA gene silencing that may be crucial for synthesis of specific proteins.

De novo assembly of plant body plan: a step ahead of Deadpool

OpenAIRE

Kareem, Abdul; Radhakrishnan, Dhanya; Sondhi, Yash; Aiyaz, Mohammed; Roy, Merin V.; Sugimoto, Kaoru; Prasad, Kalika

2016-01-01

Abstract While in the movie Deadpool it is possible for a human to recreate an arm from scratch, in reality plants can even surpass that. Not only can they regenerate lost parts, but also the whole plant body can be reborn from a few existing cells. Despite the decades old realization that plant cells possess the ability to regenerate a complete?shoot and root system, it is only now that the underlying mechanisms are being unraveled. De novo plant regeneration involves the initiation of regen...

Hemolysin coregulated protein 1 as a molecular gluing unit for the assembly of nanoparticle hybrid structures

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Tuan Anh Pham

2016-03-01

Full Text Available Hybrid nanoparticle (NP structures containing organic building units such as polymers, peptides, DNA and proteins have great potential in biosensor and electronic applications. The nearly free modification of the polymer chain, the variation of the protein and DNA sequence and the implementation of functional moieties provide a great platform to create inorganic structures of different morphology, resulting in different optical and magnetic properties. Nevertheless, the design and modification of a protein structure with functional groups or sequences for the assembly of biohybrid materials is not trivial. This is mainly due to the sensitivity of its secondary, tertiary and quaternary structure to the changes in the interaction (e.g., hydrophobic, hydrophilic, electrostatic, chemical groups between the protein subunits and the inorganic material. Here, we use hemolysin coregulated protein 1 (Hcp1 from Pseudomonas aeruginosa as a building and gluing unit for the formation of biohybrid structures by implementing cysteine anchoring points at defined positions on the protein rim (Hcp1_cys3. We successfully apply the Hcp1_cys3 gluing unit for the assembly of often linear, hybrid structures of plasmonic gold (Au NP, magnetite (Fe3O4 NP, and cobalt ferrite nanoparticles (CoFe2O4 NP. Furthermore, the assembly of Au NPs into linear structures using Hcp1_cys3 is investigated by UV–vis spectroscopy, TEM and cryo-TEM. One key parameter for the formation of Au NP assembly is the specific ionic strength in the mixture. The resulting network-like structure of Au NPs is characterized by Raman spectroscopy, showing surface-enhanced Raman scattering (SERS by a factor of 8·104 and a stable secondary structure of the Hcp1_cys3 unit. In order to prove the catalytic performance of the gold hybrid structures, they are used as a catalyst in the reduction reaction of 4-nitrophenol showing similar catalytic activity as the pure Au NPs. To further extend the

Simulation of the spherical experimental assemblies for the mixed neutron-gamma reference fields implementation

International Nuclear Information System (INIS)

Kluson, J.; Jansky, B.

2009-01-01

Reference mixed neutron-gamma fields are used for test and calibration of dosimetric and spectrometric systems, intercomparison measurements, and benchmark tests and represent experimental base for reactor studies. Set of the spherical experimental assemblies for the mixed neutron-gamma reference fields implementation was build in the NRI Rez. Extended sets of measurements and simulation calculations were done to describe the reference mixed field dosimetry and spectral characteristics with best achievable precision. The Monte Carlo technique was used for different experimental setups models description, comparison and verification and field characteristics simulation. Effects (hardly distinguishable experimentally) were also studied ( contributions from individual parts of experimental setup, field individual components and next effects as shadow shield cones transparency, etc.). Some results and main conclusions of these studies and calculations are presented and discussed. (authors)

Simulation of the spherical experimental assemblies for the mixed neutron-gamma reference fields implementation

International Nuclear Information System (INIS)

Kluson, J.; Jansky, B.

2008-01-01

Reference mixed neutron-gamma fields are used for test and calibration of dosimetric and spectrometric systems, intercomparison measurements, and benchmark tests and represent experimental base for reactor studies. Set of the spherical experimental assemblies for the mixed neutron-gamma reference fields implementation was build in the NRI Rez. Extended sets of measurements and simulation calculations were done to describe the reference mixed field dosimetry and spectral characteristics with best achievable precision. The Monte Carlo technique was used for different experimental setups models description, comparison and verification and field characteristics simulation. Effects (hardly distinguishable experimentally) were also studied ( contributions from individual parts of experimental setup, field individual components and next effects as shadow shield cones transparency, etc.). Some results and main conclusions of these studies and calculations are presented and discussed. (authors)

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

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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.

Assembling the Setaria italica L. Beauv. genome into nine chromosomes and insights into regions affecting growth and drought tolerance.

Science.gov (United States)

Tsai, Kevin J; Lu, Mei-Yeh Jade; Yang, Kai-Jung; Li, Mengyun; Teng, Yuchuan; Chen, Shihmay; Ku, Maurice S B; Li, Wen-Hsiung

2016-10-13

The diploid C 4 plant foxtail millet (Setaria italica L. Beauv.) is an important crop in many parts of Africa and Asia for the vast consumption of its grain and ability to grow in harsh environments, but remains understudied in terms of complete genomic architecture. To date, there have been only two genome assembly and annotation efforts with neither assembly reaching over 86% of the estimated genome size. We have combined de novo assembly with custom reference-guided improvements on a popular cultivar of foxtail millet and have achieved a genome assembly of 477 Mbp in length, which represents over 97% of the estimated 490 Mbp. The assembly anchors over 98% of the predicted genes to the nine assembled nuclear chromosomes and contains more functional annotation gene models than previous assemblies. Our annotation has identified a large number of unique gene ontology terms related to metabolic activities, a region of chromosome 9 with several growth factor proteins, and regions syntenic with pearl millet or maize genomic regions that have been previously shown to affect growth. The new assembly and annotation for this important species can be used for detailed investigation and future innovations in growth for millet and other grains.

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.

Analysis of insecticide resistance-related genes of the Carmine spider mite Tetranychus cinnabarinus based on a de novo assembled transcriptome.

Directory of Open Access Journals (Sweden)

Zhifeng Xu

Full Text Available The carmine spider mite (CSM, Tetranychus cinnabarinus, is an important pest mite in agriculture, because it can develop insecticide resistance easily. To gain valuable gene information and molecular basis for the future insecticide resistance study of CSM, the first transcriptome analysis of CSM was conducted. A total of 45,016 contigs and 25,519 unigenes were generated from the de novo transcriptome assembly, and 15,167 unigenes were annotated via BLAST querying against current databases, including nr, SwissProt, the Clusters of Orthologous Groups (COGs, Kyoto Encyclopedia of Genes and Genomes (KEGG and Gene Ontology (GO. Aligning the transcript to Tetranychus urticae genome, the 19255 (75.45% of the transcripts had significant (e-value <10-5 matches to T. urticae DNA genome, 19111 sequences matched to T. urticae proteome with an average protein length coverage of 42.55%. Core Eukaryotic Genes Mapping Approach (CEGMA analysis identified 435 core eukaryotic genes (CEGs in the CSM dataset corresponding to 95% coverage. Ten gene categories that relate to insecticide resistance in arthropod were generated from CSM transcriptome, including 53 P450-, 22 GSTs-, 23 CarEs-, 1 AChE-, 7 GluCls-, 9 nAChRs-, 8 GABA receptor-, 1 sodium channel-, 6 ATPase- and 12 Cyt b genes. We developed significant molecular resources for T. cinnabarinus putatively involved in insecticide resistance. The transcriptome assembly analysis will significantly facilitate our study on the mechanism of adapting environmental stress (including insecticide in CSM at the molecular level, and will be very important for developing new control strategies against this pest mite.

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

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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.

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).

Electroactive nanoparticle directed assembly of functionalized graphene nanosheets into hierarchical structures with hybrid compositions for flexible supercapacitors

Science.gov (United States)

Choi, Bong Gill; Huh, Yun Suk; Hong, Won Hi; Erickson, David; Park, Ho Seok

2013-04-01

Hierarchical structures of hybrid materials with the controlled compositions have been shown to offer a breakthrough for energy storage and conversion. Here, we report the integrative assembly of chemically modified graphene (CMG) building blocks into hierarchical complex structures with the hybrid composition for high performance flexible pseudocapacitors. The formation mechanism of hierarchical CMG/Nafion/RuO2 (CMGNR) microspheres, which is triggered by the cooperative interplay during the in situ synthesis of RuO2 nanoparticles (NPs), was extensively investigated. In particular, the hierarchical CMGNR microspheres consisting of the aggregates of CMG/Nafion (CMGN) nanosheets and RuO2 NPs provided large surface area and facile ion accessibility to storage sites, while the interconnected nanosheets offered continuous electron pathways and mechanical integrity. The synergistic effect of CMGNR hybrids on the supercapacitor (SC) performance was derived from the hybrid composition of pseudocapacitive RuO2 NPs with the conductive CMGNs as well as from structural features. Consequently, the CMGNR-SCs showed a specific capacitance as high as 160 F g-1, three-fold higher than that of conventional graphene SCs, and a capacitance retention of >95% of the maximum value even after severe bending and 1000 charge-discharge tests due to the structural and compositional features.Hierarchical structures of hybrid materials with the controlled compositions have been shown to offer a breakthrough for energy storage and conversion. Here, we report the integrative assembly of chemically modified graphene (CMG) building blocks into hierarchical complex structures with the hybrid composition for high performance flexible pseudocapacitors. The formation mechanism of hierarchical CMG/Nafion/RuO2 (CMGNR) microspheres, which is triggered by the cooperative interplay during the in situ synthesis of RuO2 nanoparticles (NPs), was extensively investigated. In particular, the hierarchical CMGNR

Analysis of 62 hybrid assembled human Y chromosomes exposes rapid structural changes and high rates of gene conversion

DEFF Research Database (Denmark)

Gonzalez-Izarzugaza, Jose Maria; Skov, Laurits; Maretty, Lasse

2017-01-01

The human Y-chromosome does not recombine across its male-specific part and is therefore an excellent marker of human migrations. It also plays an important role in male fertility. However, its evolution is difficult to fully understand because of repetitive sequences, inverted repeats and the po......The human Y-chromosome does not recombine across its male-specific part and is therefore an excellent marker of human migrations. It also plays an important role in male fertility. However, its evolution is difficult to fully understand because of repetitive sequences, inverted repeats...... and the potentially large role of gene conversion. Here we perform an evolutionary analysis of 62 Y-chromosomes of Danish descent sequenced using a wide range of library insert sizes and high coverage, thus allowing large regions of these chromosomes to be well assembled. These include 17 father-son pairs, which we...... use to validate variation calling. Using a recent method that can integrate variants based on both mapping and de novo assembly, we genotype 10898 SNVs and 2903 indels (max length of 27241 bp) in our sample and show by father-son concordance and experimental validation that the non-recurrent SNP...

Identification and assembly of genomes and genetic elements in complex metagenomic samples without using reference genomes

DEFF Research Database (Denmark)

Nielsen, Henrik Bjørn; Almeida, Mathieu; Juncker, Agnieszka

2014-01-01

of microbial genomes without the need for reference sequences. We demonstrate the method on data from 396 human gut microbiome samples and identify 7,381 co-abundance gene groups (CAGs), including 741 metagenomic species (MGS). We use these to assemble 238 high-quality microbial genomes and identify...

Hybrid methyl green/cobalt-polyoxotungstate nanostructured films: Self-assembly, electrochemical and electrocatalytic properties

Energy Technology Data Exchange (ETDEWEB)

Novais, Hugo C.; Fernandes, Diana M., E-mail: diana.fernandes@fc.up.pt; Freire, Cristina, E-mail: acfreire@fc.up.pt

2015-08-30

Graphical abstract: Hybrid {MG/Co(PW9)2}{sub n} multilayer films were successfully prepared and exhibit W-based electrocatalytic activity towards reduction of nitrite and iodate anions. - Highlights: • Layer-by-layer hybrid films {MG/Co(PW_9)_2}{sub n} were sucessfully prepared. • UV–vis was used to monitor film build-up and showed regular stepwise film growth. • XPS confirmed sucessfull {MG/Co(PW_9)_2}{sub n} film fabrication. • Films showed excellent electrocatalytic activity towards nitrite and iodate reduction. - Abstract: Hybrid multilayer films were prepared by alternately depositing cationic dye methyl green (MG) and anionic sandwich-type polyoxometalate K{sub 10}[Co{sub 4}(H{sub 2}O){sub 2}(PW{sub 9}O{sub 34}){sub 2}] (Co(PW{sub 9}){sub 2}) via electrostatic layer-by-layer (LbL) self-assembly method. Film build-up was monitored by UV–vis spectroscopy which showed a regular stepwise growth. X-ray photoelectron spectroscopy data confirmed the successful fabrication of the hybrid films with MG-Co(PW{sub 9}){sub 2} composition and scanning electron microscopy images revealed a completely covered surface with a non-uniform distribution of the molecular species. Electrochemical characterization of films by cyclic voltammetry revealed two tungsten-based reduction processes in the potential range between −0.9 and −0.5 V due to W{sup VI} → W{sup V} in Co(PW{sub 9}){sub 2}. Studies with the redox probes, [Fe(CN){sub 6}]{sup 3−/4−} and [Ru(NH{sub 3}){sub 6}]{sup 3+/2+}, revealed that not only the electrostatic attractions or repulsions have effects on the kinetics of the probe reactions, but also the film thickness. Additionally, the {MG/Co(PW_9)_2}{sub n} multilayer films exhibit efficient W-based electrocatalytic activity towards reduction of nitrite and iodate.

Comparison of de novo assembly statistics of Cucumis sativus L.

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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.

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.

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.

Self-assembled organic-inorganic magnetic hybrid adsorbent ferrite based on cyclodextrin nanoparticles.

Science.gov (United States)

Denadai, Angelo M L; De Sousa, Frederico B; Passos, Joel J; Guatimosim, Fernando C; Barbosa, Kirla D; Burgos, Ana E; de Oliveira, Fernando Castro; da Silva, Jeann C; Neves, Bernardo R A; Mohallem, Nelcy D S; Sinisterra, Rubén D

2012-01-01

Organic-inorganic magnetic hybrid materials (MHMs) combine a nonmagnetic and a magnetic component by means of electrostatic interactions or covalent bonds, and notable features can be achieved. Herein, we describe an application of a self-assembled material based on ferrite associated with β-cyclodextrin (Fe-Ni/Zn/βCD) at the nanoscale level. This MHM and pure ferrite (Fe-Ni/Zn) were used as an adsorbent system for Cr(3+) and Cr(2)O(7) (2-) ions in aqueous solutions. Prior to the adsorption studies, both ferrites were characterized in order to determine the particle size distribution, morphology and available binding sites on the surface of the materials. Microscopy analysis demonstrated that both ferrites present two different size domains, at the micro- and nanoscale level, with the latter being able to self-assemble into larger particles. Fe-Ni/Zn/βCD presented smaller particles and a more homogeneous particle size distribution. Higher porosity for this MHM compared to Fe-Ni/Zn was observed by Brunauer-Emmett-Teller isotherms and positron-annihilation-lifetime spectroscopy. Based on the pKa values, potentiometric titrations demonstrated the presence of βCD in the inorganic matrix, indicating that the lamellar structures verified by transmission electronic microscopy can be associated with βCD assembled structures. Colloidal stability was inferred as a function of time at different pH values, indicating the sedimentation rate as a function of pH. Zeta potential measurements identified an amphoteric behavior for the Fe-Ni/Zn/βCD, suggesting its better capability to remove ions (cations and anions) from aqueous solutions compared to that of Fe-Ni/Zn.

Self-assembled organic–inorganic magnetic hybrid adsorbent ferrite based on cyclodextrin nanoparticles

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Ângelo M. L. Denadai

2012-11-01

Full Text Available Organic–inorganic magnetic hybrid materials (MHMs combine a nonmagnetic and a magnetic component by means of electrostatic interactions or covalent bonds, and notable features can be achieved. Herein, we describe an application of a self-assembled material based on ferrite associated with β-cyclodextrin (Fe-Ni/Zn/βCD at the nanoscale level. This MHM and pure ferrite (Fe-Ni/Zn were used as an adsorbent system for Cr3+ and Cr2O72− ions in aqueous solutions. Prior to the adsorption studies, both ferrites were characterized in order to determine the particle size distribution, morphology and available binding sites on the surface of the materials. Microscopy analysis demonstrated that both ferrites present two different size domains, at the micro- and nanoscale level, with the latter being able to self-assemble into larger particles. Fe-Ni/Zn/βCD presented smaller particles and a more homogeneous particle size distribution. Higher porosity for this MHM compared to Fe-Ni/Zn was observed by Brunauer–Emmett–Teller isotherms and positron-annihilation-lifetime spectroscopy. Based on the pKa values, potentiometric titrations demonstrated the presence of βCD in the inorganic matrix, indicating that the lamellar structures verified by transmission electronic microscopy can be associated with βCD assembled structures. Colloidal stability was inferred as a function of time at different pH values, indicating the sedimentation rate as a function of pH. Zeta potential measurements identified an amphoteric behavior for the Fe-Ni/Zn/βCD, suggesting its better capability to remove ions (cations and anions from aqueous solutions compared to that of Fe-Ni/Zn.

