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Sample records for cas program deficiency

  1. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 8, 0.08 Mechanical, Book 1

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    System information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet too & material listing; testing methods; inspection frequency; standard system design life tables; and system work breakdown structure. Deficiency standards are given for plumbing, fire protection, heating, cooling, and special (drinking water cooling systems).

  2. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 3, 0.03 Superstructure

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented on asset determinant factor/CAS profile codes/CAS cost process; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. Deficiency standards and inspection methods are presented for beams; pre-engineered building systems; floors; roof structure; stairs; and fireproofing.

  3. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 2, 0.02 Substructure

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    System information is given for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. System assembly/component deficiencies and inspection methods are given for slabs-on-grade, columns, and column fireproofing.

  4. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 7, 0.07 Conveying

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    System information is given for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; and system work breakdown structure. Deficiency standards and inspection methods are presented for elevators and special conveyors.

  5. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 5, 0.05 Roofing

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; and system work breakdown structure. Deficiency standards and inspection methods are presented for built-up membrane; single- ply membrane; metal roofing systems; coated foam membrane; shingles; tiles; parapets; roof drainage system; roof specialties; and skylights.

  6. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 11, 0.11 Specialty systems

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. Deficiency standards and inspection methods are presented for canopies; loading dock systems; tanks; domes (bulk storage, metal framing); louvers & vents; access floors; integrated ceilings; and mezzanine structures.

  7. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 12, 0.12 Sitework

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. Deficiency standards and inspection methods are given for utility distribution systems, central heating, central cooling, electrical, utility support structures, paving roadways/walkways, and tunnels.

  8. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 6, 0.06 Interior construction

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. Deficiency standards and inspection methods are presented for conventional and specialty partitions, toilet partitions & accessories, interior doors, paint finishes/coatings/ wall covering systems; floor finishing systems; and ceiling systems.

  9. Condition Assessment Survey (CAS) Program. Deficiency standards and inspections methods manual: Volume 1, 0.01 Foundations and footings

    Energy Technology Data Exchange (ETDEWEB)

    1993-05-01

    General information is presented for asset determinant factor/CAS repair codes/CAS cost factors; guide sheet tool & material listing; testing methods; inspection frequency; standard system design life tables; system work breakdown structure; and general system/material data. Deficiency standards and inspection methods are given for footings - spread/strip/grade beams; foundation walls; foundation dampproofing/waterproofing; excavation/backfill/ and piles & caissons.

  10. CAS Readjusts Implementation of Its Talent Program

    Institute of Scientific and Technical Information of China (English)

    Guo Haiyan

    2002-01-01

    @@ CAS has decided to readjust the way of implementing its Century Program (or Hundred-Talents Program), to give more independence to research institutes in head hunting and guarantee the support for the Program recruits on a selective basis.

  11. CAS spearheads R&D program for research facilities

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ China's capacity for indigenous S&T innovation is believed to have been hampered by its lack of home- grown research facilities. To address the problem, a pilot program for the research and development of major S&T facilities has been launched at CAS. The kick-off meeting was held on 28 March in the CAS Technical Institute of Physics and Chemistry in Beijing.

  12. CAS Scientists Play an Important Role in National Basic Research Program

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ Of the 39 research projects that kicked off in 2005 with the support from the NationalBasic Research Program (dubbed the 973 Program) across the country, 12 are hosted byCAS scientists (including two under the joint leadership with other departments).

  13. Rapid generation of novel models of RAG1 deficiency by CRISPR/Cas9-induced mutagenesis in murine zygotes

    Science.gov (United States)

    de Bruin, Lisa Ott; Yang, Wei; Capuder, Kelly; Lee, Yu Nee; Antolini, Maddalena; Meyers, Robin; Gellert, Martin; Musunuru, Kiran; Manis, John; Notarangelo, Luigi

    2016-01-01

    Mutations in the Recombination Activating Gene 1 (RAG1) can cause a wide variety of clinical and immunological phenotypes in humans, ranging from absence of T and B lymphocytes to occurrence of autoimmune manifestations associated with expansion of oligoclonal T cells and production of autoantibodies. Although the mechanisms underlying this phenotypic heterogeneity remain poorly understood, some genotype-phenotype correlations can be made. Currently, mouse models of Rag deficiency are restricted to RAG1−/− mice and to knock-in models carrying severe missense mutations. The Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)/Cas9 system is a novel and powerful gene-editing strategy that permits targeted introduction of DNA double strand breaks with high efficiency through simultaneous delivery of the Cas9 endonuclease and a guide RNA (gRNA). Here, we report on CRISPR-based, single-step generation and characterization of mutant mouse models in which gene editing was attempted around residue 838 of RAG1, a region whose functional role had not been studied previously. PMID:26887046

  14. CAS-NETL-PNNL CEP Program Final Report

    Energy Technology Data Exchange (ETDEWEB)

    King, David L.; Spies, Kurt A.; Rainbolt, James E.; Zhang, Keling

    2014-03-31

    This collaborative joint research project is in the area of advanced gasification and conversion, within the CAS-NETL-PNNL Memorandum of Understanding. The goal is the development and testing of an integrated warm syngas cleanup process. This effort is focused on an advanced, integrated system for capture and removal of alkali, sulfur, PH3, AsH3, chloride, and CO2, leading to a future process demonstration at a CAS gasification facility. Syngas produced by gasification can be used for production of fuels (Fischer-Tropsch, SNG, mixed alcohols), chemicals (MeOH, NH3), and hydrogen for fuel cells and IGCC. To employ this syngas, especially for synthesis reactions, contained impurities must be removed to sub-ppmv levels [1]. Commercially available approaches to remove contaminant species suffer from inefficiencies, employing solvents at ambient or lower temperature along with backup sacrificial sorbents, whereas syngas utilization occurs at higher temperatures. The efficiency and economics syngas utilization can be significantly improved if all the contaminants and CO2 are removed at temperatures higher than the chemical synthesis reaction temperatures (> 250 °C) [2].

  15. Research undertaken by CAS scientists with support of "973 Program"

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    @@ The National Basic Research Program (dubbed as the "973 Program") is China's on-going national keystone basic research program, which was approved by the Chinese government in June 1 997 and is organized and implemented by the Ministry of Science and Technology.

  16. Evaluation of the oxidative stress modulation in Drosophila melanogaster strains deficient in endogenous antioxidants and with chronic exposure to casiopeina Cas II-gly and gamma radiation

    International Nuclear Information System (INIS)

    The casiopeinas are a family of coordination compounds with copper metallic center that have shown to have antineoplastic activity. The experimental evidences suggest that its action mechanism is through the generation of free radicals. The casiopeina (Cas II-gly) is believed to causes oxidative damage in the mitochondria, leading to the cellular death. The present study has the purpose to evaluate the antioxidant potential of the tetrapyrroles: cupro-sodica chlorophyllin (CSC), protoporphyrin-Ix (Pp-Ix) and the bilirubin (Bili) against the oxidant action of the Cas II-gly. The present study will also contribute in the characterization of the biological activity of the Cas II-gly. For this purpose is quantifies the effect of these compounds in the enzymes activity, superoxide dismutase (Sod) and catalase (Cat) in wild Drosophila melanogaster strains Canton-S and in the deficient in Sod and Cat. Two protocols were used, in the first male of 1-24 h of age were pre-treated with 0, 0.01, 0.1 and 1 m M of Cas II-gly and later on they were treated with radiation (15 Gy), and the second 69 m M of CSC, Pp-Ix or Bili, during 8 days and later they were treated with 0.1 m M of Cas II-gly during 24 h. The enzymatic activity was measured with the detection packages of enzymes Sod and Cat of Sigma. It was found that none of the three pigments increment the Sod activity but, if they diminished that of Cat (p≤0.007). The three concentrations of Cas II-gly did not increase the Sod activity significantly, only the concentration of 0.1 m M diminishes in 5.6 U the Cat activity (p <0.03) the same as the treatment with 15 Gy of gamma rays (8 U, p <0.004). The Cas II-gly combination 0.1 m M with the pigments does not modify the Sod and Cat activity. These results suggest that the proven pigments act as antioxidants, avoiding the induction of exogenous antioxidants caused by the gamma rays or the Cas II-gly. (Author)

  17. Recent CAS Achievements in Life Sciences

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    @@ Recent years have witnessed remarkable progress scored by CAS researchers along with the smooth development of the knowledge innovation program piloted at CAS. The follow-ings are just recent examples of CAS research achievements in life sciences.

  18. Deficiencies

    Data.gov (United States)

    U.S. Department of Health & Human Services — A list of all deficiencies currently listed on Nursing Home Compare, including the nursing home that received the deficiency, the associated inspection date,...

  19. [CAS General Standards 2012

    Science.gov (United States)

    Council for the Advancement of Standards in Higher Education, 2011

    2011-01-01

    The mission of the Council for the Advancement of Standards in Higher Education (CAS) is to promote the improvement of programs and services to enhance the quality of student learning and development. CAS is a consortium of professional associations who work collaboratively to develop and promulgate standards and guidelines and to encourage…

  20. JSPS-CAS Core University Program seminar on summary of 10-year collaborations in plasma and nuclear fusion research area

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on “Summary of 10-year Collaborations in Plasma and Nuclear Fusion Research Area” was held from March 9 to March 11, 2011 in the Okinawa Prefectural Art Museum, Naha city, Okinawa, Japan. The collaboration program on plasma and nuclear fusion started from 2001 under the auspices of Japanese Society of Promotion of Science (JSPS) and Chinese Academy of Sciences (CAS). This year is the last year of the CUP. This seminar was organized in the framework of the CUP. In the seminar, 29 oral talks were presented, having 14 Chinese and 30 Japanese participants. These presentations covered key topics related to the collaboration categories: (1) improvement of core plasma properties, (2) basic research on fusion reactor technologies, and (3) theory and numerical simulation. This seminar aims at summarizing the results obtained through the collaborations for 10 years, and discussing future prospects of China-Japan collaboration in plasma and nuclear fusion research areas. (author)

  1. Addressing Perceived Skill Deficiencies in Student Affairs Graduate Preparation Programs

    Science.gov (United States)

    Cooper, Jay; Mitchell, Donald, Jr.; Eckerle, Kayle; Martin, Kyle

    2016-01-01

    This article explores existing literature on perceived skill deficiencies among entry-level student affairs practitioners. Through a review of recent literature, seven perceived skill deficiencies were identified, including budgeting and financial management, strategic planning, research and assessment, legal knowledge and standards, supervision,…

  2. Economic evaluation of iodine deficiency disorder control program in Sikkim: A cost-benefit analysis

    OpenAIRE

    Pandav, Chandrakant S.

    2012-01-01

    Background: Iodine deficiency disorders (IDDs) are the most common cause of preventable brain damage globally. The strategy of prevention and control of iodine deficiency is based on iodine supplementation. Edible salt iodization and iodized oil injections are the two most commonly used vehicles for iodine supplementation. The objective of the study was to conduct a cost-benefit analysis of the two programs of iodine supplementation, i.e., iodized salt program (ISP) and iodized oil program (I...

  3. Proceeding of JSPS-CAS Core-University Program (CUP) on superconducting key technology for advanced fusion device

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on 'Superconducting Key Technology for Advanced Fusion Device' was held in Xi'an, China from October 18 to 21, 2010. This seminar was organized in the framework of the CUP in the field of plasma and nuclear fusion. This seminar honored by NIFS and ASIPP is aim to have a wide discussion on the new application and achievements on superconducting technology of nuclear fusion reactor. The superconducting technology on fusion reactor involves the fields on high current superconductor and magnet, quench protection, current control, cooling of the magnet, and reliability of large scale refrigerator. The technology on ITER high temperature superconductor current leads and the conductor test of JT-60SA are discussed in this seminar. Thirty-four oral talks and two summary talks were presented in this seminar. Total number of the participants was 34, including 12 Japanese participants. (author)

  4. Proceeding of JSPS-CAS Core University Program (CUP) seminar on modeling of theory and simulation of fusion plasmas

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on 'Modeling of Theory and Simulation of Fusion Plasmas' was held in Peking University, Beijing, China, from August 30 through September 2, 2010. This seminar was organized in the framework of the CUP in the field of plasma and nuclear fusion. This year is the final year of the CUP on Fusion and Plasma. One special talk and 26 oral talks were presented in the seminar including 11 Japanese attendees. This seminar included timely and very interesting reviews and discussions of: (1) progress in theory, simulation and integrated modeling of fusion plasmas; (2) newly experimental results closed related with present theoretical interest; (3) present status of collaboration research activities and new ideas for post-CUP collaboration. The meeting consisted of 26 oral presentations, which covered the following topical areas: (1) Turbulence and transport; (2) MHD equilibrium and stability; (3) Peripheral plasma behaviors. Several future collaboration researches were proposed in this seminar. (author)

  5. Research Facility Development at CAS

    Institute of Scientific and Technical Information of China (English)

    Tian Dongsheng; Miao Yougui; Zhang Hongsong

    2005-01-01

    @@ This article gives an introductory account on the development of research facilities at the CAS over the past six years since the initiation of the Knowledge Innovation Program in 1998 and during the period of the national 10th Five-year Plan in particular. In addition, it expounds the key points for the future work at the CAS in this regard.

  6. mCAL: A New Approach for Versatile Multiplex Action of Cas9 Using One sgRNA and Loci Flanked by a Programmed Target Sequence.

    Science.gov (United States)

    Finnigan, Gregory C; Thorner, Jeremy

    2016-01-01

    Genome editing exploiting CRISPR/Cas9 has been adopted widely in academia and in the biotechnology industry to manipulate DNA sequences in diverse organisms. Molecular engineering of Cas9 itself and its guide RNA, and the strategies for using them, have increased efficiency, optimized specificity, reduced inappropriate off-target effects, and introduced modifications for performing other functions (transcriptional regulation, high-resolution imaging, protein recruitment, and high-throughput screening). Moreover, Cas9 has the ability to multiplex, i.e., to act at different genomic targets within the same nucleus. Currently, however, introducing concurrent changes at multiple loci involves: (i) identification of appropriate genomic sites, especially the availability of suitable PAM sequences; (ii) the design, construction, and expression of multiple sgRNA directed against those sites; (iii) potential difficulties in altering essential genes; and (iv) lingering concerns about "off-target" effects. We have devised a new approach that circumvents these drawbacks, as we demonstrate here using the yeast Saccharomyces cerevisiae First, any gene(s) of interest are flanked upstream and downstream with a single unique target sequence that does not normally exist in the genome. Thereafter, expression of one sgRNA and cotransformation with appropriate PCR fragments permits concomitant Cas9-mediated alteration of multiple genes (both essential and nonessential). The system we developed also allows for maintenance of the integrated, inducible Cas9-expression cassette or its simultaneous scarless excision. Our scheme-dubbed mCAL for " M: ultiplexing of C: as9 at A: rtificial L: oci"-can be applied to any organism in which the CRISPR/Cas9 methodology is currently being utilized. In principle, it can be applied to install synthetic sequences into the genome, to generate genomic libraries, and to program strains or cell lines so that they can be conveniently (and repeatedly

  7. High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity

    OpenAIRE

    Pattanayak, Vikram; Lin, Steven; Guilinger, John P.; MA, ENBO; Doudna, Jennifer A.; Liu, David R.

    2013-01-01

    The RNA-programmable Cas9 endonuclease cleaves double-stranded DNA at sites complementary to a 20-base-pair guide RNA. The Cas9 system has been used to modify genomes in multiple cells and organisms, demonstrating its potential as a facile genome-engineering tool. We used in vitro selection and high-throughput sequencing to determine the propensity of eight Cas9:guide RNA complexes to cleave each of 10^12 potential off-target DNA sequences. The selection results predicted five off-target site...

  8. JSPS-CAS Core University Program seminar. Proceedings of Japan-China joint seminar on laser plasma and high energy density physics

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University program seminar on “Laser Plasma and High Energy Density Physics” was held in Xuhui Campus of Shanghai Jiao Tong University on October 14-16, 2010. 10 Japanese invited talks were presented while the total of 18 invited talks were given. This seminar was combined with 2nd China National Symposium on High Energy Density Physics, and another 27 oral talks and 26 posters were presented for this symposium. Totally over 90 participants including many young graduate students also attended the seminar. This workshop planned as a final in this ten years' collaboration between China and Japan. The workshop was originally planned to promote communications with young Chinese scientists and graduate students and Japanese scientists as well as Chinese scientists. Many cutting edge physics were presented from both countries and the collaboration through the JSPS-CAS Core University program was summarized by Prof. K. A. Tanaka and Prof. S. Zhu. (author)

  9. Program Model: Language Enriched English Eleven for High School Students Deficient in Language.

    Science.gov (United States)

    Kane, Janet Fenton; Arend, Paul J.

    Eighteen high school students of normal intelligence but with severely deficient language skills were given a special highly structured instructional program. Students progressed in vocabulary development, reading comprehension, and written and oral expression. Although the degree of progress was not sufficient to overcome deficits of two and…

  10. The Assessment of a Tutoring Program to Meet CAS Standards Using a SWOT Analysis and Action Plan

    Science.gov (United States)

    Fullmer, Patricia

    2009-01-01

    This article summarizes the use of SWOT (Strengths, Weaknesses, Opportunities, and Threats) analysis and subsequent action planning as a tool of self-assessment to meet CAS (Council for the Advancement of Standards in Higher Education) requirements for systematic assessment. The use of the evaluation results to devise improvements to increase the…

  11. Prevalence of micronutrient deficiency based on results obtained from the national pilot program on control of micronutrient malnutrition.

    Science.gov (United States)

    Chakravarty, Indira; Sinha, R K

    2002-05-01

    Micronutrient deficiency is a serious public health concern in most developing countries. In India, iron deficiency, vitamin A deficiency, and iodine deficiency disorder are of greatest public health significance. In addition, subclinical zinc deficiency, flourosis, and fluoride-deficient dental caries are important areas of concern. The National Pilot Program on Control of Micronutrient Malnutrition was launched in 1995 to address these problems and the Department of Biochemistry and Nutrition of the All India Institute of Hygiene and Public Health (Calcutta) was entrusted to coordinate its activities. The program presently covers one northeastern and four eastern states, namely Assam, Bihar (Jharkhand), Orissa, West Bengal, and Tripura. Baseline analyses were conducted on demographic situation, food and nutrient intake pattern, nutritional deficiency diseases (e.g., iron deficiency anemia), iodine deficiency disorder, and vitamin A deficiency. It was observed that except for cereals, the diet was deficient in all other food groups. Nutrient intake (i.e., energy, protein, vitamins, and minerals) was also deficient in almost the entire state. Anthropometric indices (e.g., weight-for-age and height-for-age data) indicated that large percentages of prevalence of anemia in various age groups was found to be high. Bitot's spot was mainly noted in the age group of 6-71 months. Nightblindness was noted in young children as well as the children 24-71 months old. High prevalence of nightblindness in pregnant women is a point of concern. Actions needed to control micronutrient deficiencies include: intervention strategies, extensive nutrition and health education through innovative IEC materials to support problem-specific programs, strengthening of various state government programs and the role of NGOs. PMID:12035859

  12. CRISPR/Cas9 For Photoactivated Localization Microscopy (PALM)

    OpenAIRE

    Zhu, Yina; Li, Pingchuan; Beuzer, Paolo; Tong, Zhisong; Watters, Robin; Lv, Dan; Murre, Cornelis; Cang, Hu

    2014-01-01

    We demonstrate that endonuclease deficient Clustered Regularly Interspaced Short Palindromic Repeats CRISPR-associated Cas9 protein (dCas9) fused to the photo-convertible fluorescence protein monomeric mEos3.1 (dCas9-mEos3) can be used to resolve sub-diffraction limited features of repetitive gene elements, thus providing a new route to investigate high-order chromatin organization at these sites.

  13. International Cooperation at CAS Since 1998

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    @@ Since 1998 when the Knowledge Innovation Program (KIP) was launched,the rapid development of CAS has offered great opportunities to forge international cooperation.Focusing on capacity building to effectively absorb global innovation resources and through organizing major innovative activities,CAS has increasingly expanded and intensified international partnerships.

  14. A Program of Nutritional Education in Schools Reduced the Prevalence of Iron Deficiency in Students

    Directory of Open Access Journals (Sweden)

    María Nieves García-Casal

    2011-01-01

    Full Text Available The objective was to determine the prevalence of iron, folates and retinol deficiencies in school children and to evaluate the changes after an intervention of nutritional education. The project was developed in 17 schools. The sample included 1,301 children (678 males and 623 females. A subsample of 480 individuals, was randomly selected for drawing blood for biochemical determinations before and after the intervention of nutritional education, which included in each school: written pre and post-intervention tests, 6 workshops, 2 participative talks, 5 game activities, 1 cooking course and 1 recipe contest. Anthropometrical and biochemical determinations included weight, height, body-mass index, nutritional status, hematocrit, serum ferritin, retinol and folate concentrations. There was high prevalence of iron (25%, folates (75% and vitamin A (43% deficiencies in school children, with a low consumption of fruit and vegetables, high consumption of soft drinks and snacks and almost no physical activity. The nutritional education intervention produced a significant reduction in iron deficiency prevalence (25 to 14%, and showed no effect on vitamin A and folates deficiencies. There was a slight improvement in nutritional status. This study shows, through biochemical determinations, that nutritional education initiatives and programs have an impact improving nutritional health in school children.

  15. MacSyFinder: a program to mine genomes for molecular systems with an application to CRISPR-Cas systems.

    Directory of Open Access Journals (Sweden)

    Sophie S Abby

    Full Text Available Biologists often wish to use their knowledge on a few experimental models of a given molecular system to identify homologs in genomic data. We developed a generic tool for this purpose.Macromolecular System Finder (MacSyFinder provides a flexible framework to model the properties of molecular systems (cellular machinery or pathway including their components, evolutionary associations with other systems and genetic architecture. Modelled features also include functional analogs, and the multiple uses of a same component by different systems. Models are used to search for molecular systems in complete genomes or in unstructured data like metagenomes. The components of the systems are searched by sequence similarity using Hidden Markov model (HMM protein profiles. The assignment of hits to a given system is decided based on compliance with the content and organization of the system model. A graphical interface, MacSyView, facilitates the analysis of the results by showing overviews of component content and genomic context. To exemplify the use of MacSyFinder we built models to detect and class CRISPR-Cas systems following a previously established classification. We show that MacSyFinder allows to easily define an accurate "Cas-finder" using publicly available protein profiles.MacSyFinder is a standalone application implemented in Python. It requires Python 2.7, Hmmer and makeblastdb (version 2.2.28 or higher. It is freely available with its source code under a GPLv3 license at https://github.com/gem-pasteur/macsyfinder. It is compatible with all platforms supporting Python and Hmmer/makeblastdb. The "Cas-finder" (models and HMM profiles is distributed as a compressed tarball archive as Supporting Information.

  16. CASKS (Computer Analysis of Storage casKS): A microcomputer based analysis system for storage cask design review. User's manual to Version 1b (including program reference)

    International Nuclear Information System (INIS)

    CASKS (Computer Analysis of Storage casKS) is a microcomputer-based system of computer programs and databases developed at the Lawrence Livermore National Laboratory (LLNL) for evaluating safety analysis reports on spent-fuel storage casks. The bulk of the complete program and this user's manual are based upon the SCANS (Shipping Cask ANalysis System) program previously developed at LLNL. A number of enhancements and improvements were added to the original SCANS program to meet requirements unique to storage casks. CASKS is an easy-to-use system that calculates global response of storage casks to impact loads, pressure loads and thermal conditions. This provides reviewers with a tool for an independent check on analyses submitted by licensees. CASKS is based on microcomputers compatible with the IBM-PC family of computers. The system is composed of a series of menus, input programs, cask analysis programs, and output display programs. All data is entered through fill-in-the-blank input screens that contain descriptive data requests

  17. Functional annotation of native enhancers with a Cas9-histone demethylase fusion.

    Science.gov (United States)

    Kearns, Nicola A; Pham, Hannah; Tabak, Barbara; Genga, Ryan M; Silverstein, Noah J; Garber, Manuel; Maehr, René

    2015-05-01

    Understanding of mammalian enhancers is limited by the lack of a technology to rapidly and thoroughly test the cell type-specific function. Here, we use a nuclease-deficient Cas9 (dCas9)-histone demethylase fusion to functionally characterize previously described and new enhancer elements for their roles in the embryonic stem cell state. Further, we distinguish the mechanism of action of dCas9-LSD1 at enhancers from previous dCas9-effectors. PMID:25775043

  18. Evaluation of the oxidative stress modulation in Drosophila melanogaster strains deficient in endogenous antioxidants and with chronic exposure to casiopeina Cas II-gly and gamma radiation; Evaluacion de la modulacion del estres oxidante en cepas de Drosophila melanogaster deficientes en antioxidantes endogenos y con exposicion cronica a casiopeina CII-gly y radiacion gamma

    Energy Technology Data Exchange (ETDEWEB)

    Jimenez V, E. R.

    2013-07-01

    The casiopeinas are a family of coordination compounds with copper metallic center that have shown to have antineoplastic activity. The experimental evidences suggest that its action mechanism is through the generation of free radicals. The casiopeina (Cas II-gly) is believed to causes oxidative damage in the mitochondria, leading to the cellular death. The present study has the purpose to evaluate the antioxidant potential of the tetrapyrroles: cupro-sodica chlorophyllin (CSC), protoporphyrin-Ix (Pp-Ix) and the bilirubin (Bili) against the oxidant action of the Cas II-gly. The present study will also contribute in the characterization of the biological activity of the Cas II-gly. For this purpose is quantifies the effect of these compounds in the enzymes activity, superoxide dismutase (Sod) and catalase (Cat) in wild Drosophila melanogaster strains Canton-S and in the deficient in Sod and Cat. Two protocols were used, in the first male of 1-24 h of age were pre-treated with 0, 0.01, 0.1 and 1 m M of Cas II-gly and later on they were treated with radiation (15 Gy), and the second 69 m M of CSC, Pp-Ix or Bili, during 8 days and later they were treated with 0.1 m M of Cas II-gly during 24 h. The enzymatic activity was measured with the detection packages of enzymes Sod and Cat of Sigma. It was found that none of the three pigments increment the Sod activity but, if they diminished that of Cat (p≤0.007). The three concentrations of Cas II-gly did not increase the Sod activity significantly, only the concentration of 0.1 m M diminishes in 5.6 U the Cat activity (p <0.03) the same as the treatment with 15 Gy of gamma rays (8 U, p <0.004). The Cas II-gly combination 0.1 m M with the pigments does not modify the Sod and Cat activity. These results suggest that the proven pigments act as antioxidants, avoiding the induction of exogenous antioxidants caused by the gamma rays or the Cas II-gly. (Author)

  19. Proceeding of JSPS-CAS Core University Program seminar on production and control of high performance plasmas with advanced plasma heating and diagnostic systems

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on 'Production and control of high performance plasmas with advanced plasma heating and diagnostic systems' took place in Guilin Bravo Hotel, Guilin, China, 1-4 November 2010. This seminar was organized in the framework of CUP in the field of plasma and nuclear fusion. Two special talks and 46 oral talks were presented in the seminar including 36 Chinese, 18 Japanese and 4 Korean attendees. Production and control of high performance plasmas is a crucial issue for realizing an advanced nuclear fusion reactor in addition to developments of advanced plasma heating and diagnostics. This seminar was motivated along the issues. Results in the field of fusion experiments obtained through CUP activities during recent two years were summarized. Possible direction of future collaboration and further encouragement of scientific activity of younger scientists were also discussed in this seminar with future experimental plans in both countries. (author)

  20. JSPS-CAS Core University Program seminar. Proceedings of Japan-China joint seminar on atomic and molecular processes in plasma

    International Nuclear Information System (INIS)

    As one of the activities of JSPS-CAS Core University Program, Japan-China Joint Seminar on Atomic and Molecular Processes in Plasma was held on March 6 - 11, 2004 in Lanzhou, China. The total number of the officially registered participants was 29, in which 17 from Japan, 10 from China, and 2 from Germany. In the nuclear fusion plasma, there are quite a variety of atomic processes such as ionization, excitation, radiative recombination, non-radiative recombination (di-electronic recombination, collisional electron transfer), cascade radiation, and cascade Auger decay over the wide range of plasma temperature. The knowledge of such the processes is indispensable for the evaluation and improvement of the plasma properties, which is desirable to be investigated by international collaboration groups. The present Japan-China Joint Seminar constitutes one of such the activities to realize the above stated aim. The 21 of the presented papers are indexed individually. (J.P.N.)

  1. Proceedings of JSPS-CAS Core University Program seminar on production and steady state confinement of high performance plasmas in magnetic confinement systems

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on 'Production and steady-state confinement of high performance plasmas in magnetic confinement systems' was held from 27 July to 29 July 2005 in Institute of Plasma Physics, the Chinese Academy of Sciences, Hefei, China. This seminar was organized in the framework of CUP in the field of plasma and nuclear fusion. About 50 persons including 20 Japanese attendees attended this seminar. Long time sustainment of high confinement and high beta plasmas is crucial for realization of an advanced nuclear fusion reactor. This seminar was motivated to summarize the results of CUP obtained in four years activities of CUP, and to extract crucial issues to be resolved near future, which must drive near and mid- term collaborations in the framework of CUP. The 32 of presented papers are indexed individually. (J.P.N.)

  2. Proceeding of JSPS-CAS core university program seminar on production and control of high performance plasmas with advanced plasma heating and diagnostic systems

    International Nuclear Information System (INIS)

    The JSPS-CAS Core University Program (CUP) seminar on 'Production and control of high performance plasmas with advanced plasma heating and diagnostic systems' took place in Shiner hotel, Lijiang, China, 4-7 November 2008. This seminar was organized in the framework of CUP in the field of plasma and nuclear fusion. One special talk and 34 oral talks were presented in the seminar including 16 Japanese attendees. Production and control of high performance plasmas is a crucial issue for realizing an advanced nuclear fusion reactor in addition to developments of advanced plasma heating and diagnostics. This seminar was motivated along the issues. Results obtained from CUP activities during recent four years were summarized. Several crucial issues to be resolved near future were also extracted in this seminar. The 31 of the papers are indexed individually. (J.P.N.)

  3. Eliciting management action: Using THERP to highlight human factors deficiencies for trip reduction programs

    International Nuclear Information System (INIS)

    Methods and criteria for performing human factors evaluations of plant systems and procedures are well developed and available. For a design review to produce a positive impact on operations, however, it is not enough to simply document deficiences and solutions. The results must be presented to management in a clear and compelling form that will direct attention to the heart of a problem and present proposed solutions in terms of explicit, quantified cost/benefits. A proactive program of trip reduction provides an excellent opportunity to accomplish human factors-related upgrades. As an evaluative context, trip reduction imposes a uniform goodness criterion on all situations: the probability of inadvertent plant trip. This in turn means that findings can be compared in terms of a common quantitative reference point: the cost of an inadvertent shutdown. To interpret human factors deficiencies in terms of trip probabilities, the Technique for Human Error Rate Prediction (THERP) can be used. THERP provides an accessible compilation of human reliability data for generic, discrete task elements. Sequences of such values are combined in standard event trees to determine the probability of failure (e.g., trip) for a given evolution. THERP is widely accepted as one of the best available alternatives for assessing human reliability

  4. Economic evaluation of iodine deficiency disorder control program in Sikkim: A cost-benefit analysis

    Directory of Open Access Journals (Sweden)

    Chandrakant S Pandav

    2012-01-01

    Full Text Available Background: Iodine deficiency disorders (IDDs are the most common cause of preventable brain damage globally. The strategy of prevention and control of iodine deficiency is based on iodine supplementation. Edible salt iodization and iodized oil injections are the two most commonly used vehicles for iodine supplementation. The objective of the study was to conduct a cost-benefit analysis of the two programs of iodine supplementation, i.e., iodized salt program (ISP and iodized oil program (IOP against no preventive program (NPP option. Materials and Methods: The study was conducted in 1990 in the state of Sikkim in India. The costs were calculated on the assumption of universal coverage of ISP and coverage of IOP among all children aged 0-14 years and women in the age group of 15-44 years. Direct and indirect cost of ISP and direct cost of IOP was computed based on the costs of year 1991. The discount rate taken was 10% and all the costs were converted to the year 2010 using wholesale price index (WPI data. Consequences in terms of health effects, Social/emotional effects, and resource use were included. Results: The discounted cost of ISP and IOP was Rs. 59,225,964 and Rs. 46,145,491, respectively. In ISP, 64.1% of the total cost was required for salt iodization, 17.6% for monitoring, and 18.3% for communication. In IOP, 50.9% of the costs were required for iodized oil; rest was for syringes and needles, manpower expenses, travel, and communication. Total resource saving was Rs. 95,566,220 for ISP and Rs. 92,177,548 for IOP. Incremental benefit for ISP was Rs. 36,340,256 and Rs. 46,032,057 for IOP. The cost-benefit ratio for ISP was 1.61 and 2.00 for IOP. Conclusion: IOP has a higher cost-benefit ratio for prevention of IDDs than ISP in the state of Sikkim, India.

  5. Programa comunitário e deficiência de G-6-PD no Brasil Community program and G-6-PD deficiency in Brazil

    Directory of Open Access Journals (Sweden)

    Mariane B. Compri

    2000-04-01

    Full Text Available A viabilidade e a eficiência da implantação de um programa comunitário relativo à deficiência de desidrogenase de 6-fosfato de glicose foram investigadas em uma população brasileira (Bragança Paulista, SP. A comunidade foi abordada a partir dos seus doadores de sangue, examinando-se 4.621 indivíduos do sexo masculino e diagnosticando-se 80 portadores da alteração genética. O índice de positividade na amostra foi de 1,7%, a taxa de aceitação à orientação genética oferecida em caráter opcional atingiu 61% e a percentagem de assimilação satisfatória dos deficientes de desidrogenase de 6-fosfato de glicose quanto às informações fornecidas foi de 81%. A análise molecular demonstrou a presença quase que exclusiva na comunidade da variante A- ou Africana de desidrogenase de 6-fosfato de glicose, relativamente benigna do ponto de vista clínico. Com um baixo custo anual, foi possível diagnosticar e orientar de forma ética um número significativo de famílias com a deficiência de desidrogenase de 6-fosfato de glicose, além de iniciar a educação da comunidade a respeito do problema.This study tested the viability and efficiency of a glucose-6-phosphate dehydrogenase deficiency program in which a Brazilian community ¾ the town of Bragança Paulista, SP (Brazil was involved. The screening was done among blood donors and the diagnosis of glucose-6-phosphate dehydrogenase deficiency was confirmed through enzymatic quantification and through electrophoresis of the enzyme. A total of 4,621 individuals were tested over a period of 36 months. A total of 80 individuals with glucose-6-phosphate dehydrogenase deficiency were detected ¾ 1,7% of the samples examined. The acceptance rate to the optional genetic orientation got to 61% and the percentage of good assimilation concerning the offered information was 81%. The molecular analysis demonstrated the presence, almost exclusive, in the community of the A- or African variant, a

  6. JSPS-CAS core university program seminar. Proceedings of Japan-China joint seminar on atomic and molecular processes in plasma

    International Nuclear Information System (INIS)

    As one of the activities of JSPS-CAS Core University Program, Japan-China Joint Seminar on Atomic and Molecular Processes in Plasma was held on October 8 - 12, 2007 in Dunhuang, China. The total number of the officially registered participants was 41, in which 12 from Japan, 25 from China, and 4 from EU. And this seminar is an extension of the last seminar that was held on March 6 - 11, 2004 in Lanzhou, China. In the nuclear fusion plasma, there are quite a variety of atomic processes such as ionization, excitation, radiative recombination, non-radiative recombination (di-electronic recombination, collisional electron transfer), cascade radiation, and cascade Auger decay over the wide range of plasma temperature. The knowledge of such processes is indispensable for the evaluation and improvement of the plasma properties, which is desirable to be investigated by international collaboration groups. The present seminar constitutes one of such activities to realize the above stated aim; especially it has given an opportunity for the collaborative workers to illustrate their achievements. The 32 of the presented papers are indexed individually. (J.P.N.)

  7. JSPS-CAS Core University Program seminar. Proceedings of Japan-China joint seminar on atomic and molecular processes in plasma

    International Nuclear Information System (INIS)

    As one of the activities of JSPS-CAS Core University Program, Japan-China Joint Seminar on Atomic and Molecular Processes in Plasma was held on October 26 - 31, 2009 in Xi'an, China. The total number of the officially registered participants was 54, in which 18 from Japan, 35 from China, and 1 from USA. And this seminar is an extension of the last two seminars that were held on March 6 - 11, 2004 in Lanzhou, China, and on October 6 - 12, 2007 in Dunhuang, China. In the nuclear fusion plasma, there are quite a variety of atomic processes such as ionization, excitation, radiative recombination, non-radiative recombination (di-electronic recombination, collisional electron transfer), cascade radiation, and cascade Auger decay over the wide range of plasma temperature. The knowledge of those processes is indispensable for the evaluation and improvement of the plasma properties. Because of the diversity of the subject, it is desirable to investigate them by international collaboration groups. The present seminar may contribute to realize the above stated aim; especially it has given an opportunity for the collaborative workers to illustrate their achievements. This seminar summarizes the collaborative researches for the last decade and propose the issues for the future prospect. The 30 of the presented papers are indexed individually. (J.P.N.)

  8. Preparing student affairs leaders for Canadian colleges and universities: Do the CAS standards for graduate preparation programs apply?

    OpenAIRE

    Howman, Cynthia Joan

    2009-01-01

    In Canada, there are currently no graduate level programs which have as a main focus the study of Student Affairs Administration. Student Affairs leaders at Canadian colleges and universities come from a wide variety of academic and career backgrounds. The purpose of this quantitative study was three fold; to gather detailed demographic information describing the current cohort of Canadian student affairs leaders, to determine, to what extent, these leaders were aware of the Council for Advan...

  9. Functional annotation of native enhancers with a Cas9 -histone demethylase fusion

    OpenAIRE

    Kearns, Nicola A; Pham, Hannah; Tabak, Barbara; Genga, Ryan M; Silverstein, Noah J; Garber, Manuel; Maehr, René

    2015-01-01

    Understanding of mammalian enhancer function is limited by the lack of a technology to rapidly and thoroughly test their cell type-specific function. Here, we use a nuclease-deficient (d)Cas9 histone demethylase fusion to functionally characterize previously described and novel enhancer elements for their roles in the embryonic stem cell state. Further, we distinguish the mechanism of action of dCas9-LSD1 at enhancers from previous dCas9-effectors.

  10. Genome Engineering of Drosophila with the CRISPR RNA-Guided Cas9 Nuclease

    OpenAIRE

    Gratz, Scott J.; Cummings, Alexander M.; Nguyen, Jennifer N.; Hamm, Danielle C.; Donohue, Laura K.; Harrison, Melissa M.; Wildonger, Jill; O’Connor-Giles, Kate M.

    2013-01-01

    We have adapted a bacterial CRISPR RNA/Cas9 system to precisely engineer the Drosophila genome and report that Cas9-mediated genomic modifications are efficiently transmitted through the germline. This RNA-guided Cas9 system can be rapidly programmed to generate targeted alleles for probing gene function in Drosophila.

  11. Regulations for CAS Visiting Professorships for Senior International Scientists

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    @@ 1.General Provisions. Article 1 These regulations are made in accordance with the Chinese Academy of Sciences (CAS) "Package Program for Talent Training & Recruitment" and "Guidelines of the Chinese Academy of Sciences for the Implementation of the Program for Attracting Overseas Scientists and Experts and Cultivating Talent through International Exchange",to guide the implementation of the "CAS Visiting Professorships for Senior International Scientists" (hereinafter referred to as the "Visiting Professorships Program").

  12. Using the CAS Standards in Assessment Projects

    Science.gov (United States)

    Dean, Laura A.

    2013-01-01

    This chapter provides an overview of the use of professional standards of practice in assessment and of the Council for the Advancement of Standards in Higher Education (CAS). It outlines a model for conducting program self-studies and discusses the importance of implementing change based on assessment results.

  13. Asteroid named after CAS scientist

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    @@ An asteroid has been named after CAS astronomy historian XI Zezong with the approval of the International Minor Planet Nomenclature Committee (IMPNC), announced China's National Astronomical Observatories at CAS (NAOC) on 17 August.