Fabrication of graphene-fullerene hybrid by self-assembly and its application as support material for methanol electrocatalytic oxidation reaction

Science.gov (United States)

Zhang, Xuan; Zhang, Jia-Wei; Xiang, Ping-Hua; Qiao, Jinli

2018-05-01

Graphene-fullerene hybrids were facilely fabricated by self-assembly of graphene oxide (GO) and multi-substituted fulleropyrrolidines (PyrC60). The hybrids (GO-PyrC60) were applied as support materials to deposit Pd nanoparticle catalyst by a simple hydrothermal co-reduction approach. The as-prepared electrocatalysts (Pd/RGO-PyrC60) were characterized by transmission electron microscopy (TEM), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS), respectively. The RGO-PyrC60 hybrid supported Pd catalyst with the optimal ratio of RGO to PyrC60, exhibited much enhanced electrocatalytic activity and stability toward methanol oxidation reaction (MOR) compared to the RGO alone supported Pd as well as commercial Pd/C. The introduction of fulleropyrrolidine as spacer between graphene layers could increase the electrocatalytic activity and improve the long-term stability. This strategy may contribute to developing graphene-fullerene hydrids as effective support materials for advanced electrocatalysts.

De novo transcriptome assembly and comparative analysis of differentially expressed genes in Prunus dulcis Mill. in response to freezing stress.

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Sadegh Mousavi

Full Text Available Almond (Prunus dulcis Mill., one of the most important nut crops, requires chilling during winter to develop fruiting buds. However, early spring chilling and late spring frost may damage the reproductive tissues leading to reduction in the rate of productivity. Despite the importance of transcriptional changes and regulation, little is known about the almond's transcriptome under the cold stress conditions. In the current research, we used RNA-seq technique to study the response of the reproductive tissues of almond (anther and ovary to frost stress. RNA sequencing resulted in more than 20 million reads from anther and ovary tissues of almond, individually. About 40,000 contigs were assembled and annotated de novo in each tissue. Profile of gene expression in ovary showed significant alterations in 5,112 genes, whereas in anther 6,926 genes were affected by freezing stress. Around two thousands of these genes were common altered genes in both ovary and anther libraries. Gene ontology indicated the involvement of differentially expressed (DE genes, responding to freezing stress, in metabolic and cellular processes. qRT-PCR analysis verified the expression pattern of eight genes randomly selected from the DE genes. In conclusion, the almond gene index assembled in this study and the reported DE genes can provide great insights on responses of almond and other Prunus species to abiotic stresses. The obtained results from current research would add to the limited available information on almond and Rosaceae. Besides, the findings would be very useful for comparative studies as the number of DE genes reported here is much higher than that of any previous reports in this plant.

De novo transcriptome assembly and comparative analysis of differentially expressed genes in Prunus dulcis Mill. in response to freezing stress.

Science.gov (United States)

Mousavi, Sadegh; Alisoltani, Arghavan; Shiran, Behrouz; Fallahi, Hossein; Ebrahimie, Esameil; Imani, Ali; Houshmand, Saadollah

2014-01-01

Almond (Prunus dulcis Mill.), one of the most important nut crops, requires chilling during winter to develop fruiting buds. However, early spring chilling and late spring frost may damage the reproductive tissues leading to reduction in the rate of productivity. Despite the importance of transcriptional changes and regulation, little is known about the almond's transcriptome under the cold stress conditions. In the current research, we used RNA-seq technique to study the response of the reproductive tissues of almond (anther and ovary) to frost stress. RNA sequencing resulted in more than 20 million reads from anther and ovary tissues of almond, individually. About 40,000 contigs were assembled and annotated de novo in each tissue. Profile of gene expression in ovary showed significant alterations in 5,112 genes, whereas in anther 6,926 genes were affected by freezing stress. Around two thousands of these genes were common altered genes in both ovary and anther libraries. Gene ontology indicated the involvement of differentially expressed (DE) genes, responding to freezing stress, in metabolic and cellular processes. qRT-PCR analysis verified the expression pattern of eight genes randomly selected from the DE genes. In conclusion, the almond gene index assembled in this study and the reported DE genes can provide great insights on responses of almond and other Prunus species to abiotic stresses. The obtained results from current research would add to the limited available information on almond and Rosaceae. Besides, the findings would be very useful for comparative studies as the number of DE genes reported here is much higher than that of any previous reports in this plant.

De novo transcriptome assembly of mangosteen (Garcinia mangostana L. fruit

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Deden Derajat Matra

2016-12-01

Full Text Available Garcinia mangostana L. (Mangosteen, of the family Clusiaceae, is one of the economically important tropical fruits in Indonesia. In the present study, we performed de novo transcriptomic analysis of Garcinia mangostana L. through RNA-Seq technology. We obtained the raw data from 12 libraries through Ion Proton System. Clean reads of 191,735,809 were obtained from 307,634,890 raw reads. The raw data obtained in this study can be accessible in DDBJ database with accession number of DRA005014 with bioproject accession number of PRJDB5091. We obtained 268,851 transcripts as well as 155,850 unigenes, having N50 value of 555 and 433 bp, respectively. Transcript/unigene length ranged from 201 to 5916 bp. The unigenes were annotated with two main databases from NCBI and UniProtKB, respectively having annotated-sequences of 73,287 and 73,107, respectively. These transcriptomic data will be beneficial for studying transcriptome of Garcinia mangostana L.

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

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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

Precise detection of de novo single nucleotide variants in human genomes.

Science.gov (United States)

Gómez-Romero, Laura; Palacios-Flores, Kim; Reyes, José; García, Delfino; Boege, Margareta; Dávila, Guillermo; Flores, Margarita; Schatz, Michael C; Palacios, Rafael

2018-05-07

The precise determination of de novo genetic variants has enormous implications across different fields of biology and medicine, particularly personalized medicine. Currently, de novo variations are identified by mapping sample reads from a parent-offspring trio to a reference genome, allowing for a certain degree of differences. While widely used, this approach often introduces false-positive (FP) results due to misaligned reads and mischaracterized sequencing errors. In a previous study, we developed an alternative approach to accurately identify single nucleotide variants (SNVs) using only perfect matches. However, this approach could be applied only to haploid regions of the genome and was computationally intensive. In this study, we present a unique approach, coverage-based single nucleotide variant identification (COBASI), which allows the exploration of the entire genome using second-generation short sequence reads without extensive computing requirements. COBASI identifies SNVs using changes in coverage of exactly matching unique substrings, and is particularly suited for pinpointing de novo SNVs. Unlike other approaches that require population frequencies across hundreds of samples to filter out any methodological biases, COBASI can be applied to detect de novo SNVs within isolated families. We demonstrate this capability through extensive simulation studies and by studying a parent-offspring trio we sequenced using short reads. Experimental validation of all 58 candidate de novo SNVs and a selection of non-de novo SNVs found in the trio confirmed zero FP calls. COBASI is available as open source at https://github.com/Laura-Gomez/COBASI for any researcher to use. Copyright © 2018 the Author(s). Published by PNAS.

Hybrid, Nanoscale Phospholipid/Block Copolymer Vesicles

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Bo Liedberg

2013-09-01

Full Text Available Hybrid phospholipid/block copolymer vesicles, in which the polymeric membrane is blended with phospholipids, display interesting self-assembly behavior, incorporating the robustness and chemical versatility of polymersomes with the softness and biocompatibility of liposomes. Such structures can be conveniently characterized by preparing giant unilamellar vesicles (GUVs via electroformation. Here, we are interested in exploring the self-assembly and properties of the analogous nanoscale hybrid vesicles (ca. 100 nm in diameter of the same composition prepared by film-hydration and extrusion. We show that the self-assembly and content-release behavior of nanoscale polybutadiene-b-poly(ethylene oxide (PB-PEO/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC hybrid phospholipid/block copolymer vesicles can be tuned by the mixing ratio of the amphiphiles. In brief, these hybrids may provide alternative tools for drug delivery purposes and molecular imaging/sensing applications and clearly open up new avenues for further investigation.

De Novo Transcriptome Assembly (NGS) of Curcuma longa L. Rhizome Reveals Novel Transcripts Related to Anticancer and Antimalarial Terpenoids

Science.gov (United States)

Jayakumar, Vasanthan; Damodaran, Anand C.; Rao, Sudha Narayana; Katta, Mohan A. V. S. K.; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C.

2013-01-01

Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa. PMID:23468859

De Novo transcriptome assembly (NGS of Curcuma longa L. rhizome reveals novel transcripts related to anticancer and antimalarial terpenoids.

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Ramasamy S Annadurai

Full Text Available Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa.

De Novo transcriptome assembly (NGS) of Curcuma longa L. rhizome reveals novel transcripts related to anticancer and antimalarial terpenoids.

Science.gov (United States)

Annadurai, Ramasamy S; Neethiraj, Ramprasad; Jayakumar, Vasanthan; Damodaran, Anand C; Rao, Sudha Narayana; Katta, Mohan A V S K; Gopinathan, Sreeja; Sarma, Santosh Prasad; Senthilkumar, Vanitha; Niranjan, Vidya; Gopinath, Ashok; Mugasimangalam, Raja C

2013-01-01

Herbal remedies are increasingly being recognised in recent years as alternative medicine for a number of diseases including cancer. Curcuma longa L., commonly known as turmeric is used as a culinary spice in India and in many Asian countries has been attributed to lower incidences of gastrointestinal cancers. Curcumin, a secondary metabolite isolated from the rhizomes of this plant has been shown to have significant anticancer properties, in addition to antimalarial and antioxidant effects. We sequenced the transcriptome of the rhizome of the 3 varieties of Curcuma longa L. using Illumina reversible dye terminator sequencing followed by de novo transcriptome assembly. Multiple databases were used to obtain a comprehensive annotation and the transcripts were functionally classified using GO, KOG and PlantCyc. Special emphasis was given for annotating the secondary metabolite pathways and terpenoid biosynthesis pathways. We report for the first time, the presence of transcripts related to biosynthetic pathways of several anti-cancer compounds like taxol, curcumin, and vinblastine in addition to anti-malarial compounds like artemisinin and acridone alkaloids, emphasizing turmeric's importance as a highly potent phytochemical. Our data not only provides molecular signatures for several terpenoids but also a comprehensive molecular resource for facilitating deeper insights into the transcriptome of C. longa.

Oxford Nanopore MinION Sequencing and Genome Assembly

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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.

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

De novo assembly of Eugenia uniflora L. transcriptome and identification of genes from the terpenoid biosynthesis pathway.

Science.gov (United States)

Guzman, Frank; Kulcheski, Franceli Rodrigues; Turchetto-Zolet, Andreia Carina; Margis, Rogerio

2014-12-01

Pitanga (Eugenia uniflora L.) is a member of the Myrtaceae family and is of particular interest due to its medicinal properties that are attributed to specialized metabolites with known biological activities. Among these molecules, terpenoids are the most abundant in essential oils that are found in the leaves and represent compounds with potential pharmacological benefits. The terpene diversity observed in Myrtaceae is determined by the activity of different members of the terpene synthase and oxidosqualene cyclase families. Therefore, the aim of this study was to perform a de novo assembly of transcripts from E. uniflora leaves and to annotation to identify the genes potentially involved in the terpenoid biosynthesis pathway and terpene diversity. In total, 72,742 unigenes with a mean length of 1048bp were identified. Of these, 43,631 and 36,289 were annotated with the NCBI non-redundant protein and Swiss-Prot databases, respectively. The gene ontology categorized the sequences into 53 functional groups. A metabolic pathway analysis with KEGG revealed 8,625 unigenes assigned to 141 metabolic pathways and 40 unigenes predicted to be associated with the biosynthesis of terpenoids. Furthermore, we identified four putative full-length terpene synthase genes involved in sesquiterpenes and monoterpenes biosynthesis, and three putative full-length oxidosqualene cyclase genes involved in the triterpenes biosynthesis. The expression of these genes was validated in different E. uniflora tissues. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

De novo sequencing, assembly, and analysis of Iris lactea var. chinensis roots' transcriptome in response to salt stress.

Science.gov (United States)

Gu, Chunsun; Xu, Sheng; Wang, Zhiquan; Liu, Liangqin; Zhang, Yongxia; Deng, Yanming; Huang, Suzhen

2018-04-01

As a halophyte, Iris lactea var. chinensis (I. lactea var. chinensis) is widely distributed and has good drought and heavy metal resistance. Moreover, it is an excellent ornamental plant. I. lactea var. chinensis has extensive application prospects owing to the global impacts of salinization. To better understand its molecular mechanism involved in salt resistance, the de novo sequencing, assembly, and analysis of I. lactea var. chinensis roots' transcriptome in response to salt-stress conditions was performed. On average, 74.17% of the clean reads were mapped to unigenes. A total of 121,093 unigenes were constructed and 56,398 (46.57%) were annotated. Among these, 13,522 differentially expressed genes (DEGs) were identified between salt-treated and control samples Compared to the transcriptional level of control, 7037 DEGs were up-regulated and 6539 down-regulated. In addition, 129 up-regulated and 1609 down-regulated genes were simultaneously detected in all three pairwise comparisons between control and salt-stressed libraries. At least 247 and 250 DEGs encoding transcription factors and transporter proteins were identified. Meanwhile, 130 DEGs regarding reactive oxygen species (ROS) scavenging system were also summarized. Based on real-time quantitative RT-PCR, we verified the changes in the expression patterns of 10 unigenes. Our study identified potential salt-responsive candidate genes and increased the understanding of halophyte responses to salinity stress. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

De Novo Assembly and Transcriptome Analysis of Bulb Onion (Allium cepa L.) during Cold Acclimation Using Contrasting Genotypes.

Science.gov (United States)

Han, Jeongsukhyeon; Thamilarasan, Senthil Kumar; Natarajan, Sathishkumar; Park, Jong-In; Chung, Mi-Young; Nou, Ill-Sup

2016-01-01

Bulb onion (Allium cepa) is the second most widely cultivated and consumed vegetable crop in the world. During winter, cold injury can limit the production of bulb onion. Genomic resources available for bulb onion are still very limited. To date, no studies on heritably durable cold and freezing tolerance have been carried out in bulb onion genotypes. We applied high-throughput sequencing technology to cold (2°C), freezing (-5 and -15°C), and control (25°C)-treated samples of cold tolerant (CT) and cold susceptible (CS) genotypes of A. cepa lines. A total of 452 million paired-end reads were de novo assembled into 54,047 genes with an average length of 1,331 bp. Based on similarity searches, these genes were aligned with entries in the public non-redundant (nr) database, as well as KEGG and COG database. Differentially expressed genes (DEGs) were identified using log10 values with the FPKM method. Among 5,167DEGs, 491 genes were differentially expressed at freezing temperature compared to the control temperature in both CT and CS libraries. The DEG results were validated with qRT-PCR. We performed GO and KEGG pathway enrichment analyses of all DEGs and iPath interactive analysis found 31 pathways including those related to metabolism of carbohydrate, nucleotide, energy, cofactors and vitamins, other amino acids and xenobiotics biodegradation. Furthermore, a large number of molecular markers were identified from the assembled genes, including simple sequence repeats (SSRs) 4,437 and SNP substitutions of transition and transversion types of CT and CS. Our study is the first to provide a transcriptome sequence resource for Allium spp. with regard to cold and freezing stress. We identified a large set of genes and determined their DEG profiles under cold and freezing conditions using two different genotypes. These data represent a valuable resource for genetic and genomic studies of Allium spp.

De Novo Assembly and Transcriptome Analysis of Bulb Onion (Allium cepa L. during Cold Acclimation Using Contrasting Genotypes.

Directory of Open Access Journals (Sweden)

Jeongsukhyeon Han

Full Text Available Bulb onion (Allium cepa is the second most widely cultivated and consumed vegetable crop in the world. During winter, cold injury can limit the production of bulb onion. Genomic resources available for bulb onion are still very limited. To date, no studies on heritably durable cold and freezing tolerance have been carried out in bulb onion genotypes. We applied high-throughput sequencing technology to cold (2°C, freezing (-5 and -15°C, and control (25°C-treated samples of cold tolerant (CT and cold susceptible (CS genotypes of A. cepa lines. A total of 452 million paired-end reads were de novo assembled into 54,047 genes with an average length of 1,331 bp. Based on similarity searches, these genes were aligned with entries in the public non-redundant (nr database, as well as KEGG and COG database. Differentially expressed genes (DEGs were identified using log10 values with the FPKM method. Among 5,167DEGs, 491 genes were differentially expressed at freezing temperature compared to the control temperature in both CT and CS libraries. The DEG results were validated with qRT-PCR. We performed GO and KEGG pathway enrichment analyses of all DEGs and iPath interactive analysis found 31 pathways including those related to metabolism of carbohydrate, nucleotide, energy, cofactors and vitamins, other amino acids and xenobiotics biodegradation. Furthermore, a large number of molecular markers were identified from the assembled genes, including simple sequence repeats (SSRs 4,437 and SNP substitutions of transition and transversion types of CT and CS. Our study is the first to provide a transcriptome sequence resource for Allium spp. with regard to cold and freezing stress. We identified a large set of genes and determined their DEG profiles under cold and freezing conditions using two different genotypes. These data represent a valuable resource for genetic and genomic studies of Allium spp.

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.