  14. Processes underlying the nutritional programming of embryonic development by iron deficiency in the rat.

    Directory of Open Access Journals (Sweden)

    Angelina Swali

    Full Text Available Poor iron status is a global health issue, affecting two thirds of the world population to some degree. It is a particular problem among pregnant women, in both developed and developing countries. Feeding pregnant rats a diet deficient in iron is associated with both hypertension and reduced nephron endowment in adult male offspring. However, the mechanistic pathway leading from iron deficiency to fetal kidney development remains elusive. This study aimed to establish the underlying processes associated with iron deficiency by assessing gene and protein expression changes in the rat embryo, focussing on the responses occurring at the time of the nutritional insult. Analysis of microarray data showed that iron deficiency in utero resulted in the significant up-regulation of 979 genes and down-regulation of 1545 genes in male rat embryos (d13. Affected processes associated with these genes included the initiation of mitosis, BAD-mediated apoptosis, the assembly of RNA polymerase II preinitiation complexes and WNT signalling. Proteomic analyses highlighted 7 proteins demonstrating significant up-regulation with iron deficiency and the down-regulation of 11 proteins. The main functions of these key proteins included cell proliferation, protein transport and folding, cytoskeletal remodelling and the proteasome complex. In line with our recent work, which identified the perturbation of the proteasome complex as a generalised response to in utero malnutrition, we propose that iron deficiency alone leads to a more specific failure in correct protein folding and transport. Such an imbalance in this delicate quality-control system can lead to cellular dysfunction and apoptosis. Therefore these findings offer an insight into the underlying mechanisms associated with the development of the embryo during conditions of poor iron status, and its health in adult life.

  15. Cas9 Functionally Opens Chromatin

    OpenAIRE

    Barkal, Amira A.; Srinivasan, Sharanya; Hashimoto, Tatsunori; Gifford, David K.; Sherwood, Richard I.

    2016-01-01

    Using a nuclease-dead Cas9 mutant, we show that Cas9 reproducibly induces chromatin accessibility at previously inaccessible genomic loci. Cas9 chromatin opening is sufficient to enable adjacent binding and transcriptional activation by the settler transcription factor retinoic acid receptor at previously unbound motifs. Thus, we demonstrate a new use for Cas9 in increasing surrounding chromatin accessibility to alter local transcription factor binding.

  16. Cas9 Functionally Opens Chromatin

    OpenAIRE

    Barkal, Amira A.; Srinivasan, Sharanya; Gifford, David K.; Sherwood, Richard I.; Hashimoto, Tatsunori Benjamin

    2015-01-01

    Using a nuclease-dead Cas9 mutant, we show that Cas9 reproducibly induces chromatin accessibility at previously inaccessible genomic loci. Cas9 chromatin opening is sufficient to enable adjacent binding and transcriptional activation by the settler transcription factor retinoic acid receptor at previously unbound motifs. Thus, we demonstrate a new use for Cas9 in increasing surrounding chromatin accessibility to alter local transcription factor binding.

  17. Cas9 Functionally Opens Chromatin.

    Directory of Open Access Journals (Sweden)

    Amira A Barkal

    Full Text Available Using a nuclease-dead Cas9 mutant, we show that Cas9 reproducibly induces chromatin accessibility at previously inaccessible genomic loci. Cas9 chromatin opening is sufficient to enable adjacent binding and transcriptional activation by the settler transcription factor retinoic acid receptor at previously unbound motifs. Thus, we demonstrate a new use for Cas9 in increasing surrounding chromatin accessibility to alter local transcription factor binding.

  18. Cas9 Functionally Opens Chromatin

    Science.gov (United States)

    Barkal, Amira A.; Srinivasan, Sharanya; Hashimoto, Tatsunori; Gifford, David K.; Sherwood, Richard I.

    2016-01-01

    Using a nuclease-dead Cas9 mutant, we show that Cas9 reproducibly induces chromatin accessibility at previously inaccessible genomic loci. Cas9 chromatin opening is sufficient to enable adjacent binding and transcriptional activation by the settler transcription factor retinoic acid receptor at previously unbound motifs. Thus, we demonstrate a new use for Cas9 in increasing surrounding chromatin accessibility to alter local transcription factor binding. PMID:27031353

  19. Nuclease Activity of Legionella pneumophila Cas2 Promotes Intracellular Infection of Amoebal Host Cells

    Science.gov (United States)

    Gunderson, Felizza F.; Mallama, Celeste A.; Fairbairn, Stephanie G.

    2014-01-01

    Legionella pneumophila, the primary agent of Legionnaires' disease, flourishes in both natural and man-made environments by growing in a wide variety of aquatic amoebae. Recently, we determined that the Cas2 protein of L. pneumophila promotes intracellular infection of Acanthamoeba castellanii and Hartmannella vermiformis, the two amoebae most commonly linked to cases of disease. The Cas2 family of proteins is best known for its role in the bacterial and archeal clustered regularly interspaced short palindromic repeat (CRISPR)–CRISPR-associated protein (Cas) system that constitutes a form of adaptive immunity against phage and plasmid. However, the infection event mediated by L. pneumophila Cas2 appeared to be distinct from this function, because cas2 mutants exhibited infectivity defects in the absence of added phage or plasmid and since mutants lacking the CRISPR array or any one of the other cas genes were not impaired in infection ability. We now report that the Cas2 protein of L. pneumophila has both RNase and DNase activities, with the RNase activity being more pronounced. By characterizing a catalytically deficient version of Cas2, we determined that nuclease activity is critical for promoting infection of amoebae. Also, introduction of Cas2, but not its catalytic mutant form, into a strain of L. pneumophila that naturally lacks a CRISPR-Cas locus caused that strain to be 40- to 80-fold more infective for amoebae, unequivocally demonstrating that Cas2 facilitates the infection process independently of any other component encoded within the CRISPR-Cas locus. Finally, a cas2 mutant was impaired for infection of Willaertia magna but not Naegleria lovaniensis, suggesting that Cas2 promotes infection of most but not all amoebal hosts. PMID:25547789

  20. Differential programming of p53-deficient embryonic cells during rotenone block

    Science.gov (United States)

    Mitochondrial dysfunction has been implicated in chemical toxicities. The present study used an in vitro model to investigate the differential expression of metabolic pathways during cellular stress in p53- efficient embryonic fibroblasts compared to p53-deficient cells. These c...

  1. Genetically Programmed Changes in Photosynthetic Cofactor Metabolism in Copper-deficient Chlamydomonas.

    Science.gov (United States)

    Strenkert, Daniela; Limso, Clariss Ann; Fatihi, Abdelhak; Schmollinger, Stefan; Basset, Gilles J; Merchant, Sabeeha S

    2016-09-01

    Genetic and genomic studies indicate that copper deficiency triggers changes in the expression of genes encoding key enzymes in various chloroplast-localized lipid/pigment biosynthetic pathways. Among these are CGL78 involved in chlorophyll biosynthesis and HPPD1, encoding 4-hydroxyphenylpyruvate dioxygenase catalyzing the committed step of plastoquinone and tocopherol biosyntheses. Copper deficiency in wild-type cells does not change the chlorophyll content, but a survey of chlorophyll protein accumulation in this situation revealed increased accumulation of LHCSR3, which is blocked at the level of mRNA accumulation when either CGL78 expression is reduced or in the crd1 mutant, which has a copper-nutrition conditional defect at the same step in chlorophyll biosynthesis. Again, like copper-deficient crd1 strains, cgl78 knock-down lines also have reduced chlorophyll content concomitant with loss of PSI-LHCI super-complexes and reduced abundance of a chlorophyll binding subunit of PSI, PSAK, which connects LHCI to PSI. For HPPD1, increased mRNA results in increased abundance of the corresponding protein in copper-deficient cells concomitant with CRR1-dependent increased accumulation of γ-tocopherols, but not plastoquinone-9 nor total tocopherols. In crr1 mutants, where increased HPPD1 expression is blocked, plastochromanol-8, derived from plastoquinone-9 and purported to also have an antioxidant function, is found instead. Although not previously found in algae, this metabolite may occur only in stress conditions. PMID:27440043

  2. Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification

    OpenAIRE

    Guilinger, John P.; Thompson, David B.; Liu, David R.

    2014-01-01

    Genome editing by Cas9, which cleaves double-stranded DNA at a sequence programmed by a short single-guide RNA (sgRNA), can result in off-target DNA modification that may be detrimental in some applications. To improve DNA cleavage specificity, we generated fusions of catalytically inactive Cas9 and FokI nuclease (fCas9). DNA cleavage by fCas9 requires association of two fCas9 monomers that simultaneously bind target sites ~15 or 25 base pairs apart. In human cells, fCas9 modified target DNA ...

  3. Nucleosome breathing and remodeling constrain CRISPR-Cas9 function.

    OpenAIRE

    Isaac, RS; F. Jiang; Doudna, JA; Lim, WA; Narlikar, GJ; Almeida, R

    2016-01-01

    eLife digest CRISPR is a method of editing the genetic material inside living cells and has enabled dramatic advances in a broad variety of research fields in recent years. The method relies on a bacterial enzyme called Cas9 that can be programmed, via short guide molecules made from RNA, to target specific sites in the cell’s DNA. Once bound to its target, the Cas9 enzyme cuts the DNA molecule; this often leads to changes in the DNA sequence. In nature, bacteria use the CRISPR-Cas9 system to...

  4. Chemical and Biophysical Modulation of Cas9 for Tunable Genome Engineering.

    Science.gov (United States)

    Nuñez, James K; Harrington, Lucas B; Doudna, Jennifer A

    2016-03-18

    The application of the CRISPR-Cas9 system for genome engineering has revolutionized the ability to interrogate genomes of mammalian cells. Programming the Cas9 endonuclease to induce DNA breaks at specified sites is achieved by simply modifying the sequence of its cognate guide RNA. Although Cas9-mediated genome editing has been shown to be highly specific, cleavage events at off-target sites have also been reported. Minimizing, and eventually abolishing, unwanted off-target cleavage remains a major goal of the CRISPR-Cas9 technology before its implementation for therapeutic use. Recent efforts have turned to chemical biology and biophysical approaches to engineer inducible genome editing systems for controlling Cas9 activity at the transcriptional and protein levels. Here, we review recent advancements to modulate Cas9-mediated genome editing by engineering split-Cas9 constructs, inteins, small molecules, protein-based dimerizing domains, and light-inducible systems. PMID:26857072

  5. Zinc Deficiency in Childhood and Pregnancy: Evidence for Intervention Effects and Program Responses.

    Science.gov (United States)

    Lamberti, Laura M; Fischer Walker, Christa L; Black, Robert E

    2016-01-01

    Zinc is a key micronutrient of particular importance during childhood and pregnancy. Zinc deficiency has been linked to increased infection and stunting among children and is a risk factor for adverse pregnancy outcomes and preterm delivery. Targeted interventions have the potential to alleviate the adverse effects of zinc deficiency via therapeutic and preventive supplementation, fortification and biofortification, but implementation is challenging. A growing number of low- and middle- income countries have introduced national policies for zinc treatment of diarrhea among children under 5 years in response to mounting evidence of reduced episode duration and severity as well as reduced incidence in the ensuing months, but coverage remains low in the absence of effective scale-up efforts. Implementation of preventive zinc supplementation in young children has also been slow, despite evidence linking routine daily supplementation and treatment regimens with reductions in stunting and the incidence of diarrhea and pneumonia. Acceptance of other zinc interventions, including traditional fortification, fortification with micronutrient powders and biofortification, is hindered by unclear evidence on efficacy. Additional research is therefore warranted to ascertain the efficacy of delivering zinc through fortified and biofortified foods and in combination with other micronutrients in supplements or powders. Operations research is also necessary to establish best practices for scale-up of therapeutic zinc supplementation for diarrhea. PMID:27198901

  6. Engineering of Isogenic Cells Deficient for MR1 with a CRISPR/Cas9 Lentiviral System: Tools To Study Microbial Antigen Processing and Presentation to Human MR1-Restricted T Cells.

    Science.gov (United States)

    Laugel, Bruno; Lloyd, Angharad; Meermeier, Erin W; Crowther, Michael D; Connor, Thomas R; Dolton, Garry; Miles, John J; Burrows, Scott R; Gold, Marielle C; Lewinsohn, David M; Sewell, Andrew K

    2016-08-01

    The nonclassical HLA molecule MHC-related protein 1 (MR1) presents metabolites of the vitamin B synthesis pathways to mucosal-associated invariant T (MAIT) cells and other MR1-restricted T cells. This new class of Ags represents a variation on the classical paradigm of self/non-self discrimination because these T cells are activated through their TCR by small organic compounds generated during microbial vitamin B2 synthesis. Beyond the fundamental significance, the invariant nature of MR1 across the human population is a tantalizing feature for the potential development of universal immune therapeutic and diagnostic tools. However, many aspects of MR1 Ag presentation and MR1-restricted T cell biology remain unknown, and the ubiquitous expression of MR1 across tissues and cell lines can be a confounding factor for experimental purposes. In this study, we report the development of a novel CRISPR/Cas9 genome editing lentiviral system and its use to efficiently disrupt MR1 expression in A459, THP-1, and K562 cell lines. We generated isogenic MR1(-/-) clonal derivatives of the A549 lung carcinoma and THP-1 monocytic cell lines and used these to study T cell responses to intracellular pathogens. We confirmed that MAIT cell clones were unable to respond to MR1(-/-) clones infected with bacteria whereas Ag presentation by classical and other nonclassical HLAs was unaffected. This system represents a robust and efficient method to disrupt the expression of MR1 and should facilitate investigations into the processing and presentation of MR1 Ags as well as into the biology of MAIT cells. PMID:27307560

  7. A newly discovered Bordetella species carries a transcriptionally active CRISPR-Cas with a small Cas9 endonuclease

    OpenAIRE

    Ivanov, Yury V.; Shariat, Nikki; Karen B Register; Linz, Bodo; Rivera, Israel; Hu, Kai; Dudley, Edward G.; Harvill, Eric T.

    2015-01-01

    Background Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated genes (cas) are widely distributed among bacteria. These systems provide adaptive immunity against mobile genetic elements specified by the spacer sequences stored within the CRISPR. Methods The CRISPR-Cas system has been identified using Basic Local Alignment Search Tool (BLAST) against other sequenced and annotated genomes and confirmed via CRISPRfinder program. Using Polymerase Chain Reactio...

  8. Carnitine levels in 26,462 individuals from the nationwide screening program for primary carnitine deficiency in the Faroe Islands

    DEFF Research Database (Denmark)

    Rasmussen, Jan; Nielsen, Olav W; Janzen, Nils; Duno, Morten; Gislason, Hannes; Køber, Lars; Steuerwald, Ulrike; Lund, Allan M

    2014-01-01

    BACKGROUND: Primary carnitine deficiency (PCD) is an autosomal recessive disorder of fatty acid oxidation and has been associated to episodes of sudden death in the Faroe Islands. Data are presented from the nationwide population based Faroese screening program to find people with low carnitine...... levels indicating PCD. METHODS: Whole blood samples from dried blood spots were analysed by tandem mass spectrometry with and without butylation. Genetic analyses were performed in all people with non-butylated free carnitine (fC0) below 7 μmol/L. RESULTS: 55 % (n = 26,462) of the entire population was...... screened and 89 PCD patients were identified, yielding an overall prevalence of 1:297 of PCD in the Faroe Islands. Carnitine levels were positively correlated to age in both males and females (p < 0.003) although levels decreased in females when reaching fertile age. The gender difference in mean carnitine...

  9. Optimization of Genome Engineering Approaches with the CRISPR/Cas9 System

    OpenAIRE

    Li, Kai; Wang, Gang; Andersen, Troels; Zhou, Pingzhu; Pu, William T.

    2014-01-01

    Designer nucleases such as TALENS and Cas9 have opened new opportunities to scarlessly edit the mammalian genome. Here we explored several parameters that influence Cas9-mediated scarless genome editing efficiency in murine embryonic stem cells. Optimization of transfection conditions and enriching for transfected cells are critical for efficiently recovering modified clones. Paired gRNAs and wild-type Cas9 efficiently create programmed deletions, which facilitate identification of targeted c...

  10. CAS School in Germany

    CERN Multimedia

    CERN Accelerator School

    The CERN Accelerator School (CAS), the Helmholtz Centre for Heavy Ion Research GmbH (GSI) and the Technische Universität Darmstadt (TU Darmstadt) jointly organised a course on General Accelerator Physics, at intermediate level, at TU Darmstadt from 27 September to 9 October 2009.   Participants in the CERN Accelerator School in Darmstadt, Germany. The Intermediate-level course followed established practice, with lectures on core topics in the mornings and specialised courses in the afternoons. The latter provided "hands-on" education and experience in the three selected topics: "RF Measurement Techniques", "Beam Instrumentation and Diagnostics" and "Optics Design and Correction". These proved to be highly successful, with participants choosing one course and following the topic throughout the school. Guided studies, tutorials, seminars and a poster session completed the programme. A visit to GSI and the F...

  11. Dual nuclease activity of a Cas2 protein in CRISPR-Cas subtype I-B of Leptospira interrogans.

    Science.gov (United States)

    Dixit, Bhuvan; Ghosh, Karukriti Kaushik; Fernandes, Gary; Kumar, Pankaj; Gogoi, Prerana; Kumar, Manish

    2016-04-01

    Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130 carries a set of cas genes associated with CRISPR-Cas subtype I-B. Herein, we report for the first time active transcription of a set of cas genes (cas1 to cas8) of L. interrogans where cas4, cas1, cas2 and cas6, cas3, cas8, cas7, cas5 are clustered together in two independent operons. As an initial step toward comprehensive understanding of CRISPR-Cas system in spirochete, the biochemical study of one of the core Leptospira Cas2 proteins (Lep_Cas2) showed nuclease activity on both DNA and RNA in a nonspecific manner. Additionally, unlike other known Cas2 proteins, Lep_Cas2 showed metal-independent RNase activity and preferential activity on RNA over DNA. These results provide insight for understanding Cas2 diversity existing in the prokaryotic adaptive immune system. PMID:26950513

  12. CAS Mathematician Receives Shaw Prize

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ WU Wentsun (WU Wenjun), a CAS Member from the Academy of Mathematics and System Sciences, has been honored with the 2006 Shaw Prize in Mathematical Sciences for his contributions to the new interdisciplinary field of mathematics and mechanization.

  13. T-bet regulates differentiation of forkhead box protein 3+ regulatory T cells in programmed cell death-1-deficient mice.

    Science.gov (United States)

    Tahara, M; Kondo, Y; Yokosawa, M; Tsuboi, H; Takahashi, S; Shibayama, S; Matsumoto, I; Sumida, T

    2015-02-01

    Programmed cell death-1 (PD-1) plays an important role in peripheral T cell tolerance, but whether or not it affects the differentiation of helper T cell subsets remains elusive. Here we describe the importance of PD-1 in the control of T helper type 1 (Th1) cell activation and development of forkhead box protein 3 (FoxP3(+)) regulatory T cells (Tr(egs)). PD-1-deficient T cell-specific T-bet transgenic (P/T) mice showed growth retardation, and the majority died within 10 weeks. P/T mice showed T-bet over-expression, increased interferon (IFN)-γ production by CD4(+) T cells and significantly low FoxP3(+) T(reg) cell percentage. P/T mice developed systemic inflammation, which was probably induced by augmented Th1 response and low FoxP3(+) T(reg) count. The study identified a unique, previously undescribed role for PD-1 in Th1 and T(reg) differentiation, with potential implication in the development of Th1 cell-targeted therapy. PMID:25219397

  14. Programa comunitário e deficiência de G-6-PD no Brasil Community program and G-6-PD deficiency in Brazil

    OpenAIRE

    Mariane B. Compri; Newton C. Polimeno; Maria J.A. Vieira; Sara T. O. Saad; Antonio S. Ramalho

    2000-01-01

    A viabilidade e a eficiência da implantação de um programa comunitário relativo à deficiência de desidrogenase de 6-fosfato de glicose foram investigadas em uma população brasileira (Bragança Paulista, SP). A comunidade foi abordada a partir dos seus doadores de sangue, examinando-se 4.621 indivíduos do sexo masculino e diagnosticando-se 80 portadores da alteração genética. O índice de positividade na amostra foi de 1,7%, a taxa de aceitação à orientação genética oferecida em caráter opcional...

  15. CAS Action Plan for the Development of Western China Progressing Smoothly

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ Since CAS is charged with the key task of carrying out strategic, fundamental and forward-looking studies for this country, it is incumbent upon CAS scientists to take an active part in the National Program for the Development of Western China. This is also an important part of the national Knowledge Innovation Program (KIP) spearheaded by CAS. Over the past two years, CAS has made a series of remarkable advances in Western China with regard to the development of research bases, exploration of crucial scientific problems, the training of talented people, and providing consultation services for decision makers.With the furtherance of the national drive to develop the country's western region, CAS will step up its efforts and increase its funding, to make more and greater contributions to the overall development of the region.

  16. Microarray data on altered transcriptional program of Phgdh-deficient mouse embryonic fibroblasts caused by ʟ-serine depletion.

    Science.gov (United States)

    Hamano, Momoko; Sayano, Tomoko; Kusada, Wataru; Kato, Hisanori; Furuya, Shigeki

    2016-06-01

    Inherent ʟ-Ser deficiency culminates in intrauterine growth retardation, severe malformation of multiple organs particularly the central nervous system, and perinatal or early postnatal death in human and mouse. To uncover the molecular mechanisms underlying the growth-arrested phenotypes of l-Ser deficiency, we compared gene expression profiles of mouse embryonic fibroblasts deficient in 3-phosphoglycerate dehydrogenase (Phgdh), the first enzyme of de novo ʟ-Ser synthetic pathway, between ʟ-Ser-depleted and -supplemented conditions. The datasets (CEL and CHP files) from this study are publicly available on the Gene Expression Omnibus repository (accession number GEO: GSE55687). PMID:27222860

  17. Transcriptional regulatory program in wild-type and retinoblastoma gene-deficient mouse embryonic fibroblasts during adipocyte differentiation

    Directory of Open Access Journals (Sweden)

    Hansen Jacob B

    2011-05-01

    Full Text Available Abstract Background Although many molecular regulators of adipogenesis have been identified a comprehensive catalogue of components is still missing. Recent studies showed that the retinoblastoma protein (pRb was expressed in the cell cycle and late cellular differentiation phase during adipogenesis. To investigate this dual role of pRb in the early and late stages of adipogenesis we used microarrays to perform a comprehensive systems-level analysis of the common transcriptional program of the classic 3T3-L1 preadipocyte cell line, wild-type mouse embryonic fibroblasts (MEFs, and retinoblastoma gene-deficient MEFs (Rb-/- MEFs. Findings Comparative analysis of the expression profiles of 3T3-L1 cells and wild-type MEFs revealed genes involved specifically in early regulation of the adipocyte differentiation as well as secreted factors and signaling molecules regulating the later phase of differentiation. In an attempt to identify transcription factors regulating adipogenesis, bioinformatics analysis of the promoters of coordinately and highly expressed genes was performed. We were able to identify a number of high-confidence target genes for follow-up experimental studies. Additionally, combination of experimental data and computational analyses pinpointed a feedback-loop between Pparg and Foxo1. To analyze the effects of the retinoblastoma protein at the transcriptional level we chose a perturbated system (Rb-/- MEFs for comparison to the transcriptional program of wild-type MEFs. Gene ontology analysis of 64 deregulated genes showed that the Rb-/- MEF model exhibits a brown(-like adipocyte phenotype. Additionally, the analysis results indicate a different or additional role for pRb family member involvement in the lineage commitment. Conclusion In this study a number of commonly modulated genes during adipogenesis in 3T3-L1 cells and MEFs, potential transcriptional regulation mechanisms, and differentially regulated targets during adipocyte

  18. CAS Scientist Receives Trieste Science Prize

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ RAO Zihe, director of the CAS Institute of Biophysics and CAS Member, has been awarded the 2006Trieste Science Prize for "his worldclass contributions to structural biology and his studies of viruses responsible for human diseases."

  19. Research Progress in the CAS Action Plan for the Development of Western China

    Institute of Scientific and Technical Information of China (English)

    Feng Renguo

    2005-01-01

    @@ To speed up the regional development in central and western China is a strategic decision made by the Chinese government at the turn of the century. For CAS research professionals, active participation into the campaign is a solemn historic commitment and a major task of the CAS-piloted national Knowledge Innovation Program. In early 2000, the CAS leadership formulated an Action Plan for Western China Development and initiated a research program aiming at the environmental evolution,ecological restoration and the sustainable exploitation of the local resources in the region.

  20. CRISPR/Cas9-mediated targeted gene mutagenesis in Spodoptera litura.

    Science.gov (United States)

    Bi, Hong-Lun; Xu, Jun; Tan, An-Jiang; Huang, Yong-Ping

    2016-06-01

    Custom-designed nuclease technologies such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated (Cas) system provide attractive genome editing tools for insect functional genetics. The targeted gene mutagenesis mediated by the CRISPR/Cas9 system has been achieved in several insect orders including Diptera, Lepidoptera and Coleoptera. However, little success has been reported in agricultural pests due to the lack of genomic information and embryonic microinjection techniques in these insect species. Here we report that the CRISPR/Cas9 system induced efficient gene mutagenesis in an important Lepidopteran pest Spodoptera litura. We targeted the S. litura Abdominal-A (Slabd-A) gene which is an important embryonic development gene and plays a significant role in determining the identities of the abdominal segments of insects. Direct injection of Cas9 messenger RNA and Slabd-A-specific single guide RNA (sgRNA) into S. litura embryos successfully induced the typical abd-A deficient phenotype, which shows anomalous segmentation and ectopic pigmentation during the larval stage. A polymerase chain reaction-based analysis revealed that the Cas9/sgRNA complex effectively induced a targeted mutagenesis in S. litura. These results demonstrate that the CRISPR/Cas9 system is a powerful tool for genome manipulation in Lepidopteran pests such as S. litura. PMID:27061764

  1. Multiplex gene editing of the Yarrowia lipolytica genome using the CRISPR-Cas9 system.

    Science.gov (United States)

    Gao, Shuliang; Tong, Yangyang; Wen, Zhiqiang; Zhu, Li; Ge, Mei; Chen, Daijie; Jiang, Yu; Yang, Sheng

    2016-08-01

    Yarrowia lipolytica is categorized as a generally recognized as safe (GRAS) organism and is a heavily documented, unconventional yeast that has been widely incorporated into multiple industrial fields to produce valuable biochemicals. This study describes the construction of a CRISPR-Cas9 system for genome editing in Y. lipolytica using a single plasmid (pCAS1yl or pCAS2yl) to transport Cas9 and relevant guide RNA expression cassettes, with or without donor DNA, to target genes. Two Cas9 target genes, TRP1 and PEX10, were repaired by non-homologous end-joining (NHEJ) or homologous recombination, with maximal efficiencies in Y. lipolytica of 85.6 % for the wild-type strain and 94.1 % for the ku70/ku80 double-deficient strain, within 4 days. Simultaneous double and triple multigene editing was achieved with pCAS1yl by NHEJ, with efficiencies of 36.7 or 19.3 %, respectively, and the pCASyl system was successfully expanded to different Y. lipolytica breeding strains. This timesaving method will enable and improve synthetic biology, metabolic engineering and functional genomic studies of Y. lipolytica. PMID:27349768

  2. Generation of knock-in primary human T cells using Cas9 ribonucleoproteins.

    Science.gov (United States)

    Schumann, Kathrin; Lin, Steven; Boyer, Eric; Simeonov, Dimitre R; Subramaniam, Meena; Gate, Rachel E; Haliburton, Genevieve E; Ye, Chun J; Bluestone, Jeffrey A; Doudna, Jennifer A; Marson, Alexander

    2015-08-18

    T-cell genome engineering holds great promise for cell-based therapies for cancer, HIV, primary immune deficiencies, and autoimmune diseases, but genetic manipulation of human T cells has been challenging. Improved tools are needed to efficiently "knock out" genes and "knock in" targeted genome modifications to modulate T-cell function and correct disease-associated mutations. CRISPR/Cas9 technology is facilitating genome engineering in many cell types, but in human T cells its efficiency has been limited and it has not yet proven useful for targeted nucleotide replacements. Here we report efficient genome engineering in human CD4(+) T cells using Cas9:single-guide RNA ribonucleoproteins (Cas9 RNPs). Cas9 RNPs allowed ablation of CXCR4, a coreceptor for HIV entry. Cas9 RNP electroporation caused up to ∼40% of cells to lose high-level cell-surface expression of CXCR4, and edited cells could be enriched by sorting based on low CXCR4 expression. Importantly, Cas9 RNPs paired with homology-directed repair template oligonucleotides generated a high frequency of targeted genome modifications in primary T cells. Targeted nucleotide replacement was achieved in CXCR4 and PD-1 (PDCD1), a regulator of T-cell exhaustion that is a validated target for tumor immunotherapy. Deep sequencing of a target site confirmed that Cas9 RNPs generated knock-in genome modifications with up to ∼20% efficiency, which accounted for up to approximately one-third of total editing events. These results establish Cas9 RNP technology for diverse experimental and therapeutic genome engineering applications in primary human T cells. PMID:26216948

  3. Genome modification by CRISPR/Cas9.

    Science.gov (United States)

    Ma, Yuanwu; Zhang, Lianfeng; Huang, Xingxu

    2014-12-01

    Clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas)9-mediated genome modification enables us to edit the genomes of a variety of organisms rapidly and efficiently. The advantages of the CRISPR-Cas9 system have made it an increasingly popular genetic engineering tool for biological and therapeutic applications. Moreover, CRISPR-Cas9 has been employed to recruit functional domains that repress/activate gene expression or label specific genomic loci in living cells or organisms, in order to explore developmental mechanisms, gene expression regulation, and animal behavior. One major concern about this system is its specificity; although CRISPR-Cas9-mediated off-target mutation has been broadly studied, more efforts are required to further improve the specificity of CRISPR-Cas9. We will also discuss the potential applications of CRISPR-Cas9. PMID:25315507

  4. Expanding CRISPR/Cas9 Genome Editing Capacity in Zebrafish Using SaCas9

    Science.gov (United States)

    Feng, Yan; Chen, Cheng; Han, Yuxiang; Chen, Zelin; Lu, Xiaochan; Liang, Fang; Li, Song; Qin, Wei; Lin, Shuo

    2016-01-01

    The type II CRISPR/Cas9 system has been used widely for genome editing in zebrafish. However, the requirement for the 5′-NGG-3′ protospacer-adjacent motif (PAM) of Cas9 from Streptococcus pyogenes (SpCas9) limits its targeting sequences. Here, we report that a Cas9 ortholog from Staphylococcus aureus (SaCas9), and its KKH variant, successfully induced targeted mutagenesis with high frequency in zebrafish. Confirming previous findings, the SpCas9 variant, VQR, can also induce targeted mutations in zebrafish. Bioinformatics analysis of these new Cas targets suggests that the number of available target sites in the zebrafish genome can be greatly expanded. Collectively, the expanded target repertoire of Cas9 in zebrafish should further facilitate the utility of this organism for genetic studies of vertebrate biology. PMID:27317783

  5. CAS - CERN Accelerator School: Course on Digital Signal Processing

    CERN Document Server

    Digital Signal Processing; CAS 2007

    2008-01-01

    These proceedings present the lectures given at the twenty-first specialized course organized by the CERN Accelerator School (CAS), the topic being Digital Signal Processing. The course was held in Sigtuna, Sweden, from 31 May–9 June 2007. This is the first time this topic has been selected for a specialized course. Taking into account the number of related applications currently in use in accelerators around the world, it was recognized that such a topic should definitively be incorporated into the CAS series of specialized courses. The specific aim of the course was to introduce the participants to the use and programming of Digital Signal Processors (DSPs) and Field Programmable Gate Arrays (FPGAs) evaluation boards. The course consisted of lectures in the mornings covering fundamental background knowledge in mathematics, controls theory, design tools, programming hardware platforms, and implementation details. In the afternoons the students split into two groups with people working in pairs. One group w...

  6. Impact of a physician-supervised exercise-nutrition program with testosterone substitution in partial androgen-deficient middle-aged obese men

    Institute of Scientific and Technical Information of China (English)

    Ernst R Schwarz; Robert D Willix Jr

    2011-01-01

    Background Partial androgen deficiency syndrome in the aging male is associated with signs of aging such as a development of abdominal obesity,sexual dysfunction,increase body fat,weight gain and the development of cardiac disease.Objective We assessed the outcome of a commercially available physician supervised nutrition and exercise program with concomitant testosterone replacement therapy in middle age obese men with partial androgen deficiency in order to reduce cardiac risks factors.Methods Fifty-six self referred men without diabetes mellitus,hypertension,or cardiovascular disease (ages 52.3 ± 7.8 years) were randomly selected from a large cohort.Baseline weight,body fat composition,fasting glucose,hemoglobin Alc and fasting lipid levels,as well as free and total testosterone levels were assessed.All patients were assessed and followed 6-18 months after initiation of the program.The program consisted of a low glycemic load balanced nutrition diet,a recommended structured daily exercise program of 30-60 minutes,as well as once to twice weekly intramuscular testosterone injections (113.0 ± 27.8 mg).Results At follow up,weight was reduced from 233.9 ± 30.0 pounds (lbs) to 221.3 ± 25.1 lbs (P < 0.001),BMI was reduced from 33.2 ± 3.3 kg/m2 to 31.3 ± 2.8 kg/m2 (P < 0.0001).Total body fat was 27.1% ± 5.2% vs.34.3% ± 5.7% at baseline (P < 0.0001).Fasting glucose was reduced from 95.3 ± 14.4 mg/dL to 87.5 ± 12.6 mg/dL (P < 0.0001).Total cholesterol was reduced from 195.4 ± 33.0 mg/dL to 172.7 ± 35.0 mg/dL (P < 0.005).No clinically significant adverse events were recorded.Conclusions Testosterone replacement therapy in middle aged obese men with partial androgen deficiency appeared safe and might have promoted the effects of a weight reduction diet and daily exercise program as long as an adequate physician supervision and follow up was granted.The combination therapy significantly reduced coronary risk factors such as glucose intolerance and

  7. f-divergences: Sufficiency, deficiency and testing of hypotheses

    Czech Academy of Sciences Publication Activity Database

    Vajda, Igor; Liese, F.

    Toronto : Nova Publishers, 2008 - (Barnett, N.; Dragomir, S.), s. 113-149 ISBN 1-60021-943-8. - (Advances in Mathematical Inequalities) R&D Projects: GA MŠk(CZ) 1M0572 Institutional research plan: CEZ:AV0Z10750506 Keywords : divergence * sufficiency * testing errors * information gain * deficiency * exponential rate Subject RIV: BB - Applied Statistics, Operational Research http://library.utia.cas.cz/separaty/2009/SI/vajda-fdivergences sufficiency deficiency and testing of hypotheses.pdf

  8. Exploiting CRISPR/Cas systems for biotechnology

    OpenAIRE

    Sampson, Timothy R.; Weiss, David S.

    2014-01-01

    The Cas9 endonuclease is the central component of the Type II CRISPR/Cas system, a prokaryotic adaptive restriction system against invading nucleic acids, such as those originating from bacteriophages and plasmids. Recently, this RNA-directed DNA endonuclease has been harnessed to target DNA sequences of interest. Here, we review the development of Cas9 as an important tool to not only edit the genomes of a number of different prokaryotic and eukaryotic species, but also as an efficient syste...

  9. Servei de Central Authentication Server (CAS)& Single Sign On (SSO): Cas pràctic

    OpenAIRE

    Marcos Rodríguez, Carlos

    2016-01-01

    Implementació d'un cas pràctic de Single Sign On mitjançant un servidor d'autenticació CAS (Central Authentication Server) Implementación de un caso práctico de Single Sign On mediante un servidor de autenticación CAS (Central Authentication Server) Implementation of a case study using a Single Sign On authentication server CAS (Central Authentication Server)

  10. Semiotic and discursive variables in cas-based didactical engineering

    DEFF Research Database (Denmark)

    Winsløw, Carl

    CAS, didactical engeneering, Maple, semiotics, undergraduate teaching, mathematics, education, didactics......CAS, didactical engeneering, Maple, semiotics, undergraduate teaching, mathematics, education, didactics...

  11. CAS Researcher Wins International Catalysis Award

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ Prof. Li Can, a CAS member and director of the State Key Laboratory of Catalysis at the CAS Dalian Institute of Chemical Physics, received the International Catalysis Award at the 13th International Catalysis Congress (ICC)held from July 11 to 16 in Paris.

  12. Smash and DASH with Cas9

    OpenAIRE

    Ramani, Vijay; Shendure, Jay

    2016-01-01

    DeRisi and colleagues present a creative application for Cas9 in vitro, using it to deplete unwanted sequence from DNA libraries. It seems plausible that the in vitro use of CRISPR/Cas9 has unrealized potential to revolutionize the practice of molecular biology well beyond genome editing.

  13. CAS paleoichthyologist gives Artedi Lecture in Sweden

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    @@ Prof. ZHANG Miman (CHANG Mee-mann), a CAS Member from the CAS Institute of Vertebrate Paleontology and Paleoanthropology, was invited to give a talk at the Artedi Lectures at the Royal Swedish Academy of Sciences in Stockholm, Sweden, on 5 December, 2008.

  14. Center established at CAS for innovation research

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    @@ The inaugural meeting for the CAS Center for Innovation and Development (CID) was held on 14 February in Beijing. Vice Chairman of the National Development and Reform Commission (NDRC) ZHANG Xiaoqiang and Member of CAS Presidium FANG Xin jointly unveiled the nameplate for the new center.

  15. 51 CAS Members Elected in 2005

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ Atotal of 51 prominent Chinese scientists were elected CAS members in 2005. The announce ment was made at a press conference held on Dec. 16 in Beijing. CAS President LU Yongxiang was present at the conference to give a briefing on the election and the new members.

  16. Iron deficiency

    DEFF Research Database (Denmark)

    Schou, Morten; Bosselmann, Helle; Gaborit, Freja;

    2015-01-01

    BACKGROUND: Both iron deficiency (ID) and cardiovascular biomarkers are associated with a poor outcome in heart failure (HF). The relationship between different cardiovascular biomarkers and ID is unknown, and the true prevalence of ID in an outpatient HF clinic is probably overlooked. OBJECTIVES...... understand iron metabolism in elderly HF patients....

  17. VLCAD deficiency

    DEFF Research Database (Denmark)

    Boneh, A; Andresen, B S; Gregersen, N; Ibrahim, M; Tzanakos, N; Peters, H; Yaplito-Lee, J; Pitt, J J

    2006-01-01

    We diagnosed six newborn babies with very long-chain acyl-CoA dehydrogenase deficiency (VLCADD) through newborn screening in three years in Victoria (prevalence rate: 1:31,500). We identified seven known and two new mutations in our patients (2/6 homozygotes; 4/6 compound heterozygotes). Blood sa...

  18. CAS at the Cockcroft Institute

    CERN Multimedia

    2007-01-01

    The CERN Accelerator School (CAS) and the UK’s new centre for accelerator science, the Cockcroft Institute, jointly organised a course on General Accelerator Physics, at intermediate level, at the Cockcroft Institute, Daresbury, UK, from 16 to 28 September 2007. The venue took advantage of the excellent new facilities in the Institute and the existing infrastructure of the adjacent Daresbury Laboratory. The intermediate level course followed established practice, with lectures on core topics in the mornings and specialised courses in the afternoons. The latter provided "hands-on" education and experience in the three selected topics: "RF Measurements Techniques", "Beam Instrumentation and Diagnostics" and "Optics Design and Correction". These proved to be highly successful, with participants choosing one course and following the topic throughout the school. Guided studies, tutorials, seminars and a poster session completed the prog...

  19. 48 CFR 9903.201-1 - CAS applicability.

    Science.gov (United States)

    2010-10-01

    ...-materials, and labor-hour contracts and subcontracts for the acquisition of commercial items. (7) Contracts...) shall be subject to CAS. A CAS-covered contract may be subject to full, modified or other types of CAS coverage. The rules for determining the applicable type of CAS coverage are in 9903.201-2. (b)...

  20. Committee set up at CAS to address scientific integrity

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    @@ On 30 April, the CAS Committee for Scientific Integrity held its first conference in Beijing. Chaired by CAS Vice President LI Jinghai, the Committee is composed of high-ranking CAS officials, S&T celebrities, and legal and policy experts.Its office is affiliated to the CAS Bureau of Supervision and Auditing.

  1. In vivo genome editing using Staphylococcus aureus Cas9

    OpenAIRE

    Ran, F Ann; Cong, Le; Yan, Winston X.; Scott, David A.; Gootenberg, Jonathan S.; Kriz, Andrea J.; Zetsche, Bernd; Shalem, Ophir; Wu, Xuebing; Makarova, Kira S.; Koonin, Eugene; Sharp, Phillip A.; Zhang, Feng

    2015-01-01

    The RNA-guided endonuclease Cas9 has emerged as a versatile genome-editing platform. However, the size of the commonly used Cas9 from Streptococcus pyogenes (SpCas9) limits its utility for basic research and therapeutic applications that employ the highly versatile adeno-associated virus (AAV) delivery vehicle. Here, we characterize six smaller Cas9 orthologs and show that Cas9 from Staphylococcus aureus (SaCas9) can edit the genome with efficiencies similar to those of SpCas9, while being >1...