Transcription Activator-Like Effectors (TALEs) Hybrid Nucleases for Genome Engineering Application

KAUST Repository

Wibowo, Anjar

2011-06-06

Gene targeting is a powerful genome engineering tool that can be used for a variety of biotechnological applications. Genomic double-strand DNA breaks generated by engineered site-specific nucleases can stimulate gene targeting. Hybrid nucleases are composed of DNA binding module and DNA cleavage module. Zinc Finger Nucleases were used to generate double-strand DNA breaks but it suffers from failures and lack of reproducibility. The transcription activator–like effectors (TALEs) from plant pathogenic Xanthomonas contain a unique type of DNA-binding domain that bind specific DNA targets. The purpose of this study is to generate novel sequence specific nucleases by fusing a de novo engineered Hax3 TALE-based DNA binding domain to a FokI cleavage domain. Our data show that the de novo engineered TALE nuclease can bind to its target sequence and create double-strand DNA breaks in vitro. We also show that the de novo engineered TALE nuclease is capable of generating double-strand DNA breaks in its target sequence in vivo, when transiently expressed in Nicotiana benthamiana leaves. In conclusion, our data demonstrate that TALE-based hybrid nucleases can be tailored to bind a user-selected DNA sequence and generate site-specific genomic double-strand DNA breaks. TALE-based hybrid nucleases hold much promise as powerful molecular tools for gene targeting applications.

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.

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.

Why close a bacterial genome? The plasmid of Alteromonas macleodii HOT1A3 is a vector for inter-specific transfer of a flexible genomic island

Directory of Open Access Journals (Sweden)

Eduard eFadeev

2016-03-01

Full Text Available Genome sequencing is rapidly becoming a staple technique in environmental and clinical microbiology, yet computational challenges still remain, leading to many draft genomes which are typically fragmented into many contigs. We sequenced and completely assembled the genome of a marine heterotrophic bacterium, Alteromonas macleodii HOT1A3, and compared its full genome to several draft genomes obtained using different reference-based and de-novo methods. In general, the de-novo assemblies clearly outperformed the reference-based or hybrid ones, covering>99% of the genes and representing essentially all of the gene functions. However, only the fully closed genome (~4.5Mbp allowed us to identify the presence of a large, 148 kbp plasmid, pAM1A3. While HOT1A3 belongs to Alteromonas macleodii, typically found in surface waters (surface ecotype, this plasmid consists of an almost complete flexible genomic island, containing many genes involved in metal resistance previously identified in the genomes of Alteromonas mediterranea (deep ecotype. Indeed, similar to A. mediterranea, A. macleodii HOT1A3 grows at concentrations of zinc, mercury and copper that are inhibitory for other A. macleodii strains. The presence of a plasmid encoding almost an entire flexible genomic island suggests that wholesale genomic exchange between heterotrophic marine bacteria belonging to related but ecologically different populations is not uncommon.

Self-assembled nanostructures

CERN Document Server

Zhang, Jin Z; Liu, Jun; Chen, Shaowei; Liu, Gang-yu

2003-01-01

Nanostructures refer to materials that have relevant dimensions on the nanometer length scales and reside in the mesoscopic regime between isolated atoms and molecules in bulk matter. These materials have unique physical properties that are distinctly different from bulk materials. Self-Assembled Nanostructures provides systematic coverage of basic nanomaterials science including materials assembly and synthesis, characterization, and application. Suitable for both beginners and experts, it balances the chemistry aspects of nanomaterials with physical principles. It also highlights nanomaterial-based architectures including assembled or self-assembled systems. Filled with in-depth discussion of important applications of nano-architectures as well as potential applications ranging from physical to chemical and biological systems, Self-Assembled Nanostructures is the essential reference or text for scientists involved with nanostructures.

DeNovoGUI: an open source graphical user interface for de novo sequencing of tandem mass spectra.

Science.gov (United States)

Muth, Thilo; Weilnböck, Lisa; Rapp, Erdmann; Huber, Christian G; Martens, Lennart; Vaudel, Marc; Barsnes, Harald

2014-02-07

De novo sequencing is a popular technique in proteomics for identifying peptides from tandem mass spectra without having to rely on a protein sequence database. Despite the strong potential of de novo sequencing algorithms, their adoption threshold remains quite high. We here present a user-friendly and lightweight graphical user interface called DeNovoGUI for running parallelized versions of the freely available de novo sequencing software PepNovo+, greatly simplifying the use of de novo sequencing in proteomics. Our platform-independent software is freely available under the permissible Apache2 open source license. Source code, binaries, and additional documentation are available at http://denovogui.googlecode.com .

Directed assembly of hybrid nanostructures using optically resonant nanotweezers

Energy Technology Data Exchange (ETDEWEB)

Erickson, David [Cornell Univ., Ithaca, NY (United States)

2015-09-09

This represents the final report for this project. Over the course of the project we have made significant progress in photonically driven nano-assembly including: (1) demonstrating the first direct optical tweezer based manipulation of proteins, (2) the ability to apply optical angular torques to microtubuals and other rod-shaped microparticles, (3) direct assembly of hybrid nanostructures comprising of polymeric nanoparticles and carbon nanotubes and, (4) the ability to drive biological reactions (specifically protein aggregation) that are thermodynamically unfavorable by applying localized optical work. These advancements are described in the list of papers provided in section 2.0 of the below. Summary details are provided in prior year annual reports. We have two additional papers which will be submitted shortly based on the work done under this award. An updated publication list will be provided to the program manager when those are accepted. In this report, we report on a new advancement made in the final project year, which uses the nanotweezer technology to perform direct measurements of particle-surface interactions. Briefly, these measurements are important for characterizing the stability and behavior of colloidal and nanoparticle suspensions and current techniques are limited in their ability to measure piconewton scale interaction forces on sub-micrometer particles due to signal detection limits and thermal noise. In this project year we developed a new technique called “Nanophotonic Force Microscopy” which uses the localized region of exponentially decaying, near-field, light to confine small particles close to a surface. From the statistical distribution of the light intensity scattered by the particle the technique maps out the potential well of the trap and directly quantify the repulsive force between the nanoparticle and the surface. The major advantage of the technique is that it can measure forces and energy wells below the thermal noise

A draft de novo genome assembly for the northern bobwhite (Colinus virginianus reveals evidence for a rapid decline in effective population size beginning in the Late Pleistocene.

Directory of Open Access Journals (Sweden)

Yvette A Halley

Full Text Available Wild populations of northern bobwhites (Colinus virginianus; hereafter bobwhite have declined across nearly all of their U.S. range, and despite their importance as an experimental wildlife model for ecotoxicology studies, no bobwhite draft genome assembly currently exists. Herein, we present a bobwhite draft de novo genome assembly with annotation, comparative analyses including genome-wide analyses of divergence with the chicken (Gallus gallus and zebra finch (Taeniopygia guttata genomes, and coalescent modeling to reconstruct the demographic history of the bobwhite for comparison to other birds currently in decline (i.e., scarlet macaw; Ara macao. More than 90% of the assembled bobwhite genome was captured within 14,000 unique genes and proteins. Bobwhite analyses of divergence with the chicken and zebra finch genomes revealed many extremely conserved gene sequences, and evidence for lineage-specific divergence of noncoding regions. Coalescent models for reconstructing the demographic history of the bobwhite and the scarlet macaw provided evidence for population bottlenecks which were temporally coincident with human colonization of the New World, the late Pleistocene collapse of the megafauna, and the last glacial maximum. Demographic trends predicted for the bobwhite and the scarlet macaw also were concordant with how opposing natural selection strategies (i.e., skewness in the r-/K-selection continuum would be expected to shape genome diversity and the effective population sizes in these species, which is directly relevant to future conservation efforts.

Transcriptomic resources for the medicinal legume Mucuna pruriens: de novo transcriptome assembly, annotation, identification and validation of EST-SSR markers.

Science.gov (United States)

Sathyanarayana, N; Pittala, Ranjith Kumar; Tripathi, Pankaj Kumar; Chopra, Ratan; Singh, Heikham Russiachand; Belamkar, Vikas; Bhardwaj, Pardeep Kumar; Doyle, Jeff J; Egan, Ashley N

2017-05-25

The medicinal legume Mucuna pruriens (L.) DC. has attracted attention worldwide as a source of the anti-Parkinson's drug L-Dopa. It is also a popular green manure cover crop that offers many agronomic benefits including high protein content, nitrogen fixation and soil nutrients. The plant currently lacks genomic resources and there is limited knowledge on gene expression, metabolic pathways, and genetics of secondary metabolite production. Here, we present transcriptomic resources for M. pruriens, including a de novo transcriptome assembly and annotation, as well as differential transcript expression analyses between root, leaf, and pod tissues. We also develop microsatellite markers and analyze genetic diversity and population structure within a set of Indian germplasm accessions. One-hundred ninety-one million two hundred thirty-three thousand two hundred forty-two bp cleaned reads were assembled into 67,561 transcripts with mean length of 626 bp and N50 of 987 bp. Assembled sequences were annotated using BLASTX against public databases with over 80% of transcripts annotated. We identified 7,493 simple sequence repeat (SSR) motifs, including 787 polymorphic repeats between the parents of a mapping population. 134 SSRs from expressed sequenced tags (ESTs) were screened against 23 M. pruriens accessions from India, with 52 EST-SSRs retained after quality control. Population structure analysis using a Bayesian framework implemented in fastSTRUCTURE showed nearly similar groupings as with distance-based (neighbor-joining) and principal component analyses, with most of the accessions clustering per geographical origins. Pair-wise comparison of transcript expression in leaves, roots and pods identified 4,387 differentially expressed transcripts with the highest number occurring between roots and leaves. Differentially expressed transcripts were enriched with transcription factors and transcripts annotated as belonging to secondary metabolite pathways. The M

De novo assembly and comparative analysis of the transcriptome of embryogenic callus formation in bread wheat (Triticum aestivum L.).

Science.gov (United States)

Chu, Zongli; Chen, Junying; Sun, Junyan; Dong, Zhongdong; Yang, Xia; Wang, Ying; Xu, Haixia; Zhang, Xiaoke; Chen, Feng; Cui, Dangqun

2017-12-19

During asexual reproduction the embryogenic callus can differentiate into a new plantlet, offering great potential for fostering in vitro culture efficiency in plants. The immature embryos (IMEs) of wheat (Triticum aestivum L.) are more easily able to generate embryogenic callus than mature embryos (MEs). To understand the molecular process of embryogenic callus formation in wheat, de novo transcriptome sequencing was used to generate transcriptome sequences from calli derived from IMEs and MEs after 3d, 6d, or 15d of culture (DC). In total, 155 million high quality paired-end reads were obtained from the 6 cDNA libraries. Our de novo assembly generated 142,221 unigenes, of which 59,976 (42.17%) were annotated with a significant Blastx against nr, Pfam, Swissprot, KOG, KEGG, GO and COG/KOG databases. Comparative transcriptome analysis indicated that a total of 5194 differentially expressed genes (DEGs) were identified in the comparisons of IME vs. ME at the three stages, including 3181, 2085 and 1468 DEGs at 3, 6 and 15 DC, respectively. Of them, 283 overlapped in all the three comparisons. Furthermore, 4731 DEGs were identified in the comparisons between stages in IMEs and MEs. Functional analysis revealed that 271transcription factor (TF) genes (10 overlapped in all 3 comparisons of IME vs. ME) and 346 somatic embryogenesis related genes (SSEGs; 35 overlapped in all 3 comparisons of IME vs. ME) were differentially expressed in at least one comparison of IME vs. ME. In addition, of the 283 overlapped DEGs in the 3 comparisons of IME vs. ME, excluding the SSEGs and TFs, 39 possessed a higher rate of involvement in biological processes relating to response to stimuli, in multi-organism processes, reproductive processes and reproduction. Furthermore, 7 were simultaneously differentially expressed in the 2 comparisons between the stages in IMEs, but not MEs, suggesting that they may be related to embryogenic callus formation. The expression levels of genes, which

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.

Improving amphibian genomic resources: a multitissue reference transcriptome of an iconic invader.

Science.gov (United States)

Richardson, Mark F; Sequeira, Fernando; Selechnik, Daniel; Carneiro, Miguel; Vallinoto, Marcelo; Reid, Jack G; West, Andrea J; Crossland, Michael R; Shine, Richard; Rollins, Lee A

2018-01-01

Cane toads (Rhinella marina) are an iconic invasive species introduced to 4 continents and well utilized for studies of rapid evolution in introduced environments. Despite the long introduction history of this species, its profound ecological impacts, and its utility for demonstrating evolutionary principles, genetic information is sparse. Here we produce a de novo transcriptome spanning multiple tissues and life stages to enable investigation of the genetic basis of previously identified rapid phenotypic change over the introduced range. Using approximately 1.9 billion reads from developing tadpoles and 6 adult tissue-specific cDNA libraries, as well as a transcriptome assembly pipeline encompassing 100 separate de novo assemblies, we constructed 62 202 transcripts, of which we functionally annotated ∼50%. Our transcriptome assembly exhibits 90% full-length completeness of the Benchmarking Universal Single-Copy Orthologs data set. Robust assembly metrics and comparisons with several available anuran transcriptomes and genomes indicate that our cane toad assembly is one of the most complete anuran genomic resources available. This comprehensive anuran transcriptome will provide a valuable resource for investigation of genes under selection during invasion in cane toads, but will also greatly expand our general knowledge of anuran genomes, which are underrepresented in the literature. The data set is publically available in NCBI and GigaDB to serve as a resource for other researchers. © The Authors 2017. Published by Oxford University Press.

Hybrid reduced order modeling for assembly calculations

Energy Technology Data Exchange (ETDEWEB)

Bang, Youngsuk, E-mail: ysbang00@fnctech.com [FNC Technology, Co. Ltd., Yongin-si (Korea, Republic of); Abdel-Khalik, Hany S., E-mail: abdelkhalik@purdue.edu [Purdue University, West Lafayette, IN (United States); Jessee, Matthew A., E-mail: jesseema@ornl.gov [Oak Ridge National Laboratory, Oak Ridge, TN (United States); Mertyurek, Ugur, E-mail: mertyurek@ornl.gov [Oak Ridge National Laboratory, Oak Ridge, TN (United States)

2015-12-15

Highlights: • Reducing computational cost in engineering calculations. • Reduced order modeling algorithm for multi-physics problem like assembly calculation. • Non-intrusive algorithm with random sampling. • Pattern recognition in the components with high sensitive and large variation. - Abstract: While the accuracy of assembly calculations has considerably improved due to the increase in computer power enabling more refined description of the phase space and use of more sophisticated numerical algorithms, the computational cost continues to increase which limits the full utilization of their effectiveness for routine engineering analysis. Reduced order modeling is a mathematical vehicle that scales down the dimensionality of large-scale numerical problems to enable their repeated executions on small computing environment, often available to end users. This is done by capturing the most dominant underlying relationships between the model's inputs and outputs. Previous works demonstrated the use of the reduced order modeling for a single physics code, such as a radiation transport calculation. This manuscript extends those works to coupled code systems as currently employed in assembly calculations. Numerical tests are conducted using realistic SCALE assembly models with resonance self-shielding, neutron transport, and nuclides transmutation/depletion models representing the components of the coupled code system.

Hybrid reduced order modeling for assembly calculations

International Nuclear Information System (INIS)

Bang, Youngsuk; Abdel-Khalik, Hany S.; Jessee, Matthew A.; Mertyurek, Ugur

2015-01-01

Highlights: • Reducing computational cost in engineering calculations. • Reduced order modeling algorithm for multi-physics problem like assembly calculation. • Non-intrusive algorithm with random sampling. • Pattern recognition in the components with high sensitive and large variation. - Abstract: While the accuracy of assembly calculations has considerably improved due to the increase in computer power enabling more refined description of the phase space and use of more sophisticated numerical algorithms, the computational cost continues to increase which limits the full utilization of their effectiveness for routine engineering analysis. Reduced order modeling is a mathematical vehicle that scales down the dimensionality of large-scale numerical problems to enable their repeated executions on small computing environment, often available to end users. This is done by capturing the most dominant underlying relationships between the model's inputs and outputs. Previous works demonstrated the use of the reduced order modeling for a single physics code, such as a radiation transport calculation. This manuscript extends those works to coupled code systems as currently employed in assembly calculations. Numerical tests are conducted using realistic SCALE assembly models with resonance self-shielding, neutron transport, and nuclides transmutation/depletion models representing the components of the coupled code system.