  2. Transcriptional regulatory program in wild-type and retinoblastoma gene-deficient mouse embryonic fibroblasts during adipocyte differentiation

    DEFF Research Database (Denmark)

    Hakim-Weber, Robab; Krogsdam, Anne-M; Jørgensen, Claus;

    2011-01-01

    Although many molecular regulators of adipogenesis have been identified a comprehensive catalogue of components is still missing. Recent studies showed that the retinoblastoma protein (pRb) was expressed in the cell cycle and late cellular differentiation phase during adipogenesis. To investigate...... this dual role of pRb in the early and late stages of adipogenesis we used microarrays to perform a comprehensive systems-level analysis of the common transcriptional program of the classic 3T3-L1 preadipocyte cell line, wild-type mouse embryonic fibroblasts (MEFs), and retinoblastoma gene...... experimental data and computational analyses pinpointed a feedback-loop between Pparg and Foxo1.To analyze the effects of the retinoblastoma protein at the transcriptional level we chose a perturbated system (Rb-/- MEFs) for comparison to the transcriptional program of wild-type MEFs. Gene ontology analysis of...

  3. Cas9 specifies functional viral targets during CRISPR-Cas adaptation

    OpenAIRE

    Heler, Robert; Samai, Poulami; Modell, Joshua W.; Weiner, Catherine; Goldberg, Gregory W.; Bikard, David; Marraffini, Luciano A.

    2015-01-01

    Clustered, regularly interspaced, short palindromic repeat (CRISPR) loci and their associated (Cas) proteins provide adaptive immunity against viral infection in prokaryotes. Upon infection, short phage sequences known as spacers integrate between CRISPR repeats and are transcribed into small RNA guides that identify the viral targets (protospacers) of the Cas9 nuclease. Streptococcus pyogenes Cas9 cleavage of the viral genome requires the presence of an NGG protospacer adjacent motif (PAM) s...

  4. Cas9 function and host genome sampling in Type II-A CRISPR–Cas adaptation

    OpenAIRE

    Wei, Yunzhou; Terns, Rebecca M.; Terns, Michael P.

    2015-01-01

    Wei et al. found that Cas9, previously identified as the nuclease responsible for ultimate invader destruction, is also essential for adaptation in Streptococcus thermophilus. Cas9 nuclease activity is dispensable for adaptation. Wei et al. also revealed extensive, unbiased acquisition of the self-targeting host genome sequence by the CRISPR–Cas system that is masked in the presence of active target destruction.

  5. CasA mediates Cas3-catalyzed target degradation during CRISPR RNA-guided interference

    OpenAIRE

    Hochstrasser, Megan L.; Taylor, David W; Bhat, Prashant; Guegler, Chantal K.; Sternberg, Samuel H.; Nogales, Eva; Doudna, Jennifer A.

    2014-01-01

    Bacteria use clustered regularly interspaced short palindromic repeats (CRISPRs) together with CRISPR-associated (Cas) proteins to defend themselves against viral infection. The CRISPR locus contains short segments acquired from viral genomes, and RNAs derived from these segments assemble with Cas proteins into programmable DNA-binding complexes that target DNA molecules complementary to the guide RNA for cleavage. In type I CRISPR-Cas systems, the CRISPR-associated complex for antiviral defe...

  6. A newly discovered Bordetella species carries a transcriptionally active CRISPR-Cas with a small Cas9 endonuclease

    Science.gov (United States)

    The Cas9 endonuclease of the Type II-a clustered regularly interspersed short palindromic repeats (CRISPR), of Streptococcus pyogenes (SpCas9) has been adapted as a widely used tool for genome editing and genome engineering. Herein, we describe a gene encoding a novel Cas9 ortholog (BpsuCas9) and th...

  7. Classification and evolution of type II CRISPR-Cas systems

    OpenAIRE

    Chylinski, Krzysztof; Makarova, Kira S.; Charpentier, Emmanuelle; Eugene V Koonin

    2014-01-01

    The CRISPR-Cas systems of archaeal and bacterial adaptive immunity are classified into three types that differ by the repertoires of CRISPR-associated (cas) genes, the organization of cas operons and the structure of repeats in the CRISPR arrays. The simplest among the CRISPR-Cas systems is type II in which the endonuclease activities required for the interference with foreign deoxyribonucleic acid (DNA) are concentrated in a single multidomain protein, Cas9, and are guided by a co-processed ...

  8. CRISPR-Cas Functional Module Exchange in Escherichia coli

    OpenAIRE

    Almendros, Cristóbal; Mojica, Francisco J. M.; Díez-Villaseñor, César; Guzmán, Noemí M.; García-Martínez, Jesús

    2014-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (cas) genes constitute the CRISPR-Cas systems found in the Bacteria and Archaea domains. At least in some strains they provide an efficient barrier against transmissible genetic elements such as plasmids and viruses. Two CRISPR-Cas systems have been identified in Escherichia coli, pertaining to subtypes I-E (cas-E genes) and I-F (cas-F genes), respectively. In order to unveil the evolutionary dynamics of ...

  9. CAS physicist receives Richard Geller Prize

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    @@ Dr. SUN Liangting from the CAS Institute of Modern Physics has been awarded the Richard Geller Prize for his outstanding contributions to the development of Electron Cyclotron Resonance (ECR) ion sources, especially an all-permanent magnetic one.

  10. Heaviest yet LBO monocrystal grown at CAS

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ After three years of hard work, a research team led by Prof. HU Zhanggui with the CAS Technical Institute of Physics and Chemistry has made breakthrough progress in the studies of lithium triborate (LBO), a nonlinear optical crystal.

  11. AKRO/SF: Catch Accounting System (CAS)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Catch Accounting System (CAS) creates total catch estimates for the groundfish fisheries in the Bering Sea/Aleutian Islands and Gulf of Alaska. Each year,...

  12. Mega-science projects at CAS

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ With the support form Chinese government,CAS has been undertaking several national megascience projects in line with both the requirements of the national socioeconomic development and cuttingedge development of science and technology in the world.

  13. CRISPR/Cas systems in archaea

    OpenAIRE

    Gophna, Uri; Brodt, Avital

    2012-01-01

    CRISPR (Clustered, Regularly, Interspaced, Short, Palindromic Repeats) loci have been shown to provide prokaryotes with an adaptive immunity against viruses and plasmids. CRISPR arrays are transcribed and processed into small CRISPR RNA molecules, which base-pair with invading DNA or RNA and lead to its degradation by CRISPR-associated (Cas) protein complexes. New spacers can be acquired by active CRISPR/Cas systems, and thus the sequences of these spacers provide a record of the past “infect...

  14. A short-term arm-crank exercise program improved testosterone deficiency in adults with chronic spinal cord injury

    Directory of Open Access Journals (Sweden)

    Manuel Rosety-Rodriguez

    2014-06-01

    Full Text Available Purpose To determine the influence of arm-crank exercise in reproductive hormone levels in adults with chronic SCI. Further objectives were to assess the influence of arm-crank exercise on muscle strength and body composition. Materials and Methods Seventeen male adults with complete SCI at or below the 5th thoracic level (T5 volunteered for this study. Participants were randomly allocated to the intervention (n = 9 or control group (n = 8 using a concealed method. The participants in the intervention group performed a 12-week arm-crank exercise program, 3 sessions/week, consisting of warming-up (10-15 min followed by a main part in arm-crank (20-30 min [increasing 2 min and 30 seconds each three weeks] at a moderate work intensity of 50-65% of heart rate reserve (HRR (starting at 50% and increasing 5% each three weeks and by a cooling-down period (5-10 min. Serum follicle-stimulating hormone (FSH, luteinizing hormone (LH, testosterone and estradiol were determined by ELISA. Muscle strength (handgrip and body composition (waist circumference [WC] were assessed. Results After the completion of the training program, testosterone level was significantly increased (p = 0.0166;d = 1.14. Furthermore, maximal handgrip and WC were significantly improved. Lastly, a significant inverse correlation was found between WC and testosterone (r =- 0.35; p = 0.0377. Conclusion The arm-crank exercise improved reproductive hormone profile by increasing testosterone levels in adults with chronic SCI. A secondary finding was that it also significantly improved muscle strength and body composition in this group.

  15. Optimization of genome engineering approaches with the CRISPR/Cas9 system.

    Directory of Open Access Journals (Sweden)

    Kai Li

    Full Text Available Designer nucleases such as TALENS and Cas9 have opened new opportunities to scarlessly edit the mammalian genome. Here we explored several parameters that influence Cas9-mediated scarless genome editing efficiency in murine embryonic stem cells. Optimization of transfection conditions and enriching for transfected cells are critical for efficiently recovering modified clones. Paired gRNAs and wild-type Cas9 efficiently create programmed deletions, which facilitate identification of targeted clones, while paired gRNAs and the Cas9D10A nickase generated smaller targeted indels with lower chance of off-target mutagenesis. Genome editing is also useful for programmed introduction of exogenous DNA sequences at a target locus. Increasing the length of the homology arms of the homology-directed repair template strongly enhanced targeting efficiency, while increasing the length of the DNA insert reduced it. Together our data provide guidance on optimal design of scarless gene knockout, modification, or knock-in experiments using Cas9 nuclease.

  16. Optimization of Genome Engineering Approaches with the CRISPR/Cas9 System

    Science.gov (United States)

    Li, Kai; Wang, Gang; Andersen, Troels; Zhou, Pingzhu; Pu, William T.

    2014-01-01

    Designer nucleases such as TALENS and Cas9 have opened new opportunities to scarlessly edit the mammalian genome. Here we explored several parameters that influence Cas9-mediated scarless genome editing efficiency in murine embryonic stem cells. Optimization of transfection conditions and enriching for transfected cells are critical for efficiently recovering modified clones. Paired gRNAs and wild-type Cas9 efficiently create programmed deletions, which facilitate identification of targeted clones, while paired gRNAs and the Cas9D10A nickase generated smaller targeted indels with lower chance of off-target mutagenesis. Genome editing is also useful for programmed introduction of exogenous DNA sequences at a target locus. Increasing the length of the homology arms of the homology-directed repair template strongly enhanced targeting efficiency, while increasing the length of the DNA insert reduced it. Together our data provide guidance on optimal design of scarless gene knockout, modification, or knock-in experiments using Cas9 nuclease. PMID:25166277

  17. Hypomorphic phenotype of Foxn1 gene-modified rats by CRISPR/Cas9 system.

    Science.gov (United States)

    Goto, Teppei; Hara, Hiromasa; Nakauchi, Hiromitsu; Hochi, Shinichi; Hirabayashi, Masumi

    2016-08-01

    The Foxn1 gene is known as a critical factor for the differentiation of thymic and skin epithelial cells. This study was designed to examine the phenotype of Foxn1-modified rats generated by the CRISPR/Cas9 system. Guide-RNA designed for first exon of the Foxn1 and mRNA of Cas9 were co-injected into the pronucleus of Crlj:WI zygotes. Transfer of 158 injected zygotes resulted in the birth of 50 offspring (32 %), and PCR identified five (10 %) as Foxn1-edited. Genomic sequencing revealed the deletion of 44 or 60 bp from and/or insertion of 4 bp into the Foxn1 gene in a single allele. The number of T-cells in the peripheral blood lymphocytes of mutant rats decreased markedly. While homozygous deleted mutant rats had no thymus, the mutant rats were not completely hairless and showed normal performance in delivery and nursing. Splicing variants of the indel-mutation in the Foxn1 gene may cause hypomorphic allele, resulting in the phenotype of thymus deficiency and incomplete hairless. In conclusion, the mutant rats in Foxn1 gene edited by the CRISPR/Cas9 system showed the phenotype of thymus deficiency and incomplete hairless which was characterized by splicing variants. PMID:26931321

  18. Joint energy program makes progress

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    @@ "Clean Energies Facing the Future," a cooperative research program jointly organized by CAS and the BP Group, has made encouraging progress, say experts at an annual sum-up workshop held on 31 July and I August at Tsinghua University in Beijing. The CAS Dalian Institute of Chemical Physics(DICP) has been entrusted as coordinator of the cooperative program between the two sides.

  19. Genetics Home Reference: isolated growth hormone deficiency

    Science.gov (United States)

    ... deficiency dwarfism, pituitary growth hormone deficiency dwarfism isolated GH deficiency isolated HGH deficiency isolated human growth hormone deficiency isolated somatotropin deficiency isolated somatotropin deficiency disorder ...

  20. Inducible in vivo genome editing with CRISPR/Cas9

    OpenAIRE

    Dow, Lukas E.; Fisher, Jonathan; O'Rourke, Kevin P; Muley, Ashlesha; Kastenhuber, Edward R.; Livshits, Geulah; Tschaharganeh, Darjus F.; Socci, Nicholas D.; Lowe, Scott W.

    2015-01-01

    CRISPR/Cas9-based genome editing enables the rapid genetic manipulation of any genomic locus without the need for gene targeting by homologous recombination. Here we describe a conditional transgenic approach that allows temporal control of CRISPR/Cas9 activity for inducible genome editing in adult mice. We show that doxycycline-regulated Cas9 induction enables widespread gene disruption in multiple tissues and that limiting the duration of Cas9 expression or using a Cas9D10A (Cas9n) variant,...

  1. A study of the H2O maser emission from R Cas

    CERN Document Server

    Brand, J; Engels, D

    2001-01-01

    We describe the monitoring program we have been carrying out during the past 10 years on a sample of late-type stars, with the Medicina 32-m and Effelsberg 100-m antennas. We discuss the observations on the Mira-variable R Cas in some detail.

  2. Phylogeny of Cas9 determines functional exchangeability of dual-RNA and Cas9 among orthologous type II CRISPR-Cas systems.

    OpenAIRE

    Fonfara, Ines; Le Rhun, Anaïs; Chylinski, Krzysztof; Makarova, Kira S; Lécrivain, Anne-Laure; Bzdrenga, Janek; Koonin, Eugene V.; Charpentier, Emmanuelle

    2013-01-01

    The CRISPR-Cas-derived RNA-guided Cas9 endonuclease is the key element of an emerging promising technology for genome engineering in a broad range of cells and organisms. The DNA-targeting mechanism of the type II CRISPR-Cas system involves maturation of tracrRNA: crRNA duplex (dual-RNA), which directs Cas9 to cleave invading DNA in a sequence-specific manner, dependent on the presence of a Protospacer Adjacent Motif (PAM) on the target. We show that evolution of dual-RNA and Cas9 in bacteria...

  3. CRISPR/Cas9 for plant genome editing: accomplishments, problems and prospects.

    Science.gov (United States)

    Paul, Joseph W; Qi, Yiping

    2016-07-01

    The increasing burden of the world population on agriculture requires the development of more robust crops. Dissecting the basic biology that underlies plant development and stress responses will inform the design of better crops. One powerful tool for studying plants at the molecular level is the RNA-programmed genome editing system composed of a clustered regularly interspaced short palindromic repeats (CRISPR)-encoded guide RNA and the nuclease Cas9. Here, some of the recent advances in CRISPR/Cas9 technology that have profound implications for improving the study of plant biology are described. These tools are also paving the way towards new horizons for biotechnologies and crop development. PMID:27114166

  4. CAS enacts rules to guide management of its institutes

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ CAS issued Regulations for Comprehensive Administration of Research Institutes at the annual work conference opened on 24 March in Beijing. It is regarded as another cardinal document for CAS management after its Constitution, which was decreed in 2006.

  5. Northeast Institute of Geography and Agricultural Ecology, CAS

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ The Northeast Institute of Geography and Agricultural Ecology, a new CAS research body inaugurated in March 2002,comes from the merger of two former CAS research affiliates:Changchun Institute of Geography and Heilongjiang Institute of Agricultural Modernization in Harbin.

  6. The tracrRNA and Cas9 families of type II CRISPR-Cas immunity systems.

    OpenAIRE

    Chylinski, Krzysztof; Le Rhun, Anaïs; Charpentier, Emmanuelle

    2013-01-01

    CRISPR-Cas is a rapidly evolving RNA-mediated adaptive immune system that protects bacteria and archaea against mobile genetic elements. The system relies on the activity of short mature CRISPR RNAs (crRNAs) that guide Cas protein(s) to silence invading nucleic acids. A set of CRISPR-Cas, type II, requires a trans-activating small RNA, tracrRNA, for maturation of precursor crRNA (pre-crRNA) and interference with invading sequences. Following co-processing of tracrRNA and pre-crRNA by RNase II...

  7. Cas6 specificity and CRISPR RNA loading in a complex CRISPR-Cas system

    OpenAIRE

    Sokolowski, Richard David; Graham, Shirley; White, Malcolm F.

    2014-01-01

    This research was funded in part by Biotechnology and Biological Sciences Research Council [BB/K000314/1]. The APC was paid through RCUK OA block grant funds. CRISPR-Cas is an adaptive prokaryotic immune system, providing protection against viruses and other mobile genetic elements. In type I and type III CRISPR-Cas systems, CRISPR RNA (crRNA) is generated by cleavage of a primary transcript by the Cas6 endonuclease and loaded into multisubunit surveillance/effector complexes, allowing hom...

  8. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Deficiency Anemia Explore Iron-Deficiency Anemia What Is... CAUSES WHO IS AT RISK SIGNS & SYMPTOMS DIAGNOSIS TREATMENTS ... less hemoglobin than normal. Iron-deficiency anemia can cause fatigue (tiredness), shortness of breath, chest pain, and ...

  9. The 10th Session of the Fourth CAS Presidium Held in Beijing

    Institute of Scientific and Technical Information of China (English)

    Guo Haiyan

    2002-01-01

    @@ Chaired by Prof. LuYongxiang, executive chairman of the CAS Presidium and president of CAS, the 10th session of the fourth CAS Presidium was held on September 2 in Beijing. The CAS Presidium is the standing leading body of the CAS General Assembly, the highest organ of the CAS Academic Divisions (CASAD).

  10. Nucleosome breathing and remodeling constrain CRISPR-Cas9 function.

    OpenAIRE

    Isaac, RS; Jiang, F; Doudna, JA; Lim, WA; Narlikar, GJ; De Almeida, R

    2016-01-01

    The CRISPR-Cas9 bacterial surveillance system has become a versatile tool for genome editing and gene regulation in eukaryotic cells, yet how CRISPR-Cas9 contends with the barriers presented by eukaryotic chromatin is poorly understood. Here we investigate how the smallest unit of chromatin, a nucleosome, constrains the activity of the CRISPR-Cas9 system. We find that nucleosomes assembled on native DNA sequences are permissive to Cas9 action. However, the accessibility of nucleosomal DNA to ...

  11. Repurposing CRISPR/Cas9 for in situ functional assays

    OpenAIRE

    Malina, Abba; Mills, John R.; Cencic, Regina; YAN, YIFEI; Fraser, James; Schippers, Laura M.; Paquet, Marilène; Dostie, Josée; Pelletier, Jerry

    2013-01-01

    Malina et al. readapted the CRISPR/Cas9 genome-editing system for targeted gene disruption positive selection assays. They generated “all-in-one” lentiviral and retroviral delivery vectors heterologously expressing both a codon-optimized Cas9 and its synthetic guide RNA (sgRNA) and linked Cas9 expression to GFP fluorescence for in vivo applications. These results establish Cas9 genome editing as a powerful and practical approach for positive in situ genetic screens.

  12. Cas9-mediated targeting of viral RNA in eukaryotic cells

    OpenAIRE

    Price, Aryn A.; Sampson, Timothy R.; Ratner, Hannah K.; Grakoui, Arash; Weiss, David S

    2015-01-01

    The clustered, regularly interspaced, short palindromic repeats associated endonuclease, Cas9, has quickly become a revolutionary tool in genome engineering. Utilizing small guiding RNAs, Cas9 can be targeted to specific DNA sequences of interest, where it catalyzes DNA cleavage. We now demonstrate that Cas9 from the Gram-negative bacterium Francisella novicida (FnCas9) can be reprogrammed to target a specific RNA substrate, the genome of the +ssRNA virus, hepatitis C virus, in eukaryotic cel...

  13. Cas9 as a versatile tool for engineering biology

    OpenAIRE

    Mali, Prashant; Esvelt, Kevin M.; Church, George M.

    2013-01-01

    RNA-guided Cas9 nucleases derived from clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems have dramatically transformed our ability to edit the genomes of diverse organisms. We believe tools and techniques based on Cas9, a single unifying factor capable of colocalizing RNA, DNA and protein, will grant unprecedented control over cellular organization, regulation and behavior. Here we describe the Cas9 targeting methodology, detail current and prospective engineering...

  14. Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems

    OpenAIRE

    DiCarlo, James; Norville, Julie; Mali, Prashant; Rios Villanueva, Xavier; Aach, John Dennis; Church, George McDonald

    2013-01-01

    Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated (Cas) systems in bacteria and archaea use RNA-guided nuclease activity to provide adaptive immunity against invading foreign nucleic acids. Here, we report the use of type II bacterial CRISPR-Cas system in Saccharomyces cerevisiae for genome engineering. The CRISPR-Cas components, Cas9 gene and a designer genome targeting CRISPR guide RNA (gRNA), show robust and specific RNA-guided endonuclease activity a...

  15. Engineered CRISPR-Cas9 nucleases with altered PAM specificities

    Science.gov (United States)

    Kleinstiver, Benjamin P.; Prew, Michelle S.; Tsai, Shengdar Q.; Topkar, Ved; Nguyen, Nhu T.; Zheng, Zongli; Gonzales, Andrew P.W.; Li, Zhuyun; Peterson, Randall T.; Yeh, Jing-Ruey Joanna; Aryee, Martin J.; Joung, J. Keith

    2015-01-01

    Although CRISPR-Cas9 nucleases are widely used for genome editing1, 2, the range of sequences that Cas9 can recognize is constrained by the need for a specific protospacer adjacent motif (PAM)3–6. As a result, it can often be difficult to target double-stranded breaks (DSBs) with the precision that is necessary for various genome editing applications. The ability to engineer Cas9 derivatives with purposefully altered PAM specificities would address this limitation. Here we show that the commonly used Streptococcus pyogenes Cas9 (SpCas9) can be modified to recognize alternative PAM sequences using structural information, bacterial selection-based directed evolution, and combinatorial design. These altered PAM specificity variants enable robust editing of endogenous gene sites in zebrafish and human cells not currently targetable by wild-type SpCas9, and their genome-wide specificities are comparable to wild-type SpCas9 as judged by GUIDE-Seq analysis7. In addition, we identified and characterized another SpCas9 variant that exhibits improved specificity in human cells, possessing better discrimination against off-target sites with non-canonical NAG and NGA PAMs and/or mismatched spacers. We also found that two smaller-size Cas9 orthologues, Streptococcus thermophilus Cas9 (St1Cas9) and Staphylococcus aureus Cas9 (SaCas9), function efficiently in the bacterial selection systems and in human cells, suggesting that our engineering strategies could be extended to Cas9s from other species. Our findings provide broadly useful SpCas9 variants and, more importantly, establish the feasibility of engineering a wide range of Cas9s with altered and improved PAM specificities. PMID:26098369

  16. The 11th CAS General Assembly Held in Beijing

    Institute of Scientific and Technical Information of China (English)

    Guo Haiyan; Zhao Baohua

    2002-01-01

    @@ As their first plenary meetings in the new century, the 11th General Assembly of the Chinese Academy of Sciences (CAS) and the Sixth General Assembly of the Chinese Academy of Engineering (CAE) were convened in Beijing from May 28 to June 1. A total of 490 CAS members and three foreign members attended the CAS conference.

  17. Structural plasticity and in vivo activity of Cas1 from the type I-F CRISPR-Cas system.

    Science.gov (United States)

    Wilkinson, Max E; Nakatani, Yoshio; Staals, Raymond H J; Kieper, Sebastian N; Opel-Reading, Helen K; McKenzie, Rebecca E; Fineran, Peter C; Krause, Kurt L

    2016-04-15

    CRISPR-Cas systems are adaptive immune systems in prokaryotes that provide protection against viruses and other foreign DNA. In the adaptation stage, foreign DNA is integrated into CRISPR (clustered regularly interspaced short palindromic repeat) arrays as new spacers. These spacers are used in the interference stage to guide effector CRISPR associated (Cas) protein(s) to target complementary foreign invading DNA. Cas1 is the integrase enzyme that is central to the catalysis of spacer integration. There are many diverse types of CRISPR-Cas systems, including type I-F systems, which are typified by a unique Cas1-Cas2-3 adaptation complex. In the present study we characterize the Cas1 protein of the potato phytopathogen Pectobacterium atrosepticum, an important model organism for understanding spacer acquisition in type I-F CRISPR-Cas systems. We demonstrate by mutagenesis that Cas1 is essential for adaptation in vivo and requires a conserved aspartic acid residue. By X-ray crystallography, we show that although P. atrosepticum Cas1 adopts a fold conserved among other Cas1 proteins, it possesses remarkable asymmetry as a result of structural plasticity. In particular, we resolve for the first time a flexible, asymmetric loop that may be unique to type I-F Cas1 proteins, and we discuss the implications of these structural features for DNA binding and enzymatic activity. PMID:26929403

  18. Boosting plant immunity with CRISPR/Cas

    OpenAIRE

    Chaparro-Garcia, Angela; Kamoun, Sophien; Nekrasov, Vladimir

    2015-01-01

    CRISPR/Cas has recently been transferred to plants to make them resistant to geminiviruses, a damaging family of DNA viruses. We discuss the potential and the limitations of this method. See related Research: http://www.genomebiology.com/2015/16/1/238

  19. Boosting plant immunity with CRISPR/Cas

    OpenAIRE

    Chaparro-Garcia, Angela; Kamoun, Sophien; Nekrasov, Vladimir

    2015-01-01

    CRISPR/Cas has recently been transferred to plants to make them resistant to geminiviruses, a damaging family of DNA viruses. We discuss the potential and the limitations of this method.See related Research: http://www.genomebiology.com/2015/16/1/238.

  20. Cas9-triggered chain ablation of cas9 as a gene drive brake

    OpenAIRE

    Wu, Bing; Luo, Liqun; Gao, Xiaojing J

    2016-01-01

    With the advent of clustered, regularly interspaced, short palindromic repeats (CRISPR)–CRISPR-associated protein 9 (Cas9) technology, researchers can construct gene drives that can bias the inheritance of edited alleles to alter entire populations. As demonstrated with the mutagenic chain reaction in Drosophila4, the CRISPR-Cas9 system can propagate genomic modification together with the genome-editing machinery itself. Although gene drives might have the potential to control insect-borne di...

  1. CasHRA (Cas9-facilitated Homologous Recombination Assembly) method of constructing megabase-sized DNA.

    Science.gov (United States)

    Zhou, Jianting; Wu, Ronghai; Xue, Xiaoli; Qin, Zhongjun

    2016-08-19

    Current DNA assembly methods for preparing highly purified linear subassemblies require complex and time-consuming in vitro manipulations that hinder their ability to construct megabase-sized DNAs (e.g. synthetic genomes). We have developed a new method designated 'CasHRA (Cas9-facilitated Homologous Recombination Assembly)' that directly uses large circular DNAs in a one-step in vivo assembly process. The large circular DNAs are co-introduced into Saccharomyces cerevisiae by protoplast fusion, and they are cleaved by RNA-guided Cas9 nuclease to release the linear DNA segments for subsequent assembly by the endogenous homologous recombination system. The CasHRA method allows efficient assembly of multiple large DNA segments in vivo; thus, this approach should be useful in the last stage of genome construction. As a proof of concept, we combined CasHRA with an upstream assembly method (Gibson procedure of genome assembly) and successfully constructed a 1.03 Mb MGE-syn1.0 (Minimal Genome of Escherichia coli) that contained 449 essential genes and 267 important growth genes. We expect that CasHRA will be widely used in megabase-sized genome constructions. PMID:27220470

  2. Protein engineering of Cas9 for enhanced function

    OpenAIRE

    Oakes, Benjamin L.; Nadler, Dana C.; Savage, David F.

    2014-01-01

    CRISPR/Cas systems act to protect the cell from invading nucleic acids in many bacteria and archaea. The bacterial immune protein Cas9 is a component of one of these CRISPR/Cas systems and has recently been adapted as a tool for genome editing. Cas9 is easily targeted to bind and cleave a DNA sequence via a complimentary RNA; this straightforward programmability has gained Cas9 rapid acceptance in the field of genetic engineering. While this technology has developed quickly, a number of chall...

  3. DNA targeting specificity of RNA-guided Cas9 nucleases

    OpenAIRE

    Hsu, Patrick D.; Scott, David A.; Weinstein, Joshua A.; Ran, F Ann; Konermann, Silvana; Agarwala, Vineeta; Li, Yinqing; Fine, Eli J; Wu, Xuebing; Shalem, Ophir; Cradick, Thomas J.; Marraffini, Luciano A.; Bao, Gang; Zhang, Feng

    2013-01-01

    The Streptococcus pyogenes Cas9 (SpCas9) nuclease can be efficiently targeted to genomic loci by means of singleguide RNAs (sgRNAs) to enable genome editing1–10. Here, we characterize SpCas9 targeting specificity in human cells to inform the selection of target sites and avoid off-target effects. Our study evaluates >700 guide RNA variants and SpCas9-induced indel mutation levels at >100 predicted genomic off-target loci in 293T and 293FT cells. We find that SpCas9 tolerates mismatches betwee...

  4. CRISPR–Cas9-assisted recombineering in Lactobacillus reuteri

    OpenAIRE

    Oh, Jee-Hwan; van Pijkeren, Jan-Peter

    2014-01-01

    Clustered regularly interspaced palindromic repeats (CRISPRs) and the CRISPR-associated (Cas) nuclease protect bacteria and archeae from foreign DNA by site-specific cleavage of incoming DNA. Type-II CRISPR–Cas systems, such as the Streptococcus pyogenes CRISPR–Cas9 system, can be adapted such that Cas9 can be guided to a user-defined site in the chromosome to introduce double-stranded breaks. Here we have developed and optimized CRISPR–Cas9 function in the lactic acid bacterium Lactobacillus...

  5. Controlling UCAVs by JTACs in CAS missions

    Science.gov (United States)

    Kumaş, A. E.

    2014-06-01

    By means of evolving technology, capabilities of UAVs (Unmanned Aerial Vehicle)s are increasing rapidly. This development provides UAVs to be used in many different areas. One of these areas is CAS (Close Air Support) mission. UAVs have several advantages compared to manned aircraft, however there are also some problematic areas. The remote controlling of these vehicles from thousands of nautical miles away via satellite may lead to various problems both ethical and tactical aspects. Therefore, CAS missions require a good level of ALI (Air-Land Integration), a high SA (situational awareness) and precision engagement. In fact, there is an aware friendly element in the target area in CAS missions, unlike the other UAV operations. This element is an Airman called JTAC (Joint Terminal Attack Controller). Unlike the JTAC, UAV operators are too far away from target area and use the limited FOV (Field of View) provided by camera and some other sensor data. In this study, target area situational awareness of a UAV operator and a JTAC, in a high-risk mission for friendly ground forces and civilians such as CAS, are compared. As a result of this comparison, answer to the question who should control the UCAV (Unmanned Combat Aerial Vehicle) in which circumstances is sought. A literature review is made in UAV and CAS fields and recent air operations are examined. The control of UCAV by the JTAC is assessed by SWOT analysis and as a result it is deduced that both control methods can be used in different situations within the framework of the ROE (Rules Of Engagement) is reached.

  6. Structure and Engineering of Francisella novicida Cas9.

    Science.gov (United States)

    Hirano, Hisato; Gootenberg, Jonathan S; Horii, Takuro; Abudayyeh, Omar O; Kimura, Mika; Hsu, Patrick D; Nakane, Takanori; Ishitani, Ryuichiro; Hatada, Izuho; Zhang, Feng; Nishimasu, Hiroshi; Nureki, Osamu

    2016-02-25

    The RNA-guided endonuclease Cas9 cleaves double-stranded DNA targets complementary to the guide RNA and has been applied to programmable genome editing. Cas9-mediated cleavage requires a protospacer adjacent motif (PAM) juxtaposed with the DNA target sequence, thus constricting the range of targetable sites. Here, we report the 1.7 Å resolution crystal structures of Cas9 from Francisella novicida (FnCas9), one of the largest Cas9 orthologs, in complex with a guide RNA and its PAM-containing DNA targets. A structural comparison of FnCas9 with other Cas9 orthologs revealed striking conserved and divergent features among distantly related CRISPR-Cas9 systems. We found that FnCas9 recognizes the 5'-NGG-3' PAM, and used the structural information to create a variant that can recognize the more relaxed 5'-YG-3' PAM. Furthermore, we demonstrated that the FnCas9-ribonucleoprotein complex can be microinjected into mouse zygotes to edit endogenous sites with the 5'-YG-3' PAM, thus expanding the target space of the CRISPR-Cas9 toolbox. PMID:26875867

  7. Photoactivatable CRISPR-Cas9 for optogenetic genome editing.

    Science.gov (United States)

    Nihongaki, Yuta; Kawano, Fuun; Nakajima, Takahiro; Sato, Moritoshi

    2015-07-01

    We describe an engineered photoactivatable Cas9 (paCas9) that enables optogenetic control of CRISPR-Cas9 genome editing in human cells. paCas9 consists of split Cas9 fragments and photoinducible dimerization domains named Magnets. In response to blue light irradiation, paCas9 expressed in human embryonic kidney 293T cells induces targeted genome sequence modifications through both nonhomologous end joining and homology-directed repair pathways. Genome editing activity can be switched off simply by extinguishing the light. We also demonstrate activation of paCas9 in spatial patterns determined by the sites of irradiation. Optogenetic control of targeted genome editing should facilitate improved understanding of complex gene networks and could prove useful in biomedical applications. PMID:26076431

  8. Unification of Cas protein families and a simple scenario for the origin and evolution of CRISPR-Cas systems

    Directory of Open Access Journals (Sweden)

    Wolf Yuri I

    2011-07-01

    Full Text Available Abstract Background The CRISPR-Cas adaptive immunity systems that are present in most Archaea and many Bacteria function by incorporating fragments of alien genomes into specific genomic loci, transcribing the inserts and using the transcripts as guide RNAs to destroy the genome of the cognate virus or plasmid. This RNA interference-like immune response is mediated by numerous, diverse and rapidly evolving Cas (CRISPR-associated proteins, several of which form the Cascade complex involved in the processing of CRISPR transcripts and cleavage of the target DNA. Comparative analysis of the Cas protein sequences and structures led to the classification of the CRISPR-Cas systems into three Types (I, II and III. Results A detailed comparison of the available sequences and structures of Cas proteins revealed several unnoticed homologous relationships. The Repeat-Associated Mysterious Proteins (RAMPs containing a distinct form of the RNA Recognition Motif (RRM domain, which are major components of the CRISPR-Cas systems, were classified into three large groups, Cas5, Cas6 and Cas7. Each of these groups includes many previously uncharacterized proteins now shown to adopt the RAMP structure. Evidence is presented that large subunits contained in most of the CRISPR-Cas systems could be homologous to Cas10 proteins which contain a polymerase-like Palm domain and are predicted to be enzymatically active in Type III CRISPR-Cas systems but inactivated in Type I systems. These findings, the fact that the CRISPR polymerases, RAMPs and Cas2 all contain core RRM domains, and distinct gene arrangements in the three types of CRISPR-Cas systems together provide for a simple scenario for origin and evolution of the CRISPR-Cas machinery. Under this scenario, the CRISPR-Cas system originated in thermophilic Archaea and subsequently spread horizontally among prokaryotes. Conclusions Because of the extreme diversity of CRISPR-Cas systems, in-depth sequence and structure

  9. Carnitine Deficiency and Pregnancy

    OpenAIRE

    Anouk de Bruyn; Yves Jacquemyn; Kristof Kinget; François Eyskens

    2015-01-01

    We present two cases of carnitine deficiency in pregnancy. In our first case, systematic screening revealed L-carnitine deficiency in the first born of an asymptomatic mother. In the course of her second pregnancy, maternal carnitine levels showed a deficiency as well. In a second case, a mother known with carnitine deficiency under supplementation was followed throughout her pregnancy. Both pregnancies had an uneventful outcome. Because carnitine deficiency can have serious complications, su...

  10. CRISPR-Cas9-assisted recombineering in Lactobacillus reuteri.

    Science.gov (United States)

    Oh, Jee-Hwan; van Pijkeren, Jan-Peter

    2014-01-01

    Clustered regularly interspaced palindromic repeats (CRISPRs) and the CRISPR-associated (Cas) nuclease protect bacteria and archeae from foreign DNA by site-specific cleavage of incoming DNA. Type-II CRISPR-Cas systems, such as the Streptococcus pyogenes CRISPR-Cas9 system, can be adapted such that Cas9 can be guided to a user-defined site in the chromosome to introduce double-stranded breaks. Here we have developed and optimized CRISPR-Cas9 function in the lactic acid bacterium Lactobacillus reuteri ATCC PTA 6475. We established proof-of-concept showing that CRISPR-Cas9 selection combined with single-stranded DNA (ssDNA) recombineering is a realistic approach to identify at high efficiencies edited cells in a lactic acid bacterium. We show for three independent targets that subtle changes in the bacterial genome can be recovered at efficiencies ranging from 90 to 100%. By combining CRISPR-Cas9 and recombineering, we successfully applied codon saturation mutagenesis in the L. reuteri chromosome. Also, CRISPR-Cas9 selection is critical to identify low-efficiency events such as oligonucleotide-mediated chromosome deletions. This also means that CRISPR-Cas9 selection will allow identification of recombinant cells in bacteria with low recombineering efficiencies, eliminating the need for ssDNA recombineering optimization procedures. We envision that CRISPR-Cas genome editing has the potential to change the landscape of genome editing in lactic acid bacteria, and other Gram-positive bacteria. PMID:25074379

  11. CRISPR–Cas9-assisted recombineering in Lactobacillus reuteri

    Science.gov (United States)

    Oh, Jee-Hwan; van Pijkeren, Jan-Peter

    2014-01-01

    Clustered regularly interspaced palindromic repeats (CRISPRs) and the CRISPR-associated (Cas) nuclease protect bacteria and archeae from foreign DNA by site-specific cleavage of incoming DNA. Type-II CRISPR–Cas systems, such as the Streptococcus pyogenes CRISPR–Cas9 system, can be adapted such that Cas9 can be guided to a user-defined site in the chromosome to introduce double-stranded breaks. Here we have developed and optimized CRISPR–Cas9 function in the lactic acid bacterium Lactobacillus reuteri ATCC PTA 6475. We established proof-of-concept showing that CRISPR–Cas9 selection combined with single-stranded DNA (ssDNA) recombineering is a realistic approach to identify at high efficiencies edited cells in a lactic acid bacterium. We show for three independent targets that subtle changes in the bacterial genome can be recovered at efficiencies ranging from 90 to 100%. By combining CRISPR–Cas9 and recombineering, we successfully applied codon saturation mutagenesis in the L. reuteri chromosome. Also, CRISPR–Cas9 selection is critical to identify low-efficiency events such as oligonucleotide-mediated chromosome deletions. This also means that CRISPR–Cas9 selection will allow identification of recombinant cells in bacteria with low recombineering efficiencies, eliminating the need for ssDNA recombineering optimization procedures. We envision that CRISPR–Cas genome editing has the potential to change the landscape of genome editing in lactic acid bacteria, and other Gram-positive bacteria. PMID:25074379

  12. CAS - Great success for the DSP course

    CERN Multimedia

    2007-01-01

    The CERN Accelerator School (CAS) and the Uppsala University jointly organized a specialized school on "Digital Signal Processing" in Sigtuna, Sweden from 1-9 June, 2007. This course was a "première" in many ways: firstly the topic had never been addressed by CAS, and secondly the structure of the course differed from the usual specialized courses in the sense that it was composed of 32 hours of theoretical lectures in the mornings and 16 hours "hands-on" courses in the afternoons. The latter, which have been designed by CERN experts, had some logistic implications in transporting computers and circuit boards (DSP and FPGA) to Sweden. The principle of this new approach was extremely well received by the accelerator community and 97 participants representing 23 different nationalities (80% of the participants originating from the CERN Member States) attended the course. As illustrated by the very positive feedback received from th...

  13. CAS Introduction to Accelerator Physics in Bulgaria

    CERN Multimedia

    CERN Bulletin

    2010-01-01

    The CERN Accelerator School (CAS) and the Institute for Nuclear Research & Nuclear Energy (INRNE – Bulgarian Academy of Sciences) jointly organised a course on Introduction to Accelerators, at the Grand Hotel Varna, Bulgaria, from 19 September to 1 October, 2010.   CERN Accelerator School group photo. The course was extremely well attended with 109 participants representing 34 different nationalities, coming from countries as far away as Australia, Canada and Vietnam. The intensive programme comprised 39 lectures, 3 seminars, 4 tutorials where the students were split into three groups, a poster session where students could present their own work, and 7 hours of guided and private study. Feedback from the participants was extremely positive, praising the expertise and enthusiasm of the lecturers, as well as the high standard and excellent quality of their lectures. For the first time at CAS, the CERN Director-General, Rolf Heuer, visited the school and presented a seminar entitled...