Selection of reference genes for expression analysis of Kumamoto and Portuguese oysters and their hybrid

Science.gov (United States)

Yan, Lulu; Su, Jiaqi; Wang, Zhaoping; Yan, Xiwu; Yu, Ruihai

2017-12-01

Quantitative real-time polymerase chain reaction (qRT-PCR) is a rapid and reliable technique which has been widely used to quantifying gene transcripts (expression analysis). It is also employed for studying heterosis, hybridization breeding and hybrid tolerability of oysters, an ecologically and economically important taxonomic group. For these studies, selection of a suitable set of housekeeping genes as references is crucial for correct interpretation of qRT-PCR data. To identify suitable reference genes for oysters during low temperature and low salinity stresses, we analyzed twelve genes from the gill tissue of Crassostrea sikamea (SS), Crassostrea angulata (AA) and their hybrid (SA), which included three ribosomal genes, 28S ribosomal protein S5 ( RPS5), ribosomal protein L35 ( RPL35), and 60S ribosomal protein L29 ( RPL29); three structural genes, tubulin gamma ( TUBγ), annexin A6 and A7 ( AA6 and AA7); three metabolic pathway genes, ornithine decarboxylase ( OD), glyceraldehyde-3-phosphate dehydrogenase ( GAPDH) and glutathione S-transferase P1 ( GSP); two transcription factors, elongation factor 1 alpha and beta ( EF1α and EF1β); and one protein synthesis gene (ubiquitin ( UBQ). Primers specific for these genes were successfully developed for the three groups of oysters. Three different algorithms, geNorm, NormFinder and BestKeeper, were used to evaluate the expression stability of these candidate genes. BestKeeper program was found to be the most reliable. Based on our analysis, we found that the expression of RPL35 and EF1α was stable under low salinity stress, and the expression of OD, GAPDH and EF1α was stable under low temperature stress in hybrid (SA) oyster; the expression of RPS5 and GAPDH was stable under low salinity stress, and the expression of RPS5, UBQ, GAPDH was stable under low temperature stress in SS oyster; the expression of RPS5, GAPDH, EF1β and AA7 was stable under low salinity stress, and the expression of RPL35, EF1α, GAPDH

HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

Science.gov (United States)

Guo, Fang; Zhao, Qiong; Sheraz, Muhammad; Cheng, Junjun; Qi, Yonghe; Su, Qing; Cuconati, Andrea; Wei, Lai; Du, Yanming; Li, Wenhui; Chang, Jinhong; Guo, Ju-Tao

2017-09-01

Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

Directory of Open Access Journals (Sweden)

Fang Guo

2017-09-01

Full Text Available Hepatitis B virus (HBV core protein assembles viral pre-genomic (pg RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs and sulfamoylbenzamides (SBAs, have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways

Science.gov (United States)

Guo, Fang; Zhao, Qiong; Cheng, Junjun; Qi, Yonghe; Su, Qing; Wei, Lai; Li, Wenhui; Chang, Jinhong

2017-01-01

Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or “empty” capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B. PMID:28945802

Conformational assembly and biological properties of collagen mimetic peptides and their thermally responsive polymer conjugates

Science.gov (United States)

Krishna, Ohm Divyam

2011-12-01

Collagens are one of the most abundant proteins found in body tissues and organs, endowing structural integrity, mechanical strength, and multiple biological functions. Destabilized collagen inside human body leads to various degenerative diseases (ex. osteoarthritis) and ageing. This has continued to motivate the design of synthetic peptides and bio-synthetic polypeptides to closely mimic the native collagens in terms of triple helix structure and stability, potential for higher order assembly, and biological properties. However, the widespread application of de novo collagens has been limited in part by the need for hydroxylated proline in the formation of stable triple helical structures. To address this continued need, a hydroxyproline-free, thermally stable collagen-mimetic peptide (CLP-Cys) was rationally designed via the incorporation of electrostatically stabilized amino acid triplets. CLP-Cys was synthesized via solid phase peptide synthesis. The formation and stability of the triple helical structure were indicated via circular dichroism (CD) experiments and confirmed via differential scanning calorimetry (DSC) results. CLP-Cys also self-assembled into nano-rods and micro-fibrils, as evidenced via a combination of dynamic light scattering and transmission electron microscopy. Given the high thermal stability and its propensity for higher-order assembly, CLP-Cys was further functionalized at both the ends with a thermally responsive polymer, poly(diethylene glycol methyl ether methacrylate), (PDEGMEMA) to synthesize a biohybrid triblock copolymer. The CD results indicated that the triple helical form is retained, the thermal unfolding is sustained and helix to coil transition is reversible in the triblock hybrid context. The LCST of PDEGMEMA homopolymer (26 °C) is increased (to 35 °C) upon conjugation to the hydrophilic collagen peptide domain. Further, a combination of static light scattering, Cryo-SEM, TEM and confocal microscopy elucidated that the

De Novo Construction of Redox Active Proteins.

Science.gov (United States)

Moser, C C; Sheehan, M M; Ennist, N M; Kodali, G; Bialas, C; Englander, M T; Discher, B M; Dutton, P L

2016-01-01

Relatively simple principles can be used to plan and construct de novo proteins that bind redox cofactors and participate in a range of electron-transfer reactions analogous to those seen in natural oxidoreductase proteins. These designed redox proteins are called maquettes. Hydrophobic/hydrophilic binary patterning of heptad repeats of amino acids linked together in a single-chain self-assemble into 4-alpha-helix bundles. These bundles form a robust and adaptable frame for uncovering the default properties of protein embedded cofactors independent of the complexities introduced by generations of natural selection and allow us to better understand what factors can be exploited by man or nature to manipulate the physical chemical properties of these cofactors. Anchoring of redox cofactors such as hemes, light active tetrapyrroles, FeS clusters, and flavins by His and Cys residues allow cofactors to be placed at positions in which electron-tunneling rates between cofactors within or between proteins can be predicted in advance. The modularity of heptad repeat designs facilitates the construction of electron-transfer chains and novel combinations of redox cofactors and new redox cofactor assisted functions. Developing de novo designs that can support cofactor incorporation upon expression in a cell is needed to support a synthetic biology advance that integrates with natural bioenergetic pathways. © 2016 Elsevier Inc. All rights reserved.

A SNP resource for Douglas-fir: de novo transcriptome assembly and SNP detection and validation.

Science.gov (United States)

Howe, Glenn T; Yu, Jianbin; Knaus, Brian; Cronn, Richard; Kolpak, Scott; Dolan, Peter; Lorenz, W Walter; Dean, Jeffrey F D

2013-02-28

Douglas-fir (Pseudotsuga menziesii), one of the most economically and ecologically important tree species in the world, also has one of the largest tree breeding programs. Although the coastal and interior varieties of Douglas-fir (vars. menziesii and glauca) are native to North America, the coastal variety is also widely planted for timber production in Europe, New Zealand, Australia, and Chile. Our main goal was to develop a SNP resource large enough to facilitate genomic selection in Douglas-fir breeding programs. To accomplish this, we developed a 454-based reference transcriptome for coastal Douglas-fir, annotated and evaluated the quality of the reference, identified putative SNPs, and then validated a sample of those SNPs using the Illumina Infinium genotyping platform. We assembled a reference transcriptome consisting of 25,002 isogroups (unique gene models) and 102,623 singletons from 2.76 million 454 and Sanger cDNA sequences from coastal Douglas-fir. We identified 278,979 unique SNPs by mapping the 454 and Sanger sequences to the reference, and by mapping four datasets of Illumina cDNA sequences from multiple seed sources, genotypes, and tissues. The Illumina datasets represented coastal Douglas-fir (64.00 and 13.41 million reads), interior Douglas-fir (80.45 million reads), and a Yakima population similar to interior Douglas-fir (8.99 million reads). We assayed 8067 SNPs on 260 trees using an Illumina Infinium SNP genotyping array. Of these SNPs, 5847 (72.5%) were called successfully and were polymorphic. Based on our validation efficiency, our SNP database may contain as many as ~200,000 true SNPs, and as many as ~69,000 SNPs that could be genotyped at ~20,000 gene loci using an Infinium II array-more SNPs than are needed to use genomic selection in tree breeding programs. Ultimately, these genomic resources will enhance Douglas-fir breeding and allow us to better understand landscape-scale patterns of genetic variation and potential responses to

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.

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.

Hybrid intermediaries

OpenAIRE

Cetorelli, Nicola

2014-01-01

I introduce the concept of hybrid intermediaries: financial conglomerates that control a multiplicity of entity types active in the "assembly line" process of modern financial intermediation, a system that has become known as shadow banking. The complex bank holding companies of today are the best example of hybrid intermediaries, but I argue that financial firms from the "nonbank" space can just as easily evolve into conglomerates with similar organizational structure, thus acquiring the cap...

Mismatch discrimination of lipidated DNA and LNA-probes (LiNAs) in hybridization-controlled liposome assembly

DEFF Research Database (Denmark)

Jakobsen, Ulla; Vogel, Stefan

2016-01-01

Assays for mismatch discrimination and detection of single nucleotide variations by hybridization-controlled assembly of liposomes, which do not require tedious surface chemistry, are versatile for both DNA and RNA targets. We report herein a comprehensive study on different DNA and LNA (locked...... assay in the context of mismatch discrimination and SNP detection are presented. The advantages of membrane-anchored LiNA-probes compared to chemically attached probes on solid nanoparticles (e.g. gold nanoparticles) are described. Key functionalities such as non-covalent attachment of LiNA probes...... without the need for long spacers and the inherent mobility of membrane-anchored probes in lipid-bilayer membranes will be described for several different probe designs....

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

Electrostatic Self-Assembly Enabling Integrated Bulk and Interfacial Sodium Storage in 3D Titania-Graphene Hybrid.

Science.gov (United States)

Xu, Gui-Liang; Xiao, Lisong; Sheng, Tian; Liu, Jianzhao; Hu, Yi-Xin; Ma, Tianyuan; Amine, Rachid; Xie, Yingying; Zhang, Xiaoyi; Liu, Yuzi; Ren, Yang; Sun, Cheng-Jun; Heald, Steve M; Kovacevic, Jasmina; Sehlleier, Yee Hwa; Schulz, Christof; Mattis, Wenjuan Liu; Sun, Shi-Gang; Wiggers, Hartmut; Chen, Zonghai; Amine, Khalil

2018-01-10

Room-temperature sodium-ion batteries have attracted increased attention for energy storage due to the natural abundance of sodium. However, it remains a huge challenge to develop versatile electrode materials with favorable properties, which requires smart structure design and good mechanistic understanding. Herein, we reported a general and scalable approach to synthesize three-dimensional (3D) titania-graphene hybrid via electrostatic-interaction-induced self-assembly. Synchrotron X-ray probe, transmission electron microscopy, and computational modeling revealed that the strong interaction between titania and graphene through comparably strong van der Waals forces not only facilitates bulk Na + intercalation but also enhances the interfacial sodium storage. As a result, the titania-graphene hybrid exhibits exceptional long-term cycle stability up to 5000 cycles, and ultrahigh rate capability up to 20 C for sodium storage. Furthermore, density function theory calculation indicated that the interfacial Li + , K + , Mg 2+, and Al 3+ storage can be enhanced as well. The proposed general strategy opens up new avenues to create versatile materials for advanced battery systems.

Electrostatic Self-Assembly Enabling Integrated Bulk and Interfacial Sodium Storage in 3D Titania-Graphene Hybrid

Energy Technology Data Exchange (ETDEWEB)

Xu, Gui-Liang [Chemical; Xiao, Lisong [Center; Sheng, Tian [Collaborative; Liu, Jianzhao [Chemical; Hu, Yi-Xin [Chemical; Department; Ma, Tianyuan [Chemical; Amine, Rachid [Materials; Xie, Yingying [Chemical; Zhang, Xiaoyi [X-ray Science; Liu, Yuzi [Nanoscience; Ren, Yang [X-ray Science; Sun, Cheng-Jun [X-ray Science; Heald, Steve M. [X-ray Science; Kovacevic, Jasmina [Center; Sehlleier, Yee Hwa [Center; Schulz, Christof [Center; Mattis, Wenjuan Liu [Microvast Power Solutions, 12603; Sun, Shi-Gang [Collaborative; Wiggers, Hartmut [Center; Chen, Zonghai [Chemical; Amine, Khalil [Chemical

2017-12-15

Room temperature sodium-ion batteries have attracted increased attention for energy storage due to the natural abundance of sodium. However, it remains a huge challenge to develop versatile electrode materials with favorable properties, which requires smart structure design and good mechanistic understanding. Herein, we reported a general and scalable approach to synthesize 3D titania-graphene hybrid via electrostatic-interaction-induced self-assembly. Synchrotron X-ray probe, transmission electron microscopy and computational modeling revealed that the strong interaction between Titania and graphene through comparably strong van-der-Waals forces not only facilitates bulk Na+ intercalation but also enhances the interfacial sodium storage. As a result, the titania-graphene hybrid exhibits exceptional long-term cycle stability up to 5000 cycles, and ultrahigh rate capability up to 20 C for sodium storage. Furthermore, density function theory calculation indicated that the interfacial Li+, K+, Mg2+ and Al3+ storage can be enhanced as well. The proposed general strategy opens up new avenues to create versatile materials for advanced battery systems.

Real-Time Observation of Exciton-Phonon Coupling Dynamics in Self-Assembled Hybrid Perovskite Quantum Wells.

Science.gov (United States)

Ni, Limeng; Huynh, Uyen; Cheminal, Alexandre; Thomas, Tudor H; Shivanna, Ravichandran; Hinrichsen, Ture F; Ahmad, Shahab; Sadhanala, Aditya; Rao, Akshay

2017-11-28

Self-assembled hybrid perovskite quantum wells have attracted attention due to their tunable emission properties, ease of fabrication, and device integration. However, the dynamics of excitons in these materials, especially how they couple to phonons, remains an open question. Here, we investigate two widely used materials, namely, butylammonium lead iodide (CH 3 (CH 2 ) 3 NH 3 ) 2 PbI 4 and hexylammonium lead iodide (CH 3 (CH 2 ) 5 NH 3 ) 2 PbI 4 , both of which exhibit broad photoluminescence tails at room temperature. We performed femtosecond vibrational spectroscopy to obtain a real-time picture of the exciton-phonon interaction and directly identified the vibrational modes that couple to excitons. We show that the choice of the organic cation controls which vibrational modes the exciton couples to. In butylammonium lead iodide, excitons dominantly couple to a 100 cm -1 phonon mode, whereas in hexylammonium lead iodide, excitons interact with phonons with frequencies of 88 and 137 cm -1 . Using the determined optical phonon energies, we analyzed photoluminescence broadening mechanisms. At low temperatures (photoluminescence line shape observed in hybrid perovskite quantum wells and provide insights into the mechanism of exciton-phonon coupling in these materials.

Structural design principles for self-assembled coordination polygons and polyhedra.

Science.gov (United States)

Young, Neil J; Hay, Benjamin P

2013-02-18

Strategies for the design of ligands that combine with metal ions to form high-symmetry coordination assemblies are reviewed. Evaluation of crystal structure evidence reveals that prior design approaches, based on the concept of complementary bonding vector angles, fail to predict the majority of known examples. After explaining the reasons for this failure, it is shown how an alternative approach, de novo structure-based design, provides a practical method that predicts a much wider range of component shapes encoded to direct the formation of such assemblies.

De Novo whole genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 from blueberry in Georgia, USA

Science.gov (United States)

This study reports a de novo assembled draft genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 causing blueberry bacterial leaf scorch in Georgia, USA. The BB01 genome is 2,517,579 bp with a G+C content of 51.8% and 2,943 open reading frames (ORFs) and 48 RNA genes....

Self-assembly of silver nanoparticles and bacteriophage

Directory of Open Access Journals (Sweden)

Santi Scibilia

2016-03-01

Full Text Available Biohybrid nanostructured materials, composed of both inorganic nanoparticles and biomolecules, offer prospects for many new applications in extremely diverse fields such as chemistry, physics, engineering, medicine and nanobiotechnology. In the recent years, Phage display technique has been extensively used to generate phage clones displaying surface peptides with functionality towards organic materials. Screening and selection of phage displayed material binding peptides has attracted great interest because of their use for development of hybrid materials with multiple functionalities. Here, we present a self-assembly approach for the construction of hybrid nanostructured networks consisting of M13 P9b phage clone, specific for Pseudomonas aeruginosa, selected by Phage display technology, directly assembled with silver nanoparticles (AgNPs, previously prepared by pulsed laser ablation. These networks are characterized by UV–vis optical spectroscopy, scanning/transmission electron microscopies and Raman spectroscopy. We investigated the influence of different ions and medium pH on self-assembly by evaluating different phage suspension buffers. The assembly of these networks is controlled by electrostatic interactions between the phage pVIII major capsid proteins and the AgNPs. The formation of the AgNPs-phage networks was obtained only in two types of tested buffers at a pH value near the isoelectric point of each pVIII proteins displayed on the surface of the clone. This systematic study allowed to optimize the synthesis procedure to assembly AgNPs and bacteriophage. Such networks find application in the biomedical field of advanced biosensing and targeted gene and drug delivery. Keywords: Phage display, Silver nanoparticles, Self-assembly, Hybrid architecture, Raman spectroscopy

De Novo Assembly and Characterization of the Transcriptome of the Parasitic Weed Dodder Identifies Genes Associated with Plant Parasitism1[C][W][OPEN

Science.gov (United States)

Ranjan, Aashish; Ichihashi, Yasunori; Farhi, Moran; Zumstein, Kristina; Townsley, Brad; David-Schwartz, Rakefet; Sinha, Neelima R.