  14. CAS - CERN Accelerator School: RF for Accelerators

    CERN Document Server

    2012-01-01

    These proceedings present the lectures given at the twenty-fourth specialized course organized by the CERN Accelerator School (CAS). The course was held in Ebeltoft, Denmark, from 8-17 June, 2010 in collaboration with Aarhus University, with the topic 'RF for Accelerators' While this topic has been covered by CAS previously, early in the 1990s and again in 2000, it was recognized that recent advances in the field warranted an updated course. Following introductory courses covering the background physics, the course attempted to cover all aspects of RF for accelerators; from RF power generation and transport, through cavity and coupler design, electronics and low level control, to beam diagnostics and RF gymnastics. The lectures were supplemented with several sessions of exercises, which were completed by discussion sessions on the solutions.

  15. Efficient Mitochondrial Genome Editing by CRISPR/Cas9

    Directory of Open Access Journals (Sweden)

    Areum Jo

    2015-01-01

    Full Text Available The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR/Cas9 system has been widely used for nuclear DNA editing to generate mutations or correct specific disease alleles. Despite its flexible application, it has not been determined if CRISPR/Cas9, originally identified as a bacterial defense system against virus, can be targeted to mitochondria for mtDNA editing. Here, we show that regular FLAG-Cas9 can localize to mitochondria to edit mitochondrial DNA with sgRNAs targeting specific loci of the mitochondrial genome. Expression of FLAG-Cas9 together with gRNA targeting Cox1 and Cox3 leads to cleavage of the specific mtDNA loci. In addition, we observed disruption of mitochondrial protein homeostasis following mtDNA truncation or cleavage by CRISPR/Cas9. To overcome nonspecific distribution of FLAG-Cas9, we also created a mitochondria-targeted Cas9 (mitoCas9. This new version of Cas9 localizes only to mitochondria; together with expression of gRNA targeting mtDNA, there is specific cleavage of mtDNA. MitoCas9-induced reduction of mtDNA and its transcription leads to mitochondrial membrane potential disruption and cell growth inhibition. This mitoCas9 could be applied to edit mtDNA together with gRNA expression vectors without affecting genomic DNA. In this brief study, we demonstrate that mtDNA editing is possible using CRISPR/Cas9. Moreover, our development of mitoCas9 with specific localization to the mitochondria should facilitate its application for mitochondrial genome editing.

  16. A Sweeping Reshuffle at CAS Institutes

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ Reorganization of CAS institutes was conducted over the past four years. For example, with the merger of the former Institute of Mathematics,Institute of Applied Mathematics, Institute of Systems Science and Institute of Computational Mathematics & Scientific/Engineering Computing, an Academy of Mathematics & System Sciences was formed. In the Shanghai area, the eight former biological institutes was regrouped into four institutes and one center,which formed the newly established Shanghai Institutes for Biological Sciences.

  17. Modeling Human Severe Combined Immunodeficiency and Correction by CRISPR/Cas9-Enhanced Gene Targeting

    Directory of Open Access Journals (Sweden)

    Chia-Wei Chang

    2015-09-01

    Full Text Available Mutations of the Janus family kinase JAK3 gene cause severe combined immunodeficiency (SCID. JAK3 deficiency in humans is characterized by the absence of circulating T cells and natural killer (NK cells with normal numbers of poorly functioning B cells (T–B+NK–. Using SCID patient-specific induced pluripotent stem cells (iPSCs and a T cell in vitro differentiation system, we demonstrate a complete block in early T cell development of JAK3-deficient cells. Correction of the JAK3 mutation by CRISPR/Cas9-enhanced gene targeting restores normal T cell development, including the production of mature T cell populations with a broad T cell receptor (TCR repertoire. Whole-genome sequencing of corrected cells demonstrates no CRISPR/Cas9 off-target modifications. These studies describe an approach for the study of human lymphopoiesis and provide a foundation for gene correction therapy in humans with immunodeficiencies.

  18. Generation of mutant mice via the CRISPR/Cas9 system using FokI-dCas9.

    Science.gov (United States)

    Hara, Satoshi; Tamano, Moe; Yamashita, Satoshi; Kato, Tomoko; Saito, Takeshi; Sakuma, Tetsushi; Yamamoto, Takashi; Inui, Masafumi; Takada, Shuji

    2015-01-01

    Genome editing, which introduces mutations in genes of interest using artificial DNA nucleases such as the ZFN, TALEN, and CRISPR/Cas9 systems in living cells, is a useful tool for generating mutant animals. Although CRISPR/Cas9 provides advantages over the two other systems, such as an easier vector construction and high efficiency of genome editing, it raises concerns of off-target effects when single guide RNA (gRNA) is used. Recently, FokI-dCas9 (fCas9), a fusion protein comprised of the inactivated mutant form of Cas9 and the DNA nuclease domain of FokI, has been developed. It enables genome editing with reduced risks of off-target effects in mammalian cultured cell lines, as fCas9 requires gRNAs to bind opposite strands with an appropriate distance between them. Here, we demonstrated that fCas9 efficiently generates living mutant mice through microinjection of its mRNA and gRNAs into zygotes. A comparison of the relative efficiencies of genome editing using fCas9 and other modified Cas9s showed that these mutagenesis efficiencies are similar when the targets of two gRNAs are separated by an appropriate distance, suggesting that in addition to the ease of vector construction, fCas9 exhibit high efficiency in producing mutant mice and in reducing risks of off-target effects. PMID:26057433

  19. CRISPR-Cas9 Based Engineering of Actinomycetal Genomes

    DEFF Research Database (Denmark)

    Tong, Yaojun; Charusanti, Pep; Zhang, Lixin;

    2015-01-01

    facilitate the genetic manipulation of actinomycetes, we developed a highly efficient CRISPR-Cas9 system to delete gene(s) or gene cluster(s), implement precise gene replacements, and reversibly control gene expression in actinomycetes. We demonstrate our system by targeting two genes, actIORF1 (SCO5087) and...... actVB (SCO5092), from the actinorhodin biosynthetic gene cluster in Streptomyces coelicolor A3(2). Our CRISPR-Cas9 system successfully inactivated the targeted genes. When no templates for homology-directed repair (HDR) were present, the site-specific DNA double-strand breaks (DSBs) introduced by Cas9....... Moreover, we developed a system to efficiently and reversibly control expression of target genes, deemed CRISPRi, based on a catalytically dead variant of Cas9 (dCas9). The CRISPR-Cas9 based system described here comprises a powerful and broadly applicable set of tools to manipulate actinomycetal genomes....

  20. Development of the CAS-LIBB single-particle microbeam for localized irradiation of living cells

    Institute of Scientific and Technical Information of China (English)

    WANG Xufei; XU Mingliang; WU Lijun; WANG Shaohu; FENG Huiyun; ZHAN Furu; PENG Shixiang; HU Chundong; ZHANG Shuqing; CHENG Jianjun; SHI Zhongtao; WANG Xiaohua; YUAN Hang; YUAN Haitao; YU Zengliang; CHEN Lianyun; HU Zhiwen; LI Jun; WU Yu; CHEN Bin; HU Suhua; ZHANG Jun

    2004-01-01

    A single-particle microbeam facility has been constructed at the Key Laboratory of Ion Beam Bioengineering (LIBB), Chinese Academy of Sciences (CAS). The system was designed to deliver a defined numbers of hydrogen ions, produced by a van de Graaff accelerator, in an energy range of 2.0-3.0 MeV, into an area smaller than that of the nucleus of an individual living cell. The beam is collimated by a borosilicate glass capillary that forms the beam-line exit. An integrated computer program recognizes the cells and locates them one by one over the microbeam exit for irradiation. We present technical details of the CAS-LIBB microbeam facility, particularly on the collimator, hardware, control program, as well as cell irradiation protocols available. Various factors contributing to the targeting and positioning precision are discussed along with accuracy measurement results.

  1. Effect of a mandatory iodization program on thyroid gland volume based on individuals' age, gender, and preceding severity of dietary iodine deficiency: A prospective, population-based study

    DEFF Research Database (Denmark)

    Vejbjerg, Pernille; Knudsen, Nils; Perrild, Hans; Carle, Allan; Laurberg, Peter; Pedersen, Inge Bulow; Rasmussen, Lone Banke; Ovesen, Lars; Jorgensen, Torben

    2007-01-01

    = 3570) the iodization in year 2000 in two areas with mild and moderate iodine deficiency. Women aged 18-22, 25-30, 40-45, and 60-65 yr and men aged 60-65 yr were examined. Thyroid ultrasonography was performed. Results: A lower median thyroid volume was seen in all age groups after iodization. The...... volume in the age groups younger than 45 yr. When adjusted for confounders, a lower mean volume was seen among those with multiple nodules in both areas and in the group with diffuse structure in the area with moderate iodine deficiency. Before the iodization, 17.6% of the total cross-section had thyroid...... enlargement; after the iodization, 10.9% of the cross-section had thyroid enlargement. Conclusion: In this prospective study, we demonstrated a lower thyroid volume in all age groups after iodization of salt. The decline was largest in the area with former, moderate iodine deficiency. The equal volumes in the...

  2. Factor VII deficiency

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/000548.htm Factor VII deficiency To use the sharing features on this page, please enable JavaScript. Factor VII (seven) deficiency is a disorder caused by a ...

  3. Folate-deficiency anemia

    Science.gov (United States)

    ... medlineplus.gov/ency/article/000551.htm Folate-deficiency anemia To use the sharing features on this page, please enable JavaScript. Folate-deficiency anemia is a decrease in red blood cells (anemia) ...

  4. Leukocyte Adhesion Deficiency (LAD)

    Science.gov (United States)

    ... Content Marketing Share this: Main Content Area Leukocyte Adhesion Deficiency (LAD) LAD is an immune deficiency in ... are slow to heal also may have LAD. Treatment and Research Doctors prescribe antibiotics to prevent and ...

  5. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Deficiency Anemia What Is... CAUSES WHO IS AT RISK SIGNS & SYMPTOMS DIAGNOSIS TREATMENTS PREVENTION LIVING WITH CLINICAL ... and women are the two groups at highest risk for iron-deficiency anemia. Outlook Doctors usually can ...

  6. Factor V deficiency

    Science.gov (United States)

    Factor V deficiency is a condition that is passed down through families, which affects the ability of the blood ... These proteins are called blood coagulation factors. Factor V deficiency is caused by a lack of Factor ...

  7. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... and women are the two groups at highest risk for iron-deficiency anemia. Outlook Doctors usually can successfully ... With and Managing Iron-Deficiency Anemia 05/18/2011 This video— ...

  8. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... This Content: NEXT >> Featured Video Living With and Managing Iron-Deficiency Anemia 05/18/2011 This video— ... treatment. For more information about living with and managing iron-deficiency anemia, go to the Health Topics ...

  9. Familial lipoprotein lipase deficiency

    Science.gov (United States)

    ... medlineplus.gov/ency/article/000408.htm Familial lipoprotein lipase deficiency To use the sharing features on this page, please enable JavaScript. Familial lipoprotein lipase deficiency is a group of rare genetic disorders ...

  10. CRISPR/Cas9 advances engineering of microbial cell factories

    DEFF Research Database (Denmark)

    Jakociunas, Tadas; Jensen, Michael Krogh; Keasling, Jay D.

    2016-01-01

    interspaced palindromic repeats (CRISPR) and its associated proteins (Cas) have become the method of choice for precision genome engineering in many organisms due to their orthogonality, versatility and efficacy. Here we review the strategies adopted for implementation of RNA-guided CRISPR/Cas9 genome editing...... be highlighted. Finally, this review will provide a perspective on the immediate challenges and opportunities foreseen by the use of CRISPR/Cas9 genome engineering and regulation in the context of metabolic engineering....

  11. Multiplex Genome Engineering Using CRISPR/Cas Systems

    OpenAIRE

    Cong, Le; Ran, F. Ann; Cox, David; Lin, Shuailiang; Barretto, Robert; Habib, Naomi; Hsu, Patrick D.; Wu, Xuebing; Jiang, Wenyan; Marraffini, Luciano A.; Zhang, Feng

    2013-01-01

    Functional elucidation of causal genetic variants and elements requires precise genome editing technologies. The type II prokaryotic CRISPR (clustered regularly interspaced short palindromic repeats)/Cas adaptive immune system has been shown to facilitate RNA-guided site-specific DNA cleavage. We engineered two different type II CRISPR/Cas systems and demonstrate that Cas9 nucleases can be directed by short RNAs to induce precise cleavage at endogenous genomic loci in human and mouse cells. C...

  12. Targeted mutagenesis in chicken using CRISPR/Cas9 system

    OpenAIRE

    Isao Oishi; Kyoko Yoshii; Daichi Miyahara; Hiroshi Kagami; Takahiro Tagami

    2016-01-01

    The CRISPR/Cas9 system is a simple and powerful tool for genome editing in various organisms including livestock animals. However, the system has not been applied to poultry because of the difficulty in accessing their zygotes. Here we report the implementation of CRISPR/Cas9-mediated gene targeting in chickens. Two egg white genes, ovalbumin and ovomucoid, were efficiently (>90%) mutagenized in cultured chicken primordial germ cells (PGCs) by transfection of circular plasmids encoding Cas9, ...

  13. Control of gene expression by CRISPR-Cas systems

    OpenAIRE

    Bikard, David; Marraffini, Luciano A.

    2013-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) loci and their associated cas (CRISPR-associated) genes provide adaptive immunity against viruses (phages) and other mobile genetic elements in bacteria and archaea. While most of the early work has largely been dominated by examples of CRISPR-Cas systems directing the cleavage of phage or plasmid DNA, recent studies have revealed a more complex landscape where CRISPR-Cas loci might be involved in gene regulation. In this revi...

  14. Iron deficiency and cognition

    OpenAIRE

    Hulthén, Lena

    2003-01-01

    Iron deficiency is the most prevalent nutritional disorder in the world. One of the most worrying consequences of iron deficiency in children is the alteration of behaviour and cognitive performance. In iron-deficient children, striking behavioural changes are observed, such as reduced attention span, reduced emotional responsiveness and low scores on tests of intelligence. Animal studies on nutritional iron deficiency show effects on learning ability that parallel the human studies. Despite ...

  15. Emerging pratices for recordging written solutions in a cas classroom

    OpenAIRE

    Ball, Lynda

    2003-01-01

    Written solutions for mathematical problems have the potential to be very different in a CAS classroom compared to a non-CAS classroom. New opportunities in a classroom where CAS is the normal technology may require a rethinking of what constitutes a good written record of a solution for a problem. This paper reports on the approaches for modelling good written records in class examples that were used by three teachers teaching year 12 mathematics with CAS for the first time. It was found tha...

  16. Recent Advances in Genome Editing Using CRISPR/Cas9

    OpenAIRE

    Ding, Yuduan; Li, Hong; Chen, Ling-Ling; Xie, Kabin

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9 (CRISPR-associated nuclease 9) system is a versatile tool for genome engineering that uses a guide RNA (gRNA) to target Cas9 to a specific sequence. This simple RNA-guided genome-editing technology has become a revolutionary tool in biology and has many innovative applications in different fields. In this review, we briefly introduce the Cas9-mediated genome-editing method, summarize the recent advances in CRISPR/Cas9 ...

  17. The 6th Meeting of CAS Board of Executive Directors

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ The 6th meeting of the board of executive directors of China Association for Standardization (CAS) (the 2nd chairman meeting in 2005) was held in Beijing on January 19th, 2006. The attendees were Mr. Li Zhonghai, President of CAS, who presided over the meeting, Ms. Li Rui and Mr. Li Chuntian, Advisers of CAS, Sun Xiaokang and Shi Baoquan, Vice Directors of SAC (Standardization Administration of China), Jiao Yunqi, President of CNIS (China National Institute of Standardization), Zhang Jianquan, Director of SPC (the Standards Press of China), over 20executive directors and some CAS officers.

  18. Optical Control of CRISPR/Cas9 Gene Editing

    Science.gov (United States)

    Hemphill, James; Borchardt, Erin K.; Brown, Kalyn; Asokan, Aravind; Deiters, Alexander

    2016-01-01

    The CRISPR/Cas9 system has emerged as an important tool in biomedical research for a wide range of applications, with significant potential for genome engineering and gene therapy. In order to achieve conditional control of the CRISPR/Cas9 system, a genetically encoded light-activated Cas9 was engineered through the site-specific installation of a caged lysine amino acid. Several potential lysine residues were identified as viable caging sites that can be modified to optically control Cas9 function, as demonstrated through optical activation and deactivation of both exogenous and endogenous gene function. PMID:25905628

  19. CRISPR-Cas9-guided Genome Engineering in C. elegans

    Science.gov (United States)

    Kim, Hyun-Min; Colaiácovo, Monica P.

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) system is successfully being used for efficient and targeted genome editing in various organisms including the nematode C. elegans. Recent studies developed various CRISPR-Cas9 approaches to enhance genome engineering via two major DNA double-strand break repair pathways: non-homologous end joining and homologous recombination. Here we describe a protocol for Cas9-mediated C. elegans genome editing together with single guide RNA (sgRNA) and repair template cloning and injection methods required for delivering Cas9, sgRNAs and repair template DNA into the C. elegans germline. PMID:27366893

  20. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... the NHLBI on Twitter. What Is Iron-Deficiency Anemia? Español Iron-deficiency anemia is a common, easily ... Featured Video Living With and Managing Iron-Deficiency Anemia 05/18/2011 This video—presented by the ...

  1. Iron-Deficiency Anemia

    Science.gov (United States)

    ... the NHLBI on Twitter. What Is Iron-Deficiency Anemia? Español Iron-deficiency anemia is a common, easily ... Featured Video Living With and Managing Iron-Deficiency Anemia 05/18/2011 This video—presented by the ...

  2. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... page from the NHLBI on Twitter. What Is Iron-Deficiency Anemia? Español Iron-deficiency anemia is a ... Content: NEXT >> Featured Video Living With and Managing Iron-Deficiency Anemia 05/18/2011 This video—presented ...

  3. Generation of mutant mice via the CRISPR/Cas9 system using FokI-dCas9

    OpenAIRE

    Satoshi Hara; Moe Tamano; Satoshi Yamashita; Tomoko Kato; Takeshi Saito; Tetsushi Sakuma; Takashi Yamamoto; Masafumi Inui; Shuji Takada

    2015-01-01

    Genome editing, which introduces mutations in genes of interest using artificial DNA nucleases such as the ZFN, TALEN, and CRISPR/Cas9 systems in living cells, is a useful tool for generating mutant animals. Although CRISPR/Cas9 provides advantages over the two other systems, such as an easier vector construction and high efficiency of genome editing, it raises concerns of off-target effects when single guide RNA (gRNA) is used. Recently, FokI-dCas9 (fCas9), a fusion protein comprised of the ...

  4. High Prevalence of Vitamin D Deficiency in Native versus Migrant Mothers and Newborns in the North of Italy: A Call to Act with a Stronger Prevention Program

    OpenAIRE

    Cadario, Francesco; Savastio, Silvia; Magnani, Corrado; Cena, Tiziana; Pagliardini, Veronica; Bellomo, Giorgio; Bagnati, Marco; Vidali, Matteo; Pozzi, Erica; Pamparana, Stella; Zaffaroni, Mauro; Genoni, Giulia; Bona, Gianni

    2015-01-01

    Background Vitamin D status during pregnancy is related to neonatal vitamin D status. Vitamin D deficiency has been associated with an increased risk of rickets in children and osteomalacia in adults. Aim of this study was to investigate 25OHD levels in maternal serum and in neonatal blood spots in native and migrant populations living in Novara (North Italy, 45°N latitude). Methods and Findings We carried out a cross sectional study from April 1st 2012 to March 30th 2013, in a tertiary Care ...

  5. Recent Advances in Genome Editing Using CRISPR/Cas9

    Science.gov (United States)

    Ding, Yuduan; Li, Hong; Chen, Ling-Ling; Xie, Kabin

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9 (CRISPR-associated nuclease 9) system is a versatile tool for genome engineering that uses a guide RNA (gRNA) to target Cas9 to a specific sequence. This simple RNA-guided genome-editing technology has become a revolutionary tool in biology and has many innovative applications in different fields. In this review, we briefly introduce the Cas9-mediated genome-editing method, summarize the recent advances in CRISPR/Cas9 technology, and discuss their implications for plant research. To date, targeted gene knockout using the Cas9/gRNA system has been established in many plant species, and the targeting efficiency and capacity of Cas9 has been improved by optimizing its expression and that of its gRNA. The CRISPR/Cas9 system can also be used for sequence-specific mutagenesis/integration and transcriptional control of target genes. We also discuss off-target effects and the constraint that the protospacer-adjacent motif (PAM) puts on CRISPR/Cas9 genome engineering. To address these problems, a number of bioinformatic tools are available to help design specific gRNAs, and new Cas9 variants and orthologs with high fidelity and alternative PAM specificities have been engineered. Owing to these recent efforts, the CRISPR/Cas9 system is becoming a revolutionary and flexible tool for genome engineering. Adoption of the CRISPR/Cas9 technology in plant research would enable the investigation of plant biology at an unprecedented depth and create innovative applications in precise crop breeding. PMID:27252719

  6. CAS and NSFC Strengthen Partnership in Strategy Studies

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    On April 5, 2012, CAS and the National Natural Science Foundation of China (NSFC) signed a cooperative framework agreement on the strategic research of academic disciplinary development at the launching ceremony of the book series China's Discipline Development Strategy in the Next Ten Years at the CAS Headquarters in Beijing.

  7. Recent Progress in CRISPR/Cas9 Technology.

    Science.gov (United States)

    Mei, Yue; Wang, Yan; Chen, Huiqian; Sun, Zhong Sheng; Ju, Xing-Da

    2016-02-20

    The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system, a simple and efficient tool for genome editing, has experienced rapid progress in its technology and applicability in the past two years. Here, we review the recent advances in CRISPR/Cas9 technology and the ways that have been adopted to expand our capacity for precise genome manipulation. First, we introduce the mechanism of CRISPR/Cas9, including its biochemical and structural implications. Second, we highlight the latest improvements in the CRISPR/Cas9 system, especially Cas9 protein modifications for customization. Third, we review its current applications, in which the versatile CRISPR/Cas9 system was employed to edit the genome, epigenome, or RNA of various organisms. Although CRISPR/Cas9 allows convenient genome editing accompanied by many benefits, we should not ignore the significant ethical and biosafety concerns that it raises. Finally, we discuss the prospective applications and challenges of several promising techniques adapted from CRISPR/Cas9. PMID:26924689

  8. CAS, MPG Set up New Partner Groups in Shanghai

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ CAS and the Max Planck Society (MPS) for the Advancement of Science have established two more partner groups in China. On August 3, CAS President LU Yongxiang and MPS President Peter Gruss were present at the inaugural ceremony of the groups in Shanghai.

  9. CAS conducts ex-post evaluation on research projects

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ Under the auspices of the CAS Center for Project Supervision and Evaluation, a panel of experts recently conducted an ex-post evaluation on a research project entitled "the electrical system for an electric vehicle." The appraisal is the first of its kind in project management at CAS.

  10. CRISPR-Cas9 Genome Engineering in Saccharomyces cerevisiae Cells.

    Science.gov (United States)

    Ryan, Owen W; Poddar, Snigdha; Cate, Jamie H D

    2016-01-01

    This protocol describes a method for CRISPR-Cas9-mediated genome editing that results in scarless and marker-free integrations of DNA into Saccharomyces cerevisiae genomes. DNA integration results from cotransforming (1) a single plasmid (pCAS) that coexpresses the Cas9 endonuclease and a uniquely engineered single guide RNA (sgRNA) expression cassette and (2) a linear DNA molecule that is used to repair the chromosomal DNA damage by homology-directed repair. For target specificity, the pCAS plasmid requires only a single cloning modification: replacing the 20-bp guide RNA sequence within the sgRNA cassette. This CRISPR-Cas9 protocol includes methods for (1) cloning the unique target sequence into pCAS, (2) assembly of the double-stranded DNA repair oligonucleotides, and (3) cotransformation of pCAS and linear repair DNA into yeast cells. The protocol is technically facile and requires no special equipment. It can be used in any S. cerevisiae strain, including industrial polyploid isolates. Therefore, this CRISPR-Cas9-based DNA integration protocol is achievable by virtually any yeast genetics and molecular biology laboratory. PMID:27250940

  11. First International Student Obtains His PhD at CAS

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    @@ Mr. Ghulam Rasul, the first international student formally enrolled by the Graduate University of CAS (GUCAS) has received his PhD degree this summer after having passed his dissertation defense on May 21 at the CAS Institute of Atmospheric Physics (IAP).

  12. CAS Scientist Awarded Tyler Prize for Environment Achievement

    Institute of Scientific and Technical Information of China (English)

    Guo Haiyan; Zhao Baohua

    2002-01-01

    @@ Prof. Liu Dongsheng (Tungsheng Liu), a CAS member and professor at CAS Institute of Geology and Geophysics, has been awarded the 2002 Tyler Prize for Environment Achievement. He shared a cash prize of $200,000 with the other laureate, Dr. Wallace S. Broecker from Columbia University, the US, and received a gold Tyler Prize medallion on April 12.

  13. CAS issues its 2007 prizes for outstanding achievements

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ One individual and three research groups have been honored with the prestigious CAS Awards for Outstanding S&T Achievements in 2007. CAS President LU Yongxiang conferred the awards on the laureates at the Academy's annual work conference opened on 24 March in Beijing.

  14. Diversity of CRISPR-Cas immune systems and molecular machines

    OpenAIRE

    Barrangou, Rodolphe

    2015-01-01

    Bacterial adaptive immunity hinges on CRISPR-Cas systems that provide DNA-encoded, RNA-mediated targeting of exogenous nucleic acids. A plethora of CRISPR molecular machines occur broadly in prokaryotic genomes, with a diversity of Cas nucleases that can be repurposed for various applications.

  15. Teaching Undergraduate Mathematics Using CAS Technology: Issues and Prospects

    Science.gov (United States)

    Tobin, Patrick C.; Weiss, Vida

    2016-01-01

    The use of handheld CAS technology in undergraduate mathematics courses in Australia is paradoxically shrinking under sustained disapproval or disdain from the professional mathematics community. Mathematics education specialists argue with their mathematics colleagues over a range of issues in course development and this use of CAS or even…

  16. 48 CFR 9903.201-2 - Types of CAS coverage.

    Science.gov (United States)

    2010-10-01

    ... coverage. Full coverage requires that the business unit comply with all of the CAS specified in part 9904... later award of a CAS-covered contract. Full coverage applies to contractor business units that— (1... covered contract of less than $50 million awarded to a business unit that received less than $50...

  17. [CI] 492 GHz mapping toward Cas A

    CERN Document Server

    Mookerjea, B; Roshi, D A; Masur, M

    2006-01-01

    We have mapped the [C I] emission at 492 GHz toward the supernova remnant Cas A. We detect [C I] emission from the periphery of the diffuse Photon Dominated Region (PDR) covering the disk of Cas A, as traced by the carbon recombination lines, as well as from the denser PDRs associated with the molecular clouds towards the south-east. [C I] emission is detected from both the Perseus and Orion arm molecular clouds, with the -47 km s^-1 Perseus arm feature being strong enough to be detected at all positions. We estimate the C/CO relative abundance to be 0.2 at the position of the identified CO clouds and >1 for most of the cloud. Here we show that the distribution of [C I] emitting regions compared to the C+ region and molecular cloud is consistent with a scenario involving PDRs. Using physical models for PDRs we constrain the physical properties of the [C I] line-forming regions. We estimate the densities of the [C I] emitting regions to be between 10^2 and 10^3 cm^-3. Based on rather high volume filling factor...

  18. CAS CERN Accelerator School vacuum technology. Proceedings

    International Nuclear Information System (INIS)

    These proceedings present the lectures given at the twelfth specialized course organized by the CERN Accelerator School (CAS), the topic this time being 'Vacuum Technology'. Despite the importance of vacuum technology in the design and operation of particle accelerators at CERN and at the many other accelerators already installed around the world, this was the first time that CAS has organized a course devoted entirely to this topic. Perhaps this reflects the facts that vacuum has become one of the more critical aspects of future accelerators, and that many of the pioneers in the accelerator field are being replaced by new, younger personnel. The lectures start with the basic concepts of the physics and technology of vacuum followed by detailed descriptions of the many different types of gas-pumping devices and methods to measure the pressures achieved. The outgassing characteristics of the different materials used in the construction of vacuum systems and the optimisation of cleaning methods to reduce this outgassing are then explained together with the effects of the residual gases on the particle beams. Then follow chapters on leak detection, materials and vacuum system engineering. Finally, seminars are presented on designing vacuum systems, the history of vacuum devices, the LHC (large hadron collider) vacuum system, vacuum systems for electron storage rings, and quality assurance for vacuum. (orig.)

  19. Trichobezoar gastrique - à propos de deux cas

    Directory of Open Access Journals (Sweden)

    Karim Ibn Majdoub Hassani

    2010-09-01

    Full Text Available Le trichobezoar gastrique est une affection rare (un peu plus d’une dizaine de cas dans la littérature, qui affecte essentiellement des jeunes filles perturbées par des désordres psychologiques. Les auteurs rapportent deux cas de jeunes filles, hospitalisées pour volumineuse masse épigastrique. La fibroscopie gastrique a posé le diagnostic de trichobezoar. Une exérèse chirurgicale a été réalisée à travers une gastrotomie, sans complications. Un suivi psychiatrique des deux patientes a été recommandé. Le trichobezoar gastrique désigne l’accumulation inhabituelle de cheveux au niveau de l’estomac. Son diagnostic est facile en présence d’un contexte de trichophagie évocateur. La fibroscopie œsogastroduodénale est l’examen de référence permettant la visualisation du trichobezoar dont le traitement est essentiellement chirurgical.

  20. Implementing the CAS Standards: The Implementation of the CAS Standards in Student Affairs as a Comprehensive Assessment Approach

    Science.gov (United States)

    Dorman, Jesse A.

    2012-01-01

    The increasing use of the CAS standards as a comprehensive assessment approach in divisions of student affairs necessitates a more in-depth understanding of how the CAS standards are being implemented in these settings. In response to increasing calls for improvement, accountability and professionalism in student affairs (Bresciani, 2006; Cooper…

  1. CRISPR/Cas9系统的脱靶效应%Off-target of CRISPR/Cas9 System

    Institute of Scientific and Technical Information of China (English)

    尹珅; 贺桂芳; 赖方秾; 谢凤云; 马俊宇

    2016-01-01

    在细菌中发现的免疫系统CRISPR/Cas9,已经成为最有效的基因工程编辑工具,甚至大有希望可以治疗人类遗传性疾病。但是CRISPR/Cas9系统在使用时会产生严重的脱靶问题,导致假表型和错误的解释。提高与靶点结合的高效率,同时减少脱靶效应,将是今后CRISPR/Cas9技术的挑战。综述关注与CRISPR/Cas9脱靶效应相关的内容,总结了影响其靶点专一性的因素,减少CRISPR/Cas9脱靶效应的可行性方法和设计工具等,供大家学习讨论。%As an adaptive immune system discovered in bacteria,clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)technology has been used as the most effective genome-editing tool. By now,CRISPR/Cas9 has shown a great promise in healing human genetic diseases,however,off-target is a critical issue while applying it. The off-target of CRISPR/Cas9 leads to the false phenotype and wrong interpretation. Optimization of on-target activity and minimizing off-target activity will be challenges while CRISPR/Cas9 is applied in the future. In this review,we focused on the off-target related issues,including factors involved in the target specificity,strategies and tools of minimizing the off-target of CRISPR/Cas9.

  2. Vitamin D deficiency in Europe

    DEFF Research Database (Denmark)

    Cashman, Kevin D.; Dowling, Kirsten G; Škrabáková, Zuzana;

    2016-01-01

    BACKGROUND: Vitamin D deficiency has been described as being pandemic, but serum 25-hydroxyvitamin D [25(OH)D] distribution data for the European Union are of very variable quality. The NIH-led international Vitamin D Standardization Program (VDSP) has developed protocols for standardizing existing...... 25(OH)D values from national health/nutrition surveys. OBJECTIVE: This study applied VDSP protocols to serum 25(OH)D data from representative childhood/teenage and adult/older adult European populations, representing a sizable geographical footprint, to better quantify the prevalence of vitamin D...... deficiency in Europe. DESIGN: The VDSP protocols were applied in 14 population studies [reanalysis of subsets of serum 25(OH)D in 11 studies and complete analysis of all samples from 3 studies that had not previously measured it] by using certified liquid chromatography-tandem mass spectrometry on biobanked...

  3. Pilomatricome: étude de 22 cas

    Science.gov (United States)

    Nasreddine, Fatima Zahra; Hali, Fouzia; Chiheb, Soumiya

    2016-01-01

    Le pilomatricome est une tumeur cutanée fréquente et bénigne du follicule pileux chez l'enfant. C'est une tumeur annexielle souvent méconnue et confondue avec d'autres lésions cutanées. Les localisations habituelles sont la tête et le cou. Le but de ce travail est de rapporter une série de 22 cas comportant des formes inhabituelles colligées au service de dermatologie sur une période allant de Janvier 2006 jusqu'au Mai 2015. L’étude a concerné 16 femmes et 6 hommes. La moyenne d’âge était de 23,3 ans (4-80 ans). La localisation cervico faciale a été observée dans 12 cas, 2 patients avaient des localisations multiples, un garçon de 4ans avait une localisation au niveau fronto-temporal et une fillette de 14 ans avait une localisation au niveau du visage et de l'avant-bras, et un patient de 48 ans avait une localisation sous unguéale. L'aspect clinique était typique dans tous les cas avec des nodules sous cutanés de consistance pierreuse. Tous les patients ont bénéficié d'une exérèse des nodules sous anesthésie locale. L’étude histologique était en faveur d'un épithélioma momifié de Malherbe d'exérèse complète sans signes de malignité. Aucun patient n'a présenté de rechute. L'originalité de notre étude réside dans la présence de localisations exceptionnelles au niveau latéro-vertébral, des membres et sous-unguéale, l’âge de survenue inhabituel à 80 ans et la présence de localisations multiples signalées chez 2 enfants. PMID:27516819

  4. CRISPR/Cas9-mediated viral interference in plants

    KAUST Repository

    Ali, Zahir

    2015-11-11

    Background The CRISPR/Cas9 system provides bacteria and archaea with molecular immunity against invading phages and conjugative plasmids. Recently, CRISPR/Cas9 has been used for targeted genome editing in diverse eukaryotic species. Results In this study, we investigate whether the CRISPR/Cas9 system could be used in plants to confer molecular immunity against DNA viruses. We deliver sgRNAs specific for coding and non-coding sequences of tomato yellow leaf curl virus (TYLCV) into Nicotiana benthamiana plants stably overexpressing the Cas9 endonuclease, and subsequently challenge these plants with TYLCV. Our data demonstrate that the CRISPR/Cas9 system targeted TYLCV for degradation and introduced mutations at the target sequences. All tested sgRNAs exhibit interference activity, but those targeting the stem-loop sequence within the TYLCV origin of replication in the intergenic region (IR) are the most effective. N. benthamiana plants expressing CRISPR/Cas9 exhibit delayed or reduced accumulation of viral DNA, abolishing or significantly attenuating symptoms of infection. Moreover, this system could simultaneously target multiple DNA viruses. Conclusions These data establish the efficacy of the CRISPR/Cas9 system for viral interference in plants, thereby extending the utility of this technology and opening the possibility of producing plants resistant to multiple viral infections.

  5. CAS Accelerator Physics held in Erice, Italy

    CERN Multimedia

    CERN Accelerator School

    2013-01-01

    The CERN Accelerator School (CAS) recently organised a specialised course on Superconductivity for Accelerators, held at the Ettore Majorana Foundation and Centre for Scientific Culture in Erice, Italy from 24 April-4 May, 2013.   Photo courtesy of Alessandro Noto, Ettore Majorana Foundation and Centre for Scientific Culture. Following a handful of summary lectures on accelerator physics and the fundamental processes of superconductivity, the course covered a wide range of topics related to superconductivity and highlighted the latest developments in the field. Realistic case studies and topical seminars completed the programme. The school was very successful with 94 participants representing 23 nationalities, coming from countries as far away as Belorussia, Canada, China, India, Japan and the United States (for the first time a young Ethiopian lady, studying in Germany, attended this course). The programme comprised 35 lectures, 3 seminars and 7 hours of case study. The case studies were p...

  6. CAS Accelerator Physics (Ion Sources) in Slovakia

    CERN Multimedia

    CAS School

    2012-01-01

    The CERN Accelerator School (CAS) and the Slovak University of Technology jointly organised a specialised course on ion sources, held at the Hotel Senec, Senec, Slovakia, from 29 May to 8 June, 2012.   Following some background lectures on accelerator physics and the fundamental processes of atomic and plasma physics, the course covered a wide range of topics related to ion sources and highlighted the latest developments in the field. Realistic case studies and topical seminars completed the programme. The school was very successful, with 69 participants representing 25 nationalities. Feedback from the participants was extremely positive, reflecting the high standard of the lectures. The case studies were performed with great enthusiasm and produced some excellent results. In addition to the academic programme, the participants were able to take part in a one-day excursion consisting of a guided tour of Bratislava and free time. A welcome event was held at the Hotel Senec, with s...

  7. CAS course on Plasma Wake Acceleration

    CERN Multimedia

    CERN Accelerator School

    2015-01-01

    The CERN Accelerator School (CAS) recently organised a specialised course on Plasma Wake Acceleration, held at CERN, Geneva, Switzerland, from 23 to 29 November 2014.    Following a number of introductory lectures on laser and plasma physics, as well as an overview of conventional accelerators and their limitations, the course covered a large number of aspects of plasma wake acceleration schemes: the creation of plasma by high power lasers or particle beams, a description of the plasma creation process through simulations and the characteristics of the accelerated particle beams, including results of the latest achievements. Lectures on beam diagnostics, the applications of plasma accelerated beams, and topical seminars completed the programme.  The course was very successful, with 109 students of 26 nationalities attending; most participants coming from European counties, but also from the US, Israel, India, South Korea, Russia and Ukraine. Feedback from the participants was...

  8. Hernie de Spiegel: a propos dun cas

    OpenAIRE

    Karim Ibn Majdoub Hassani; Fatimzohra Zahid; Hicham Anoune; Imane Toughrai; Said Ait Laalim; Khalid Mazaz

    2010-01-01

    La hernie de Spiegel ou hernie ventrale latérale est une déhiscence inhabituelle apparaissant sur la ligne ou fascia semi-lunaire de Spiegel. C’est une entité clinique rare, représente 0.10 à 1 pourcent des hernies. Aussi, nous a-t-il paru opportun de rapporter ce cas colligé dans le service de chirurgie B du CHU Hassan II de Fès. Nous rapportons l’observation d’une patiente âgée de 60 ans, sans antécédent particulier qui présentais une tuméfaction para ombilicale gauche augmentant progressiv...

  9. CAS Introduction to Accelerator Physics in Spain

    CERN Multimedia

    CERN Bulletin

    2012-01-01

    The CERN Accelerator School (CAS) and the University of Granada jointly organised a course called "Introduction to Accelerator Physics" in Granada, Spain, from 28 October to 9 November, 2012.   The course attracted over 200 applicants, of whom 139 were selected to attend. The students were of 25 different nationalities, coming from countries as far away as Australia, China, Guatemala and India. The intensive programme comprised 38 lectures, 3 seminars, 4 tutorials where the students were split into three groups, a poster session and 7 hours of guided and private study. Feedback from the students was very positive, praising the expertise of the lecturers, as well as the high standard and quality of their lectures. CERN's Director-General, Rolf Heuer, gave a public lecture at the Parque de las Ciencias entitled "The Large Hadron Collider: Unveiling the Universe". In addition to the academic programme, the students had the opportunity to visit the well...

  10. Evaluation of a professional social skills program for unemployed people with physical disability Avaliação de um programa de habilidades sociais profissionais para pessoas com deficiência física desempregadas

    Directory of Open Access Journals (Sweden)

    Camila de Sousa Pereira-Guizzo

    2012-01-01

    Full Text Available The literature on labor points out the importance of social skills for employability. This study analyses, on a multiple probe design with two separate groups, the efficiency and effectiveness of a Professional Social Skills Training Program designed for unemployed people with physical disabilities. The sample consisted of 16 people with physical disabilities, aged from 18 to 36, forming two intervention groups. They were assessed by quantitative and qualitative instruments. Results indicated gains in social skills after the intervention, maintenance of acquisitions in the follow-up and generalization of learned skills to the natural environment for both groups. Such results suggest benefits of the Program on the interpersonal and professional development of unemployed people with physical disability.A literatura sobre trabalho aponta a importância das habilidades sociais para a empregabilidade. Este estudo avalia, sob delineamento de grupo com sondagens múltiplas, a eficácia e efetividade de um Programa de Desenvolvimento de Habilidades Sociais para o Trabalho junto a pessoas com deficiência física desempregadas. Participaram 16 pessoas, com idade entre 18 e 36 anos, que formaram dois grupos de intervenção e foram avaliados por meio de instrumentos quantitativos e qualitativos. Os resultados indicaram ganhos de habilidades sociais após a intervenção, manutenção das aquisições nas avaliações de seguimento e generalização das habilidades aprendidas para o ambiente natural em ambos os grupos. Tais resultados sugerem os benefícios do Programa para o desenvolvimento interpessoal e profissional de pessoas com deficiência física desempregadas.

  11. G6PD Deficiency

    Science.gov (United States)

    Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a genetic disorder that is most common in males. About 1 in 10 African American males in the United States has it. G6PD deficiency mainly affects red blood cells, which carry oxygen ...

  12. Deficiently Extremal Gorenstein Algebras

    Indian Academy of Sciences (India)

    Pavinder Singh

    2011-08-01

    The aim of this article is to study the homological properties of deficiently extremal Gorenstein algebras. We prove that if / is an odd deficiently extremal Gorenstein algebra with pure minimal free resolution, then the codimension of / must be odd. As an application, the structure of pure minimal free resolution of a nearly extremal Gorenstein algebra is obtained.

  13. Iron deficiency anemia

    Science.gov (United States)

    Anemia - iron deficiency ... iron from old red blood cells. Iron deficiency anemia develops when your body's iron stores run low. ... You may have no symptoms if the anemia is mild. Most of the time, ... slowly. Symptoms may include: Feeling weak or tired more often ...

  14. Nutritional iron deficiency

    NARCIS (Netherlands)

    Zimmermann, M.B.; Hurrell, R.F.

    2007-01-01

    Iron deficiency is one of the leading risk factors for disability and death worldwide, affecting an estimated 2 billion people. Nutritional iron deficiency arises when physiological requirements cannot be met by iron absorption from diet. Dietary iron bioavailability is low in populations consuming

  15. Vitamin deficiencies and excesses

    Science.gov (United States)

    Vitamins are essential nutrients that must be supplied exogenously either as part of a well balanced diet or as supplements. Deficiency states are uncommon in developed countries except, perhaps, among some food insecure families. In contrast, deficiency states are quite common in many developing ...

  16. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Blood Tests Blood Transfusion Restless Legs Syndrome Send a link to NHLBI to someone by E-MAIL | ... Iron-Deficiency Anemia? Español Iron-deficiency anemia is a common, easily treated condition that occurs if you ...

  17. Muscle phosphorylase kinase deficiency

    DEFF Research Database (Denmark)

    Preisler, N; Orngreen, M C; Echaniz-Laguna, A; Laforet, P; Lonsdorfer-Wolf, E; Doutreleau, S; Geny, B; Akman, H O; Dimauro, S; Vissing, J

    2012-01-01

    To examine metabolism during exercise in 2 patients with muscle phosphorylase kinase (PHK) deficiency and to further define the phenotype of this rare glycogen storage disease (GSD).......To examine metabolism during exercise in 2 patients with muscle phosphorylase kinase (PHK) deficiency and to further define the phenotype of this rare glycogen storage disease (GSD)....

  18. Lipoprotein lipase deficiency.

    OpenAIRE

    Shankar K; Bava H; Shetty J; Joshi M

    1997-01-01

    A rare case of a 3 month old child with lipoprotein lipase deficiency who presented with bronchopneumonia is reported. After noticing lipaemic serum and lipaemia retinalis, a diagnosis of hyperlipoproteinaemia was considered. Lipoprotein lipase deficiency was confirmed with post heparin lipoprotein lipase enzyme activity estimation.

  19. Cas9 gRNA engineering for genome editing, activation and repression

    OpenAIRE

    Kiani, Samira; Chavez, Alejandro; Tuttle, Marcelle; Hall, Richard N; Chari, Raj; Ter-Ovanesyan, Dmitry; Qian, Jason; Pruitt, Benjamin W.; Beal, Jacob; Vora, Suhani; Buchthal, Joanna; Kowal, Emma J K; Ebrahimkhani, Mohammad R.; James J Collins; Weiss, Ron

    2015-01-01

    We demonstrate that by altering the length of Cas9-associated guide RNA(gRNA) we were able to control Cas9 nuclease activity and simultaneously perform genome editing and transcriptional regulation with a single Cas9 protein. We exploited these principles to engineer mammalian synthetic circuits with combined transcriptional regulation and kill functions governed by a single multifunctional Cas9 protein.

  20. Programmable RNA recognition and cleavage by CRISPR/Cas9

    OpenAIRE

    O’Connell, Mitchell R.; Oakes, Benjamin L.; Sternberg, Samuel H.; East-Seletsky, Alexandra; Kaplan, Matias; Doudna, Jennifer A.

    2014-01-01

    The CRISPR-associated protein Cas9 is an RNA-guided DNA endonuclease that uses RNA:DNA complementarity to identify target sites for sequence-specific doublestranded DNA (dsDNA) cleavage 1-5 . In its native context, Cas9 acts on DNA substrates exclusively because both binding and catalysis require recognition of a short DNA sequence, the protospacer adjacent motif (PAM), next to and on the strand opposite the 20-nucleotide target site in dsDNA 4-7 . Cas9 has proven to be a versatile tool for g...

  1. Rationally engineered Cas9 nucleases with improved specificity

    OpenAIRE

    Slaymaker, Ian M.; Gao, Linyi; Zetsche, Bernd; Scott, David A.; Yan, Winston X.; Zhang, Feng

    2015-01-01

    The RNA-guided endonuclease Cas9 is a versatile genome editing tool with a broad range of applications from therapeutics to functional annotation of genes. Cas9 creates double-strand breaks (DSBs) at targeted genomic loci complementary to a short RNA guide. However, Cas9 can cleave off-target sites that are not fully complementary to the guide, which poses a major challenge for genome editing. Here, we use structure-guided protein engineering to improve the specificity of Streptococcus pyogen...

  2. Engineered CRISPR-Cas9 nucleases with altered PAM specificities

    OpenAIRE

    Kleinstiver, Benjamin P.; Prew, Michelle S.; Tsai, Shengdar Q.; Topkar, Ved; Nguyen, Nhu T.; Zheng, Zongli; Gonzales, Andrew P.W.; Li, Zhuyun; Peterson, Randall T.; Yeh, Jing-Ruey Joanna; Aryee, Martin J.; Joung, J. Keith

    2015-01-01

    Although CRISPR-Cas9 nucleases are widely used for genome editing1, 2, the range of sequences that Cas9 can recognize is constrained by the need for a specific protospacer adjacent motif (PAM)3–6. As a result, it can often be difficult to target double-stranded breaks (DSBs) with the precision that is necessary for various genome editing applications. The ability to engineer Cas9 derivatives with purposefully altered PAM specificities would address this limitation. Here we show that the commo...

  3. Engineered CRISPR-Cas9 nucleases with altered PAM specificities

    OpenAIRE

    Kleinstiver, Benjamin P.; Prew, Michelle S.; Tsai, Shengdar Q.; Topkar, Ved; Nguyen, Nhu T.; Zheng, Zongli; Gonzales, Andrew P.W.; Li, Zhuyun; Peterson, Randall T.; Yeh, Jing-Ruey Joanna; Aryee, Martin J.; Joung, J. Keith

    2015-01-01

    Although CRISPR-Cas9 nucleases are widely used for genome editing 1, 2 , the range of sequences that Cas9 can recognize is constrained by the need for a specific protospacer adjacent motif (PAM) 3–6 . As a result, it can often be difficult to target double-stranded breaks (DSBs) with the precision that is necessary for various genome editing applications. The ability to engineer Cas9 derivatives with purposefully altered PAM specificities would address this limitation. Here we show that the c...

  4. Rationally engineered Cas9 nucleases with improved specificity

    OpenAIRE

    Slaymaker, Ian; Gao, Linyi; Zetsche, Bernd; Scott, David Arthur; Yan, Winston Xia; Zhang, Feng

    2015-01-01

    The RNA-guided endonuclease Cas9 is a versatile genome-editing tool with a broad range of applications from therapeutics to functional annotation of genes. Cas9 creates double-strand breaks (DSBs) at targeted genomic loci complementary to a short RNA guide. However, Cas9 can cleave off-target sites that are not fully complementary to the guide, which poses a major challenge for genome editing. Here, we use structure-guided protein engineering to improve the specificity of Streptococcus pyogen...

  5. CRISPR-Cas9-Mediated Genome Editing in Leishmania donovani

    OpenAIRE

    Zhang, Wen-Wei; Matlashewski, Greg

    2015-01-01

    ABSTRACT The prokaryotic CRISPR (clustered regularly interspaced short palindromic repeat)-Cas9, an RNA-guided endonuclease, has been shown to mediate efficient genome editing in a wide variety of organisms. In the present study, the CRISPR-Cas9 system has been adapted to Leishmania donovani, a protozoan parasite that causes fatal human visceral leishmaniasis. We introduced the Cas9 nuclease into L. donovani and generated guide RNA (gRNA) expression vectors by using the L. donovani rRNA promo...

  6. The 13th CAS General Assembly Convenes in Beijing

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    @@ The 13th General Assembly of CAS was held from June 5 to 8 in Beijing. Top State leaders were present at the joint opening session of the 13th CAS General Assembly and the Eighth General Assembly of the Chinese Academy of Engineering (CAE)held on the morning of June 5 at the Great Hall of the People. Chinese President HU Jintao delivered an important speech at the ceremony. State Councilor CHEN Zhili made a special report about the Framework for the National Long- and Mid-term Plan for S&T Development in China at the joint plenary session of CAS and CAE assemblies on June 6.

  7. Efficient Mitochondrial Genome Editing by CRISPR/Cas9

    OpenAIRE

    Areum Jo; Sangwoo Ham; Gum Hwa Lee; Yun-Il Lee; SangSeong Kim; Yun-Song Lee; Joo-Ho Shin; Yunjong Lee

    2015-01-01

    The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 system has been widely used for nuclear DNA editing to generate mutations or correct specific disease alleles. Despite its flexible application, it has not been determined if CRISPR/Cas9, originally identified as a bacterial defense system against virus, can be targeted to mitochondria for mtDNA editing. Here, we show that regular FLAG-Cas9 can localize to mitochondria to edit mitochondrial DNA with sgRNAs targeting s...

  8. CRISPR/Cas9-mediated somatic correction of a novel coagulator factor IX gene mutation ameliorates hemophilia in mouse.

    Science.gov (United States)

    Guan, Yuting; Ma, Yanlin; Li, Qi; Sun, Zhenliang; Ma, Lie; Wu, Lijuan; Wang, Liren; Zeng, Li; Shao, Yanjiao; Chen, Yuting; Ma, Ning; Lu, Wenqing; Hu, Kewen; Han, Honghui; Yu, Yanhong; Huang, Yuanhua; Liu, Mingyao; Li, Dali

    2016-01-01

    The X-linked genetic bleeding disorder caused by deficiency of coagulator factor IX, hemophilia B, is a disease ideally suited for gene therapy with genome editing technology. Here, we identify a family with hemophilia B carrying a novel mutation, Y371D, in the human F9 gene. The CRISPR/Cas9 system was used to generate distinct genetically modified mouse models and confirmed that the novel Y371D mutation resulted in a more severe hemophilia B phenotype than the previously identified Y371S mutation. To develop therapeutic strategies targeting this mutation, we subsequently compared naked DNA constructs versus adenoviral vectors to deliver Cas9 components targeting the F9 Y371D mutation in adult mice. After treatment, hemophilia B mice receiving naked DNA constructs exhibited correction of over 0.56% of F9 alleles in hepatocytes, which was sufficient to restore hemostasis. In contrast, the adenoviral delivery system resulted in a higher corrective efficiency but no therapeutic effects due to severe hepatic toxicity. Our studies suggest that CRISPR/Cas-mediated in situ genome editing could be a feasible therapeutic strategy for human hereditary diseases, although an efficient and clinically relevant delivery system is required for further clinical studies. PMID:26964564

  9. A Guild of 45 CRISPR-Associated (Cas Protein Families and Multiple CRISPR/Cas Subtypes Exist in Prokaryotic Genomes.

    Directory of Open Access Journals (Sweden)

    2005-11-01

    Full Text Available Clustered regularly interspaced short palindromic repeats (CRISPRs are a family of DNA direct repeats found in many prokaryotic genomes. Repeats of 21-37 bp typically show weak dyad symmetry and are separated by regularly sized, nonrepetitive spacer sequences. Four CRISPR-associated (Cas protein families, designated Cas1 to Cas4, are strictly associated with CRISPR elements and always occur near a repeat cluster. Some spacers originate from mobile genetic elements and are thought to confer "immunity" against the elements that harbor these sequences. In the present study, we have systematically investigated uncharacterized proteins encoded in the vicinity of these CRISPRs and found many additional protein families that are strictly associated with CRISPR loci across multiple prokaryotic species. Multiple sequence alignments and hidden Markov models have been built for 45 Cas protein families. These models identify family members with high sensitivity and selectivity and classify key regulators of development, DevR and DevS, in Myxococcus xanthus as Cas proteins. These identifications show that CRISPR/cas gene regions can be quite large, with up to 20 different, tandem-arranged cas genes next to a repeat cluster or filling the region between two repeat clusters. Distinctive subsets of the collection of Cas proteins recur in phylogenetically distant species and correlate with characteristic repeat periodicity. The analyses presented here support initial proposals of mobility of these units, along with the likelihood that loci of different subtypes interact with one another as well as with host cell defensive, replicative, and regulatory systems. It is evident from this analysis that CRISPR/cas loci are larger, more complex, and more heterogeneous than previously appreciated.

  10. A guild of 45 CRISPR-associated (Cas protein families and multiple CRISPR/Cas subtypes exist in prokaryotic genomes.

    Directory of Open Access Journals (Sweden)

    Daniel H Haft

    2005-11-01

    Full Text Available Clustered regularly interspaced short palindromic repeats (CRISPRs are a family of DNA direct repeats found in many prokaryotic genomes. Repeats of 21-37 bp typically show weak dyad symmetry and are separated by regularly sized, nonrepetitive spacer sequences. Four CRISPR-associated (Cas protein families, designated Cas1 to Cas4, are strictly associated with CRISPR elements and always occur near a repeat cluster. Some spacers originate from mobile genetic elements and are thought to confer "immunity" against the elements that harbor these sequences. In the present study, we have systematically investigated uncharacterized proteins encoded in the vicinity of these CRISPRs and found many additional protein families that are strictly associated with CRISPR loci across multiple prokaryotic species. Multiple sequence alignments and hidden Markov models have been built for 45 Cas protein families. These models identify family members with high sensitivity and selectivity and classify key regulators of development, DevR and DevS, in Myxococcus xanthus as Cas proteins. These identifications show that CRISPR/cas gene regions can be quite large, with up to 20 different, tandem-arranged cas genes next to a repeat cluster or filling the region between two repeat clusters. Distinctive subsets of the collection of Cas proteins recur in phylogenetically distant species and correlate with characteristic repeat periodicity. The analyses presented here support initial proposals of mobility of these units, along with the likelihood that loci of different subtypes interact with one another as well as with host cell defensive, replicative, and regulatory systems. It is evident from this analysis that CRISPR/cas loci are larger, more complex, and more heterogeneous than previously appreciated.

  11. Harnessing heterologous and endogenous CRISPR-Cas machineries for efficient markerless genome editing in Clostridium

    OpenAIRE

    Pyne, Michael E.; Mark R. Bruder; Murray Moo-Young; Chung, Duane A; C. Perry Chou

    2016-01-01

    Application of CRISPR-Cas9 systems has revolutionized genome editing across all domains of life. Here we report implementation of the heterologous Type II CRISPR-Cas9 system in Clostridium pasteurianum for markerless genome editing. Since 74% of species harbor CRISPR-Cas loci in Clostridium, we also explored the prospect of co-opting host-encoded CRISPR-Cas machinery for genome editing. Motivation for this work was bolstered from the observation that plasmids expressing heterologous cas9 resu...

  12. Engineering Translational Activators with CRISPR-Cas System.

    Science.gov (United States)

    Du, Pei; Miao, Chensi; Lou, Qiuli; Wang, Zefeng; Lou, Chunbo

    2016-01-15

    RNA parts often serve as critical components in genetic engineering. Here we report a design of translational activators which is composed of an RNA endoribonuclease (Csy4) and two exchangeable RNA modules. Csy4, a member of Cas endoribonuclease, cleaves at a specific recognition site; this cleavage releases a cis-repressive RNA module (crRNA) from the masked ribosome binding site (RBS), which subsequently allows the downstream translation initiation. Unlike small RNA as a translational activator, the endoribonuclease-based activator is able to efficiently unfold the perfect RBS-crRNA pairing. As an exchangeable module, the crRNA-RBS duplex was forwardly and reversely engineered to modulate the dynamic range of translational activity. We further showed that Csy4 and its recognition site, together as a module, can also be replaced by orthogonal endoribonuclease-recognition site homologues. These modularly structured, high-performance translational activators would endow the programming of gene expression in the translation level with higher feasibility. PMID:26414660

  13. EU Commissioner Delivers a Speech at CAS Institute

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ European Union (EU) Environment Commissioner Margot Wallstrom gave a talk on cooperation between the EU and China on environmental issues during her trip to the CAS Institute of Geographic Sciences and Natural Resource Research (IGSNRR) on Nov. 13.

  14. CRISPR-Cas9-Guided Genome Engineering in C. elegans.

    Science.gov (United States)

    Kim, Hyun-Min; Colaiácovo, Monica P

    2016-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) system is successfully being used for efficient and targeted genome editing in various organisms, including the nematode C. elegans. Recent studies have developed various CRISPR-Cas9 approaches to enhance genome engineering via two major DNA double-strand break repair pathways: non-homologous end joining and homologous recombination. Here we describe a protocol for Cas9-mediated C. elegans genome editing together with single guide RNA (sgRNA) and repair template cloning, as well as injection methods required for delivering Cas9, sgRNAs, and repair template DNA into the C. elegans germline. © 2016 by John Wiley & Sons, Inc. PMID:27366893

  15. Methods for Optimizing CRISPR-Cas9 Genome Editing Specificity.

    Science.gov (United States)

    Tycko, Josh; Myer, Vic E; Hsu, Patrick D

    2016-08-01

    Advances in the development of delivery, repair, and specificity strategies for the CRISPR-Cas9 genome engineering toolbox are helping researchers understand gene function with unprecedented precision and sensitivity. CRISPR-Cas9 also holds enormous therapeutic potential for the treatment of genetic disorders by directly correcting disease-causing mutations. Although the Cas9 protein has been shown to bind and cleave DNA at off-target sites, the field of Cas9 specificity is rapidly progressing, with marked improvements in guide RNA selection, protein and guide engineering, novel enzymes, and off-target detection methods. We review important challenges and breakthroughs in the field as a comprehensive practical guide to interested users of genome editing technologies, highlighting key tools and strategies for optimizing specificity. The genome editing community should now strive to standardize such methods for measuring and reporting off-target activity, while keeping in mind that the goal for specificity should be continued improvement and vigilance. PMID:27494557

  16. CAS-The Prospect of Work in 2006

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    @@ Mission Statement China Association for Standardization (CAS) is to act as a bridge linking nationwide specialists in standardization with social communities. To meet the needs of government,society, market and enterprises, CAS endeavors to disseminate and popularize the standardization knowledge, organize academic forums and seminars, provide technical consultation on standardization, promote domestic and international cooperation and exchanges,and push forward the standardization process in China.

  17. Comparison of Cas9 activators in multiple species.

    Science.gov (United States)

    Chavez, Alejandro; Tuttle, Marcelle; Pruitt, Benjamin W; Ewen-Campen, Ben; Chari, Raj; Ter-Ovanesyan, Dmitry; Haque, Sabina J; Cecchi, Ryan J; Kowal, Emma J K; Buchthal, Joanna; Housden, Benjamin E; Perrimon, Norbert; Collins, James J; Church, George

    2016-07-01

    Several programmable transcription factors exist based on the versatile Cas9 protein, yet their relative potency and effectiveness across various cell types and species remain unexplored. Here, we compare Cas9 activator systems and examine their ability to induce robust gene expression in several human, mouse, and fly cell lines. We also explore the potential for improved activation through the combination of the most potent activator systems, and we assess the role of cooperativity in maximizing gene expression. PMID:27214048

  18. High-throughput functional genomics using CRISPR-Cas9

    OpenAIRE

    Shalem, Ophir; Sanjana, Neville E.; Zhang, Feng

    2015-01-01

    Forward genetic screens are powerful tools for the discovery and functional annotation of genetic elements. Recently, the RNA-guided CRISPR (clustered regularly interspaced short palindromic repeat)-associated Cas9 nuclease has been combined with genome-scale guide RNA libraries for unbiased, phenotypic screening. In this Review, we describe recent advances using Cas9 for genome-scale screens, including knockout approaches that inactivate genomic loci and strategies that modulate transcriptio...

  19. CRISPR-Cas9: A Revolutionary Tool for Cancer Modelling

    OpenAIRE

    Raul Torres-Ruiz; Sandra Rodriguez-Perales

    2015-01-01

    The cancer-modelling field is now experiencing a conversion with the recent emergence of the RNA-programmable CRISPR-Cas9 system, a flexible methodology to produce essentially any desired modification in the genome. Cancer is a multistep process that involves many genetic mutations and other genome rearrangements. Despite their importance, it is difficult to recapitulate the degree of genetic complexity found in patient tumors. The CRISPR-Cas9 system for genome editing has been proven as a ...

  20. Targeted genomic rearrangements using CRISPR/Cas technology

    OpenAIRE

    Choi, Peter S.; Meyerson, Matthew

    2014-01-01

    Genomic rearrangements are frequently observed in cancer cells but have been difficult to generate in a highly specific manner for functional analysis. Here we report the application of CRISPR/Cas technology to successfully generate several types of chromosomal rearrangements implicated as driver events in lung cancer, including the CD74-ROS1 translocation event and the EML4-ALK and KIF5B-RET inversion events. Our results demonstrate that Cas9-induced DNA breaks promote efficient rearrangemen...

  1. Target specificity of the CRISPR-Cas9 system

    OpenAIRE

    Wu, Xuebing; Kriz, Andrea J.; Sharp, Phillip A.

    2014-01-01

    The CRISPR-Cas9 system, naturally a defense mechanism in prokaryotes, has been repurposed as an RNA-guided DNA targeting platform. It has been widely used for genome editing and transcriptome modulation, and has shown great promise in correcting mutations in human genetic diseases. Off-target effects are a critical issue for all of these applications. Here we review the current status on the target specificity of the CRISPR-Cas9 system.

  2. CAS sets up an award to promote international cooperation

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ Established in 2007, the CAS Award for International Cooperation in Science and Technology is to honor those eminent international experts who have made outstanding contributions to China's global cooperation in science and technology, so as to encourage more efforts in this respect that will lead to the enhancement of CAS innovation capacity and the improvement of its research performance, education and training, management and reputations among the international community.

  3. Winners of 2008 CAS Award for international Scientific Cooperation

    Institute of Scientific and Technical Information of China (English)

    XIN Ling

    2009-01-01

    @@ On 14 January, 2009, CAS honored three foreign scientists for their lasting and fruitful contributions to the collaboration between CAS and the world: Akito Arima, president of the Japan Science Foundation, Yuen-Ron Shen, professor of physics with the University of California at Berkley, and Michel Che, catalyst expert and chair professor at Universite Pierre et Marie Curie. Their association with the Chinese scientific community can be dated back to some three decades ago.

  4. The Structural Biology of CRISPR-Cas Systems

    OpenAIRE

    Jiang, Fuguo; Doudna, Jennifer A.

    2015-01-01

    Prokaryotic CRISPR-Cas genomic loci encode RNA-mediated adaptive immune systems that bear some functional similarities with eukaryotic RNA interference. Acquired and heritable immunity against bacteriophage and plasmids begins with integration of ~30 base pair foreign DNA sequences into the host genome. CRISPR-derived transcripts assemble with CRISPR-associated (Cas) proteins to target complementary nucleic acids for degradation. Here we review recent advances in the structural biology of the...

  5. CAS Held the 2nd Yearly Official Business Conference

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ On July 15th 2004, China Association For Standardization (CAS) held official business conference the second time of this year in Beijing, which is its 5th Council Meeting. The conference was moderated by Li Rui, President of CAS's Board of Directors. Vice-presidents Li- Zhonghai, JiaoYunqi and Jing Xiaodong, Secretary-General Ma Lincong and Zhang Wei, and other cadres totally more than 20delegates attended the conference.

  6. Potential pitfalls of CRISPR/Cas9-mediated genome editing.

    Science.gov (United States)

    Peng, Rongxue; Lin, Guigao; Li, Jinming

    2016-04-01

    Recently, a novel technique named the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas)9 system has been rapidly developed. This genome editing tool has improved our ability tremendously with respect to exploring the pathogenesis of diseases and correcting disease mutations, as well as phenotypes. With a short guide RNA, Cas9 can be precisely directed to target sites, and functions as an endonuclease to efficiently produce breaks in DNA double strands. Over the past 30 years, CRISPR has evolved from the 'curious sequences of unknown biological function' into a promising genome editing tool. As a result of the incessant development in the CRISPR/Cas9 system, Cas9 co-expressed with custom guide RNAs has been successfully used in a variety of cells and organisms. This genome editing technology can also be applied to synthetic biology, functional genomic screening, transcriptional modulation and gene therapy. However, although CRISPR/Cas9 has a broad range of action in science, there are several aspects that affect its efficiency and specificity, including Cas9 activity, target site selection and short guide RNA design, delivery methods, off-target effects and the incidence of homology-directed repair. In the present review, we highlight the factors that affect the utilization of CRISPR/Cas9, as well as possible strategies for handling any problems. Addressing these issues will allow us to take better advantage of this technique. In addition, we also review the history and rapid development of the CRISPR/Cas system from the time of its initial discovery in 2012. PMID:26535798

  7. Thiamine deficiency and delirium.

    Science.gov (United States)

    Osiezagha, Kenneth; Ali, Shahid; Freeman, C; Barker, Narviar C; Jabeen, Shagufta; Maitra, Sarbani; Olagbemiro, Yetunde; Richie, William; Bailey, Rahn K

    2013-04-01

    Thiamine is an essential vitamin that plays an important role in cellular production of energy from ingested food and enhances normal neuronal actives. Deficiency of this vitamin leads to a very serious clinical condition known as delirium. Studies performed in the United States and other parts of the world have established the link between thiamine deficiency and delirium. This literature review examines the physiology, pathophysiology, predisposing factors, clinical manifestations (e.g., Wernicke's encephalopathy, Wernicke-Korsakoff syndrome, structural and functional brain injuries) and diagnosis of thiamine deficiency and delirium. Current treatment practices are also discussed that may improve patient outcome, which ultimately may result in a reduction in healthcare costs. PMID:23696956

  8. Internal guide RNA interactions interfere with Cas9-mediated cleavage

    Science.gov (United States)

    Thyme, Summer B.; Akhmetova, Laila; Montague, Tessa G.; Valen, Eivind; Schier, Alexander F.

    2016-01-01

    The CRISPR/Cas system uses guide RNAs (gRNAs) to direct sequence-specific DNA cleavage. Not every gRNA elicits cleavage and the mechanisms that govern gRNA activity have not been resolved. Low activity could result from either failure to form a functional Cas9–gRNA complex or inability to recognize targets in vivo. Here we show that both phenomena influence Cas9 activity by comparing mutagenesis rates in zebrafish embryos with in vitro cleavage assays. In vivo, our results suggest that genomic factors such as CTCF inhibit mutagenesis. Comparing near-identical gRNA sequences with different in vitro activities reveals that internal gRNA interactions reduce cleavage. Even though gRNAs containing these structures do not yield cleavage-competent complexes, they can compete with active gRNAs for binding to Cas9. These results reveal that both genomic context and internal gRNA interactions can interfere with Cas9-mediated cleavage and illuminate previously uncharacterized features of Cas9–gRNA complex formation. PMID:27282953

  9. CAS Presents '09 Outstanding S&T Achievement Awards

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    @@ CAS announced winners of its 2009 Science & Technology Achievement Awards on January27,2010 in Beijing. Five research groups were conferred on the awards,namely,the group for the heavy ion research facility in Lanzhou-cooler storage ring(HIRFL-CSR)with the CAS Institute of Modern Physics(IMP),the group for theoretical research into regional space development and practical research into major planning in China with the CAS Institute of Geographic Sciences and Natural Resources Research,the group for the world's first"synthesis glycol from coal"technology with the CAS Fujian Institute of Research on the Structure of Matter,the group for the concomitant satellite for"Shenzhou-Ⅶ"-China's third manned spacecraft-with the CAS Shanghai Engineering Center for Microsatellites,and the group for the visualized casting-forging technology with the CAS Institute of Metal Research.The biennial awards were founded in 2002 to honor individuals and research groups with significant research achievements in the latest five years.Less than 10 individuals or research groups are awarded each time.

  10. Characterization of Staphylococcus aureus Cas9: a smaller Cas9 for all-in-one adeno-associated virus delivery and paired nickase applications

    OpenAIRE

    Friedland, Ari E.; Baral, Reshica; Singhal, Pankhuri; Loveluck, Katherine; Shen, Shen; Sanchez, Minerva; Marco, Eugenio; Gotta, Gregory M.; Maeder, Morgan L.; Kennedy, Edward M.; Kornepati, Anand V. R.; Sousa, Alexander; Collins, McKensie A.; Jayaram, Hari; Cullen, Bryan R.

    2015-01-01

    Background CRISPR-Cas systems have been broadly embraced as effective tools for genome engineering applications, with most studies to date utilizing the Streptococcus pyogenes Cas9. Here we characterize and manipulate the smaller, 1053 amino acid nuclease Staphylococcus aureus Cas9. Results We find that the S. aureus Cas9 recognizes an NNGRRT protospacer adjacent motif (PAM) and cleaves target DNA at high efficiency with a variety of guide RNA (gRNA) spacer lengths. When directed against geno...

  11. The prevalence of clinical goiter and urinary iodine concentration in 8-12 years old students of Semirom in 2004 (15 years after execution of national program for eliminationof iodine deficiency disorders

    Directory of Open Access Journals (Sweden)

    A. Kachouei

    2006-01-01

    Full Text Available Background and purpose: With respect to the high prevalence of goiter in Semirom before the initiation of National Program for Elimination of Iodine Deficiency Disorders (NPEIDD, the impact of the program on the prevalence of goiter in 8-12 years old students of Semirom was investigated in this study, after 15 years of sucessful execution of NPEIDD.Materials and Methods: After excluding students with any clinical evidence of hyper- or hypo-thyroidism, 1817 students aged 8-12 years were recruited based on a cluster sampling method. The participants were examined for the presence of goiter by experiencd endocrinologists. Urinary iodine was measured in 182 examined students, too. Chi- Square statistical method was used to analyze the data.Results: Goiter was detected in 36.7% of the students (grade 1: 30.4%, grade 2: 6.3%. The median urinary iodine was 18.5 and mean was 19.37 ± 9.22 μg/dl in collected specimens.Conclusion: The adequacy of urinary iodine in students of Semirom confirms the success of NPEIDD, however, the higher than expected prevalence of goiter in the region suggests the probable role of other goitrogenous or autoimmune factors. Further studies should be performed to investigate the possible causes.

  12. Highly specific targeted mutagenesis in plants using Staphylococcus aureus Cas9.

    Science.gov (United States)

    Kaya, Hidetaka; Mikami, Masafumi; Endo, Akira; Endo, Masaki; Toki, Seiichi

    2016-01-01

    The CRISPR/Cas9 system is an efficient and convenient tool for genome editing in plants. Cas9 nuclease derived from Streptococcus pyogenes (Sp) is commonly used in this system. Recently, Staphylococcus aureus Cas9 (SaCas9)-mediated genome editing was reported in human cells and Arabidopsis. Because SaCas9 (1053 a.a.) is smaller than SpCas9 (1368 a.a.), SaCas9 could have substantial advantages for delivering and expressing Cas9 protein, especially using virus vectors. Since the protospacer adjacent motif (PAM) sequence of SaCas9 (5'-NNGRRT-3') differs from that of SpCas9 (5'-NGG-3'), the use of this alternative Cas9 nuclease could expand the selectivity at potential cleavage target sites of the CRISPR/Cas9 system. Here we show that SaCas9 can mutagenize target sequences in tobacco and rice with efficiencies similar to those of SpCas9. We also analyzed the base preference for 'T' at the 6th position of the SaCas9 PAM. Targeted mutagenesis efficiencies in target sequences with non-canonical PAMs (5'-NNGRRV-3') were much lower than those with a canonical PAM (5'-NNGRRT-3'). The length of target sequence recognized by SaCas9 is one or two nucleotides longer than that recognized by SpCas9. Taken together, our results demonstrate that SaCas9 has higher sequence recognition capacity than SpCas9 and is useful for reducing off-target mutations in crop. PMID:27226350

  13. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... symptoms. Severe iron-deficiency anemia can lead to heart problems, infections, problems with growth and development in ... 18/2011 This video—presented by the National Heart, Lung, and Blood Institute, part of the National ...

  14. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Entire Site Health Topics News & Resources Intramural Research Public Health Topics Education & Awareness Resources Contact The Health ... Severe iron-deficiency anemia can lead to heart problems, infections, problems with growth and development in children, ...

  15. Folate-deficiency anemia

    Science.gov (United States)

    Folate-deficiency anemia is a decrease in red blood cells (anemia) due to a lack of folate. Folate is a type ... B vitamin. It is also called folic acid. Anemia is a condition in which the body does ...

  16. Sleep Deprivation and Deficiency

    Science.gov (United States)

    ... page from the NHLBI on Twitter. What Are Sleep Deprivation and Deficiency? Sleep deprivation (DEP-rih-VA- ... Rate This Content: NEXT >> Updated: February 22, 2012 Sleep Infographic Sleep Disorders & Insufficient Sleep: Improving Health through ...

  17. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Digg. Share this page from the NHLBI on Facebook. Add this link to the NHLBI to my ... such as tiredness, poor skin tone, dizziness, and depression. After her doctor diagnosed her with iron-deficiency ...

  18. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Topics Anemia Blood Tests Blood Transfusion Restless Legs Syndrome Send a link to NHLBI to someone by ... symptoms. Severe iron-deficiency anemia can lead to heart problems, infections, problems with growth and development in ...

  19. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... CAUSES WHO IS AT RISK SIGNS & SYMPTOMS DIAGNOSIS TREATMENTS PREVENTION LIVING WITH CLINICAL TRIALS LINKS Related Topics ... Doctors usually can successfully treat iron-deficiency anemia. Treatment will depend on the cause and severity of ...

  20. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... iron-rich protein that carries oxygen from the lungs to the rest of the body. Iron-deficiency ... 2011 This video—presented by the National Heart, Lung, and Blood Institute, part of the National Institutes ...

  1. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Health Topics Education & Awareness Resources Contact The Health Information Center Health Professionals Systematic Evidence Reviews & Clinical Practice ... and see the benefits of treatment. For more information about living with and managing iron-deficiency anemia, ...

  2. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... Alerts E-Newsletters About NHLBI Organization NHLBI Director Budget, Planning, & Legislative Advisory Committees Contact Us FAQs Home » ... severity of the condition. Treatments may include dietary changes, medicines, and surgery. Severe iron-deficiency anemia may ...

  3. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... intravenous iron therapy. Rate This Content: NEXT >> Featured Video Living With and Managing Iron-Deficiency Anemia 05/18/2011 This video—presented by the National Heart, Lung, and Blood ...

  4. Clinical significance of complement deficiencies.

    Science.gov (United States)

    Pettigrew, H David; Teuber, Suzanne S; Gershwin, M Eric

    2009-09-01

    The complement system is composed of more than 30 serum and membrane-bound proteins, all of which are needed for normal function of complement in innate and adaptive immunity. Historically, deficiencies within the complement system have been suspected when young children have had recurrent and difficult-to-control infections. As our understanding of the complement system has increased, many other diseases have been attributed to deficiencies within the complement system. Generally, complement deficiencies within the classical pathway lead to increased susceptibility to encapsulated bacterial infections as well as a syndrome resembling systemic lupus erythematosus. Complement deficiencies within the mannose-binding lectin pathway generally lead to increased bacterial infections, and deficiencies within the alternative pathway usually lead to an increased frequency of Neisseria infections. However, factor H deficiency can lead to membranoproliferative glomerulonephritis and hemolytic uremic syndrome. Finally, deficiencies within the terminal complement pathway lead to an increased incidence of Neisseria infections. Two other notable complement-associated deficiencies are complement receptor 3 and 4 deficiency, which result from a deficiency of CD18, a disease known as leukocyte adhesion deficiency type 1, and CD59 deficiency, which causes paroxysmal nocturnal hemoglobinuria. Most inherited deficiencies of the complement system are autosomal recessive, but properidin deficiency is X-linked recessive, deficiency of C1 inhibitor is autosomal dominant, and mannose-binding lectin and factor I deficiencies are autosomal co-dominant. The diversity of clinical manifestations of complement deficiencies reflects the complexity of the complement system. PMID:19758139

  5. Adult growth hormone deficiency

    OpenAIRE

    Vishal Gupta

    2011-01-01

    Adult growth hormone deficiency (AGHD) is being recognized increasingly and has been thought to be associated with premature mortality. Pituitary tumors are the commonest cause for AGHD. Growth hormone deficiency (GHD) has been associated with neuropsychiatric-cognitive, cardiovascular, neuromuscular, metabolic, and skeletal abnormalities. Most of these can be reversed with growth hormone therapy. The insulin tolerance test still remains the gold standard dynamic test to diagnose AGHD. Growth...

  6. Iron deficiency anemia Review

    OpenAIRE

    Yıldız, İnci

    2009-01-01

    Iron deficiency anemia is the most frequent and widespread anemia around the world Its prevalence is increased in infants and adolescent girls The etiologic factors may vary but anemia is essentially related to iron deficient nutrition blood loss and malabsorption Children may have paleness cardiovascular and neurologic impacts of anemia pica epithelial changes as koilonychia glossitis angular stomatitis Treatment is by oral or parenteral supplementation of iron Turk Arch Ped 2009; 44 Suppl: ...

  7. Dramatic Enhancement of Genome Editing by CRISPR/Cas9 Through Improved Guide RNA Design

    OpenAIRE

    Farboud, B; Meyer, BJ

    2015-01-01

    Success with genome editing by the RNA-programmed nuclease Cas9 has been limited by the inability to predict effective guide RNAs and DNA target sites. Not all guide RNAs have been successful, and even those that were, varied widely in their efficacy. Here we describe and validate a strategy for Caenorhabditis elegans that reliably achieved a high frequency of genome editing for all targets tested in vivo. The key innovation was to design guide RNAs with a GG motif at the 3′ end of their targ...

  8. The CRISPR/Cas Immune System Is an Operon Regulated by LeuO, H-NS, and Leucine-Responsive Regulatory Protein in Salmonella enterica Serovar Typhi ▿ †

    OpenAIRE

    Medina-Aparicio, L.; Rebollar-Flores, J. E.; Gallego-Hernández, A. L.; A. Vázquez; Olvera, L.; Gutiérrez-Ríos, R. M.; Calva, E; Hernández-Lucas, I.

    2011-01-01

    Prokaryotes have developed multiple strategies to survive phage attack and invasive DNA. Recently, a novel genetic program denominated the CRISPR/Cas system was demonstrated to have a role in these biological processes providing genetic immunity. This defense mechanism is widespread in the Archaea and Bacteria, suggesting an ancient origin. In the last few years, progress has been made regarding the functionality of the CRISPR/Cas system; however, many basic aspects of the system remain unkno...