2014-01-01

Parasitic flowering plants are one of the most destructive agricultural pests and have major impact on crop yields throughout the world. Being dependent on finding a host plant for growth, parasitic plants penetrate their host using specialized organs called haustoria. Haustoria establish vascular connections with the host, which enable the parasite to steal nutrients and water. The underlying molecular and developmental basis of parasitism by plants is largely unknown. In order to investigate the process of parasitism, RNAs from different stages (i.e. seed, seedling, vegetative strand, prehaustoria, haustoria, and flower) were used to de novo assemble and annotate the transcriptome of the obligate plant stem parasite dodder (Cuscuta pentagona). The assembled transcriptome was used to dissect transcriptional dynamics during dodder development and parasitism and identified key gene categories involved in the process of plant parasitism. Host plant infection is accompanied by increased expression of parasite genes underlying transport and transporter categories, response to stress and stimuli, as well as genes encoding enzymes involved in cell wall modifications. By contrast, expression of photosynthetic genes is decreased in the dodder infective stages compared with normal stem. In addition, genes relating to biosynthesis, transport, and response of phytohormones, such as auxin, gibberellins, and strigolactone, were differentially expressed in the dodder infective stages compared with stems and seedlings. This analysis sheds light on the transcriptional changes that accompany plant parasitism and will aid in identifying potential gene targets for use in controlling the infestation of crops by parasitic weeds. PMID:24399359

Hybrid reduced order modeling for assembly calculations

Energy Technology Data Exchange (ETDEWEB)

Bang, Y.; Abdel-Khalik, H. S. [North Carolina State University, Raleigh, NC (United States); Jessee, M. A.; Mertyurek, U. [Oak Ridge National Laboratory, Oak Ridge, TN (United States)

2013-07-01

While the accuracy of assembly calculations has considerably improved due to the increase in computer power enabling more refined description of the phase space and use of more sophisticated numerical algorithms, the computational cost continues to increase which limits the full utilization of their effectiveness for routine engineering analysis. Reduced order modeling is a mathematical vehicle that scales down the dimensionality of large-scale numerical problems to enable their repeated executions on small computing environment, often available to end users. This is done by capturing the most dominant underlying relationships between the model's inputs and outputs. Previous works demonstrated the use of the reduced order modeling for a single physics code, such as a radiation transport calculation. This manuscript extends those works to coupled code systems as currently employed in assembly calculations. Numerical tests are conducted using realistic SCALE assembly models with resonance self-shielding, neutron transport, and nuclides transmutation/depletion models representing the components of the coupled code system. (authors)

Exploratory Topology Modelling of Form-Active Hybrid Structures

DEFF Research Database (Denmark)

Holden Deleuran, Anders; Pauly, Mark; Tamke, Martin

2016-01-01

The development of novel form-active hybrid structures (FAHS) is impeded by a lack of modelling tools that allow for exploratory topology modelling of shaped assemblies. We present a flexible and real-time computational design modelling pipeline developed for the exploratory modelling of FAHS...... that enables designers and engineers to iteratively construct and manipulate form-active hybrid assembly topology on the fly. The pipeline implements Kangaroo2's projection-based methods for modelling hybrid structures consisting of slender beams and cable networks. A selection of design modelling sketches...

Detection of an inversion in the Ty-2 region between S. lycopersicum and S. habrochaites by a combination of de novo genome assembly and BAC cloning.

Science.gov (United States)

Wolters, Anne-Marie A; Caro, Myluska; Dong, Shufang; Finkers, Richard; Gao, Jianchang; Visser, Richard G F; Wang, Xiaoxuan; Du, Yongchen; Bai, Yuling

2015-10-01

A chromosomal inversion associated with the tomato Ty - 2 gene for TYLCV resistance is the cause of severe suppression of recombination in a tomato Ty - 2 introgression line. Among tomato and its wild relatives inversions are often observed, which result in suppression of recombination. Such inversions hamper the transfer of important traits from a related species to the crop by introgression breeding. Suppression of recombination was reported for the TYLCV resistance gene, Ty-2, which has been introgressed in cultivated tomato (Solanum lycopersicum) from the wild relative S. habrochaites accession B6013. Ty-2 was mapped to a 300-kb region on the long arm of chromosome 11. The suppression of recombination in the Ty-2 region could be caused by chromosomal rearrangements in S. habrochaites compared with S. lycopersicum. With the aim of visualizing the genome structure of the Ty-2 region, we compared the draft de novo assembly of S. habrochaites accession LYC4 with the sequence of cultivated tomato ('Heinz'). Furthermore, using populations derived from intraspecific crosses of S. habrochaites accessions, the order of markers in the Ty-2 region was studied. Results showed the presence of an inversion of approximately 200 kb in the Ty-2 region when comparing S. lycopersicum and S. habrochaites. By sequencing a BAC clone from the Ty-2 introgression line, one inversion breakpoint was identified. Finally, the obtained results are discussed with respect to introgression breeding and the importance of a priori de novo sequencing of the species involved.

New lager yeast strains generated by interspecific hybridization.

Science.gov (United States)

Krogerus, Kristoffer; Magalhães, Frederico; Vidgren, Virve; Gibson, Brian

2015-05-01

The interspecific hybrid Saccharomyces pastorianus is the most commonly used yeast in brewery fermentations worldwide. Here, we generated de novo lager yeast hybrids by mating a domesticated and strongly flocculent Saccharomyces cerevisiae ale strain with the Saccharomyces eubayanus type strain. The hybrids were characterized with respect to the parent strains in a wort fermentation performed at temperatures typical for lager brewing (12 °C). The resulting beers were analysed for sugar and aroma compounds, while the yeasts were tested for their flocculation ability and α-glucoside transport capability. These hybrids inherited beneficial properties from both parent strains (cryotolerance, maltotriose utilization and strong flocculation) and showed apparent hybrid vigour, fermenting faster and producing beer with higher alcohol content (5.6 vs 4.5 % ABV) than the parents. Results suggest that interspecific hybridization is suitable for production of novel non-GM lager yeast strains with unique properties and will help in elucidating the evolutionary history of industrial lager yeast.

Pesquisa de novos elementos Pesquisa de novos elementos

Directory of Open Access Journals (Sweden)

Gil Mário de Macedo Grassi

1978-11-01

Full Text Available The present study deals with the discovery of new elements synthesized by man. The introduction discusses in general the theories about nuclear transmutation, which is the method employed in these syntheses. The study shows the importance of the Periodical Table since it is through this table that one can reach a prevision of new elements and its, properties. The discoveries of the transuranic elements, together wich the data of their first preparations are also tabulated The stability of these elements is also discussed, and future speculations are showedNeste trabalho estuda-se, teoricamente, a descoberta de novos elementos sintetizados pelo homem Na introdução apresentamos um apanhado geral sobre as teorias a respeito da transmutação nuclear, que é o método utilizado nestas sínteses. Em seguida, mostramos a importância da Tabela Periódica, pois é através dela que se chega à previsão dos novos elementos e de suas propriedades. As descobertas dos transurânicos, Já realizadas com êxito, juntamente com os dados de suas primeiras preparações são tabelados. A estabilidade destes novos elementos também é discutida, e apresentadas futuras especulações.

Proximity hybridization-regulated catalytic DNA hairpin assembly for electrochemical immunoassay based on in situ DNA template-synthesized Pd nanoparticles

International Nuclear Information System (INIS)

Zhou, Fuyi; Yao, Yao; Luo, Jianjun; Zhang, Xing; Zhang, Yu; Yin, Dengyang; Gao, Fenglei; Wang, Po

2017-01-01

Novel hybridization proximity-regulated catalytic DNA hairpin assembly strategy has been proposed for electrochemical immunoassay based on in situ DNA template-synthesized Pd nanoparticles as signal label. The DNA template-synthesized Pd nanoparticles were characterized with atomic force microscopic and X-ray photoelectron spectroscopy. The highly efficient electrocatalysis by DNA template synthesized Pd nanoparticles for NaBH 4 oxidation produced an intense detection signal. The label-free electrochemical method achieved the detection of carcinoembryonic antigen (CEA) with a linear range from 10 −15 to 10 −11  g mL −1 and a detection limit of 0.43 × 10 −15  g mL −1 . Through introducing a supersandwich reaction to increase the DNA length, the electrochemical signal was further amplified, leading to a detection limit of 0.52 × 10 −16  g mL −1 . And it rendered satisfactory analytical performance for the determination of CEA in serum samples. Furthermore, it exhibited good reproducibility and stability; meanwhile, it also showed excellent specificity due to the specific recognition of antigen by antibody. Therefore, the DNA template synthesized Pd nanoparticles based signal amplification approach has great potential in clinical applications and is also suitable for quantification of biomarkers at ultralow level. - Graphical abstract: A novel label-free and enzyme-free electrochemical immunoassay based on proximity hybridization-regulated catalytic DNA hairpin assemblies for recycling of the CEA. - Highlights: • A novel enzyme-free electrochemical immunosensor was developed for detection of CEA. • The signal amplification was based on catalytic DNA hairpin assembly and DNA-template-synthesized Pd nanoparticles. • The biosensor could detect CEA down to 0.52 × 10 −16  g mL −1 level with a dynamic range spanning 5 orders of magnitude.

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.

De novo assembly and functional annotation of Myrciaria dubia fruit transcriptome reveals multiple metabolic pathways for L-ascorbic acid biosynthesis.

Science.gov (United States)

Castro, Juan C; Maddox, J Dylan; Cobos, Marianela; Requena, David; Zimic, Mirko; Bombarely, Aureliano; Imán, Sixto A; Cerdeira, Luis A; Medina, Andersson E

2015-11-24

Myrciaria dubia is an Amazonian fruit shrub that produces numerous bioactive phytochemicals, but is best known by its high L-ascorbic acid (AsA) content in fruits. Pronounced variation in AsA content has been observed both within and among individuals, but the genetic factors responsible for this variation are largely unknown. The goals of this research, therefore, were to assemble, characterize, and annotate the fruit transcriptome of M. dubia in order to reconstruct metabolic pathways and determine if multiple pathways contribute to AsA biosynthesis. In total 24,551,882 high-quality sequence reads were de novo assembled into 70,048 unigenes (mean length = 1150 bp, N50 = 1775 bp). Assembled sequences were annotated using BLASTX against public databases such as TAIR, GR-protein, FB, MGI, RGD, ZFIN, SGN, WB, TIGR_CMR, and JCVI-CMR with 75.2 % of unigenes having annotations. Of the three core GO annotation categories, biological processes comprised 53.6 % of the total assigned annotations, whereas cellular components and molecular functions comprised 23.3 and 23.1 %, respectively. Based on the KEGG pathway assignment of the functionally annotated transcripts, five metabolic pathways for AsA biosynthesis were identified: animal-like pathway, myo-inositol pathway, L-gulose pathway, D-mannose/L-galactose pathway, and uronic acid pathway. All transcripts coding enzymes involved in the ascorbate-glutathione cycle were also identified. Finally, we used the assembly to identified 6314 genic microsatellites and 23,481 high quality SNPs. This study describes the first next-generation sequencing effort and transcriptome annotation of a non-model Amazonian plant that is relevant for AsA production and other bioactive phytochemicals. Genes encoding key enzymes were successfully identified and metabolic pathways involved in biosynthesis of AsA, anthocyanins, and other metabolic pathways have been reconstructed. The identification of these genes and pathways is in agreement with

Carbon nanotube-templated assembly of regioregular poly(3-alkylthiophene) in solution

Science.gov (United States)

Zhu, Jiahua; Stevens, Eric; He, Youjun; Hong, Kunlun; Ivanov, Ilia

2016-09-01

Control of structural heterogeneity by rationally encoding of the molecular assemblies is a key enabling design of hierarchical, multifunctional materials of the future. Here we report the strategies to gain such control using solution- based assembly to construct a hybrid nano-assembly and a network hybrid structure of regioregular poly(3- alkylthiophene) - carbon nanotube (P3AT-CNT). The opto-electronic performance of conjugated polymer (P3AT) is defined by the structure of the aggregate in solution and in the solid film. Control of P3AT aggregation would allow formation of broad range of morphologies with very distinct electro-optical. We utilize interactive templating to confine the assembly behavior of conjugated polymers, replacing poorly controlled solution processing approach. Perfect crystalline surface of the single-walled and multi-walled carbon nanotube (SWCNT/MWCNT) acts as a template, seeding P3AT aggregation of the surface of the nanotube. The seed continues directional growth through pi-pi stacking leading to the formation of to well-defined P3AT-CNT morphologies, including comb-like nano-assemblies, super- structures and gel networks. Interconnected, highly-branched network structure of P3AT-CNT hybrids is of particular interest to enable efficient, long-range, balanced charge carrier transport. The structure and opto-electionic function of the intermediate assemblies and networks of P3AT/CNT hybrids are characterized by transmission election microscopy and UV-vis absorption.

De novo Assembly and Analysis of the Chilean Pencil Catfish Trichomycterus areolatus Transcriptome

Science.gov (United States)

Schulze, Thomas T.; Ali, Jonathan M.; Bartlett, Maggie L.; McFarland, Madalyn M.; Clement, Emalie J.; Won, Harim I.; Sanford, Austin G.; Monzingo, Elyssa B.; Martens, Matthew C.; Hemsley, Ryan M.; Kumar, Sidharta; Gouin, Nicolas; Kolok, Alan S.; Davis, Paul H.

2016-01-01

Trichomycterus areolatus is an endemic species of pencil catfish that inhabits the riffles and rapids of many freshwater ecosystems of Chile. Despite its unique adaptation to Chile's high gradient watersheds and therefore potential application in the investigation of ecosystem integrity and environmental contamination, relatively little is known regarding the molecular biology of this environmental sentinel. Here, we detail the assembly of the Trichomycterus areolatus transcriptome, a molecular resource for the study of this organism and its molecular response to the environment. RNA-Seq reads were obtained by next-generation sequencing with an Illumina® platform and processed using PRINSEQ. The transcriptome assembly was performed using TRINITY assembler. Transcriptome validation was performed by functional characterization with KOG, KEGG, and GO analyses. Additionally, differential expression analysis highlights sex-specific expression patterns, and a list of endocrine and oxidative stress related transcripts are included. PMID:27672404

DNA-directed self-assembly of gold nanoparticles into binary and ternary nanostructures

International Nuclear Information System (INIS)

Yao Hui; Yi Changqing; Tzang Chihung; Zhu Junjie; Yang Mengsu

2007-01-01

The assembly and characterization of gold nanoparticle-based binary and ternary structures are reported. Two strategies were used to assemble gold nanoparticles into ordered nanoscale architectures: in strategy 1, gold nanoparticles were functionalized with single-strand DNA (ssDNA) first, and then hybridized with complementary ssDNA-labelled nanoparticles to assemble designed architectures. In strategy 2, the designed architectures were constructed through hybridization between complementary ssDNA first, then by assembling gold nanoparticles to the scaffolding through gold-sulfur bonds. Both TEM measurements and agarose gel electrophoresis confirmed that the latter strategy is more efficient in generating the designed nanostructures

Highly active bidirectional electron transfer by a self-assembled electroactive reduced-graphene-oxide-hybridized biofilm.

Science.gov (United States)

Yong, Yang-Chun; Yu, Yang-Yang; Zhang, Xinhai; Song, Hao

2014-04-22

Low extracellular electron transfer performance is often a bottleneck in developing high-performance bioelectrochemical systems. Herein, we show that the self-assembly of graphene oxide and Shewanella oneidensis MR-1 formed an electroactive, reduced-graphene-oxide-hybridized, three-dimensional macroporous biofilm, which enabled highly efficient bidirectional electron transfers between Shewanella and electrodes owing to high biomass incorporation and enhanced direct contact-based extracellular electron transfer. This 3D electroactive biofilm delivered a 25-fold increase in the outward current (oxidation current, electron flux from bacteria to electrodes) and 74-fold increase in the inward current (reduction current, electron flux from electrodes to bacteria) over that of the naturally occurring biofilms. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

De novo assembly and characterization of tissue specific transcriptomes in the emerald notothen, Trematomus bernacchii.

Science.gov (United States)

Huth, Troy J; Place, Sean P

2013-11-20

The notothenioids comprise a diverse group of fishes that rapidly radiated after isolation by the Antarctic Circumpolar Current approximately 14-25 million years ago. Given that evolutionary adaptation has led to finely tuned traits with narrow physiological limits in these organisms, this system provides a unique opportunity to examine physiological trade-offs and limits of adaptive responses to environmental perturbation. As such, notothenioids have a rich history with respect to studies attempting to understand the vulnerability of polar ecosystems to the negative impacts associated with global climate change. Unfortunately, despite being a model system for understanding physiological adaptations to extreme environments, we still lack fundamental molecular tools for much of the Nototheniidae family. Specimens of the emerald notothen, Trematomus bernacchii, were acclimated for 28 days in flow-through seawater tanks maintained near ambient seawater temperatures (-1.5°C) or at +4°C. Following acclimation, tissue specific cDNA libraries for liver, gill and brain were created by pooling RNA from n = 5 individuals per temperature treatment. The tissue specific libraries were bar-coded and used for 454 pyrosequencing, which yielded over 700 thousand sequencing reads. A de novo assembly and annotation of these reads produced a functional transcriptome library of T. bernacchii containing 30,107 unigenes, 13,003 of which possessed significant homology to a known protein product. Digital gene expression analysis of these extremely cold adapted fish reinforced the loss of an inducible heat shock response and allowed the preliminary exploration into other elements of the cellular stress response. Preliminary exploration of the transcriptome of T. bernacchii under elevated temperatures enabled a semi-quantitative comparison to prior studies aimed at characterizing the thermal response of this endemic fish whose size, abundance and distribution has established it as a

Self-assembled hybrid materials based on conjugated polymers and semiconductors nano-crystals for plastic solar cells

International Nuclear Information System (INIS)

Girolamo, J. de

2007-11-01

This work is devoted to the elaboration of self-assembled hybrid materials based on poly(3- hexyl-thiophene) and CdSe nano-crystals for photovoltaic applications. For that, complementary molecular recognition units were introduced as side chain groups on the polymer and at the nano-crystals' surface. Diamino-pyrimidine groups were introduced by post-functionalization of a precursor copolymer, namely poly(3-hexyl-thiophene-co-3- bromo-hexyl-thiophene) whereas thymine groups were introduced at the nano-crystals' surface by a ligand exchange reaction with 1-(6-mercapto-hexyl)thymine. However, due to their different solubility, the mixing of the two components by solution processes is difficult. A 'one-pot' procedure was developed, but this method led to insoluble aggregates without control of the hybrid composition. To overcome the solubility problem, the layer-by-layer method was used to prepare the films. This method allows a precise control of the deposition process. Experimental parameters were tested in order to evaluate their impact on the resulting film. The films morphology was investigated by microscopy and X-Ray diffraction techniques. These analyses reveal an interpenetrated structure of nano-crystals within the polymer matrix rather than a multilayered structure. Electrochemical and spectro electrochemical studies were performed on the hybrid material deposited by the LBL process. Finally the materials were tested in a solar cell configuration and the I=f(V) curves reveals a clear photovoltaic behaviour. (author)

Post-assembly transformations of porphyrin-containing metal-organic framework (MOF) films fabricated via automated layer-by-layer coordination

KAUST Repository

So, Monica; Beyzavi, M. Hassan; Sawhney, Rohan; Shekhah, Osama; Eddaoudi, Mohamed; Al-Juaid, Salih Salem; Hupp, Joseph T.; Farha, Omar K.

2015-01-01

Herein, we demonstrate the robustness of layer-by-layer (LbL)-assembled, pillared-paddlewheel-type MOF films toward conversion to new or modified MOFs via solvent-assisted linker exchange (SALE) and post-assembly linker metalation. Further, we show that LbL synthesis can afford MOFs that have proven inaccessible through other de novo strategies.

InteraqCT Comparison on Assemblies - Reference Measurements

DEFF Research Database (Denmark)

Stolfi, Alessandro; De Chiffre, Leonardo

2016-01-01

The InteraqCT comparison on assemblies is as an activity within the Marie Curie ESR Project INTERAQCT - International Network for the Training of Early stage Researchers on Advanced Quality control by Computed Tomography funded by the European Commission's 7th Framework Programme FP7-PEOPLE - Under...

A reference genome of the European beech (Fagus sylvatica L.).

Science.gov (United States)

Mishra, Bagdevi; Gupta, Deepak K; Pfenninger, Markus; Hickler, Thomas; Langer, Ewald; Nam, Bora; Paule, Juraj; Sharma, Rahul; Ulaszewski, Bartosz; Warmbier, Joanna; Burczyk, Jaroslaw; Thines, Marco

2018-06-01

The European beech is arguably the most important climax broad-leaved tree species in Central Europe, widely planted for its valuable wood. Here, we report the 542 Mb draft genome sequence of an up to 300-year-old individual (Bhaga) from an undisturbed stand in the Kellerwald-Edersee National Park in central Germany. Using a hybrid assembly approach, Illumina reads with short- and long-insert libraries, coupled with long Pacific Biosciences reads, we obtained an assembled genome size of 542 Mb, in line with flow cytometric genome size estimation. The largest scaffold was of 1.15 Mb, the N50 length was 145 kb, and the L50 count was 983. The assembly contained 0.12% of Ns. A Benchmarking with Universal Single-Copy Orthologs (BUSCO) analysis retrieved 94% complete BUSCO genes, well in the range of other high-quality draft genomes of trees. A total of 62,012 protein-coding genes were predicted, assisted by transcriptome sequencing. In addition, we are reporting an efficient method for extracting high-molecular-weight DNA from dormant buds, by which contamination by environmental bacteria and fungi was kept at a minimum. The assembled genome will be a valuable resource and reference for future population genomics studies on the evolution and past climate change adaptation of beech and will be helpful for identifying genes, e.g., involved in drought tolerance, in order to select and breed individuals to adapt forestry to climate change in Europe. A continuously updated genome browser and download page can be accessed from beechgenome.net, which will include future genome versions of the reference individual Bhaga, as new sequencing approaches develop.

NATpipe: an integrative pipeline for systematical discovery of natural antisense transcripts (NATs) and phase-distributed nat-siRNAs from de novo assembled transcriptomes

Science.gov (United States)

Yu, Dongliang; Meng, Yijun; Zuo, Ziwei; Xue, Jie; Wang, Huizhong

2016-01-01

Nat-siRNAs (small interfering RNAs originated from natural antisense transcripts) are a class of functional small RNA (sRNA) species discovered in both plants and animals. These siRNAs are highly enriched within the annealed regions of the NAT (natural antisense transcript) pairs. To date, great research efforts have been taken for systematical identification of the NATs in various organisms. However, developing a freely available and easy-to-use program for NAT prediction is strongly demanded by researchers. Here, we proposed an integrative pipeline named NATpipe for systematical discovery of NATs from de novo assembled transcriptomes. By utilizing sRNA sequencing data, the pipeline also allowed users to search for phase-distributed nat-siRNAs within the perfectly annealed regions of the NAT pairs. Additionally, more reliable nat-siRNA loci could be identified based on degradome sequencing data. A case study on the non-model plant Dendrobium officinale was performed to illustrate the utility of NATpipe. Finally, we hope that NATpipe would be a useful tool for NAT prediction, nat-siRNA discovery, and related functional studies. NATpipe is available at www.bioinfolab.cn/NATpipe/NATpipe.zip. PMID:26858106

Hybrid systems, optimal control and hybrid vehicles theory, methods and applications

CERN Document Server

Böhme, Thomas J

2017-01-01

This book assembles new methods showing the automotive engineer for the first time how hybrid vehicle configurations can be modeled as systems with discrete and continuous controls. These hybrid systems describe naturally and compactly the networks of embedded systems which use elements such as integrators, hysteresis, state-machines and logical rules to describe the evolution of continuous and discrete dynamics and arise inevitably when modeling hybrid electric vehicles. They can throw light on systems which may otherwise be too complex or recondite. Hybrid Systems, Optimal Control and Hybrid Vehicles shows the reader how to formulate and solve control problems which satisfy multiple objectives which may be arbitrary and complex with contradictory influences on fuel consumption, emissions and drivability. The text introduces industrial engineers, postgraduates and researchers to the theory of hybrid optimal control problems. A series of novel algorithmic developments provides tools for solving engineering pr...

Automated de novo phasing and model building of coiled-coil proteins.

Science.gov (United States)

Rämisch, Sebastian; Lizatović, Robert; André, Ingemar

2015-03-01

Models generated by de novo structure prediction can be very useful starting points for molecular replacement for systems where suitable structural homologues cannot be readily identified. Protein-protein complexes and de novo-designed proteins are examples of systems that can be challenging to phase. In this study, the potential of de novo models of protein complexes for use as starting points for molecular replacement is investigated. The approach is demonstrated using homomeric coiled-coil proteins, which are excellent model systems for oligomeric systems. Despite the stereotypical fold of coiled coils, initial phase estimation can be difficult and many structures have to be solved with experimental phasing. A method was developed for automatic structure determination of homomeric coiled coils from X-ray diffraction data. In a benchmark set of 24 coiled coils, ranging from dimers to pentamers with resolutions down to 2.5 Å, 22 systems were automatically solved, 11 of which had previously been solved by experimental phasing. The generated models contained 71-103% of the residues present in the deposited structures, had the correct sequence and had free R values that deviated on average by 0.01 from those of the respective reference structures. The electron-density maps were of sufficient quality that only minor manual editing was necessary to produce final structures. The method, named CCsolve, combines methods for de novo structure prediction, initial phase estimation and automated model building into one pipeline. CCsolve is robust against errors in the initial models and can readily be modified to make use of alternative crystallographic software. The results demonstrate the feasibility of de novo phasing of protein-protein complexes, an approach that could also be employed for other small systems beyond coiled coils.

EAST machine assembly and its measurement system

International Nuclear Information System (INIS)

Wu, S.T.

2005-01-01

The EAST (HT-7U) superconducting tokamak consists of a superconducting poloidal field magnet system, a toroidal field magnet system, a vacuum vessel and in-vessel components, thermal shields and a cryostat vessel. The main parts of the machine have been delivered to ASIPP (Institute of Plasma Physics, Chinese Academy of Sciences) successionally from 2003. For its complicated constitution and precise requirement, a reasonable assembly procedure and measurement technique should be defined carefully. Before the assembly procedure, a reference frame has been set up with reference fiducial targets on the wall of the test hall by an industrial measurement system. After the torus of TF coils is formed, a new reference frame will be set up from the position of the TF torus. The vacuum vessel with all inner parts will be installed with reference of the new reference frame. The big size and mass of components, special configuration of the superconducting machine with tight installation tolerances of the HT-7U (EAST) machine result in complicated assembly procedure. The procedure had begun with the installation of the support frame and the base of cryostat vessel last year. In this paper, the requirements of the assembly precise for some key components of the machine are described. The reference frame for the assembly and maintenance is explained. The assembly procedure is introduced

Gene-enriched draft genome of the cattle tick Rhipicephalus microplus: assembly by the hybrid Pacific Biosciences/Illumina approach enabled analysis of the highly repetitive genome.

Science.gov (United States)

Barrero, Roberto A; Guerrero, Felix D; Black, Michael; McCooke, John; Chapman, Brett; Schilkey, Faye; Pérez de León, Adalberto A; Miller, Robert J; Bruns, Sara; Dobry, Jason; Mikhaylenko, Galina; Stormo, Keith; Bell, Callum; Tao, Quanzhou; Bogden, Robert; Moolhuijzen, Paula M; Hunter, Adam; Bellgard, Matthew I

2017-08-01

The genome of the cattle tick Rhipicephalus microplus, an ectoparasite with global distribution, is estimated to be 7.1Gbp in length and consists of approximately 70% repetitive DNA. We report the draft assembly of a tick genome that utilized a hybrid sequencing and assembly approach to capture the repetitive fractions of the genome. Our hybrid approach produced an assembly consisting of 2.0Gbp represented in 195,170 scaffolds with a N50 of 60,284bp. The Rmi v2.0 assembly is 51.46% repetitive with a large fraction of unclassified repeats, short interspersed elements, long interspersed elements and long terminal repeats. We identified 38,827 putative R. microplus gene loci, of which 24,758 were protein coding genes (≥100 amino acids). OrthoMCL comparative analysis against 11 selected species including insects and vertebrates identified 10,835 and 3,423 protein coding gene loci that are unique to R. microplus or common to both R. microplus and Ixodes scapularis ticks, respectively. We identified 191 microRNA loci, of which 168 have similarity to known miRNAs and 23 represent novel miRNA families. We identified the genomic loci of several highly divergent R. microplus esterases with sequence similarity to acetylcholinesterase. Additionally we report the finding of a novel cytochrome P450 CYP41 homolog that shows similar protein folding structures to known CYP41 proteins known to be involved in acaricide resistance. Copyright © 2017 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

Generation of Hybrid Peptide-Silver Nanoparticles for Antibacterial and Antifouling Applications

KAUST Repository

Seferji, Kholoud

2018-05-01

An alarming increase of antibiotic-resistant bacterial strains has made the demand for novel antibacterial agents, for example, more effective antibiotics, highly crucial. One of the oldest antimicrobial agents is elementary silver which has been used for thousands of years. Even in our days, elementary silver is used for medical purposes, such as for burns, wounds, and microbial infections. We have taken the effectiveness of elementary silver into consideration to generate novel antibacterial and antifouling agents. Our innovative antibacterial agents are hybrid peptide silver nanoparticles (CH-01-AgNPs) that are created de novo and in situ from a silver nitrate solution (AgNO3) in the presence of ultrashort self-assembling peptides compounds. The nucleation of CH-01-AgNPs is initiated by irradiating the peptide solution mixed with the AgNO3 solution using ultraviolet (UV) light at a wavelength of 254 nm, in the absence of any reducing or capping agents. Obviously, the peptide itself serves as the reducing agent. The ultrashort peptides are four amino acids in length with an innate ability to self-assemble into nanofibrous scaffolds. Using these ultrashort peptides CH-01 we were able to create hybrid peptide silver nanoparticles CH-01-AgNPs with a diameter of 4-6 nm. The synthesized CH-01-AgNPs were further characterized using ultraviolet-visible spectroscopy, transmission electron microscopy, dynamic light scattering, and X-ray photoelectron spectroscopy. The antibacterial and antifouling activity of CH-01-AgNPs were then investigated using either gram-negative bacteria, such as antibiotic-resistant Top10 Escherichia coli and Pseudomonas aeruginosa PDO300, or gram-positive bacteria, such as Staphylococcus aureus CECT 976. The hybrid nanoparticles demonstrated very promising antibacterial and antifouling activity with higher antibacterial and antifouling activity as commercial silver nanoparticles. Quantitative Polymerase Chain Reaction (qPCR) results showed

Solid-State Densification of Spun-Cast Self-Assembled Monolayers for Use in Ultra-Thin Hybrid Dielectrics.

Science.gov (United States)

Hutchins, Daniel O; Acton, Orb; Weidner, Tobias; Cernetic, Nathan; Baio, Joe E; Castner, David G; Ma, Hong; Jen, Alex K-Y

2012-11-15

Ultra-thin self-assembled monolayer (SAM)-oxide hybrid dielectrics have gained significant interest for their application in low-voltage organic thin film transistors (OTFTs). A [8-(11-phenoxy-undecyloxy)-octyl]phosphonic acid (PhO-19-PA) SAM on ultrathin AlO x (2.5 nm) has been developed to significantly enhance the dielectric performance of inorganic oxides through reduction of leakage current while maintaining similar capacitance to the underlying oxide structure. Rapid processing of this SAM in ambient conditions is achieved by spin coating, however, as-cast monolayer density is not sufficient for dielectric applications. Thermal annealing of a bulk spun-cast PhO-19-PA molecular film is explored as a mechanism for SAM densification. SAM density, or surface coverage, and order are examined as a function of annealing temperature. These SAM characteristics are probed through atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and near edge X-ray absorption fine structure spectroscopy (NEXAFS). It is found that at temperatures sufficient to melt the as-cast bulk molecular film, SAM densification is achieved; leading to a rapid processing technique for high performance SAM-oxide hybrid dielectric systems utilizing a single wet processing step. To demonstrate low-voltage devices based on this hybrid dielectric (with leakage current density of 7.7×10 -8 A cm -2 and capacitance density of 0.62 µF cm -2 at 3 V), pentacene thin-film transistors (OTFTs) are fabricated and yield sub 2 V operation and charge carrier mobilites of up to 1.1 cm 2 V -1 s -1 .

Hybrid platform. Economical hybrid drive for commercial vehicles; Hybrid Plattform. Wirtschaftlicher Hybridantrieb fuer Nutzfahrzeuge

Energy Technology Data Exchange (ETDEWEB)

Wallner, S.; Lamke, M.; Mohr, M.; Sedlacek, M.; Speck, F.D. [ZF Friedrichshafen AG, Friedrichshafen (Germany)

2011-07-01

Up to now, hybrid systems have been adapted to their specific requirements in the various applications for trucks, buses as well as mobile and building machines. From a technical point of view, this does indeed result in optimized hybrid drives for each single vehicle application, but due to small volumes, such single developments are critical from a business point of view. ZF Friedrichshafen AG is providing a solution to the technical and economical requirements of the cost-sensitive CV segment in the form of a modular CV parallel hybrid platform composed of a hybrid module system, an inverter, a battery system, and a hybrid software integrated into the overall vehicle. Thanks to the intelligent combination of assemblies and the use of as many identical parts as possible, platforms are realized which cover power ranges between 60 and 120 kW, voltage ranges between 350 and 650 V, and battery capacities between 2 and 4 kWh. The dimensions of the platform elements are such that integration into the diverse commercial vehicle applications is made easy. The hybrid software required for the vehicle-specific functions is also configurable for the mentioned CV applications. (orig.)

Assembling and auditing a comprehensive DNA barcode reference library for European marine fishes.

Science.gov (United States)

Oliveira, L M; Knebelsberger, T; Landi, M; Soares, P; Raupach, M J; Costa, F O

2016-12-01

A large-scale comprehensive reference library of DNA barcodes for European marine fishes was assembled, allowing the evaluation of taxonomic uncertainties and species genetic diversity that were otherwise hidden in geographically restricted studies. A total of 4118 DNA barcodes were assigned to 358 species generating 366 Barcode Index Numbers (BIN). Initial examination revealed as much as 141 BIN discordances (more than one species in each BIN). After implementing an auditing and five-grade (A-E) annotation protocol, the number of discordant species BINs was reduced to 44 (13% grade E), while concordant species BINs amounted to 271 (78% grades A and B) and 14 other had insufficient data (grade D). Fifteen species displayed comparatively high intraspecific divergences ranging from 2·6 to 18·5% (grade C), which is biologically paramount information to be considered in fish species monitoring and stock assessment. On balance, this compilation contributed to the detection of 59 European fish species probably in need of taxonomic clarification or re-evaluation. The generalized implementation of an auditing and annotation protocol for reference libraries of DNA barcodes is recommended. © 2016 The Fisheries Society of the British Isles.

de novo'' aneurysms following endovascular procedures

International Nuclear Information System (INIS)

Briganti, F.; Cirillo, S.; Caranci, F.; Esposito, F.; Maiuri, F.