  9. A non-inheritable maternal Cas9-based multiple-gene editing system in mice.

    Science.gov (United States)

    Sakurai, Takayuki; Kamiyoshi, Akiko; Kawate, Hisaka; Mori, Chie; Watanabe, Satoshi; Tanaka, Megumu; Uetake, Ryuichi; Sato, Masahiro; Shindo, Takayuki

    2016-01-01

    The CRISPR/Cas9 system is capable of editing multiple genes through one-step zygote injection. The preexisting method is largely based on the co-injection of Cas9 DNA (or mRNA) and guide RNAs (gRNAs); however, it is unclear how many genes can be simultaneously edited by this method, and a reliable means to generate transgenic (Tg) animals with multiple gene editing has yet to be developed. Here, we employed non-inheritable maternal Cas9 (maCas9) protein derived from Tg mice with systemic Cas9 overexpression (Cas9 mice). The maCas9 protein in zygotes derived from mating or in vitro fertilization of Tg/+ oocytes and +/+ sperm could successfully edit the target genome. The efficiency of such maCas9-based genome editing was comparable to that of zygote microinjection-based genome editing widely used at present. Furthermore, we demonstrated a novel approach to create "Cas9 transgene-free" gene-modified mice using non-Tg (+/+) zygotes carrying maCas9. The maCas9 protein in mouse zygotes edited nine target loci simultaneously after injection with nine different gRNAs alone. Cas9 mouse-derived zygotes have the potential to facilitate the creation of genetically modified animals carrying the Cas9 transgene, enabling repeatable genome engineering and the production of Cas9 transgene-free mice. PMID:26817415

  10. Iron deficiency in Europe.

    Science.gov (United States)

    Hercberg, S; Preziosi, P; Galan, P

    2001-04-01

    In Europe, iron deficiency is considered to be one of the main nutritional deficiency disorders affecting large fractions of the population, particularly such physiological groups as children, menstruating women and pregnant women. Some factors such as type of contraception in women, blood donation or minor pathological blood loss (haemorrhoids, gynaecological bleeding...) considerably increase the difficulty of covering iron needs. Moreover, women, especially adolescents consuming low-energy diets, vegetarians and vegans are at high risk of iron deficiency. Although there is no evidence that an absence of iron stores has any adverse consequences, it does indicate that iron nutrition is borderline, since any further reduction in body iron is associated with a decrease in the level of functional compounds such as haemoglobin. The prevalence of iron-deficient anaemia has slightly decreased in infants and menstruating women. Some positive factors may have contributed to reducing the prevalence of iron-deficiency anaemia in some groups of population: the use of iron-fortified formulas and iron-fortified cereals; the use of oral contraceptives and increased enrichment of iron in several countries; and the use of iron supplements during pregnancy in some European countries. It is possible to prevent and control iron deficiency by counseling individuals and families about sound iron nutrition during infancy and beyond, and about iron supplementation during pregnancy, by screening persons on the basis of their risk for iron deficiency, and by treating and following up persons with presumptive iron deficiency. This may help to reduce manifestations of iron deficiency and thus improve public health. Evidence linking iron status with risk of cardiovascular disease or cancer is unconvincing and does not justify changes in food fortification or medical practice, particularly because the benefits of assuring adequate iron intake during growth and development are well established

  11. Glucose-6-phosphate dehydrogenase deficiency

    Science.gov (United States)

    G-6-PD deficiency; Hemolytic anemia due to G6PD deficiency; Anemia - hemolytic due to G6PD deficiency ... G6PD deficiency occurs when a person is missing or doesn't have enough of an enzyme called glucose- ...

  12. Some deficiencies and solutions in gamma ray spectrometry

    International Nuclear Information System (INIS)

    A number of problems in high-resolution gamma ray spectrometry as well as some deficiencies of existing computer programs for the quantitative evaluation of spectra are discussed and some practical solutions are proposed. (author)

  13. Glucose-6-Phosphate Dehydrogenase Deficiency and Neonatal Hyperbilirubinemia

    Directory of Open Access Journals (Sweden)

    Ezzat Khodashenas

    2015-09-01

    Conclusion: Based on the findings, establishment of an early G6PD screening program, which can prevent further complications in neonates, seems essential, particularly in countries such as Iran where G6PD deficiency is highly prevalent.

  14. SD-CAS: Spin Dynamics by Computer Algebra System.

    Science.gov (United States)

    Filip, Xenia; Filip, Claudiu

    2010-11-01

    A computer algebra tool for describing the Liouville-space quantum evolution of nuclear 1/2-spins is introduced and implemented within a computational framework named Spin Dynamics by Computer Algebra System (SD-CAS). A distinctive feature compared with numerical and previous computer algebra approaches to solving spin dynamics problems results from the fact that no matrix representation for spin operators is used in SD-CAS, which determines a full symbolic character to the performed computations. Spin correlations are stored in SD-CAS as four-entry nested lists of which size increases linearly with the number of spins into the system and are easily mapped into analytical expressions in terms of spin operator products. For the so defined SD-CAS spin correlations a set of specialized functions and procedures is introduced that are essential for implementing basic spin algebra operations, such as the spin operator products, commutators, and scalar products. They provide results in an abstract algebraic form: specific procedures to quantitatively evaluate such symbolic expressions with respect to the involved spin interaction parameters and experimental conditions are also discussed. Although the main focus in the present work is on laying the foundation for spin dynamics symbolic computation in NMR based on a non-matrix formalism, practical aspects are also considered throughout the theoretical development process. In particular, specific SD-CAS routines have been implemented using the YACAS computer algebra package (http://yacas.sourceforge.net), and their functionality was demonstrated on a few illustrative examples. PMID:20843716

  15. Displacement of p130Cas from focal adhesions links actomyosin contraction to cell migration.

    Science.gov (United States)

    Machiyama, Hiroaki; Hirata, Hiroaki; Loh, Xia Kun; Kanchi, Madhu Mathi; Fujita, Hideaki; Tan, Song Hui; Kawauchi, Keiko; Sawada, Yasuhiro

    2014-08-15

    Cell adhesion complexes provide platforms where cell-generated forces are transmitted to the extracellular matrix (ECM). Tyrosine phosphorylation of focal adhesion proteins is crucial for cells to communicate with the extracellular environment. However, the mechanisms that transmit actin cytoskeletal motion to the extracellular environment to drive cell migration are poorly understood. We find that the movement of p130Cas (Cas, also known as BCAR1), a mechanosensor at focal adhesions, correlates with actin retrograde flow and depends upon actomyosin contraction and phosphorylation of the Cas substrate domain (CasSD). This indicates that CasSD phosphorylation underpins the physical link between Cas and the actin cytoskeleton. Fluorescence recovery after photobleaching (FRAP) experiments reveal that CasSD phosphorylation, as opposed to the association of Cas with Src, facilitates Cas displacement from adhesion complexes in migrating cells. Furthermore, the stabilization of Src-Cas binding and inhibition of myosin II, both of which sustain CasSD phosphorylation but mitigate Cas displacement from adhesion sites, retard cell migration. These results indicate that Cas promotes cell migration by linking actomyosin contractions to the adhesion complexes through a dynamic interaction with Src as well as through the phosphorylation-dependent association with the actin cytoskeleton. PMID:24928898

  16. Antepartum ornithine transcarbamylase deficiency.

    Science.gov (United States)

    Nakajima, Hitoshi; Sasaki, Yosuke; Maeda, Tadashi; Takeda, Masako; Hara, Noriko; Nakanishi, Kazushige; Urita, Yoshihisa; Hattori, Risa; Miura, Ken; Taniguchi, Tomoko

    2014-01-01

    Ornithine transcarbamylase deficiency (OTCD) is the most common type urea cycle enzyme deficiencies. This syndrome results from a deficiency of the mitochondrial enzyme ornithine transcarbamylase, which catalyzes the conversion of ornithine and carbamoyl phosphate to citrullin. Our case was a 28-year-old female diagnosed with OTCD following neurocognitive deficit during her first pregnancy. Although hyperammonemia was suspected as the cause of the patient's mental changes, there was no evidence of chronic liver disease. Plasma amino acid and urine organic acid analysis revealed OTCD. After combined modality treatment with arginine, sodium benzoate and hemodialysis, the patient's plasma ammonia level stabilized and her mental status returned to normal. At last she recovered without any damage left. PMID:25759629

  17. Study of Eclipsing Binary and Multiple Systems in OB Associations IV: Cas OB6 Member DN Cas

    CERN Document Server

    Bakis, V; Bilir, S; Eker, Z

    2016-01-01

    An early-type, massive, short-period ($P_{\\rm orb}=2^{\\rm d}.310951$) eclipsing spectroscopic binary DN Cas has been re-visited with new spectral and photometric data. The masses and radii of the components have been obtained as $M_1=19.04\\pm0.07 M_\\odot$, $M_2=13.73\\pm 0.05M_\\odot$ and $R_1=7.22\\pm0.06 R_\\odot$, $R_2=5.79\\pm0.06R_\\odot$, respectively. Both components present synchronous rotation ($V_{\\rm rot1}=160\\,$km s$^{-1}$, $V_{\\rm rot2}=130\\,$km s$^{-1}$) with their orbit. Orbital period analysis yielded a physically bound additional component in the system with a minimum mass of $M_3=0.88 M_\\odot$ orbiting in an eccentric orbit ($e=0.37\\pm0.2$) with an orbital period of $P_{12}=42\\pm9$ yrs. High precision absolute parameters of the system allowed us to derive a distance to DN Cas as 1.7$\\pm$0.2 kpc which locates the system within the borders of the Cas OB6 association (d=1.8 kpc; Mel'nik & Dambis, 2009). The space velocities and the age of DN Cas are in agreement with those of Cas OB6. The age of ...

  18. CRISPR-Cas9: A Revolutionary Tool for Cancer Modelling

    Directory of Open Access Journals (Sweden)

    Raul Torres-Ruiz

    2015-09-01

    Full Text Available The cancer-modelling field is now experiencing a conversion with the recent emergence of the RNA-programmable CRISPR-Cas9 system, a flexible methodology to produce essentially any desired modification in the genome. Cancer is a multistep process that involves many genetic mutations and other genome rearrangements. Despite their importance, it is difficult to recapitulate the degree of genetic complexity found in patient tumors. The CRISPR-Cas9 system for genome editing has been proven as a robust technology that makes it possible to generate cellular and animal models that recapitulate those cooperative alterations rapidly and at low cost. In this review, we will discuss the innovative applications of the CRISPR-Cas9 system to generate new models, providing a new way to interrogate the development and progression of cancers.

  19. Exploiting CRISPR/Cas: Interference Mechanisms and Applications

    Directory of Open Access Journals (Sweden)

    André Plagens

    2013-07-01

    Full Text Available The discovery of biological concepts can often provide a framework for the development of novel molecular tools, which can help us to further understand and manipulate life. One recent example is the elucidation of the prokaryotic adaptive immune system, clustered regularly interspaced short palindromic repeats (CRISPR/CRISPR-associated (Cas that protects bacteria and archaea against viruses or conjugative plasmids. The immunity is based on small RNA molecules that are incorporated into versatile multi-domain proteins or protein complexes and specifically target viral nucleic acids via base complementarity. CRISPR/Cas interference machines are utilized to develop novel genome editing tools for different organisms. Here, we will review the latest progress in the elucidation and application of prokaryotic CRISPR/Cas systems and discuss possible future approaches to exploit the potential of these interference machineries.

  20. Targeted Heritable Mutation and Gene Conversion by Cas9-CRISPR in Caenorhabditis elegans

    OpenAIRE

    Katic, Iskra; Großhans, Helge

    2013-01-01

    We have achieved targeted heritable genome modification in Caenorhabditis elegans by injecting mRNA of the nuclease Cas9 and Cas9 guide RNAs. This system rapidly creates precise genomic changes, including knockouts and transgene-instructed gene conversion.

  1. Costo-beneficio de un programa preventivo y terapéutico para reducir la deficiencia de hierro en Argentina Cost-benefit of a prevention and treatment program to reduce iron deficiency in Argentina

    Directory of Open Access Journals (Sweden)

    Ignacio Drake

    2009-01-01

    Full Text Available OBJETIVOS: Estimar los costos directos e indirectos de la deficiencia de hierro (DH y la anemia ferropénica (AF en Argentina y compararlos con los costos de un programa específico para su prevención y tratamiento. MÉTODOS: Análisis de escenario previo (ex ante de la relación costo-beneficio de un programa de prevención y tratamiento de la AF dirigido a todos los niños y las embarazadas pobres y sin cobertura social de Argentina. Las consecuencias económicas de la DH y la AF se estimaron a partir de los costos directos - gastos vinculados a la atención de un parto prematuro - e indirectos - pérdidas en la productividad futura de los niños por su peor desarrollo cognitivo debido a la DH y la menor productividad de los adultos por la AF - mediante la metodología específica desarrollada por The Micronutrient Initiative. Las intervenciones se definieron según las Guías de Práctica Clínica vigentes en Argentina y los costos de los componentes se tomaron de los precios de las licitaciones del Ministerio de Salud de la Nación. RESULTADOS: Cada US$ 1,00 invertido en un programa de prevención y tratamiento de la DH y la AF, con una cobertura de 90% de la población de lactantes y embarazadas pobres sin seguro explícito de salud, representaría un ahorro de US$ 33,40 por la prevención de las pérdidas económicas debidas a estas enfermedades. CONCLUSIÓN: Las intervenciones para enfrentar la DH no solo mejoran significativamente el estado de salud de la población, sino que representan un ahorro considerable de recursos.OBJECTIVES: To estimate the direct and indirect cost of iron deficiency (ID and iron-deficiency anemia (IDA in Argentina and compare it with the cost of a prevention and treatment program. METHODS: Analysis of a prior scenario to gage the relative cost-benefit of an IDA prevention and treatment program for all low-income children and expectant mothers without social coverage/benefits in Argentina. The economic

  2. Performance of the Cas9 Nickase System in Drosophila melanogaster

    OpenAIRE

    Ren, Xingjie; Yang, Zhihao; Mao, Decai; Chang, Zai; Qiao, Huan-Huan; Wang, Xia; Sun, Jin; Hu, Qun; Cui, Yan; Liu, Lu-Ping; Ji, Jun-Yuan; Xu, Jiang; Ni, Jian-Quan

    2014-01-01

    Recent studies of the Cas9/sgRNA system in Drosophila melanogaster genome editing have opened new opportunities to generate site-specific mutant collections in a high-throughput manner. However, off-target effects of the system are still a major concern when analyzing mutant phenotypes. Mutations converting Cas9 to a DNA nickase have great potential for reducing off-target effects in vitro. Here, we demonstrated that injection of two plasmids encoding neighboring offset sgRNAs into transgenic...

  3. Applications of CRISPR-Cas9 mediated genome engineering

    Institute of Scientific and Technical Information of China (English)

    Xiao Yang

    2015-01-01

    Targeted mutagenesis based on homologous recombination has been a powerful tool for understanding the mechanisms underlying development, normal physiology, and disease. A recent breakthrough in genome engineering technology based on the class of RNA-guided endonucleases, such as clustered regularly interspaced short palindromic repeats (CRISPR)-associated Cas9, is further revolutionizing biology and medical studies. The simplicity of the CRISPR-Cas9 system has enabled its widespread applications in generating germline animal models, somatic genome engineering, and functional genomic screening and in treating genetic and infectious diseases. This technology will likely be used in all fields of biomedicine, ranging from basic research to human gene therapy.

  4. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... the body. Iron-deficiency anemia usually develops over time if your body doesn't have enough iron ... Institutes of Health—shows how Susan, a full-time worker and student, has coped with having iron- ...

  5. Iron-Deficiency Anemia

    Medline Plus

    Full Text Available ... body. Low iron levels usually are due to blood loss, poor diet, or an inability to absorb enough iron from food. Overview Iron-deficiency anemia is a common type of anemia . The term "anemia" usually refers to ...

  6. Alpha1-antitrypsin deficiency

    DEFF Research Database (Denmark)

    Stolk, Jan; Seersholm, Niels; Kalsheker, Noor

    2006-01-01

    biennially to exchange views and research findings. The fourth biennial meeting was held in Copenhagen, Denmark, on 2-3 June 2005. This review covers the wide range of AAT deficiency-related topics that were addressed encompassing advances in genetic characterization, risk factor identification, clinical...... epidemiology, inflammatory and signalling processes, therapeutic advances, and lung imaging techniques....

  7. MCAD deficiency in Denmark

    DEFF Research Database (Denmark)

    Andresen, Brage Storstein; Lund, Allan Meldgaard; Hougaard, David Michael; Christensen, Ernst; Gahrn, Birthe; Christensen, Mette; Bross, Peter; Vested, Anne; Simonsen, Henrik; Skogstrand, Kristin; Olpin, Simon; Brandt, Niels Jacob; Skovby, Flemming; Nørgaard-Pedersen, Bent; Gregersen, Niels

    2012-01-01

    Medium-chain acyl-CoA dehydrogenase deficiency (MCADD) is the most common defect of fatty acid oxidation. Many countries have introduced newborn screening for MCADD, because characteristic acylcarnitines can easily be identified in filter paper blood spot samples by tandem mass spectrometry (MS/M...

  8. Carnitine deficiency disorders in children.

    Science.gov (United States)

    Stanley, Charles A

    2004-11-01

    acylcarnitines has been helpful in expanded newborn screening programs using tandem mass-spectrometry of blood spot acylcarnitine profiles to detect genetic fatty acid oxidation defects in neonates. Carnitine-deficient diets (vegetarian) do not have much effect on carnitine pools in adults. A modest 50% reduction in carnitine levels is associated with hyperalimentation in newborn infants, but is of doubtful significance. The above considerations indicate that carnitine does not become rate-limiting unless extremely low; testing the benefits of nutritional supplements may require invasive endurance studies of fasting ketogenesis or muscle and cardiovascular work. PMID:15591002

  9. Small Molecule-Triggered Cas9 Protein with Improved Genome-Editing Specificity

    OpenAIRE

    Davis, Kevin M.; Pattanayak, Vikram; Thompson, David B.; Zuris, John A.; Liu, David R.

    2015-01-01

    Directly modulating the activity of genome-editing proteins has the potential to increase their specificity by reducing activity following target locus modification. We developed Cas9 nucleases that are activated by the presence of a cell-permeable small molecule by inserting an evolved 4-hydroxytamoxifen (4-HT)-responsive intein at specific positions in Cas9. In human cells, conditionally active Cas9s modify target genomic sites with up to 25-fold higher specificity than wild-type Cas9.

  10. A simplified and efficient germline-specific CRISPR/Cas9 system for Drosophila genomic engineering

    OpenAIRE

    Sebo, Zachary L; Lee, Han B.; Peng, Ying; Guo, Yi

    2013-01-01

    The type II CRISPR/Cas9 system (clustered regularly interspaced short palindromic repeats/CRISPR-associated) has recently emerged as an efficient and simple tool for site-specific engineering of eukaryotic genomes. To improve its applications in Drosophila genome engineering, we simplified the standard two-component CRISPR/Cas9 system by generating a stable transgenic fly line expressing the Cas9 endonuclease in the germline (Vasa-Cas9 line). By injecting vectors expressing engineered target-...

  11. DNA and RNA interference mechanisms by CRISPR-Cas surveillance complexes

    OpenAIRE

    Plagens, Andre; Richter, Hagen; Charpentier, Emmanuelle; Randau, Lennart

    2015-01-01

    The CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) adaptive immune systems use small guide RNAs, the CRISPR RNAs (crRNAs), to mark foreign genetic material, e.g. viral nucleic acids, for degradation. Archaea and bacteria encode a large variety of Cas proteins that bind crRNA molecules and build active ribonucleoprotein surveillance complexes. The evolution of CRISPR-Cas systems has resulted in a diversification of cas genes and a classification of t...

  12. Occurrence and Diversity of CRISPR-Cas Systems in the Genus Bifidobacterium

    OpenAIRE

    Briner, Alexandra E.; Lugli, Gabriele Andrea; Milani, Christian; Duranti, Sabrina; Turroni, Francesca; Gueimonde, Miguel; Margolles, Abelardo; van Sinderen, Douwe; Ventura, Marco; Barrangou, Rodolphe

    2015-01-01

    CRISPR-Cas systems constitute adaptive immune systems for antiviral defense in bacteria. We investigated the occurrence and diversity of CRISPR-Cas systems in 48 Bifidobacterium genomes to gain insights into the diversity and co-evolution of CRISPR-Cas systems within the genus and investigate CRISPR spacer content. We identified the elements necessary for the successful targeting and inference of foreign DNA in select Type II CRISPR-Cas systems, including the tracrRNA and target PAM sequence....

  13. Cleavage of Phage DNA by the Streptococcus thermophilus CRISPR3-Cas System

    OpenAIRE

    Magadán, Alfonso H.; Marie-Ève Dupuis; Manuela Villion; Sylvain Moineau

    2012-01-01

    Streptococcus thermophilus, similar to other Bacteria and Archaea, has developed defense mechanisms to protect cells against invasion by foreign nucleic acids, such as virus infections and plasmid transformations. One defense system recently described in these organisms is the CRISPR-Cas system (Clustered Regularly Interspaced Short Palindromic Repeats loci coupled to CRISPR-associated genes). Two S. thermophilus CRISPR-Cas systems, CRISPR1-Cas and CRISPR3-Cas, have been shown to actively blo...

  14. CRISPR/Cas-Mediated Site-Specific Mutagenesis in Arabidopsis thaliana Using Cas9 Nucleases and Paired Nickases.

    Science.gov (United States)

    Schiml, Simon; Fauser, Friedrich; Puchta, Holger

    2016-01-01

    The CRISPR/Cas system has recently become the most important tool for genome engineering due to its simple architecture that allows for rapidly changing the target sequence and its applicability to organisms throughout all kingdoms of life. The need for an easy-to-use and reliable nuclease is especially high in plant research, as precise genome modifications are almost impossible to achieve by Agrobacterium-mediated transformation and the regeneration of plants from protoplast cultures is very labor intensive. Here, we describe the application of the Cas9 nuclease to Arabidopsis thaliana for the induction of heritable targeted mutations, which may also be used for other plant species. To cover the concern for off-target activity, we also describe the generation of stable mutants using paired Cas9 nickases. PMID:27557689

  15. 41 CFR 102-33.435 - What CAS cost and utilization data must we report?

    Science.gov (United States)

    2010-07-01

    ... 41 Public Contracts and Property Management 3 2010-07-01 2010-07-01 false What CAS cost and utilization data must we report? 102-33.435 Section 102-33.435 Public Contracts and Property Management... Services (cas) Cost and Utilization Data § 102-33.435 What CAS cost and utilization data must we...

  16. On the Integration of Computer Algebra Systems (CAS) by Canadian Mathematicians: Results of a National Survey

    Science.gov (United States)

    Buteau, Chantal; Jarvis, Daniel H.; Lavicza, Zsolt

    2014-01-01

    In this article, we outline the findings of a Canadian survey study (N = 302) that focused on the extent of computer algebra systems (CAS)-based technology use in postsecondary mathematics instruction. Results suggest that a considerable number of Canadian mathematicians use CAS in research and teaching. CAS use in research was found to be the…

  17. Guide RNAs: A Glimpse at the Sequences that Drive CRISPR-Cas Systems.

    Science.gov (United States)

    Briner, Alexandra E; Barrangou, Rodolphe

    2016-01-01

    CRISPR-Cas systems provide adaptive immunity in bacteria and archaea. Although there are two main classes of CRISPR-Cas systems defined by gene content, interfering RNA biogenesis, and effector proteins, Type II systems have recently been exploited on a broad scale to develop next-generation genetic engineering and genome-editing tools. Conveniently, Type II systems are streamlined and rely on a single protein, Cas9, and a guide RNA molecule, comprised of a CRISPR RNA (crRNA) and trans-acting CRISPR RNA (tracrRNA), to achieve effective and programmable nucleic acid targeting and cleavage. Currently, most commercially available Cas9-based genome-editing tools use the CRISPR-Cas system from Streptococcus pyogenes (SpyCas9), although many orthogonal Type II systems are available for diverse and multiplexable genome engineering applications. Here, we discuss the biological significance of Type II CRISPR-Cas elements, including the tracrRNA, crRNA, Cas9, and protospacer-adjacent motif (PAM), and look at the native function of these elements to understand how they can be engineered, enhanced, and optimized for genome editing applications. Additionally, we discuss the basis for orthogonal Cas9 and guide RNA systems that would allow researchers to concurrently use multiple Cas9-based systems for different purposes. Understanding the native function of endogenous Type II CRISPR-Cas systems can lead to new Cas9 tool development to expand the genetic manipulation toolbox. PMID:27371605

  18. 78 FR 40665 - Cost Accounting Standards: CAS 413 Pension Adjustments for Extraordinary Events

    Science.gov (United States)

    2013-07-08

    ... Notice of Proposed Rulemaking (NPRM) on pension harmonization (the CAS Pension Harmonizatoin Rule, 75 FR... Harmonization Rule was published as a Final Rule (76 FR 81296, December 27, 2011), the CAS Board summarized and... BUDGET Office of Federal Procurement Policy 48 CFR Part 9904 Cost Accounting Standards: CAS 413...

  19. High Prevalence of Vitamin D Deficiency in Cambodian Women: A Common Deficiency in a Sunny Country.

    Science.gov (United States)

    Smith, Geoffry; Wimalawansa, Sunil J; Laillou, Arnaud; Sophonneary, Prak; Un, Samoeurn; Hong, Rathavuth; Poirot, Etienne; Kuong, Khov; Chamnan, Chhoun; De Los Reyes, Francisco N; Wieringa, Frank T

    2016-01-01

    Recent studies have shown that in spite of being generally close to the equator; vitamin D deficiency is common in South East Asian countries. In order to quantify micronutrient status for women and children in Cambodia; a nationally-representative survey was conducted in 2014 linked to the Cambodian Demographic Health Survey. The countrywide median of 25(OH)D was, respectively, 64.9 and 91.1 nmol/L for mothers and children. Based on The Endocrine Society cutoffs (>50children had plasma vitamin D concentrations indicating insufficiency or deficiency. For deficiency alone, 29% of the mothers were found to be vitamin D deficient, but only 13.4% of children. Children who live in urban areas had a 43% higher rate of vitamin D insufficiency versus those who live in rural areas (OR; 1.434; 95% CI: 1.007; 2.041). However, such differences were not observed in their mothers. The high prevalence of vitamin D deficiency is likely in part due to lifestyle choices, including sun avoidance, increasingly predominant indoor work, and covered transport. These survey findings support the need for a broader national Cambodian study incorporating testing of adult men, adolescents and the elderly, and encompassing other parameters such as skeletal health. However, the data presented in this study already show significant deficiencies which need to be addressed and we discuss the benefit of establishing nationally-mandated food fortification programs to enhance the intake of vitamin D. PMID:27187456

  20. Glucose-6-phosphate dehydrogenase deficiency

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/000528.htm Glucose-6-phosphate dehydrogenase deficiency To use the sharing features on this page, please enable JavaScript. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a condition ...

  1. Transient partial growth hormone deficiency due to zinc deficiency.

    Science.gov (United States)

    Nishi, Y; Hatano, S; Aihara, K; Fujie, A; Kihara, M

    1989-04-01

    We present here a 13-year-old boy with partial growth hormone deficiency due to chronic mild zinc deficiency. When zinc administration was started, his growth rate, growth hormone levels, and plasma zinc concentrations increased significantly. His poor dietary intake resulted in chronic mild zinc deficiency, which in turn could be the cause of a further loss of appetite and growth retardation. There was also a possibility of renal zinc wasting which may have contributed to zinc deficiency. Zinc deficiency should be carefully ruled out in patients with growth retardation. PMID:2708733

  2. Iron deficiency and cognitive functions

    Directory of Open Access Journals (Sweden)

    Jáuregui-Lobera I

    2014-11-01

    Full Text Available Ignacio Jáuregui-Lobera Department of Nutrition and Bromatology, Pablo de Olavide University, Seville, Spain Abstract: Micronutrient deficiencies, especially those related to iodine and iron, are linked to different cognitive impairments, as well as to potential long-term behavioral changes. Among the cognitive impairments caused by iron deficiency, those referring to attention span, intelligence, and sensory perception functions are mainly cited, as well as those associated with emotions and behavior, often directly related to the presence of iron deficiency anemia. In addition, iron deficiency without anemia may cause cognitive disturbances. At present, the prevalence of iron deficiency and iron deficiency anemia is 2%–6% among European children. Given the importance of iron deficiency relative to proper cognitive development and the alterations that can persist through adulthood as a result of this deficiency, the objective of this study was to review the current state of knowledge about this health problem. The relevance of iron deficiency and iron deficiency anemia, the distinction between the cognitive consequences of iron deficiency and those affecting specifically cognitive development, and the debate about the utility of iron supplements are the most relevant and controversial topics. Despite there being methodological differences among studies, there is some evidence that iron supplementation improves cognitive functions. Nevertheless, this must be confirmed by means of adequate follow-up studies among different groups. Keywords: iron deficiency, anemia, cognitive functions, supplementation

  3. CAS Researchers Honored by Hua Loo-Keng Mathematics Prize

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ Two mathematicians of the CAS Academy of Mathematics and System Sciences have received the 2003 Hua LooKeng Mathematics Prize. This was announced by Prof. Ma Zhiming,president of the Chinese Mathematics Society (CMS), at the Ninth CMS Congress inaugurated on October 31 at Wuhan University in central China's Hubei Province.

  4. Preparing Students to Take SOA/CAS Exam FM/2

    Science.gov (United States)

    Marchand, Richard J.

    2014-01-01

    This paper provides suggestions for preparing students to take the actuarial examination on financial mathematics, SOA/CAS Exam FM/2. It is based on current practices employed at Slippery Rock University, a small public liberal arts university. Detailed descriptions of our Theory of Interest course and subsequent Exam FM/2 prep course are provided…

  5. Maximizing mutagenesis with solubilized CRISPR-Cas9 ribonucleoprotein complexes.

    Science.gov (United States)

    Burger, Alexa; Lindsay, Helen; Felker, Anastasia; Hess, Christopher; Anders, Carolin; Chiavacci, Elena; Zaugg, Jonas; Weber, Lukas M; Catena, Raul; Jinek, Martin; Robinson, Mark D; Mosimann, Christian

    2016-06-01

    CRISPR-Cas9 enables efficient sequence-specific mutagenesis for creating somatic or germline mutants of model organisms. Key constraints in vivo remain the expression and delivery of active Cas9-sgRNA ribonucleoprotein complexes (RNPs) with minimal toxicity, variable mutagenesis efficiencies depending on targeting sequence, and high mutation mosaicism. Here, we apply in vitro assembled, fluorescent Cas9-sgRNA RNPs in solubilizing salt solution to achieve maximal mutagenesis efficiency in zebrafish embryos. MiSeq-based sequence analysis of targeted loci in individual embryos using CrispRVariants, a customized software tool for mutagenesis quantification and visualization, reveals efficient bi-allelic mutagenesis that reaches saturation at several tested gene loci. Such virtually complete mutagenesis exposes loss-of-function phenotypes for candidate genes in somatic mutant embryos for subsequent generation of stable germline mutants. We further show that targeting of non-coding elements in gene regulatory regions using saturating mutagenesis uncovers functional control elements in transgenic reporters and endogenous genes in injected embryos. Our results establish that optimally solubilized, in vitro assembled fluorescent Cas9-sgRNA RNPs provide a reproducible reagent for direct and scalable loss-of-function studies and applications beyond zebrafish experiments that require maximal DNA cutting efficiency in vivo. PMID:27130213

  6. CAS leads development of application system for spaceship

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ The spacecraft Shenzhou-VII blasted off on 25 September,carrying three astronauts to attempt China's first-ever spacewalk.It is the seventh space mission successfully launched by this country.As usual,its space application system,one of the seven major systems for this space engineering project,is developed under the leadership of CAS.

  7. Evolution and classification of the CRISPR-Cas systems

    NARCIS (Netherlands)

    Makarova, K.S.; Brouns, S.J.J.; Oost, van der J.

    2011-01-01

    The CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) modules are adaptive immunity systems that are present in many archaea and bacteria. These defence systems are encoded by operons that have an extraordinarily diverse architecture and a high rate of

  8. Duchenne muscular dystrophy: CRISPR/Cas9 treatment.

    Science.gov (United States)

    Mendell, Jerry R; Rodino-Klapac, Louise R

    2016-05-01

    A novel approach to gene correction by genome editing shows great promise as a treatment for Duchenne muscular dystrophy (DMD). CRISPR/Cas9 delivered by adeno-associated virus to a mouse model for DMD demonstrated improvement in function and histology. PMID:26926391

  9. Recent Achievements Scored by CAS Scientists in Life Sciences

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ Rice Genome Project for Oryza sativa L. Ssp. Indica After 17-month hard work, the scientists of the CAS Beijing Genomics Institute accomplished the draft sequence of the rice genome for Oryza sativa L. Ssp. Indica in October, 2001, and the data were unconditionally shared by the whole world. About 270,000 people had visited Chinese rice genome website by May 2003.

  10. Targeted mutagenesis in chicken using CRISPR/Cas9 system

    Science.gov (United States)

    Oishi, Isao; Yoshii, Kyoko; Miyahara, Daichi; Kagami, Hiroshi; Tagami, Takahiro

    2016-01-01

    The CRISPR/Cas9 system is a simple and powerful tool for genome editing in various organisms including livestock animals. However, the system has not been applied to poultry because of the difficulty in accessing their zygotes. Here we report the implementation of CRISPR/Cas9-mediated gene targeting in chickens. Two egg white genes, ovalbumin and ovomucoid, were efficiently (>90%) mutagenized in cultured chicken primordial germ cells (PGCs) by transfection of circular plasmids encoding Cas9, a single guide RNA, and a gene encoding drug resistance, followed by transient antibiotic selection. We transplanted CRISPR-induced mutant-ovomucoid PGCs into recipient chicken embryos and established three germline chimeric roosters (G0). All of the roosters had donor-derived mutant-ovomucoid spermatozoa, and the two with a high transmission rate of donor-derived gametes produced heterozygous mutant ovomucoid chickens as about half of their donor-derived offspring in the next generation (G1). Furthermore, we generated ovomucoid homozygous mutant offspring (G2) by crossing the G1 mutant chickens. Taken together, these results demonstrate that the CRISPR/Cas9 system is a simple and effective gene-targeting method in chickens. PMID:27050479

  11. CAS presents Jeoujang Jaw Award at COSPAR Assembly

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ CAS Vice President JIANG Mianheng led a Chinese delegation to attend the 37th Scientific Assembly of the International Committee on Space Research (COSPAR) held from 13 to 20 July in Montreal,Canada.At the opening ceremony of the assembly,Prof.Jiang presented the 2008 Jeoujang Jaw Award to its winner.

  12. Indispensable Manual Calculation Skills in a CAS Environment.

    Science.gov (United States)

    Herget, Wilfried; Heugl, Helmut; Kutzler, Bernhard; Lehmann, Eberhard

    Which manual calculation skills are still needed when students use graphic/symbolic calculators or computers with computer algebra systems (CAS)? What should students be able to do manually, i.e. just using paper and pencil? This text is the outcome of a two-day discussion on these questions, held by the four authors. Our answers and proposals are…

  13. Novel progress in clean energy partnership between CAS and BP

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    @@ On behalf of their respective organizations,CAS Vice President LI Jinghai and BP Group Chief Executive lain Conn recently put their names on an agreement for the partner selection and technology roadmap principle for the Clean Energy Commercialization Center (CECC),a joint venture between the two sides.

  14. CAS-potentialer realiseret som fleksibilitet i matematiske begreber

    DEFF Research Database (Denmark)

    Andresen, Mette

    2006-01-01

    skift mellem niveauerne i den model for matematisk begrebsdannelse og ?udvikling gennem matematisering som danner grundlagt for Realistic Math Education. I foredraget gives eksempler på hvordan brugen af CAS fremmer fleksibiliteten i gymnasieelevers opfattelse af differentialligninger og deres løsning...

  15. The 2008 February superoutburst of V452 Cas

    CERN Document Server

    Lloyd, Christopher; Shears, Jeremy; Miller, Ian; Boyd, David; Brady, Steve

    2008-01-01

    Observations of the 2008 February outburst of V452 Cas show that the profile, duration and magnitude at maximum were very similar to the previous superoutburst in 2007 September. Low-amplitude variations consistent with previously observed superhumps were also seen.

  16. CAS Delegation Visited the Aprica Childcare Institute,Japan

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ A delegation from the China Association of Standardization(CAS), headed by Mr. Ma Lincong, secretary-general, visited Aprica Childcare Institute, Japan from June 19 to June 26, 2004, to carry out technical investigation for the drafting of a new standard for in-car child restraints.

  17. Genome Editing with CRISPR-Cas9: Can It Get Any Better?

    Science.gov (United States)

    Haeussler, Maximilian; Concordet, Jean-Paul

    2016-05-20

    The CRISPR-Cas revolution is taking place in virtually all fields of life sciences. Harnessing DNA cleavage with the CRISPR-Cas9 system of Streptococcus pyogenes has proven to be extraordinarily simple and efficient, relying only on the design of a synthetic single guide RNA (sgRNA) and its co-expression with Cas9. Here, we review the progress in the design of sgRNA from the original dual RNA guide for S. pyogenes and Staphylococcus aureus Cas9 (SpCas9 and SaCas9). New assays for genome-wide identification of off-targets have provided important insights into the issue of cleavage specificity in vivo. At the same time, the on-target activity of thousands of guides has been determined. These data have led to numerous online tools that facilitate the selection of guide RNAs in target sequences. It appears that for most basic research applications, cleavage activity can be maximized and off-targets minimized by carefully choosing guide RNAs based on computational predictions. Moreover, recent studies of Cas proteins have further improved the flexibility and precision of the CRISPR-Cas toolkit for genome editing. Inspired by the crystal structure of the complex of sgRNA-SpCas9 bound to target DNA, several variants of SpCas9 have recently been engineered, either with novel protospacer adjacent motifs (PAMs) or with drastically reduced off-targets. Novel Cas9 and Cas9-like proteins called Cpf1 have also been characterized from other bacteria and will benefit from the insights obtained from SpCas9. Genome editing with CRISPR-Cas9 may also progress with better understanding and control of cellular DNA repair pathways activated after Cas9-induced DNA cleavage. PMID:27210042

  18. Biotin and biotinidase deficiency

    OpenAIRE

    Zempleni, Janos; Hassan, Yousef I.; Wijeratne, Subhashinee SK

    2008-01-01

    Biotin is a water-soluble vitamin that serves as an essential coenzyme for five carboxylases in mammals. Biotin-dependent carboxylases catalyze the fixation of bicarbonate in organic acids and play crucial roles in the metabolism of fatty acids, amino acids and glucose. Carboxylase activities decrease substantially in response to biotin deficiency. Biotin is also covalently attached to histones; biotinylated histones are enriched in repeat regions in the human genome and appear to play a role...

  19. Highly Specific Epigenome Editing by CRISPR/Cas9 Repressors for Silencing of Distal Regulatory Elements

    OpenAIRE

    Thakore, Pratiksha I; D’Ippolito, Anthony M.; Song, Lingyun; Safi, Alexias; Shivakumar, Nishkala K.; Kabadi, Ami M.; Reddy, Timothy E.; Crawford, Gregory E; Gersbach, Charles A

    2015-01-01

    Epigenome editing with the CRISPR/Cas9 platform is a promising technology to modulate gene expression to direct cell phenotype and to dissect the causal epigenetic mechanisms of gene regulation. Fusions of the nuclease-inactive dCas9 to the KRAB repressor (dCas9-KRAB) can silence target gene expression, but the genome-wide specificity and the extent of heterochromatin formation catalyzed by dCas9-KRAB is not known. We targeted dCas9-KRAB to the HS2 enhancer, a distal regulatory element that o...

  20. A non-inheritable maternal Cas9-based multiple-gene editing system in mice

    OpenAIRE

    Takayuki Sakurai; Akiko Kamiyoshi; Hisaka Kawate; Chie Mori; Satoshi Watanabe; Megumu Tanaka; Ryuichi Uetake; Masahiro Sato; Takayuki Shindo

    2016-01-01

    The CRISPR/Cas9 system is capable of editing multiple genes through one-step zygote injection. The preexisting method is largely based on the co-injection of Cas9 DNA (or mRNA) and guide RNAs (gRNAs); however, it is unclear how many genes can be simultaneously edited by this method, and a reliable means to generate transgenic (Tg) animals with multiple gene editing has yet to be developed. Here, we employed non-inheritable maternal Cas9 (maCas9) protein derived from Tg mice with systemic Cas9...

  1. Conditional targeting of Ispd using paired Cas9 nickase and a single DNA template in mice

    OpenAIRE

    Angus Yiu-fai Lee; Kevin C. Kent Lloyd

    2014-01-01

    CRISPR/Cas9 technology is a highly promising genome editing tool in the mouse, potentially overcoming the costs and time required for more traditional gene targeting methods in embryonic stem (ES) cells. Recently, compared to the wildtype nuclease, paired Cas9 nickase (Cas9n) combined with single guide RNA (sgRNA) molecules has been found to enhance the specificity of genome editing while reducing off-target effects. Paired Cas9n has been shown to be as efficient as Cas9 for generating insert...