2002-01-01

Two personal cases of ''de novo'' aneurysms of the anterior communicating artery (ACoA) occurring 9 and 4 years, respectively, after endovascular carotid occlusion are described. A review of the 30 reported cases (including our own two) of ''de novo'' aneurysms after occlusion of the major cerebral vessels has shown some features, including a rather long time interval after the endovascular procedure of up to 20-25 years (average 9.6 years), a preferential ACoA (36.3%) and internal carotid artery-posterior communicating artery (ICA-PCoA) (33.3%) location of the ''de novo'' aneurysms, and a 10% rate of multiple aneurysms. These data are compared with those of the group of reported spontaneous ''de novo'' aneurysms after SAH or previous aneurysm clipping. We agree that the frequency of ''de novo'' aneurysms after major-vessel occlusion (two among ten procedures in our series, or 20%) is higher than commonly reported (0 to 11%). For this reason, we suggest that patients who have been submitted to endovascular major-vessel occlusion be followed up for up to 20-25 years after the procedure, using non-invasive imaging studies such as MR angiography and high-resolution CT angiography. On the other hand, periodic digital angiography has a questionable risk-benefit ratio; it may be used when a ''de novo'' aneurysm is detected or suspected on non-invasive studies. The progressive enlargement of the ACoA after carotid occlusion, as described in our case 1, must be considered a radiological finding of risk for ''de novo'' aneurysm formation. (orig.)

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

Enabling complex nanoscale pattern customization using directed self-assembly.

Science.gov (United States)

Doerk, Gregory S; Cheng, Joy Y; Singh, Gurpreet; Rettner, Charles T; Pitera, Jed W; Balakrishnan, Srinivasan; Arellano, Noel; Sanders, Daniel P

2014-12-16

Block copolymer directed self-assembly is an attractive method to fabricate highly uniform nanoscale features for various technological applications, but the dense periodicity of block copolymer features limits the complexity of the resulting patterns and their potential utility. Therefore, customizability of nanoscale patterns has been a long-standing goal for using directed self-assembly in device fabrication. Here we show that a hybrid organic/inorganic chemical pattern serves as a guiding pattern for self-assembly as well as a self-aligned mask for pattern customization through cotransfer of aligned block copolymer features and an inorganic prepattern. As informed by a phenomenological model, deliberate process engineering is implemented to maintain global alignment of block copolymer features over arbitrarily shaped, 'masking' features incorporated into the chemical patterns. These hybrid chemical patterns with embedded customization information enable deterministic, complex two-dimensional nanoscale pattern customization through directed self-assembly.

Hybrid approaches to nanometer-scale patterning: Exploiting tailored intermolecular interactions

International Nuclear Information System (INIS)

Mullen, Thomas J.; Srinivasan, Charan; Shuster, Mitchell J.; Horn, Mark W.; Andrews, Anne M.; Weiss, Paul S.

2008-01-01

In this perspective, we explore hybrid approaches to nanometer-scale patterning, where the precision of molecular self-assembly is combined with the sophistication and fidelity of lithography. Two areas - improving existing lithographic techniques through self-assembly and fabricating chemically patterned surfaces - will be discussed in terms of their advantages, limitations, applications, and future outlook. The creation of such chemical patterns enables new capabilities, including the assembly of biospecific surfaces to be recognized by, and to capture analytes from, complex mixtures. Finally, we speculate on the potential impact and upcoming challenges of these hybrid strategies.

De novo assembly and characterization of the transcriptome, and development of SSR markers in wax gourd (Benicasa hispida.

Directory of Open Access Journals (Sweden)

Biao Jiang

Full Text Available BACKGROUND: Wax gourd is a widely used vegetable of Cucuribtaceae, and also has important medicinal and health values. However, the genomic resources of wax gourd were scarcity, and only a few nucleotide sequences could be obtained in public databases. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we examined transcriptome in wax gourd. More than 44 million of high quality reads were generated from five different tissues of wax gourd using Illumina paired-end sequencing technology. Approximately 4 Gbp data were generated, and de novo assembled into 65,059 unigenes, with an N50 of 1,132 bp. Based on sequence similarity search with known protein database, 36,070 (55.4% showed significant similarity to known proteins in Nr database, and 24,969 (38.4% had BLAST hits in Swiss-Prot database. Among the annotated unigenes, 14,994 of wax gourd unigenes were assigned to GO term annotation, and 23,977 were found to have COG classifications. In addition, a total of 18,713 unigenes were assigned to 281 KEGG pathways. Furthermore, 6,242 microsatellites (simple sequence repeats were detected as potential molecular markers in wax gourd. Two hundred primer pairs for SSRs were designed for validation of the amplification and polymorphism. The result showed that 170 of the 200 primer pairs were successfully amplified and 49 (28.8% of them exhibited polymorphisms. CONCLUSION/SIGNIFICANCE: Our study enriches the genomic resources of wax gourd and provides powerful information for future studies. The availability of this ample amount of information about the transcriptome and SSRs in wax gourd could serve as valuable basis for studies on the physiology, biochemistry, molecular genetics and molecular breeding of this important vegetable crop.

Alternative normalization methods demonstrate widespread cortical hypometabolism in untreated de novo Parkinson's disease

DEFF Research Database (Denmark)

Berti, Valentina; Polito, C; Borghammer, Per

2012-01-01

, recent studies suggested that conventional data normalization procedures may not always be valid, and demonstrated that alternative normalization strategies better allow detection of low magnitude changes. We hypothesized that these alternative normalization procedures would disclose more widespread...... metabolic alterations in de novo PD. METHODS: [18F]FDG PET scans of 26 untreated de novo PD patients (Hoehn & Yahr stage I-II) and 21 age-matched controls were compared using voxel-based analysis. Normalization was performed using gray matter (GM), white matter (WM) reference regions and Yakushev...... normalization. RESULTS: Compared to GM normalization, WM and Yakushev normalization procedures disclosed much larger cortical regions of relative hypometabolism in the PD group with extensive involvement of frontal and parieto-temporal-occipital cortices, and several subcortical structures. Furthermore...

De novo analysis of transcriptome dynamics in the migratory locust during the development of phase traits.

Directory of Open Access Journals (Sweden)

Shuang Chen

Full Text Available Locusts exhibit remarkable density-dependent phenotype (phase changes from the solitary to the gregarious, making them one of the most destructive agricultural pests. This phenotype polyphenism arises from a single genome and diverse transcriptomes in different conditions. Here we report a de novo transcriptome for the migratory locust and a comprehensive, representative core gene set. We carried out assembly of 21.5 Gb Illumina reads, generated 72,977 transcripts with N50 2,275 bp and identified 11,490 locust protein-coding genes. Comparative genomics analysis with eight other sequenced insects was carried out to identify the genomic divergence between hemimetabolous and holometabolous insects for the first time and 18 genes relevant to development was found. We further utilized the quantitative feature of RNA-seq to measure and compare gene expression among libraries. We first discovered how divergence in gene expression between two phases progresses as locusts develop and identified 242 transcripts as candidates for phase marker genes. Together with the detailed analysis of deep sequencing data of the 4(th instar, we discovered a phase-dependent divergence of biological investment in the molecular level. Solitary locusts have higher activity in biosynthetic pathways while gregarious locusts show higher activity in environmental interaction, in which genes and pathways associated with regulation of neurotransmitter activities, such as neurotransmitter receptors, synthetase, transporters, and GPCR signaling pathways, are strongly involved. Our study, as the largest de novo transcriptome to date, with optimization of sequencing and assembly strategy, can further facilitate the application of de novo transcriptome. The locust transcriptome enriches genetic resources for hemimetabolous insects and our understanding of the origin of insect metamorphosis. Most importantly, we identified genes and pathways that might be involved in locust development

Proximity hybridization-regulated catalytic DNA hairpin assembly for electrochemical immunoassay based on in situ DNA template-synthesized Pd nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Zhou, Fuyi [School of Chemistry and Chemical Engineering, Jiangsu Normal University, Xuzhou 221116 (China); Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, Department of Pharmaceutical Analysis, School of Pharmacy, Xuzhou Medical College, 221004, Xuzhou (China); Yao, Yao; Luo, Jianjun; Zhang, Xing; Zhang, Yu; Yin, Dengyang [Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, Department of Pharmaceutical Analysis, School of Pharmacy, Xuzhou Medical College, 221004, Xuzhou (China); Gao, Fenglei, E-mail: jsxzgfl@sina.com [Jiangsu Key Laboratory of New Drug Research and Clinical Pharmacy, Department of Pharmaceutical Analysis, School of Pharmacy, Xuzhou Medical College, 221004, Xuzhou (China); Wang, Po, E-mail: wangpo@jsnu.edu.cn [School of Chemistry and Chemical Engineering, Jiangsu Normal University, Xuzhou 221116 (China)

2017-05-29

Novel hybridization proximity-regulated catalytic DNA hairpin assembly strategy has been proposed for electrochemical immunoassay based on in situ DNA template-synthesized Pd nanoparticles as signal label. The DNA template-synthesized Pd nanoparticles were characterized with atomic force microscopic and X-ray photoelectron spectroscopy. The highly efficient electrocatalysis by DNA template synthesized Pd nanoparticles for NaBH{sub 4} oxidation produced an intense detection signal. The label-free electrochemical method achieved the detection of carcinoembryonic antigen (CEA) with a linear range from 10{sup −15} to 10{sup −11} g mL{sup −1} and a detection limit of 0.43 × 10{sup −15} g mL{sup −1}. Through introducing a supersandwich reaction to increase the DNA length, the electrochemical signal was further amplified, leading to a detection limit of 0.52 × 10{sup −16} g mL{sup −1}. And it rendered satisfactory analytical performance for the determination of CEA in serum samples. Furthermore, it exhibited good reproducibility and stability; meanwhile, it also showed excellent specificity due to the specific recognition of antigen by antibody. Therefore, the DNA template synthesized Pd nanoparticles based signal amplification approach has great potential in clinical applications and is also suitable for quantification of biomarkers at ultralow level. - Graphical abstract: A novel label-free and enzyme-free electrochemical immunoassay based on proximity hybridization-regulated catalytic DNA hairpin assemblies for recycling of the CEA. - Highlights: • A novel enzyme-free electrochemical immunosensor was developed for detection of CEA. • The signal amplification was based on catalytic DNA hairpin assembly and DNA-template-synthesized Pd nanoparticles. • The biosensor could detect CEA down to 0.52 × 10{sup −16} g mL{sup −1} level with a dynamic range spanning 5 orders of magnitude.

Novel brewing yeast hybrids: creation and application.

Science.gov (United States)

Krogerus, Kristoffer; Magalhães, Frederico; Vidgren, Virve; Gibson, Brian

2017-01-01

The natural interspecies Saccharomyces cerevisiae × Saccharomyces eubayanus hybrid yeast is responsible for global lager beer production and is one of the most important industrial microorganisms. Its success in the lager brewing environment is due to a combination of traits not commonly found in pure yeast species, principally low-temperature tolerance, and maltotriose utilization. Parental transgression is typical of hybrid organisms and has been exploited previously for, e.g., the production of wine yeast with beneficial properties. The parental strain S. eubayanus has only been discovered recently and newly created lager yeast strains have not yet been applied industrially. A number of reports attest to the feasibility of this approach and artificially created hybrids are likely to have a significant impact on the future of lager brewing. De novo S. cerevisiae × S. eubayanus hybrids outperform their parent strains in a number of respects, including, but not restricted to, fermentation rate, sugar utilization, stress tolerance, and aroma formation. Hybrid genome function and stability, as well as different techniques for generating hybrids and their relative merits are discussed. Hybridization not only offers the possibility of generating novel non-GM brewing yeast strains with unique properties, but is expected to aid in unraveling the complex evolutionary history of industrial lager yeast.

Hybrid molecule/superconductor assemblies

International Nuclear Information System (INIS)

McDevitt, J.T.; Haupt, S.G.; Riley, D.R.; Zhao, J.; Zhou, J.P., Jones, C.

1993-01-01

The fabrication of electronic devices from molecular materials has attracted much attention recently. Schottky diodes, molecular transistors, metal-insulator-semiconductor diodes, MIS field effect transistors and light emitting diodes have all been prepared utilizing such substances. The active elements in these devices have been constructed by depositing the molecular phase onto the surface of a metal, semiconductor or insulating substrate. With the recent discovery of high temperature superconductivity, new opportunities now exist for the study of molecule/superconductor interactions as well as for the construction of novel hybrid molecule/superconductor devices. In this paper, methods for preparing the initial two composite molecule/semiconductor devices will be reported. Consequently, light sensors based on dye-coated superconductor junctions as well as molecular switches fashioned from conductive polymer coated superconductor junctions as well as molecular switches fashioned from conductive polymer coated superconductor microbridges will be discussed. Moreover, molecule/superconductor energy and electron transfer phenomena will be illustrated also for the first time

Human Artificial Chromosomes with Alpha Satellite-Based De Novo Centromeres Show Increased Frequency of Nondisjunction and Anaphase Lag

OpenAIRE

Rudd, M. Katharine; Mays, Robert W.; Schwartz, Stuart; Willard, Huntington F.

2003-01-01

Human artificial chromosomes have been used to model requirements for human chromosome segregation and to explore the nature of sequences competent for centromere function. Normal human centromeres require specialized chromatin that consists of alpha satellite DNA complexed with epigenetically modified histones and centromere-specific proteins. While several types of alpha satellite DNA have been used to assemble de novo centromeres in artificial chromosome assays, the extent to which they fu...

A de novo designed monomeric, compact three helix bundle protein on a carbohydrate template

DEFF Research Database (Denmark)

Malik, Leila; Nygård, Jesper; Christensen, Niels Johan

2015-01-01

De novo design and chemical synthesis of proteins and of other artificial structures, which mimic them, is a central strategy for understanding protein folding and for accessing proteins with novel functions. We have previously described carbohydrates as templates for the assembly of artificial...... the template could facilitate protein folding. Here we report the design and synthesis of 3-helix bundle carboproteins on deoxy-hexopyranosides. The carboproteins were analyzed by CD, AUC, SAXS, and NMR, which revealed the formation of the first compact, and folded monomeric carboprotein distinctly different...

Leaf Transcriptome Sequencing for Identifying Genic-SSR Markers and SNP Heterozygosity in Crossbred Mango Variety ‘Amrapali’ (Mangifera indica L.)

Science.gov (United States)

Mahato, Ajay Kumar; Sharma, Nimisha; Singh, Akshay; Srivastav, Manish; Jaiprakash; Singh, Sanjay Kumar; Singh, Anand Kumar; Sharma, Tilak Raj; Singh, Nagendra Kumar

2016-01-01

Mango (Mangifera indica L.) is called “king of fruits” due to its sweetness, richness of taste, diversity, large production volume and a variety of end usage. Despite its huge economic importance genomic resources in mango are scarce and genetics of useful horticultural traits are poorly understood. Here we generated deep coverage leaf RNA sequence data for mango parental varieties ‘Neelam’, ‘Dashehari’ and their hybrid ‘Amrapali’ using next generation sequencing technologies. De-novo sequence assembly generated 27,528, 20,771 and 35,182 transcripts for the three genotypes, respectively. The transcripts were further assembled into a non-redundant set of 70,057 unigenes that were used for SSR and SNP identification and annotation. Total 5,465 SSR loci were identified in 4,912 unigenes with 288 type I SSR (n ≥ 20 bp). One hundred type I SSR markers were randomly selected of which 43 yielded PCR amplicons of expected size in the first round of validation and were designated as validated genic-SSR markers. Further, 22,306 SNPs were identified by aligning high quality sequence reads of the three mango varieties to the reference unigene set, revealing significantly enhanced SNP heterozygosity in the hybrid Amrapali. The present study on leaf RNA sequencing of mango varieties and their hybrid provides useful genomic resource for genetic improvement of mango. PMID:27736892

Leaf Transcriptome Sequencing for Identifying Genic-SSR Markers and SNP Heterozygosity in Crossbred Mango Variety 'Amrapali' (Mangifera indica L.).

Science.gov (United States)

Mahato, Ajay Kumar; Sharma, Nimisha; Singh, Akshay; Srivastav, Manish; Jaiprakash; Singh, Sanjay Kumar; Singh, Anand Kumar; Sharma, Tilak Raj; Singh, Nagendra Kumar

2016-01-01

Mango (Mangifera indica L.) is called "king of fruits" due to its sweetness, richness of taste, diversity, large production volume and a variety of end usage. Despite its huge economic importance genomic resources in mango are scarce and genetics of useful horticultural traits are poorly understood. Here we generated deep coverage leaf RNA sequence data for mango parental varieties 'Neelam', 'Dashehari' and their hybrid 'Amrapali' using next generation sequencing technologies. De-novo sequence assembly generated 27,528, 20,771 and 35,182 transcripts for the three genotypes, respectively. The transcripts were further assembled into a non-redundant set of 70,057 unigenes that were used for SSR and SNP identification and annotation. Total 5,465 SSR loci were identified in 4,912 unigenes with 288 type I SSR (n ≥ 20 bp). One hundred type I SSR markers were randomly selected of which 43 yielded PCR amplicons of expected size in the first round of validation and were designated as validated genic-SSR markers. Further, 22,306 SNPs were identified by aligning high quality sequence reads of the three mango varieties to the reference unigene set, revealing significantly enhanced SNP heterozygosity in the hybrid Amrapali. The present study on leaf RNA sequencing of mango varieties and their hybrid provides useful genomic resource for genetic improvement of mango.