  2. Development of a mono-promoter-driven CRISPR/Cas9 system in mammalian cells

    OpenAIRE

    Shin Yoshioka; Wataru Fujii; Tetsuhiro Ogawa; Koji Sugiura; Kunihiko Naito

    2015-01-01

    The CRISPR/Cas9 system has been used for spatio-temporal gene modification through the ubiquitous expression of gRNA by an RNA polymerase III promoter and the controlled expression of Cas9 using a tissue-specific or inducible promoter. However, unexpected gene disruptions indicate the necessity of a tissue-specific or inducible expression of not only Cas9 but also gRNA. In the present study, we attempted to develop a CRISPR/Cas9 system that could express functional gRNAs and Cas9 by a single ...

  3. Development of an intein-mediated split-Cas9 system for gene therapy

    OpenAIRE

    Truong, D.J.J.; Kuehner, K.; Kuehn, R.; Werfel, S.; Engelhardt, S.; WURST, W; Ortiz, O.

    2015-01-01

    Using CRISPR/Cas9, it is possible to target virtually any gene in any organism. A major limitation to its application in gene therapy is the size of Cas9 (>4 kb), impeding its efficient delivery via recombinant adeno-associated virus (rAAV). Therefore, we developed a split–Cas9 system, bypassing the packaging limit using split-inteins. Each Cas9 half was fused to the corresponding split-intein moiety and, only upon co-expression, the intein-mediated trans-splicing occurs and the full Cas9 pro...

  4. Mobile CRISPR/Cas-mediated bacteriophage resistance in Lactococcus lactis.

    Directory of Open Access Journals (Sweden)

    Anne M Millen

    Full Text Available Lactococcus lactis is a biotechnological workhorse for food fermentations and potentially therapeutic products and is therefore widely consumed by humans. It is predominantly used as a starter microbe for fermented dairy products, and specialized strains have adapted from a plant environment through reductive evolution and horizontal gene transfer as evidenced by the association of adventitious traits with mobile elements. Specifically, L. lactis has armed itself with a myriad of plasmid-encoded bacteriophage defensive systems to protect against viral predation. This known arsenal had not included CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins, which forms a remarkable microbial immunity system against invading DNA. Although CRISPR/Cas systems are common in the genomes of closely related lactic acid bacteria (LAB, none was identified within the eight published lactococcal genomes. Furthermore, a PCR-based search of the common LAB CRISPR/Cas systems (Types I and II in 383 industrial L. lactis strains proved unsuccessful. Here we describe a novel, Type III, self-transmissible, plasmid-encoded, phage-interfering CRISPR/Cas discovered in L. lactis. The native CRISPR spacers confer resistance based on sequence identity to corresponding lactococcal phage. The interference is directed at phages problematic to the dairy industry, indicative of a responsive system. Moreover, targeting could be modified by engineering the spacer content. The 62.8-kb plasmid was shown to be conjugally transferrable to various strains. Its mobility should facilitate dissemination within microbial communities and provide a readily applicable system to naturally introduce CRISPR/Cas to industrially relevant strains for enhanced phage resistance and prevention against acquisition of undesirable genes.

  5. Iron Deficiency Anemia

    Directory of Open Access Journals (Sweden)

    Hassan Ahari

    1965-01-01

    Full Text Available The object of this paper is to draw attention to iron deficiency anemia which is the most common nutritional disturbance in infants and children. Iron deficiency anemia constitutes the most prevalent form of anemia in this age group. The records of infants and children admitted to the Pediatric Department of Tehran University Puhlavi Hospital for various ailments during a one year period (Mnrch l!l63 - HHi-t were analyzed. 262 infants and children out of a total number of an5, or 7t•/., showed iron deficiency anemia detect cd by blood film studies and hemoglobin determination, The majority, 123 or 4{.!t•/., of these patients were infants and children between six months and two years of age. The etiology indicates that faulty feeding is the main cause. Infections, parnsitcs, and hemorrhage were among other causes observed. ,'('itll regard to treatment, parenteral iron was preferred because cf its ef., Icctivcncss in short periods of hospital stay. In conclusion, the routine study of blood films and hemoglobin determiualion, especially in the low socio _ economic group of medically less organized countries is advised

  6. Dramatic enhancement of genome editing by CRISPR/Cas9 through improved guide RNA design.

    Science.gov (United States)

    Farboud, Behnom; Meyer, Barbara J

    2015-04-01

    Success with genome editing by the RNA-programmed nuclease Cas9 has been limited by the inability to predict effective guide RNAs and DNA target sites. Not all guide RNAs have been successful, and even those that were, varied widely in their efficacy. Here we describe and validate a strategy for Caenorhabditis elegans that reliably achieved a high frequency of genome editing for all targets tested in vivo. The key innovation was to design guide RNAs with a GG motif at the 3' end of their target-specific sequences. All guides designed using this simple principle induced a high frequency of targeted mutagenesis via nonhomologous end joining (NHEJ) and a high frequency of precise DNA integration from exogenous DNA templates via homology-directed repair (HDR). Related guide RNAs having the GG motif shifted by only three nucleotides showed severely reduced or no genome editing. We also combined the 3' GG guide improvement with a co-CRISPR/co-conversion approach. For this co-conversion scheme, animals were only screened for genome editing at designated targets if they exhibited a dominant phenotype caused by Cas9-dependent editing of an unrelated target. Combining the two strategies further enhanced the ease of mutant recovery, thereby providing a powerful means to obtain desired genetic changes in an otherwise unaltered genome. PMID:25695951

  7. The Cas6e ribonuclease is not required for interference and adaptation by the E. coli type I-E CRISPR-Cas system

    OpenAIRE

    Semenova, Ekaterina; Kuznedelov, Konstantin; Datsenko, Kirill A.; Boudry, Pierre M.; Savitskaya, Ekaterina E.; Medvedeva, Sofia; Beloglazova, Natalia; Logacheva, Maria; Yakunin, Alexander F.; Severinov, Konstantin

    2015-01-01

    CRISPR-Cas are small RNA-based adaptive prokaryotic immunity systems protecting cells from foreign DNA or RNA. Type I CRISPR-Cas systems are composed of a multiprotein complex (Cascade) that, when bound to CRISPR RNA (crRNA), can recognize double-stranded DNA targets and recruit the Cas3 nuclease to destroy target-containing DNA. In the Escherichia coli type I-E CRISPR-Cas system, crRNAs are generated upon transcription of CRISPR arrays consisting of multiple palindromic repeats and interveni...

  8. Iron-Deficiency Anemia (For Parents)

    Science.gov (United States)

    ... Things to Know About Zika & Pregnancy Iron-Deficiency Anemia KidsHealth > For Parents > Iron-Deficiency Anemia Print A ... common nutritional deficiency in children. About Iron-Deficiency Anemia Every red blood cell in the body contains ...

  9. Anti-cas spacers in orphan CRISPR4 arrays prevent uptake of active CRISPR-Cas I-F systems.

    Science.gov (United States)

    Almendros, Cristóbal; Guzmán, Noemí M; García-Martínez, Jesús; Mojica, Francisco J M

    2016-01-01

    Archaea and bacteria harbour clustered regularly interspaced short palindromic repeats (CRISPR) loci. These arrays encode RNA molecules (crRNA), each containing a sequence of a single repeat-intervening spacer. The crRNAs guide CRISPR-associated (Cas) proteins to cleave nucleic acids complementary to the crRNA spacer, thus interfering with targeted foreign elements. Notably, pre-existing spacers may trigger the acquisition of new spacers from the target molecule by means of a primed adaptation mechanism. Here, we show that naturally occurring orphan CRISPR arrays that contain spacers matching sequences of the cognate (absent) cas genes are able to elicit both primed adaptation and direct interference against genetic elements carrying those genes. Our findings show the existence of an anti-cas mechanism that prevents the transfer of a fully equipped CRISPR-Cas system. Hence, they suggest that CRISPR immunity may be undesired by particular prokaryotes, potentially because they could limit possibilities for gaining favourable sequences by lateral transfer. PMID:27573106

  10. The CRISPR-Associated Gene cas2 of Legionella pneumophila Is Required for Intracellular Infection of Amoebae

    OpenAIRE

    Felizza F Gunderson; Cianciotto, Nicholas P.

    2013-01-01

    ABSTRACT Recent studies have shown that the clustered regularly interspaced palindromic repeats (CRISPR) array and its associated (cas) genes can play a key role in bacterial immunity against phage and plasmids. Upon analysis of the Legionella pneumophila strain 130b chromosome, we detected a subtype II-B CRISPR-Cas locus that contains cas9, cas1, cas2, cas4, and an array with 60 repeats and 58 unique spacers. Reverse transcription (RT)-PCR analysis demonstrated that the entire CRISPR-Cas loc...

  11. Programming

    International Nuclear Information System (INIS)

    The programmer's task is often taken to be the construction of algorithms, expressed in hierarchical structures of procedures: this view underlies the majority of traditional programming languages, such as Fortran. A different view is appropriate to a wide class of problem, perhaps including some problems in High Energy Physics. The programmer's task is regarded as having three main stages: first, an explicit model is constructed of the reality with which the program is concerned; second, this model is elaborated to produce the required program outputs; third, the resulting program is transformed to run efficiently in the execution environment. The first two stages deal in network structures of sequential processes; only the third is concerned with procedure hierarchies. (orig.)

  12. CRISPR/Cas9 Cleavage of Viral DNA Efficiently Suppresses Vaccinia Viruses%CRISPR/Cas9切割痘病毒DNA抑制病毒复制

    Institute of Scientific and Technical Information of China (English)

    王娇娇; 张新敏; 倪爱民; 章康健; 刘新垣; 刘锡君

    2016-01-01

    当前,CRISPR/Cas9系统靶向痘苗病毒的相关研究鲜有报道.该文采用了CRISPR/Cas9基因组定点编辑技术,以痘病毒为研究对象.在重组痘病-毒WR-EGFP中针对EGFP基因序列设计不同靶点gRNAs,转染Cas9/gRNA质粒,同时感染WR-EGFP痘病毒,随后观察EGFP荧光表达、提取病毒基因组,通过PCR进行切口鉴定并运用错配酶切法验-CRISPR/Cas9对痘病毒靶点DNA切割的切口修复情况.与单独感染WR-EGFP组相比,转染了Cas9/gRNA 174和Cas9/gRNA 175质粒的EGFP荧光强度明显降低,表明CRISPR/Cas9抑制WR-EGFP病毒中EGFP基因的表达.相对定量PCR及错配酶切实验结果显示,CRISPR/Cas9切割靶点导致WR-EGFP病毒基因组产生了切口,并且存在DNA错配修复现象.最后,通过结晶紫染色法测定痘病毒的滴度,结果显示,经CRISPR/Cas9处理组的痘病毒滴度显著下降,即CRISPR/Cas9能够使痘病毒的复制能力显著地降低.综上所述,CRISPR/Cas9切割痘病毒基因组DNA并且抑制痘病毒复制.

  13. Insights into the CRISPR/Cas system of Gardnerella vaginalis

    Directory of Open Access Journals (Sweden)

    Pleckaityte Milda

    2012-12-01

    Full Text Available Abstract Background Gardnerella vaginalis is identified as the predominant colonist of the vaginal tracts of women diagnosed with bacterial vaginosis (BV. G. vaginalis can be isolated from healthy women, and an asymptomatic BV state is also recognised. The association of G. vaginalis with different clinical phenotypes could be explained by different cytotoxicity of the strains, presumably based on disparate gene content. The contribution of horizontal gene transfer to shaping the genomes of G. vaginalis is acknowledged. The CRISPR loci of the recently discovered CRISPR/Cas microbial defence system provide a historical view of the exposure of prokaryotes to a variety of foreign genetic elements. Results The CRISPR/Cas loci were analysed using available sequence data from three G. vaginalis complete genomes and 18 G. vaginalis draft genomes in the NCBI database, as well as PCR amplicons of the genomic DNA of 17 clinical isolates. The cas genes in the CRISPR/Cas loci of G. vaginalis belong to the E. coli subtype. Approximately 20% of the spacers had matches in the GenBank database. Sequence analysis of the CRISPR arrays revealed that nearly half of the spacers matched G. vaginalis chromosomal sequences. The spacers that matched G. vaginalis chromosomal sequences were determined to not be self-targeting and were presumably neither constituents of mobile-element-associated genes nor derived from plasmids/viruses. The protospacers targeted by these spacers displayed conserved protospacer-adjacent motifs. Conclusions The CRISPR/Cas system has been identified in about one half of the analysed G. vaginalis strains. Our analysis of CRISPR sequences did not reveal a potential link between their presence and the virulence of the G. vaginalis strains. Based on the origins of the spacers found in the G. vaginalis CRISPR arrays, we hypothesise that the transfer of genetic material among G. vaginalis strains could be regulated by the CRISPR/Cas mechanism. The

  14. Degeneration of a CRISPR/Cas system and its regulatory target during the evolution of a pathogen

    OpenAIRE

    Sampson, Timothy R.; Weiss, David S.

    2013-01-01

    CRISPR/Cas systems are bacterial RNA-guided endonuclease machineries that target foreign nucleic acids. Recently, we demonstrated that the Cas protein Cas9 controls gene expression and virulence in Francisella novicida by altering the stability of the mRNA for an immunostimulatory bacterial lipoprotein (BLP). Genomic analyses, however, revealed that Francisella species with increased virulence harbor degenerated CRISPR/Cas systems. We hypothesize that CRISPR/Cas degeneration removed a barrier...

  15. Treatment of carnitine deficiency.

    Science.gov (United States)

    Winter, S C

    2003-01-01

    Carnitine deficiency is a secondary complication of many inborn errors of metabolism. Pharmacological treatment with carnitine not only corrects the deficiency, it facilitates removal of accumulating toxic acyl intermediates and the generation of mitochondrial free coenzyme A (CoA). The United States Food and Drug Administration (US FDA) approved the use of carnitine for the treatment of inborn errors of metabolism in 1992. This approval was based on retrospective chart analysis of 90 patients, with 18 in the untreated cohort and 72 in the treated cohort. Efficacy was evaluated on the basis of clinical and biochemical findings. Compelling data included increased excretion of disease-specific acylcarnitine derivatives in a dose-response relationship, decreased levels of metabolites in the blood, and improved clinical status with decreased hospitalization frequency, improved growth and significantly lower mortality rates as compared to historical controls. Complications of carnitine treatment were few, with gastrointestinal disturbances and odour being the most frequent. No laboratory or clinical safety issues were identified. Intravenous carnitine preparations were also approved for treatment of secondary carnitine deficiency. Since only 25% of enteral carnitine is absorbed and gastrointestinal tolerance of high doses is poor, parenteral carnitine treatment is an appealing alternative therapeutic approach. In 7 patients treated long term with high-dose weekly to daily venous boluses of parenteral carnitine through a subcutaneous venous port, benefits included decreased frequency of decompensations, improved growth, improved muscle strength and decreased reliance on medical foods with liberalization of protein intake. Port infections were the most troubling complication. Theoretical concerns continue to be voiced that carnitine might result in fatal arrhythmias in patients with long-chain fat metabolism defects. No published clinical studies substantiate these

  16. Carnitine palmitoyltransferase II deficiency

    Science.gov (United States)

    Roe, C R.; Yang, B-Z; Brunengraber, H; Roe, D S.; Wallace, M; Garritson, B K.

    2008-01-01

    Background: Carnitine palmitoyltransferase II (CPT II) deficiency is an important cause of recurrent rhabdomyolysis in children and adults. Current treatment includes dietary fat restriction, with increased carbohydrate intake and exercise restriction to avoid muscle pain and rhabdomyolysis. Methods: CPT II enzyme assay, DNA mutation analysis, quantitative analysis of acylcarnitines in blood and cultured fibroblasts, urinary organic acids, the standardized 36-item Short-Form Health Status survey (SF-36) version 2, and bioelectric impedance for body fat composition. Diet treatment with triheptanoin at 30% to 35% of total daily caloric intake was used for all patients. Results: Seven patients with CPT II deficiency were studied from 7 to 61 months on the triheptanoin (anaplerotic) diet. Five had previous episodes of rhabdomyolysis requiring hospitalizations and muscle pain on exertion prior to the diet (two younger patients had not had rhabdomyolysis). While on the diet, only two patients experienced mild muscle pain with exercise. During short periods of noncompliance, two patients experienced rhabdomyolysis with exercise. None experienced rhabdomyolysis or hospitalizations while on the diet. All patients returned to normal physical activities including strenuous sports. Exercise restriction was eliminated. Previously abnormal SF-36 physical composite scores returned to normal levels that persisted for the duration of the therapy in all five symptomatic patients. Conclusions: The triheptanoin diet seems to be an effective therapy for adult-onset carnitine palmitoyltransferase II deficiency. GLOSSARY ALT = alanine aminotransferase; AST = aspartate aminotransferase; ATP = adenosine triphosphate; BHP = β-hydroxypentanoate; BKP = β-ketopentanoate; BKP-CoA = β-ketopentanoyl–coenzyme A; BUN = blood urea nitrogen; CAC = citric acid cycle; CoA = coenzyme A; CPK = creatine phosphokinase; CPT II = carnitine palmitoyltransferase II; LDL = low-density lipoprotein; MCT

  17. Antithrombin deficiency in pregnancy.

    Science.gov (United States)

    Durai, Shivani; Tan, Lay Kok; Lim, Serene

    2016-01-01

    We present a case of a 39-year-old, gravida 3 para 2, Chinese female with a history of inherited type 1 Antithrombin deficiency and multiple prior episodes of venous thromboembolism. She presented at 29+4 weeks' gestation with severe pre-eclampsia complicated by haemolysis, elevated liver enzymes and low platelet (HELLP) syndrome. She subsequently underwent an emergency caesarean section for non-reassuring fetal status, which was complicated by postpartum haemorrhage secondary to uterine atony, requiring a B-Lynch suture intraoperatively. PMID:27207982

  18. Nasal Tip Deficiency.

    Science.gov (United States)

    Cerkes, Nazim

    2016-01-01

    Nasal tip deficiency can be congenital or secondary to previous nasal surgeries. Underdeveloped medial crura usually present with underprojected tip and lack of tip definition. Weakness or malposition of lateral crura causes alar rim retraction and lateral nasal wall weakness. Structural grafting of alar cartilages strengthens the tip framework, reinforces the disrupted support mechanisms, and controls the position of the nasal tip. In secondary cases, anatomic reconstruction of the weakened or interrupted alar cartilages and reconstitution of a stable nasal tip tripod must be the goal for a predictable outcome. PMID:26616702

  19. Ensuring effective prevention of iodine-deficiency disorders

    DEFF Research Database (Denmark)

    Völzke, Henry; Caron, Philippe Jean; Dahl, Lisbeth;

    2016-01-01

    BACKGROUND: Programs initiated to prevent iodine deficiency disorders (IDD) may not remain effective due to changes in government policies, commercial factors, and human behavior that may affect the efficacy of IDD prevention programs in unpredictable directions. Monitoring and outcome studies ar...

  20. An Effective Strategy for Reliably Isolating Heritable and Cas9-Free Arabidopsis Mutants Generated by CRISPR/Cas9-Mediated Genome Editing1[OPEN

    Science.gov (United States)

    Gao, Xiuhua; Chen, Jilin; Dai, Xinhua; Zhang, Da

    2016-01-01

    Mutations generated by CRISPR/Cas9 in Arabidopsis (Arabidopsis thaliana) are often somatic and are rarely heritable. Isolation of mutations in Cas9-free Arabidopsis plants can ensure the stable transmission of the identified mutations to next generations, but the process is laborious and inefficient. Here, we present a simple visual screen for Cas9-free T2 seeds, allowing us to quickly obtain Cas9-free Arabidopsis mutants in the T2 generation. To demonstrate this in principle, we targeted two sites in the AUXIN-BINDING PROTEIN1 (ABP1) gene, whose function as a membrane-associated auxin receptor has been challenged recently. We obtained many T1 plants with detectable mutations near the target sites, but only a small fraction of T1 plants yielded Cas9-free abp1 mutations in the T2 generation. Moreover, the mutations did not segregate in Mendelian fashion in the T2 generation. However, mutations identified in the Cas9-free T2 plants were stably transmitted to the T3 generation following Mendelian genetics. To further simplify the screening procedure, we simultaneously targeted two sites in ABP1 to generate large deletions, which can be easily identified by PCR. We successfully generated two abp1 alleles that contained 1,141- and 711-bp deletions in the ABP1 gene. All of the Cas9-free abp1 alleles we generated were stable and heritable. The method described here allows for effectively isolating Cas9-free heritable CRISPR mutants in Arabidopsis. PMID:27208253

  1. Velocity Curve Studies of Spectroscopic Binary Stars V380 Cygni, V401 Cyg, V523 Cas, V373 Cas and V2388 Oph

    Indian Academy of Sciences (India)

    K. Karami; R. Mohebi

    2009-09-01

    Using measured radial velocity data of five double lined spectroscopic binary systems V380 Cygni, V401 Cyg, V523 Cas, V373 Cas and V2388 Oph, we find corresponding orbital and spectroscopic elements via the method introduced by Karami & Mohebi (2007) and Karami & Teimoorinia (2007). Our numerical results are in good agreement with those obtained by others using more traditional methods.

  2. CRISPR/Cas9-induced knockout and knock-in mutations in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Shin, Sung-Eun; Lim, Jong-Min; Koh, Hyun Gi; Kim, Eun Kyung; Kang, Nam Kyu; Jeon, Seungjib; Kwon, Sohee; Shin, Won-Sub; Lee, Bongsoo; Hwangbo, Kwon; Kim, Jungeun; Ye, Sung Hyeok; Yun, Jae-Young; Seo, Hogyun; Oh, Hee-Mock; Kim, Kyung-Jin; Kim, Jin-Soo; Jeong, Won-Joong; Chang, Yong Keun; Jeong, Byeong-Ryool

    2016-01-01

    Genome editing is crucial for genetic engineering of organisms for improved traits, particularly in microalgae due to the urgent necessity for the next generation biofuel production. The most advanced CRISPR/Cas9 system is simple, efficient and accurate in some organisms; however, it has proven extremely difficult in microalgae including the model alga Chlamydomonas. We solved this problem by delivering Cas9 ribonucleoproteins (RNPs) comprising the Cas9 protein and sgRNAs to avoid cytotoxicity and off-targeting associated with vector-driven expression of Cas9. We obtained CRISPR/Cas9-induced mutations at three loci including MAA7, CpSRP43 and ChlM, and targeted mutagenic efficiency was improved up to 100 fold compared to the first report of transgenic Cas9-induced mutagenesis. Interestingly, we found that unrelated vectors used for the selection purpose were predominantly integrated at the Cas9 cut site, indicative of NHEJ-mediated knock-in events. As expected with Cas9 RNPs, no off-targeting was found in one of the mutagenic screens. In conclusion, we improved the knockout efficiency by using Cas9 RNPs, which opens great opportunities not only for biological research but also industrial applications in Chlamydomonas and other microalgae. Findings of the NHEJ-mediated knock-in events will allow applications of the CRISPR/Cas9 system in microalgae, including "safe harboring" techniques shown in other organisms. PMID:27291619

  3. RNA-guided transcriptional regulation in planta via synthetic dCas9-based transcription factors

    KAUST Repository

    Piatek, Agnieszka

    2014-11-14

    Targeted genomic regulation is a powerful approach to accelerate trait discovery and development in agricultural biotechnology. Bacteria and archaea use clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) regulatory systems for adaptive molecular immunity against foreign nucleic acids introduced by invading phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing in many cell types and organisms. A recent study used the catalytically inactive Cas9 (dCas9) protein combined with guide-RNAs (gRNAs) as a DNA-targeting platform to modulate gene expression in bacterial, yeast, and human cells. Here, we modified this DNA-targeting platform for targeted transcriptional regulation in planta by developing chimeric dCas9-based transcriptional activators and repressors. To generate transcriptional activators, we fused the dCas9 C-terminus with the activation domains of EDLL and TAL effectors. To generate a transcriptional repressor, we fused the dCas9 C-terminus with the SRDX repression domain. Our data demonstrate that dCas9 fusion with the EDLL activation domain (dCas9:EDLL) and the TAL activation domain (dCas9:TAD), guided by gRNAs complementary to selected promoter elements, induce strong transcriptional activation on Bs3

  4. CRISPR/Cas9-induced knockout and knock-in mutations in Chlamydomonas reinhardtii

    Science.gov (United States)

    Shin, Sung-Eun; Lim, Jong-Min; Koh, Hyun Gi; Kim, Eun Kyung; Kang, Nam Kyu; Jeon, Seungjib; Kwon, Sohee; Shin, Won-Sub; Lee, Bongsoo; Hwangbo, Kwon; Kim, Jungeun; Ye, Sung Hyeok; Yun, Jae-Young; Seo, Hogyun; Oh, Hee-Mock; Kim, Kyung-Jin; Kim, Jin-Soo; Jeong, Won-Joong; Chang, Yong Keun; Jeong, Byeong-ryool

    2016-01-01

    Genome editing is crucial for genetic engineering of organisms for improved traits, particularly in microalgae due to the urgent necessity for the next generation biofuel production. The most advanced CRISPR/Cas9 system is simple, efficient and accurate in some organisms; however, it has proven extremely difficult in microalgae including the model alga Chlamydomonas. We solved this problem by delivering Cas9 ribonucleoproteins (RNPs) comprising the Cas9 protein and sgRNAs to avoid cytotoxicity and off-targeting associated with vector-driven expression of Cas9. We obtained CRISPR/Cas9-induced mutations at three loci including MAA7, CpSRP43 and ChlM, and targeted mutagenic efficiency was improved up to 100 fold compared to the first report of transgenic Cas9-induced mutagenesis. Interestingly, we found that unrelated vectors used for the selection purpose were predominantly integrated at the Cas9 cut site, indicative of NHEJ-mediated knock-in events. As expected with Cas9 RNPs, no off-targeting was found in one of the mutagenic screens. In conclusion, we improved the knockout efficiency by using Cas9 RNPs, which opens great opportunities not only for biological research but also industrial applications in Chlamydomonas and other microalgae. Findings of the NHEJ-mediated knock-in events will allow applications of the CRISPR/Cas9 system in microalgae, including “safe harboring” techniques shown in other organisms. PMID:27291619

  5. CRISPR-Cas adaptive immune systems of the sulfolobales

    DEFF Research Database (Denmark)

    Garrett, Roger Antony; Shah, Shiraz Ali; Erdmann, Susanne;

    2015-01-01

    The Sulfolobales have provided good model organisms for studying CRISPR-Cas systems of the crenarchaeal kingdom of the archaea. These organisms are infected by a wide range of exceptional archaea-specific viruses and conjugative plasmids, and their CRISPR-Cas systems generally exhibit extensive...... structural and functional diversity. They carry large and multiple CRISPR loci and often multiple copies of diverse Type I and Type III interference modules as well as more homogeneous adaptation modules. These acidothermophilic organisms have recently provided seminal insights into both the adaptation...... process, the diverse modes of interference, and their modes of regulation. The functions of the adaptation and interference modules tend to be loosely coupled and the stringency of the crRNA-DNA sequence matching during DNA interference is relatively low, in contrast to some more streamlined CRISPR...

  6. MPS/CAS Partner Group on Nanostructured Materials

    Institute of Scientific and Technical Information of China (English)

    Lu Ke; Manfred Rühle

    2004-01-01

    @@ As one of the first Partner Groups which were initiated to foster scientific exchange and interaction between the Max Pianck Society (MPS) and the Chinese Academy of Sciences (CAS), the Partner Group led by Lu Ke was established on April 1, 1999.During the past five years, the group has received substantial support from the CAS for equipment and from the MPS for personnel and travel expenses. Extensive and productive collaborations between the research staff and students of the Partner Group with several professors (departments) in the Max Planck Institute for Metals Research (MPI-MF) have led to significant advances in synthesis,mechanical properties, melting and superheating of nanostructured metals. The research is well recognized internationally and a substantial number of publications in high quality, peer-reviewed journals (including Science and Physical Review Letters) have resulted.

  7. Chromosome engineering in zygotes with CRISPR/Cas9

    Science.gov (United States)

    Boroviak, Katharina; Doe, Brendan; Banerjee, Ruby; Yang, Fengtang

    2016-01-01

    SUMMARY Deletions, duplications, and inversions of large genomic regions covering several genes are an important class of disease causing variants in humans. Modeling these structural variants in mice requires multistep processes in ES cells, which has limited their availability. Mutant mice containing small insertions, deletions, and single nucleotide polymorphisms can be reliably generated using CRISPR/Cas9 directly in mouse zygotes. Large structural variants can be generated using CRISPR/Cas9 in ES cells, but it has not been possible to generate these directly in zygotes. We now demonstrate the direct generation of deletions, duplications and inversions of up to one million base pairs by zygote injection. genesis 54:78–85, 2016. © 2016 The Authors. genesis Published by Wiley Periodicals, Inc. PMID:26742453

  8. Towards Developing a CAS E-Journal Archiving System

    Institute of Scientific and Technical Information of China (English)

    ZHANG ZHIXIONG; WU ZHENXIN; ZHAO QI; LIU JIANHUA; GUO WENLI; XU JIAN

    2008-01-01

    As a part of NSTL proposed long-term preservation of digital information network,the National Science Library of CAS(NSLC)constructed a long-term preservation system of electronic journals called CAS E-Journal Archiving System,which was based on OAIS and adapted Fedora as the fundament system.The system chose a certain amount of electronic journals as pilot resources.Through three major system functional components comprising ingest,preservation management and data dissemination,the system took the long-term preservation of digital objects function into practice.The system is designed to be open and scalable.It realized digital objects management and lifecycle management.In this paper we describe the design of the system,and the issues we have learnt about long-term preservation.

  9. Maladie de Haglund: à propos de trois cas

    OpenAIRE

    Adigo, Amégninou Mawuko Yao; Gnakadja, Néille Gbèssi; Dellanh, Yaovi Yanick; Adambounou, Kokou; Djagnikpo, Oni; Agoda-Kousséma, Lama Kegdigoma; Adoko, Abikou Léon; Adjénou, Komlanvi Victor

    2015-01-01

    La maladie de Haglund est une pathologie relativement sous évaluée. Elle est liée à un conflit calcanéo-achilléen. Nous rapportons les cas de patients âgés de 40, 42 et 37 ans, révélés par des œdèmes douloureux de la cheville. Le diagnostic a été confirmé à la radiographie standard de la cheville en charge et à l’échographie chez tous les patients. Un seul patient avait bénéficié d'une exploration IRM. Le traitement, initialement médical dans tous les cas, s'est soldé par une chirurgie de rés...

  10. Next Generation Prokaryotic Engineering: The CRISPR-Cas Toolkit.

    Science.gov (United States)

    Mougiakos, Ioannis; Bosma, Elleke F; de Vos, Willem M; van Kranenburg, Richard; van der Oost, John

    2016-07-01

    The increasing demand for environmentally friendly production processes of green chemicals and fuels has stimulated research in microbial metabolic engineering. CRISPR-Cas-based tools for genome editing and expression control have enabled fast, easy, and accurate strain development for established production platform organisms, such as Escherichia coli and Saccharomyces cerevisiae. However, the growing interest in alternative production hosts, for which genome editing options are generally limited, requires further developing such engineering tools. In this review, we discuss established and emerging CRISPR-Cas-based tools for genome editing and transcription control of model and non-model prokaryotes, and we analyse the possibilities for further improvement and expansion of these tools for next generation prokaryotic engineering. PMID:26944793

  11. Application of CRISPR/Cas9 Technology to HBV

    Directory of Open Access Journals (Sweden)

    Guigao Lin

    2015-11-01

    Full Text Available More than 240 million people around the world are chronically infected with hepatitis B virus (HBV. Nucleos(tide analogs and interferon are the only two families of drugs to treat HBV currently. However, none of these anti-virals directly target the stable nuclear covalently closed circular DNA (cccDNA, which acts as a transcription template for viral mRNA and pre-genomic RNA synthesis and secures virus persistence. Thus, the fact that only a small number of patients treated achieve sustained viral response (SVR or cure, highlights the need for new therapies against HBV. The clustered regularly interspaced short palindromic repeats (CRISPR/Cas9 gene editing system can specifically target the conserved regions of the HBV genome. This results in robust viral suppression and provides a promising tool for eradicating the virus. In this review, we discuss the function and application of the CRISPR/Cas9 system as a novel therapy for HBV.

  12. Vision Guided Modeling and Simulation for CAS Rover

    Directory of Open Access Journals (Sweden)

    Hongwei Gao

    2014-01-01

    Full Text Available With the potential application of planetary exploration, the issues on vision-guided modeling and movement simulation technologies for the planetary rover of Chinese Academy of Sciences (CAS are investigated. Based on stereo vision and the virtual reality, a simulation platform is developed. The system working in an off-line teaching way can be used to ensure the safety of the rover. Firstly, a detail description of the CAS rover mechanism structure and kinematics model is given. Secondly, the implementation process of simulation platform based on OpenGL has been introduced. Finally, movement simulation on synthesis terrain and real terrain got by stereo vision are executed, and the simulation results show the validity of the whole scheme, and the scheme has the availability for future exploration operation.

  13. Glucose-6-phosphatase deficiency

    Directory of Open Access Journals (Sweden)

    Labrune Philippe

    2011-05-01

    Full Text Available Abstract Glucose-6-phosphatase deficiency (G6P deficiency, or glycogen storage disease type I (GSDI, is a group of inherited metabolic diseases, including types Ia and Ib, characterized by poor tolerance to fasting, growth retardation and hepatomegaly resulting from accumulation of glycogen and fat in the liver. Prevalence is unknown and annual incidence is around 1/100,000 births. GSDIa is the more frequent type, representing about 80% of GSDI patients. The disease commonly manifests, between the ages of 3 to 4 months by symptoms of hypoglycemia (tremors, seizures, cyanosis, apnea. Patients have poor tolerance to fasting, marked hepatomegaly, growth retardation (small stature and delayed puberty, generally improved by an appropriate diet, osteopenia and sometimes osteoporosis, full-cheeked round face, enlarged kydneys and platelet dysfunctions leading to frequent epistaxis. In addition, in GSDIb, neutropenia and neutrophil dysfunction are responsible for tendency towards infections, relapsing aphtous gingivostomatitis, and inflammatory bowel disease. Late complications are hepatic (adenomas with rare but possible transformation into hepatocarcinoma and renal (glomerular hyperfiltration leading to proteinuria and sometimes to renal insufficiency. GSDI is caused by a dysfunction in the G6P system, a key step in the regulation of glycemia. The deficit concerns the catalytic subunit G6P-alpha (type Ia which is restricted to expression in the liver, kidney and intestine, or the ubiquitously expressed G6P transporter (type Ib. Mutations in the genes G6PC (17q21 and SLC37A4 (11q23 respectively cause GSDIa and Ib. Many mutations have been identified in both genes,. Transmission is autosomal recessive. Diagnosis is based on clinical presentation, on abnormal basal values and absence of hyperglycemic response to glucagon. It can be confirmed by demonstrating a deficient activity of a G6P system component in a liver biopsy. To date, the diagnosis is most

  14. Precision Targeted Mutagenesis via Cas9 Paired Nickases in Rice

    OpenAIRE

    Mikami, Masafumi; Toki, Seiichi; Endo, Masaki

    2016-01-01

    Recent reports of CRISPR- (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) mediated heritable mutagenesis in plants highlight the need for accuracy of the mutagenesis directed by this system. Off-target mutations are an important issue when considering functional gene analysis, as well as the molecular breeding of crop plants with large genome size, i.e. with many duplicated genes, and where the whole-genome sequence is still lacking. In mammals, ...

  15. CRISPR-Cas: Development and applications for mammalian genome editing

    OpenAIRE

    Ran, Fei Ann

    2014-01-01

    The ability to introduce targeted modifications into genomes and engineer model organisms holds enormous promise for biomedical and technological applications, and has driven the development of tools such as zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs). To facilitate genome engineering in mammalian cells, we have engineered the CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 programmable nuclease systems from Streptococcus p...

  16. Burstein Moss effect in nanocrystalline CaS : Ce

    Indian Academy of Sciences (India)

    Geeta Sharma; Puja Chawla; S P Lochab; Nafa Singh

    2011-07-01

    The nanocrystalline CaS : Ce nanophosphors are synthesized by wet chemical co-precipitation method. The particles possess an average size of 10 nm as calculated using Debye–Scherrer formula. The particle size and the crystalline nature of the formed nanoparticles are confirmed by TEM micrograph. The optical studies are carried out using UV–Vis absorption spectroscopy. The absorption edge is found to show blue shift with increasing cerium concentration. The shift may be attributed to Burstein Moss effect.

  17. Mouse Genome Editing using CRISPR/Cas System

    OpenAIRE

    Harms, Donald W.; Quadros, Rolen M; Seruggia, Davide; Ohtsuka, Masato; Takahashi, Gou; Montoliu, Lluis; Gurumurthy, Channabasavaiah B

    2014-01-01

    The availability of techniques to create desired genetic mutations has enabled the laboratory mouse as an extensively used model organism in biomedical research including human genetics. A new addition to this existing technical repertoire is the CRISPR/Cas system. Specifically, this system allows editing of the mouse genome much faster than the previously used techniques and more importantly multiple mutations can be created in a single experiment. Here we provide protocols for preparation o...

  18. How specific is CRISPR/Cas9 really?

    OpenAIRE

    O'Geen, H; Yu, AS; Segal, DJ

    2015-01-01

    © 2015 The Authors. The specificity of RNA-guided nucleases has gathered considerable interest as they become broadly applied to basic research and therapeutic development. Reports of the simple generation of animal models and genome engineering of cells raised questions about targeting precision. Conflicting early reports led the field to believe that CRISPR/Cas9 system was promiscuous, leading to a variety of strategies for improving specificity and increasingly sensitive methods to detect ...

  19. Meeting report: GARNet/OpenPlant CRISPR-Cas workshop

    OpenAIRE

    Parry, Geraint; Patron, Nicola; Bastow, Ruth; Matthewman, Colette

    2016-01-01

    Targeted genome engineering has been described as a “game-changing technology” for fields as diverse as human genetics and plant biotechnology. One technique used for precise gene editing utilises the CRISPR-Cas system and is an effective method for genetic engineering in a wide variety of plants. However, many researchers remain unaware of both the technical challenges that emerge when using this technique or of its potential benefits. Therefore in September 2015, GARNet and OpenPlant organi...

  20. Targeted mutagenesis using CRISPR/Cas system in medaka

    OpenAIRE

    Satoshi Ansai; Masato Kinoshita

    2014-01-01

    ABSTRACT Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) system-based RNA-guided endonuclease (RGEN) has recently emerged as a simple and efficient tool for targeted genome editing. In this study, we showed successful targeted mutagenesis using RGENs in medaka, Oryzias latipes. Somatic and heritable mutations were induced with high efficiency at the targeted genomic sequence on the DJ-1 gene in embryos that had been injected with the single guide RNA...

  1. The CRISPR-Cas immune system : Biology, mechanisms and applications

    OpenAIRE

    Rath, Devashish; Amlinger, Lina; Rath, Archana; Lundgren, Magnus

    2015-01-01

    Viruses are a common threat to cellular life, not the least to bacteria and archaea who constitute the majority of life on Earth. Consequently, a variety of mechanisms to resist virus infection has evolved. A recent discovery is the adaptive immune system in prokaryotes, a type of system previously thought to be present only in vertebrates. The system, called CRISPR-Cas, provide sequence-specific adaptive immunity and fundamentally affect our understanding of virus host interaction. CRISPR-ba...

  2. CRISPR RNA biogenesis by a Cas6 nuclease

    OpenAIRE

    Sokolowski, Richard D.

    2015-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPRs) and associated (Cas) proteins form the basis of a prokaryotic adaptive immune system. Acquired sections of viral DNA are stored within the host genome as ‘spacers’ flanked by ‘repeat’ sequences. The CRISPR arrays are transcribed and processed to release mature CRISPR RNAs (crRNAs) – containing a single, intact spacer sequence – that are used by effector complexes to base-pair with matching hostile genetic elements and silence...

  3. Exploiting CRISPR/Cas: Interference Mechanisms and Applications

    OpenAIRE

    André Plagens; Lennart Randau; Hagen Richter

    2013-01-01

    The discovery of biological concepts can often provide a framework for the development of novel molecular tools, which can help us to further understand and manipulate life. One recent example is the elucidation of the prokaryotic adaptive immune system, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) that protects bacteria and archaea against viruses or conjugative plasmids. The immunity is based on small RNA molecules that are incorporated into ver...