Solid-state densification of spun-cast self-assembled monolayers for use in ultra-thin hybrid dielectrics

Energy Technology Data Exchange (ETDEWEB)

Hutchins, Daniel O.; Acton, Orb [Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195 (United States); Weidner, Tobias [Department of Bioengineering, University of Washington, Seattle, WA 98195 (United States); Cernetic, Nathan [Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195 (United States); Baio, Joe E. [Department of Chemical Engineering, University of Washington, Seattle, WA 98195 (United States); Castner, David G. [Department of Bioengineering, University of Washington, Seattle, WA 98195 (United States); Department of Chemical Engineering, University of Washington, Seattle, WA 98195 (United States); Ma, Hong, E-mail: hma@uw.edu [Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195 (United States); Jen, Alex K.-Y., E-mail: ajen@uw.edu [Department of Materials Science and Engineering, University of Washington, Seattle, WA 98195 (United States); Department of Chemistry, University of Washington, Seattle, WA 98195 (United States)

2012-11-15

Highlights: Black-Right-Pointing-Pointer Rapid processing of SAM in ambient conditions is achieved by spin coating. Black-Right-Pointing-Pointer Thermal annealing of a bulk spun-cast molecular film is explored as a mechanism for SAM densification. Black-Right-Pointing-Pointer High-performance SAM-oxide hybrid dielectric is obtained utilizing a single wet processing step. - Abstract: Ultra-thin self-assembled monolayer (SAM)-oxide hybrid dielectrics have gained significant interest for their application in low-voltage organic thin film transistors (OTFTs). A [8-(11-phenoxy-undecyloxy)-octyl]phosphonic acid (PhO-19-PA) SAM on ultrathin AlO{sub x} (2.5 nm) has been developed to significantly enhance the dielectric performance of inorganic oxides through reduction of leakage current while maintaining similar capacitance to the underlying oxide structure. Rapid processing of this SAM in ambient conditions is achieved by spin coating, however, as-cast monolayer density is not sufficient for dielectric applications. Thermal annealing of a bulk spun-cast PhO-19-PA molecular film is explored as a mechanism for SAM densification. SAM density, or surface coverage, and order are examined as a function of annealing temperature. These SAM characteristics are probed through atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and near edge X-ray absorption fine structure spectroscopy (NEXAFS). It is found that at temperatures sufficient to melt the as-cast bulk molecular film, SAM densification is achieved; leading to a rapid processing technique for high performance SAM-oxide hybrid dielectric systems utilizing a single wet processing step. To demonstrate low-voltage devices based on this hybrid dielectric (with leakage current density of 7.7 Multiplication-Sign 10{sup -8} A cm{sup -2} and capacitance density of 0.62 {mu}F cm{sup -2} at 3 V), pentacene thin-film transistors (OTFTs) are fabricated and yield sub 2 V operation and charge carrier mobilites of up to

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.

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/.

Reference-free SNP discovery for the Eurasian beaver from restriction site-associated DNA paired-end data.

Science.gov (United States)

Senn, Helen; Ogden, Rob; Cezard, Timothee; Gharbi, Karim; Iqbal, Zamin; Johnson, Eric; Kamps-Hughes, Nick; Rosell, Frank; McEwing, Ross

2013-06-01

In this study, we used restriction site-associated DNA (RAD) sequencing to discover SNP markers suitable for population genetic and parentage analysis with the aim of using them for monitoring the reintroduction of the Eurasian beaver (Castor fibre) to Scotland. In the absence of a reference genome for beaver, we built contigs and discovered SNPs within them using paired-end RAD data, so as to have sufficient flanking region around the SNPs to conduct marker design. To do this, we used a simple pipeline which catalogued the Read 1 data in stacks and then used the assembler cortex_var to conduct de novo assembly and genotyping of multiple samples using the Read 2 data. The analysis of around 1.1 billion short reads of sequence data was reduced to a set of 2579 high-quality candidate SNP markers that were polymorphic in Norwegian and Bavarian beaver. Both laboratory validation of a subset of eight of the SNPs (1.3% error) and internal validation by confirming patterns of Mendelian inheritance in a family group (0.9% error) confirmed the success of this approach. © 2013 John Wiley & Sons Ltd.

De novo malignancy after pancreas transplantation in Japan.

Science.gov (United States)

Tomimaru, Y; Ito, T; Marubashi, S; Kawamoto, K; Tomokuni, A; Asaoka, T; Wada, H; Eguchi, H; Mori, M; Doki, Y; Nagano, H

2015-04-01

Long-term immunosuppression is associated with an increased risk of cancer. Especially, the immunosuppression in pancreas transplantation is more intensive than that in other organ transplantation because of its strong immunogenicity. Therefore, it suggests that the risk of post-transplant de novo malignancy might increase in pancreas transplantation. However, there have been few studies of de novo malignancy after pancreas transplantation. The aim of this study was to analyze the incidence of de novo malignancy after pancreas transplantation in Japan. Post-transplant patients with de novo malignancy were surveyed and characterized in Japan. Among 107 cases receiving pancreas transplantation in Japan between 2001 and 2010, de novo malignancy developed in 9 cases (8.4%): post-transplant lymphoproliferative disorders in 6 cases, colon cancer in 1 case, renal cancer in 1 case, and brain tumor in 1 case. We clarified the incidence of de novo malignancy after pancreas transplantation in Japan. Copyright © 2015 Elsevier Inc. All rights reserved.

Analysis of SNP rs16754 of WT1 gene in a series of de novo acute myeloid leukemia patients.

Science.gov (United States)

Luna, Irene; Such, Esperanza; Cervera, Jose; Barragán, Eva; Jiménez-Velasco, Antonio; Dolz, Sandra; Ibáñez, Mariam; Gómez-Seguí, Inés; López-Pavía, María; Llop, Marta; Fuster, Óscar; Oltra, Silvestre; Moscardó, Federico; Martínez-Cuadrón, David; Senent, M Leonor; Gascón, Adriana; Montesinos, Pau; Martín, Guillermo; Bolufer, Pascual; Sanz, Miguel A

2012-12-01

The single nucleotide polymorphism (SNP) rs16754 of the WT1 gene has been previously described as a possible prognostic marker in normal karyotype acute myeloid leukemia (AML) patients. Nevertheless, the findings in this field are not always reproducible in different series. One hundred and seventy-five adult de novo AML patients were screened with two different methods for the detection of SNP rs16754: high-resolution melting (HRM) and FRET hybridization probes. Direct sequencing was used to validate both techniques. The SNP was detected in 52 out of 175 patients (30 %), both by HRM and hybridization probes. Direct sequencing confirmed that every positive sample in the screening methods had a variation in the DNA sequence. Patients with the wild-type genotype (WT1(AA)) for the SNP rs16754 were significantly younger than those with the heterozygous WT1(AG) genotype. No other difference was observed for baseline characteristic or outcome between patients with or without the SNP. Both techniques are equally reliable and reproducible as screening methods for the detection of the SNP rs16754, allowing for the selection of those samples that will need to be sequenced. We were unable to confirm the suggested favorable outcome of SNP rs16754 in de novo AML.

Aqueous cholesteric liquid crystals using uncharged rodlike polypeptides. Polypeptide vesicles by conformation-specific assembly. Ordered chiral macroporous hybrid silica-polypeptide composites

Science.gov (United States)

Bellomo, Enrico Giuseppe

2005-07-01

Aqueous cholesteric liquid crystals using uncharged rodlike polypeptides . The aqueous, lyotropic liquid-crystalline phase behavior of an alpha helical polypeptide, has been studied using optical microscopy and X-ray scattering. Solutions of optically pure polypeptide were found to form cholesteric liquid crystals at volume fractions that decreased with increasing average chain length. At very high volume fractions, the formation of a hexagonal mesophase was observed. The pitch of the cholesteric phase could be varied by a mixture of enantiomeric samples, where the pitch increased as the mixture approached equimolar. The cholesteric phases could be untwisted, using either magnetic field or shear flow, into nematic phases, which relaxed into cholesterics upon removal of field or shear. We have found that the phase diagram of this polypeptide in aqueous solution parallels that of poly(gamma-benzyl glutamate) in organic solvents, thus providing a useful system for liquid-crystal applications requiring water as solvent. Polypeptide vesicles by conformation-specific assembly. We have found that block copolymers composed of polypeptide segments provide significant advantages in controlling both the function and supramolecular structure of bioinspired self-assemblies. Incorporation of the stable chain conformations found in proteins into block copolymers was found to provide an additional element of control, beyond amphiphilicity and composition that defines self-assembled architecture. The abundance of functionality present in amino acids, and the ease by which they can be incorporated into these materials, also provides a powerful mechanism to impart block copolypeptides with function. This combination of structure and function work synergistically to enable significant advantages in the preparation of therapeutic agents as well as provide insight into design of self-assemblies beginning to approach the complexity of natural structures such as virus capsids. Ordered

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.

Nuclear reactor fuel assembly

International Nuclear Information System (INIS)

Marmonier, Pierre; Mesnage, Bernard; Nervi, J.C.

1975-01-01

This invention refers to fuel assemblies for a liquid metal cooled fast neutron reactor. Each assembly is composed of a hollow vertical casing, of regular polygonal section, containing a bundle of clad pins filled with a fissile or fertile substance. The casing is open at its upper end and has a cylindrical foot at its lower end for positioning the assembly in a housing provided in the horizontal diagrid, on which the core assembly rests. A set of flat bars located on the external surface of the casing enables it to be correctly orientated in its housing among the other core assemblies [fr

A Theoretical and Experimental Study of DNA Self-assembly

Science.gov (United States)

Chandran, Harish

providing detailed designs for local molecular computations that involve spatially contiguous molecules arranged on addressable substrates via enzyme-free DNA hybridization reaction cascades. We use the Visual DSD simulation software in conjunction with localized reaction rates obtained from biophysical modeling to create chemical reaction networks of localized hybridization circuits that are then model checked using the PRISM model checking software. We develop a DNA detection system employing the triggered self-assembly of a novel DNA dendritic nanostructure. Detection begins when a specific, single-stranded target DNA strand triggers a hybridization chain reaction between two distinct DNA hairpins. Each hairpin opens and hybridizes up to two copies of the other, and hence each layer of the growing dendritic nanostructure can in principle accommodate an exponentially increasing number of cognate molecules, generating a nanostructure with high molecular weight. We build linear activatable assemblies employing a novel protection/deprotection strategy to strictly enforce the direction of tiling assembly growth to ensure the robustness of the assembly process. Our system consists of two tiles that can form a linear co-polymer. These tiles, which are initially protected such that they do not react with each other, can be activated to form linear co-polymers via the use of a strand displacing enzyme.

KrillDB: A de novo transcriptome database for the Antarctic krill (Euphausia superba.

Directory of Open Access Journals (Sweden)

Gabriele Sales

Full Text Available Antarctic krill (Euphausia superba is a key species in the Southern Ocean with an estimated biomass between 100 and 500 million tonnes. Changes in krill population viability would have catastrophic effect on the Antarctic ecosystem. One looming threat due to elevated levels of anthropogenic atmospheric carbon dioxide (CO2 is ocean acidification (lowering of sea water pH by CO2 dissolving into the oceans. The genetics of Antarctic krill has long been of scientific interest for both for the analysis of population structure and analysis of functional genetics. However, the genetic resources available for the species are relatively modest. We have developed the most advanced genetic database on Euphausia superba, KrillDB, which includes comprehensive data sets of former and present transcriptome projects. In particular, we have built a de novo transcriptome assembly using more than 360 million Illumina sequence reads generated from larval krill including individuals subjected to different CO2 levels. The database gives access to: 1 the full list of assembled genes and transcripts; 2 their level of similarity to transcripts and proteins from other species; 3 the predicted protein domains contained within each transcript; 4 their predicted GO terms; 5 the level of expression of each transcript in the different larval stages and CO2 treatments. All references to external entities (sequences, domains, GO terms are equipped with a link to the appropriate source database. Moreover, the software implements a full-text search engine that makes it possible to submit free-form queries. KrillDB represents the first large-scale attempt at classifying and annotating the full krill transcriptome. For this reason, we believe it will constitute a cornerstone of future approaches devoted to physiological and molecular study of this key species in the Southern Ocean food web.

The plus-hybrid effect on the grain yield of two ZP maize hybrids

Directory of Open Access Journals (Sweden)

Božinović Sofija

2010-01-01

Full Text Available The combined effect of cytoplasmic male sterility and xenia on maize hybrid traits is referred to as the plus-hybrid effect. Two studied ZP hybrids differently responded to this effect for grain yield. All plus-hybrid combinations of the firstly observed hybrid had a higher yield than their fertile counterparts, but not significantly, while only one combination of the second hybrid positively responded, also without statistical significance. It seems that the observed effect mostly depended on the genotype of the female component.

Hybrid microcircuit technology handbook materials, processes, design, testing and production

CERN Document Server

Licari, James J

1998-01-01

The Hybrid Microcircuit Technology Handbook integrates the many diverse technologies used in the design, fabrication, assembly, and testing of hybrid segments crucial to the success of producing reliable circuits in high yields. Among these are: resistor trimming, wire bonding, die attachment, cleaning, hermetic sealing, and moisture analysis. In addition to thin films, thick films, and assembly processes, important chapters on substrate selections, handling (including electrostatic discharge), failure analysis, and documentation are included. A comprehensive chapter of design guidelines will

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

Origins of De Novo Genes in Human and Chimpanzee.

Science.gov (United States)

Ruiz-Orera, Jorge; Hernandez-Rodriguez, Jessica; Chiva, Cristina; Sabidó, Eduard; Kondova, Ivanela; Bontrop, Ronald; Marqués-Bonet, Tomàs; Albà, M Mar

2015-12-01

The birth of new genes is an important motor of evolutionary innovation. Whereas many new genes arise by gene duplication, others originate at genomic regions that did not contain any genes or gene copies. Some of these newly expressed genes may acquire coding or non-coding functions and be preserved by natural selection. However, it is yet unclear which is the prevalence and underlying mechanisms of de novo gene emergence. In order to obtain a comprehensive view of this process, we have performed in-depth sequencing of the transcriptomes of four mammalian species--human, chimpanzee, macaque, and mouse--and subsequently compared the assembled transcripts and the corresponding syntenic genomic regions. This has resulted in the identification of over five thousand new multiexonic transcriptional events in human and/or chimpanzee that are not observed in the rest of species. Using comparative genomics, we show that the expression of these transcripts is associated with the gain of regulatory motifs upstream of the transcription start site (TSS) and of U1 snRNP sites downstream of the TSS. In general, these transcripts show little evidence of purifying selection, suggesting that many of them are not functional. However, we find signatures of selection in a subset of de novo genes which have evidence of protein translation. Taken together, the data support a model in which frequently-occurring new transcriptional events in the genome provide the raw material for the evolution of new proteins.

Inheritance of brewing-relevant phenotypes in constructed Saccharomyces cerevisiae × Saccharomyces eubayanus hybrids.

Science.gov (United States)

Krogerus, Kristoffer; Seppänen-Laakso, Tuulikki; Castillo, Sandra; Gibson, Brian

2017-04-21

Interspecific hybridization has proven to be a potentially valuable technique for generating de novo lager yeast strains that possess diverse and improved traits compared to their parent strains. To further enhance the value of hybridization for strain development, it would be desirable to combine phenotypic traits from more than two parent strains, as well as remove unwanted traits from hybrids. One such trait, that has limited the industrial use of de novo lager yeast hybrids, is their inherent tendency to produce phenolic off-flavours; an undesirable trait inherited from the Saccharomyces eubayanus parent. Trait removal and the addition of traits from a third strain could be achieved through sporulation and meiotic recombination or further mating. However, interspecies hybrids tend to be sterile, which impedes this opportunity. Here we generated a set of five hybrids from three different parent strains, two of which contained DNA from all three parent strains. These hybrids were constructed with fertile allotetraploid intermediates, which were capable of efficient sporulation. We used these eight brewing strains to examine two brewing-relevant phenotypes: stress tolerance and phenolic off-flavour formation. Lipidomics and multivariate analysis revealed links between several lipid species and the ability to ferment in low temperatures and high ethanol concentrations. Unsaturated fatty acids, such as oleic acid, and ergosterol were shown to positively influence growth at high ethanol concentrations. The ability to produce phenolic off-flavours was also successfully removed from one of the hybrids, Hybrid T2, through meiotic segregation. The potential application of these strains in industrial fermentations was demonstrated in wort fermentations, which revealed that the meiotic segregant Hybrid T2 not only didn't produce any phenolic off-flavours, but also reached the highest ethanol concentration and consumed the most maltotriose. Our study demonstrates the

A comprehensive reference transcriptome resource for the common house spider Parasteatoda tepidariorum.