  4. CAS Will Attend the 11th IFAN International Conference

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    @@ IFAN (The International Federation of Standards Users), together with NKN (Normalisatie Kringen Nederland) and NEN (Nederlands Normalisatie-institute), will organize the 11th IFAN International Conference on the subject of "Standardization-Awareness-Compliance"in Amsterdam, the Netherlands on 11th-12th November 2004. Leaders from industry, government, standards user organizations and standards developing organizations will attend the conference. CAS (China Association for Standardization) is going to attend the conference and have an address as an invited speaker.

  5. SSFinder: High Throughput CRISPR-Cas Target Sites Prediction Tool

    OpenAIRE

    Santosh Kumar Upadhyay; Shailesh Sharma

    2014-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein (Cas) system facilitates targeted genome editing in organisms. Despite high demand of this system, finding a reliable tool for the determination of specific target sites in large genomic data remained challenging. Here, we report SSFinder, a python script to perform high throughput detection of specific target sites in large nucleotide datasets. The SSFinder is a user-friendly tool, compatible wit...

  6. Cognitive Assessment System (CAS): Psychometric studies with Portuguese children.

    OpenAIRE

    Rosário, A.; Candeias, A.A.; Roazzi, A.

    2013-01-01

    The Cognitive Assessment System (CAS) is a new measure of cognitive abilities based on the Planning, Attention, Simultaneous and Successive (PASS) Theory. This theory is derived from research in neuropsychological and cognitive Psychology with particular emphasis on the work of Luria (1973). According to Naglieri (1999) and Naglieri and Das (1997), the PASS cognitive processes are the basic building blocks of human intellectual functioning. Planning processes provide cognitive control, utiliz...

  7. CAS Academic Divisions: Advisory Services and International Cooperation

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    @@ Since its establishment more than 50 years ago,the Academic Divisions of the Chinese Academy of Sciences (CASAD) has given much important advice concerning national socioeconomic development,security and S&T progress,by giving full play to China's outstanding scientists and pooling the collective wisdom of CAS Members.These suggestions have laid a solid S&T foundation for government policy-making on major issues.

  8. Iron Deficiency and Bariatric Surgery

    OpenAIRE

    Ignacio Jáuregui-Lobera

    2013-01-01

    It is estimated that the prevalence of anaemia in patients scheduled for bariatric surgery is higher than in the general population and the prevalence of iron deficiencies (with or without anaemia) may be higher as well. After surgery, iron deficiencies and anaemia may occur in a higher percentage of patients, mainly as a consequence of nutrient deficiencies. In addition, perioperative anaemia has been related with increased postoperative morbidity and mortality and poorer quality of life aft...

  9. Iatrogenic limbal stem cell deficiency.

    OpenAIRE

    Holland, E J; Schwartz, G S

    1997-01-01

    PURPOSE: To describe a group of patients with limbal stem cell (SC) deficiency without prior diagnosis of a specific disease entity known to be causative of SC deficiency. METHODS: We performed a retrospective review of the records of all patients with ocular surface disease presenting to the University of Minnesota between 1987 and 1996. Patients were categorized according to etiology of limbal deficiency. Patients who did not have a specific diagnosis previously described as being causative...

  10. Targeted mutagenesis using CRISPR/Cas system in medaka

    Directory of Open Access Journals (Sweden)

    Satoshi Ansai

    2014-04-01

    Full Text Available Clustered regularly interspaced short palindromic repeats (CRISPR/CRISPR-associated (Cas system-based RNA-guided endonuclease (RGEN has recently emerged as a simple and efficient tool for targeted genome editing. In this study, we showed successful targeted mutagenesis using RGENs in medaka, Oryzias latipes. Somatic and heritable mutations were induced with high efficiency at the targeted genomic sequence on the DJ-1 gene in embryos that had been injected with the single guide RNA (sgRNA transcribed by a T7 promoter and capped RNA encoding a Cas9 nuclease. The sgRNAs that were designed for the target genomic sequences without the 5′ end of GG required by the T7 promoter induced the targeted mutations. This suggests that the RGEN can target any sequence adjacent to an NGG protospacer adjacent motif (PAM sequence, which occurs once every 8 bp. The off-target alterations at 2 genomic loci harboring double mismatches in the 18-bp targeting sequences were induced in the RGEN-injected embryos. However, we also found that the off-target effects could be reduced by lower dosages of sgRNA. Taken together, our results suggest that CRISPR/Cas-mediated RGENs may be an efficient and flexible tool for genome editing in medaka.

  11. CRISPR/Cas-mediated targeted mutagenesis in Daphnia magna.

    Directory of Open Access Journals (Sweden)

    Takashi Nakanishi

    Full Text Available The water flea Daphnia magna has been used as an animal model in ecology, evolution, and environmental sciences. Thanks to the recent progress in Daphnia genomics, genetic information such as the draft genome sequence and expressed sequence tags (ESTs is now available. To investigate the relationship between phenotypes and the available genetic information about Daphnia, some gene manipulation methods have been developed. However, a technique to induce targeted mutagenesis into Daphnia genome remains elusive. To overcome this problem, we focused on an emerging genome editing technique mediated by the clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas system to introduce genomic mutations. In this study, we targeted a functionally conserved regulator of eye development, the eyeless gene in D. magna. When we injected Cas9 mRNAs and eyeless-targeting guide RNAs into eggs, 18-47% of the survived juveniles exhibited abnormal eye morphology. After maturation, up to 8.2% of the adults produced progenies with deformed eyes, which carried mutations in the eyeless loci. These results showed that CRISPR/Cas system could introduce heritable mutations into the endogenous eyeless gene in D. magna. This is the first report of a targeted gene knockout technique in Daphnia and will be useful in uncovering Daphnia gene functions.

  12. The triple-mode pulsating variable V823 Cas

    CERN Document Server

    Jurcsik, J; Varadi, M; Henden, A; Hurta, Z; Lakatos, B; Posztobanyi, K; Klagyivik, P; Sodor, A; Hurta, Zs.

    2005-01-01

    Based on extended multicolour CCD photometry of the triple-mode radial pulsator V823 Cas we studied the properties of the coupling frequencies invoked by nonlinear processes. Our results support that a resonance connection as suggested by Antonello & Aikawa (1998) affects the mode coupling behaviour. The P1/P0 period ratio of V823 Cas has an "out of range" value if compared with the period ratios of the known double mode pulsators, while the P2/P1 period ratio is normal. The periods and period ratios cannot be consistently interpret without conflict with pulsation and/or evolution models. We attempt to interpret this failure by the suggestion that at present, the periods of V823 Cas are in a transient, resonance affected state, thus do not reflect the true parameters of the object. The anomalous period change behaviour of the fundamental and second overtone modes supports this idea. We have also raised the possibility that a f0 + f2 = 2f1 resonance may act in triple mode pulsators.

  13. Zinc deficiency and eating disorders.

    Science.gov (United States)

    Humphries, L; Vivian, B; Stuart, M; McClain, C J

    1989-12-01

    Decreased food intake, a cyclic pattern of eating, and weight loss are major manifestations of zinc deficiency. In this study, zinc status was evaluated in 62 patients with bulimia and 24 patients with anorexia nervosa. Forty percent of patients with bulimia and 54% of those with anorexia nervosa had biochemical evidence of zinc deficiency. The authors suggest that for a variety of reasons, such as lower dietary intake of zinc, impaired zinc absorption, vomiting, diarrhea, and binging on low-zinc foods, patients with eating disorders may develop zinc deficiency. This acquired zinc deficiency could then add to the chronicity of altered eating behavior in those patients. PMID:2600063

  14. Primary Carnitine deficiency in the Faroe Islands

    DEFF Research Database (Denmark)

    Rasmussen, Jan; Køber, Lars; Lund, Allan M; Nielsen, Olav W

    2014-01-01

    BACKGROUND: Carnitine deficiency can cause cardiomyopathy and cardiac arrhythmia. The prevalence in the Faroe Islands is the highest reported in the world (1:300). A nationwide screening program identified 76 Faroese adult patients (15-80 years) with Primary Carnitine Deficiency (PCD). We describe...... prior and current health status and symptoms in these patients, especially focusing on cardiac characteristics. METHODS: Upon identification, patients were immediately admitted for physical examination, ECG, blood tests and initiation of L-carnitine supplementation. Medical records were reviewed and...... patients were interviewed. Echocardiography and blood tests were performed in 35 patients before and after L-carnitine supplementation. RESULTS: All patients were either asymptomatic or had minor symptoms when diagnosed. Echocardiography including LVEF, global longitudinal strain and dimensions were normal...

  15. Generation of an Oocyte-Specific Cas9 Transgenic Mouse for Genome Editing.

    Directory of Open Access Journals (Sweden)

    Linlin Zhang

    Full Text Available The CRISPR/Cas9 system has been developed as an easy-handle and multiplexable approach for engineering eukaryotic genomes by zygote microinjection of Cas9 and sgRNA, while preparing Cas9 for microinjection is laborious and introducing inconsistency into the experiment. Here, we describe a modified strategy for gene targeting through using oocyte-specific Cas9 transgenic mouse. With this mouse line, we successfully achieve precise gene targeting by injection of sgRNAs only into one-cell-stage embryos. Through comprehensive analysis, we also show allele complexity and off-target mutagenesis induced by this strategy is obviously lower than Cas9 mRNA/sgRNA injection. Thus, injection of sgRNAs into oocyte-specific Cas9 transgenic mouse embryo provides a convenient, efficient and reliable approach for mouse genome editing.

  16. CRISPRscan: designing highly efficient sgRNAs for CRISPR/Cas9 targeting in vivo

    OpenAIRE

    Moreno-Mateos, Miguel A.; Vejnar, Charles E.; Beaudoin, Jean-Denis; Fernandez, Juan P.; Mis, Emily K.; Khokha, Mustafa K.; Giraldez, Antonio J.

    2015-01-01

    CRISPR/Cas9 technology provides a powerful system for genome engineering. However, variable activity across different single guide RNAs (sgRNAs) remains a significant limitation. We have analyzed the molecular features that influence sgRNA stability, activity and loading into Cas9 in vivo. We observe that guanine enrichment and adenine depletion increase sgRNA stability and activity, while loading, nucleosome positioning and Cas9 off-target binding are not major determinants. We additionally ...

  17. Efficient Gene Knockout in Goats Using CRISPR/Cas9 System

    OpenAIRE

    Ni, Wei; Qiao, Jun; Hu, Shengwei; Zhao, Xinxia; Regouski, Misha; Yang, Min; Polejaeva, Irina A.; Chen, Chuangfu

    2014-01-01

    The CRISPR/Cas9 system has been adapted as an efficient genome editing tool in laboratory animals such as mice, rats, zebrafish and pigs. Here, we report that CRISPR/Cas9 mediated approach can efficiently induce monoallelic and biallelic gene knockout in goat primary fibroblasts. Four genes were disrupted simultaneously in goat fibroblasts by CRISPR/Cas9-mediated genome editing. The single-gene knockout fibroblasts were successfully used for somatic cell nuclear transfer (SCNT) and resulted i...

  18. Efficient CRISPR/Cas9 Plasmids for Rapid and Versatile Genome Editing in Drosophila

    OpenAIRE

    Gokcezade, Joseph; Sienski, Grzegorz; Duchek, Peter

    2014-01-01

    The CRISPR-associated RNA-guided nuclease Cas9 has emerged as a powerful tool for genome engineering in a variety of organisms. To achieve efficient gene targeting rates in Drosophila, current approaches require either injection of in vitro transcribed RNAs or injection into transgenic Cas9-expressing embryos. We report a simple and versatile alternative method for CRISPR-mediated genome editing in Drosophila using bicistronic Cas9/sgRNA expression vectors. Gene targeting with this single-pla...

  19. PAM multiplicity marks genomic target sites as inhibitory to CRISPR-Cas9 editing

    OpenAIRE

    Malina, Abba; Cameron, Christopher J. F.; Robert, Francis; Blanchette, Mathieu; Dostie, Josée; Pelletier, Jerry

    2015-01-01

    In CRISPR-Cas9 genome editing, the underlying principles for selecting guide RNA (gRNA) sequences that would ensure for efficient target site modification remain poorly understood. Here we show that target sites harbouring multiple protospacer adjacent motifs (PAMs) are refractory to Cas9-mediated repair in situ. Thus we refine which substrates should be avoided in gRNA design, implicating PAM density as a novel sequence-specific feature that inhibits in vivo Cas9-driven DNA modification.

  20. CRISPR/Cas9-mediated genome engineering and the promise of designer flies on demand

    OpenAIRE

    Gratz, Scott J.; Wildonger, Jill; Harrison, Melissa M.; O'Connor-Giles, Kate M.

    2013-01-01

    The CRISPR/Cas9 system has attracted significant attention for its potential to transform genome engineering. We and others have recently shown that the RNA-guided Cas9 nuclease can be employed to engineer the Drosophila genome, and that these modifications are efficiently transmitted through the germline. A single targeting RNA can guide Cas9 to a specific genomic sequence where it induces double-strand breaks that, when imperfectly repaired, yield mutations. We have also demonstrated that 2...

  1. Prediction and Validation of Native and Engineered Cas9 Guide Sequences.

    Science.gov (United States)

    Briner, Alexandra E; Henriksen, Emily D; Barrangou, Rodolphe

    2016-01-01

    Cas9-based technologies rely on native elements of Type II CRISPR-Cas bacterial immune systems, including the trans-activating CRISPR RNA (tracrRNA), CRISPR RNA (crRNA), Cas9 protein, and protospacer-adjacent motif (PAM). The tracrRNA and crRNA form an RNA duplex that guides the Cas9 endonuclease to complementary nucleic acid sequences. Mechanistically, Cas9 initiates interactions by binding to the target PAM sequence and interrogating the target DNA in a 3'-to-5' manner. Complementarity between the guide RNA and the target DNA is key. In natural systems, precise cleavage occurs when the target DNA sequence contains a PAM flanking a sequence homologous to the crRNA spacer sequence. Currently, the majority of commercial Cas9-based genome-editing tools are derived from the Type II CRISPR-Cas system of Streptococcus pyogenes However, a diverse set of Type II CRISPR-Cas systems exist in nature that are potentially valuable for genome engineering applications. Exploitation of these systems requires prediction and validation of both native and engineered dual and single guide RNAs to drive Cas9 functionality. Here, we discuss how to identify the elements of these immune systems to develop next-generation Cas9-based genome-editing tools. We first discuss how to predict tracrRNA sequences and suggest a method for designing single guide RNAs containing only critical structural modules. We then outline how to predict the PAM sequence, which is crucial for determining potential targets for Cas9. Finally, validation of the system elements through transcriptome analysis and interference assays is essential for developing next-generation Cas9-based genome-editing tools. PMID:27371591

  2. RNA-guided Transcriptional Regulation in Plants via dCas9 Chimeric Proteins

    KAUST Repository

    Baazim, Hatoon

    2014-05-01

    Developing targeted genome regulation approaches holds much promise for accelerating trait discovery and development in agricultural biotechnology. Clustered Regularly Interspaced Palindromic Repeats (CRISPRs)/CRISPR associated (Cas) system provides bacteria and archaea with an adaptive molecular immunity mechanism against invading nucleic acids through phages and conjugative plasmids. The type II CRISPR/Cas system has been adapted for genome editing purposes across a variety of cell types and organisms. Recently, the catalytically inactive Cas9 (dCas9) protein combined with guide RNAs (gRNAs) were used as a DNA-targeting platform to modulate the expression patterns in bacterial, yeast and human cells. Here, we employed this DNA-targeting system for targeted transcriptional regulation in planta by developing chimeric dCas9-based activators and repressors. For example, we fused to the C-terminus of dCas9 with the activation domains of EDLL and TAL effectors, respectively, to generate transcriptional activators, and the SRDX repression domain to generate transcriptional repressor. Our data demonstrate that the dCas9:EDLL and dCas9:TAD activators, guided by gRNAs complementary to promoter elements, induce strong transcriptional activation on episomal targets in plant cells. Moreover, our data suggest that the dCas9:SRDX repressor and the dCas9:EDLL and dCas9:TAD activators are capable of markedly repressing or activating, respectively, the transcription of an endogenous genomic target. Our data indicate that the CRISPR/dCas9:TFs DNA targeting system can be used in plants as a functional genomic tool and for biotechnological applications.

  3. Efficient Production of Gene-Modified Mice using Staphylococcus aureus Cas9

    Science.gov (United States)

    Zhang, Xiya; Liang, Puping; Ding, Chenhui; Zhang, Zhen; Zhou, Jianwen; Xie, Xiaowei; Huang, Rui; Sun, Ying; Sun, Hongwei; Zhang, Jinran; Xu, Yanwen; Songyang, Zhou; Huang, Junjiu

    2016-01-01

    The CRISPR/Cas system is an efficient genome-editing tool to modify genes in mouse zygotes. However, only the Streptococcus pyogenes Cas9 (SpCas9) has been systematically tested for generating gene-modified mice. The protospacer adjacent motif (PAM, 5′-NGG-3′) recognized by SpCas9 limits the number of potential target sites for this system. Staphylococcus aureus Cas9 (SaCas9), with its smaller size and unique PAM (5′-NNGRRT-3′) preferences, presents an alternative for genome editing in zygotes. Here, we showed that SaCas9 could efficiently and specifically edit the X-linked gene Slx2 and the autosomal gene Zp1 in mouse zygotes. SaCas9-mediated disruption of the tyrosinase (Tyr) gene led to C57BL/6J mice with mosaic coat color. Furthermore, multiplex targeting proved efficient multiple genes disruption when we co-injected gRNAs targeting Slx2, Zp1, and Tyr together with SaCas9 mRNA. We were also able to insert a Flag tag at the C-terminus of histone H1c, when a Flag-encoding single-stranded DNA oligo was co-introduced into mouse zygotes with SaCas9 mRNA and the gRNA. These results indicate that SaCas9 can specifically cleave the target gene locus, leading to successful gene knock-out and precise knock-in in mouse zygotes, and highlight the potential of using SaCas9 for genome editing in preimplantation embryos and producing gene-modified animal models. PMID:27586692

  4. CRISPR/Cas9-Mediated Immunity to Geminiviruses: Differential Interference and Evasion

    OpenAIRE

    Zahir Ali; Shakila Ali; Manal Tashkandi; Syed Shan-e-Ali Zaidi; Mahfouz, Magdy M.

    2016-01-01

    The CRISPR/Cas9 system has recently been used to confer molecular immunity against several eukaryotic viruses, including plant DNA geminiviruses. Here, we provide a detailed analysis of the efficiencies of targeting different coding and non-coding sequences in the genomes of multiple geminiviruses. Moreover, we analyze the ability of geminiviruses to evade the CRISPR/Cas9 machinery. Our results demonstrate that the CRISPR/Cas9 machinery can efficiently target coding and non-coding sequences a...

  5. Targeted Germline Modifications in Rats using CRISPR/Cas9 and Spermatogonial Stem Cells

    OpenAIRE

    Karen M. Chapman; Gerardo A. Medrano; Priscilla Jaichander; Jaideep Chaudhary; Alexandra E. Waits; Marcelo A. Nobrega; James M. Hotaling; Carole Ober; F. Kent Hamra

    2015-01-01

    Organisms with targeted genomic modifications are efficiently produced by gene editing in embryos using CRISPR/Cas9 RNA-guided DNA endonuclease. Here, to facilitate germline editing in rats, we used CRISPR/Cas9 to catalyze targeted genomic mutations in rat spermatogonial stem cell cultures. CRISPR/Cas9-modified spermatogonia regenerated spermatogenesis and displayed long-term sperm-forming potential following transplantation into rat testes. Targeted germline mutations in Epsti1 and Erbb3 wer...

  6. CRISPR/Cas9-induced knockout and knock-in mutations in Chlamydomonas reinhardtii

    OpenAIRE

    Sung-Eun Shin; Jong-Min Lim; Hyun Gi Koh; Eun Kyung Kim; Nam Kyu Kang; Seungjib Jeon; Sohee Kwon; Won-Sub Shin; Bongsoo Lee; Kwon Hwangbo; Jungeun Kim; Sung Hyeok Ye; Jae-Young Yun; Hogyun Seo; Hee-Mock Oh

    2016-01-01

    Genome editing is crucial for genetic engineering of organisms for improved traits, particularly in microalgae due to the urgent necessity for the next generation biofuel production. The most advanced CRISPR/Cas9 system is simple, efficient and accurate in some organisms; however, it has proven extremely difficult in microalgae including the model alga Chlamydomonas. We solved this problem by delivering Cas9 ribonucleoproteins (RNPs) comprising the Cas9 protein and sgRNAs to avoid cytotoxicit...

  7. Efficient genome editing in zebrafish using a CRISPR-Cas system

    OpenAIRE

    Hwang, Woong Y.; Fu, Yanfang; Reyon, Deepak; Maeder, Morgan L.; Tsai, Shengdar Q.; Sander, Jeffry D.; Peterson, Randall T.; Yeh, J.-R. Joanna; Joung, J Keith

    2013-01-01

    In bacteria, foreign nucleic acids are silenced by clustered, regularly interspaced, short palindromic repeats (CRISPR)--CRISPR-associated (Cas) systems. Bacterial type II CRISPR systems have been adapted to create guide RNAs that direct site-specific DNA cleavage by the Cas9 endonuclease in cultured cells. Here we show that the CRISPR-Cas system functions in vivo to induce targeted genetic modifications in zebrafish embryos with efficiencies similar to those obtained using zinc finger nuclea...

  8. A new age in functional genomics using CRISPR/Cas9 in arrayed library screening

    OpenAIRE

    Agrotis, Alexander; Ketteler, Robin

    2015-01-01

    CRISPR technology has rapidly changed the face of biological research, such that precise genome editing has now become routine for many labs within several years of its initial development. What makes CRISPR/Cas9 so revolutionary is the ability to target a protein (Cas9) to an exact genomic locus, through designing a specific short complementary nucleotide sequence, that together with a common scaffold sequence, constitute the guide RNA bridging the protein and the DNA. Wild-type Cas9 cleaves...

  9. Gene targeting using the Agrobacterium tumefaciens-mediated CRISPR-Cas system in rice

    OpenAIRE

    Xu, Rongfang; Li, Hao; Qin, Ruiying; WANG, LU; LI Li; Wei, Pengcheng; Yang, Jianbo

    2014-01-01

    Background The type II clustered, regularly interspaced, short palindromic repeat (CRISPR)/ CRISPR-associated protein 9 (Cas9) system is a novel molecular tool for site-specific genome modification. The CRISPR-Cas9 system was recently introduced into plants by transient or stable transformation. Findings Here, we report gene targeting in rice via the Agrobacterium tumefaciens-mediated CRISPR-Cas9 system. Three 20-nt CRISPR RNAs were designed to pair with diverse sites followed by the protospa...

  10. Evolutionary Dynamics of the Prokaryotic Adaptive Immunity System CRISPR-Cas in an Explicit Ecological Context

    OpenAIRE

    Iranzo, Jaime; Lobkovsky, Alexander E; Wolf, Yuri I; Koonin, Eugene V

    2013-01-01

    A stochastic, agent-based mathematical model of the coevolution of the archaeal and bacterial adaptive immunity system, CRISPR-Cas, and lytic viruses shows that CRISPR-Cas immunity can stabilize the virus-host coexistence rather than leading to the extinction of the virus. In the model, CRISPR-Cas immunity does not specifically promote viral diversity, presumably because the selection pressure on each single proto-spacer is too weak. However, the overall virus diversity in the presence of CRI...

  11. The Neisseria meningitidis CRISPR-Cas9 System Enables Specific Genome Editing in Mammalian Cells

    OpenAIRE

    Lee, Ciaran M; Cradick, Thomas J.; Bao, Gang

    2016-01-01

    The clustered regularly-interspaced short palindromic repeats (CRISPR)—CRISPR-associated (Cas) system from Streptococcus pyogenes (Spy) has been successfully adapted for RNA-guided genome editing in a wide range of organisms. However, numerous reports have indicated that Spy CRISPR-Cas9 systems may have significant off-target cleavage of genomic DNA sequences differing from the intended on-target site. Here, we report the performance of the Neisseria meningitidis (Nme) CRISPR-Cas9 system that...

  12. Evolution of CRISPR RNA recognition and processing by Cas6 endonucleases

    OpenAIRE

    Niewoehner, Ole; Jinek, Martin; Doudna, Jennifer A.

    2014-01-01

    In many bacteria and archaea, small RNAs derived from clustered regularly interspaced short palindromic repeats (CRISPRs) associate with CRISPR-associated (Cas) proteins to target foreign DNA for destruction. In Type I and III CRISPR/Cas systems, the Cas6 family of endoribonucleases generates functional CRISPR-derived RNAs by site-specific cleavage of repeat sequences in precursor transcripts. CRISPR repeats differ widely in both sequence and structure, with varying propensity to form hairpin...

  13. Mathematical Modeling of CRISPR-CAS system effects on biofilm formation

    OpenAIRE

    Ali, Qasim; Wahl, Lindi

    2016-01-01

    Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), linked with CRISPR associated (CAS) genes, play a profound role in the interactions between phage and their bacterial hosts. It is now well understood that CRISPR-CAS systems can confer adaptive immunity against bacteriophage infections. However, the possibility of failure of CRISPR immunity may lead to a productive infection by the phage (cell lysis) or lysogeny. Recently, CRISPR-CAS genes have been implicated in changes to ...

  14. Application Progress of CRISPR/Cas9 System for Gene Editing in Tumor Research

    OpenAIRE

    Liu, Chao; Li, Zhiwei; Zhang, Yanqiao

    2015-01-01

    TCRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeat/CRISPR-associated nuclease 9) gene editing system is a new type of gene editing technology developed based on the immune mechanism of archaea resisting the invasion of exogenous nucleic acid. Compared with traditional gene editing system, CRISPR/Cas9 system is more efficient, easier operating, and less cytotoxic. Currently, CRISPR/Cas9 gene editing technology has been applied to many aspects of cancer research, including r...

  15. Cleft palate and gonadotrophin deficiency.

    OpenAIRE

    Gillis, P H; Peeters, R.

    1984-01-01

    A boy who had previously had a cleft lip and palate repaired and bilateral orchiopexies presented at 16 years of age with delayed puberty. Isolated gonadotrophin deficiency and testicular hyporesponsiveness to human chorionic gonadotrophin were found. The possibility of bilateral cryptorchidism due to gonadotrophin deficiency should be considered in boys with either cleft lip or palate, or both.

  16. Cleavage of phage DNA by the Streptococcus thermophilus CRISPR3-Cas system.

    Directory of Open Access Journals (Sweden)

    Alfonso H Magadán

    Full Text Available Streptococcus thermophilus, similar to other Bacteria and Archaea, has developed defense mechanisms to protect cells against invasion by foreign nucleic acids, such as virus infections and plasmid transformations. One defense system recently described in these organisms is the CRISPR-Cas system (Clustered Regularly Interspaced Short Palindromic Repeats loci coupled to CRISPR-associated genes. Two S. thermophilus CRISPR-Cas systems, CRISPR1-Cas and CRISPR3-Cas, have been shown to actively block phage infection. The CRISPR1-Cas system interferes by cleaving foreign dsDNA entering the cell in a length-specific and orientation-dependant manner. Here, we show that the S. thermophilus CRISPR3-Cas system acts by cleaving phage dsDNA genomes at the same specific position inside the targeted protospacer as observed with the CRISPR1-Cas system. Only one cleavage site was observed in all tested strains. Moreover, we observed that the CRISPR1-Cas and CRISPR3-Cas systems are compatible and, when both systems are present within the same cell, provide increased resistance against phage infection by both cleaving the invading dsDNA. We also determined that overall phage resistance efficiency is correlated to the total number of newly acquired spacers in both CRISPR loci.

  17. Harnessing heterologous and endogenous CRISPR-Cas machineries for efficient markerless genome editing in Clostridium.

    Science.gov (United States)

    Pyne, Michael E; Bruder, Mark R; Moo-Young, Murray; Chung, Duane A; Chou, C Perry

    2016-01-01

    Application of CRISPR-Cas9 systems has revolutionized genome editing across all domains of life. Here we report implementation of the heterologous Type II CRISPR-Cas9 system in Clostridium pasteurianum for markerless genome editing. Since 74% of species harbor CRISPR-Cas loci in Clostridium, we also explored the prospect of co-opting host-encoded CRISPR-Cas machinery for genome editing. Motivation for this work was bolstered from the observation that plasmids expressing heterologous cas9 result in poor transformation of Clostridium. To address this barrier and establish proof-of-concept, we focus on characterization and exploitation of the C. pasteurianum Type I-B CRISPR-Cas system. In silico spacer analysis and in vivo interference assays revealed three protospacer adjacent motif (PAM) sequences required for site-specific nucleolytic attack. Introduction of a synthetic CRISPR array and cpaAIR gene deletion template yielded an editing efficiency of 100%. In contrast, the heterologous Type II CRISPR-Cas9 system generated only 25% of the total yield of edited cells, suggesting that native machinery provides a superior foundation for genome editing by precluding expression of cas9 in trans. To broaden our approach, we also identified putative PAM sequences in three key species of Clostridium. This is the first report of genome editing through harnessing native CRISPR-Cas machinery in Clostridium. PMID:27157668

  18. Genome-wide binding of the CRISPR endonuclease Cas9 in mammalian cells

    OpenAIRE

    Wu, Xuebing; Scott, David A.; Kriz, Andrea J.; Chiu, Anthony C; Hsu, Patrick D.; Dadon, Daniel B.; Cheng, Albert W.; Trevino, Alexandro E.; Konermann, Silvana; Chen, Sidi; Jaenisch, Rudolf; Zhang, Feng; Sharp, Phillip A.

    2014-01-01

    Bacterial type II CRISPR-Cas9 systems have been widely adapted for RNA-guided genome editing and transcription regulation in eukaryotic cells, yet their in vivo target specificity is poorly understood. Here we mapped genome-wide binding sites of a catalytically inactive Cas9 (dCas9) from Streptococcus pyogenes loaded with single guide RNAs (sgRNAs) in mouse embryonic stem cells (mESCs). Each of the four sgRNAs we tested targets dCas9 to between tens and thousands of genomic sites, frequently ...

  19. DNA interrogation by the CRISPR RNA-guided endonuclease Cas9

    OpenAIRE

    Sternberg, Samuel H.; Redding, Sy; Jinek, Martin; Greene, Eric C.; Doudna, Jennifer A.

    2014-01-01

    The CRISPR-associated enzyme Cas9 is an RNA-guided endonuclease that uses RNA:DNA base-pairing to target foreign DNA in bacteria. Cas9:guide RNA complexes are also effective genome engineering agents in animals and plants. Here we use single-molecule and bulk biochemical experiments to determine how Cas9:RNA interrogates DNA to find specific cleavage sites. We show that both binding and cleavage of DNA by Cas9:RNA require recognition of a short trinucleotide protospacer adjacent motif (PAM). ...

  20. Structures of Cas9 Endonucleases Reveal RNA-Mediated Conformational Activation

    OpenAIRE

    Jinek, Martin; Jiang, Fuguo; Taylor, David W.; Sternberg, Samuel H.; Kaya, Emine; MA, ENBO; Anders, Carolin; Hauer, Michael; Zhou, Kaihong; Lin, Steven; Kaplan, Matias; Anthony T Iavarone; Charpentier, Emmanuelle; Nogales, Eva; Doudna, Jennifer A.

    2014-01-01

    Type II CRISPR (clustered regularly interspaced short palindromic repeats)–Cas (CRISPR-associated) systems use an RNA-guided DNA endonuclease, Cas9, to generate double-strand breaks in invasive DNA during an adaptive bacterial immune response. Cas9 has been harnessed as a powerful tool for genome editing and gene regulation in many eukaryotic organisms. We report 2.6 and 2.2 angstrom resolution crystal structures of two major Cas9 enzyme subtypes, revealing the structural core shared by all C...

  1. Generation of an Oocyte-Specific Cas9 Transgenic Mouse for Genome Editing

    OpenAIRE

    Linlin ZHANG; Zhou, Jiankui; Han, Jinxiong; Hu, Bian; Hou, Ningning; Shi, Yun; Huang, Xingxu; Lou, Xin

    2016-01-01

    The CRISPR/Cas9 system has been developed as an easy-handle and multiplexable approach for engineering eukaryotic genomes by zygote microinjection of Cas9 and sgRNA, while preparing Cas9 for microinjection is laborious and introducing inconsistency into the experiment. Here, we describe a modified strategy for gene targeting through using oocyte-specific Cas9 transgenic mouse. With this mouse line, we successfully achieve precise gene targeting by injection of sgRNAs only into one-cell-stage ...

  2. Targeted Gene Manipulation in Plants Using the CRISPR/Cas Technology.

    Science.gov (United States)

    Zhang, Dandan; Li, Zhenxiang; Li, Jian-Feng

    2016-05-20

    The CRISPR/Cas technology is emerging as a revolutionary genome editing tool in diverse organisms including plants, and has quickly evolved into a suite of versatile tools for sequence-specific gene manipulations beyond genome editing. Here, we review the most recent applications of the CRISPR/Cas toolkit in plants and also discuss key factors for improving CRISPR/Cas performance and strategies for reducing the off-target effects. Novel technical breakthroughs in mammalian research regarding the CRISPR/Cas toolkit will also be incorporated into this review in hope to stimulate prospective users from the plant research community to fully explore the potential of these technologies. PMID:27165865

  3. CRISPR/Cas9: Implications for Modeling and Therapy of Neurodegenerative Diseases

    OpenAIRE

    Yang, Weili; Tu, Zhuchi; Sun, Qiang; Li, Xiao-Jiang

    2016-01-01

    CRISPR/Cas9 is now used widely to genetically modify the genomes of various species. The ability of CRISPR/Cas9 to delete DNA sequences and correct DNA mutations opens up a new avenue to treat genetic diseases that are caused by DNA mutations. In this review, we describe the advantages of using CRISPR/Cas9 to engineer genomic DNAs in animal embryos, as well as in specific regions or cell types in the brain. We also discuss how to apply CRISPR/Cas9 to establish animal models of neurodegenerati...

  4. Generating Mouse Models Using CRISPR-Cas9-Mediated Genome Editing.

    Science.gov (United States)

    Qin, Wenning; Kutny, Peter M; Maser, Richard S; Dion, Stephanie L; Lamont, Jeffrey D; Zhang, Yingfan; Perry, Greggory A; Wang, Haoyi

    2016-01-01

    The CRISPR-Cas9 system in bacteria and archaea has recently been exploited for genome editing in various model organisms, including mice. The CRISPR-Cas9 reagents can be delivered directly into the mouse zygote to derive a mutant animal carrying targeted genetic modifications. The major components of the system include the guide RNA, which provides target specificity, the Cas9 nuclease that creates the DNA double-strand break, and the donor oligonucleotide or plasmid carrying the intended mutation flanked by sequences homologous to the target site. Here we describe the general considerations and experimental protocols for creating genetically modified mice using the CRISPR-Cas9 system. PMID:26928663

  5. CRISPR/Cas9基因组编辑技术及其在动物基因组定点修饰中的应用%CRISPR/Cas9 genome editing technique and its application in site-directed genome modification of animals

    Institute of Scientific and Technical Information of China (English)

    周金伟; 徐绮嫔; 姚婧; 余树民; 曹随忠

    2015-01-01

    CRISPR/Cas system, which uses CRISPR RNAs (crRNAs) to guide Cas nuclease to silence invading nucleic acids, is self-defense system against exogenous virus or plasmid in bacteria and archaea. Through molecular modification, the typeⅡCRISPR/Cas system has become a highly efficient site-directed genome editing technique, which is simpler than zinc-finger nucleases (ZFNs) and transcription activator like effector nucleases (TALENs) and easier to be designed and applied. In this review, we summarize the evolutionary history of CRISPR/Cas9 system, the working principle and modification process of type Ⅱ CRISPR/Cas and its application in animal genome modifica-tion. We also analyze the existing problems and improvement program of the CRISPR/Cas9 system as well as its ap-plication prospect combined with successful cases, which may provide innovative perspectives on improving animal traits and establishing animal models of human diseases.%CRISPR/Cas系统是细菌和古生菌中抵抗外源病毒或质粒入侵的获得性免疫系统,利用 CRISPR RNAs (crRNAs)引导Cas核酸酶沉默入侵的核酸。通过分子生物学改造使Ⅱ型CRISPR/Cas系统成为一种高效的基因组定点修饰技术,并且比锌指核酸酶(Zinc-finger nucleases, ZFNs)和 TALE 核酸酶(Transcription activator like effector nucleases, TALENs)结构更简单,更容易设计和应用。文章主要介绍了CRISPR/Cas9系统成为高效基因组定点修饰技术的发展历程、Ⅱ型CRISPR/Cas的工作原理和改造过程以及在动物基因组定点修饰的应用,剖析了该技术存在的问题和现有改进方案,并与成功案例相结合展望了 CRISPR/Cas9系统的应用前景,以期为动物性状改良和人类疾病动物模型的创立提供新思路。

  6. Iron deficiency anemia in children.

    Science.gov (United States)

    Subramaniam, Girish; Girish, Meenakshi

    2015-06-01

    Iron deficiency is not just anemia; it can be responsible for a long list of other manifestations. This topic is of great importance, especially in infancy and early childhood, for a variety of reasons. Firstly, iron need is maximum in this period. Secondly, diet in infancy is usually deficient in iron. Thirdly and most importantly, iron deficiency at this age can result in neurodevelopmental and cognitive deficits, which may not be reversible. Hypochromia and microcytosis in a complete blood count (CBC) makes iron deficiency anemia (IDA) most likely diagnosis. Absence of response to iron should make us look for other differential diagnosis like β thalassemia trait and anemia of chronic disease. Celiac disease is the most important cause of true IDA not responding to oral iron therapy. While oral ferrous sulphate is the cheapest and most effective therapy for IDA, simple nonpharmacological and pharmacological measures can go a long way in prevention of iron deficiency. PMID:25636824

  7. Cas5d Protein Processes Pre-crRNA and Assembles into a Cascade-like Interference Complex in Subtype I-C/Dvulg CRISPR-Cas System

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Ki Hyun; Haitjema, Charles; Liu, Xueqi; Ding, Fran; Wang, Hongwei; DeLisa, Matthew P.; Ke, Ailong (Yale); (Cornell); (Tsinghua)

    2012-10-10

    Clustered regularly interspaced short palindromic repeats (CRISPRs), together with an operon of CRISPR-associated (Cas) proteins, form an RNA-based prokaryotic immune system against exogenous genetic elements. Cas5 family proteins are found in several type I CRISPR-Cas systems. Here, we report the molecular function of subtype I-C/Dvulg Cas5d from Bacillus halodurans. We show that Cas5d cleaves pre-crRNA into unit length by recognizing both the hairpin structure and the 3 single stranded sequence in the CRISPR repeat region. Cas5d structure reveals a ferredoxin domain-based architecture and a catalytic triad formed by Y46, K116, and H117 residues. We further show that after pre-crRNA processing, Cas5d assembles with crRNA, Csd1, and Csd2 proteins to form a multi-sub-unit interference complex similar to Escherichia coli Cascade (CRISPR-associated complex for antiviral defense) in architecture. Our results suggest that formation of a crRNA-presenting Cascade-like complex is likely a common theme among type I CRISPR subtypes.

  8. Interactions between copper deficiency, selenium deficiency and adriamycin toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Fischer, J.; Tackett, R.; Johnson, M.A. (Univ. of Georgia, Athens (United States))

    1991-03-15

    The objective of this study was to test the hypothesis that there are interactions between copper (Cu) and selenium (Se) status, and adriamycin (ADR) toxicity. Male Sprague Dawley rats were fed Cu,Se adequate; Cu deficient, Se adequate ({minus}Cu); Cu adequate, Se deficient; or Cu,Se deficient diets for 38-41 days. ADR or saline (SAL) were administered weekly for the last 4 weeks of the study. Cu deficiency was confirmed by a 3-fold decrease in liver Cu,Zn-superoxide dismutase and liver Cu, and a 5-fold decrease in RBC Cu,Zn-SOD. Se deficiency was confirmed by a 10-fold decrease in liver glutathione peroxidase (GSH-Px). ADR, Cu deficiency and Se deficiency all caused EKG abnormalities. However, Cu and Se deficiencies did not enhance ADR's influence on EKGs. ADR increased lipid peroxidation in liver by 15% and in heart by 18% (NS). Cu deficiency decreased ADR-induced lipid peroxidation in heart tissue by 25%. ADR influenced Se status by significantly increasing heart GSH-Px, and Cu status by increasing liver Cu, plasma ceruloplasmin and liver Cu, Zn-SOD. These elevations in Cu,Zn-SOD and GSH-Px may be a consequence of the increased lipid peroxidation initiated by ADR. In {minus}Cu rats, ADR caused severe hemolytic anemia characterized by a 19% decrease in hematocrit and a 17-fold increase in splenic Fe. These data suggest that there are numerous interactions between ADR toxicity and Cu and Se status.

  9. CRISPR/Cas基因组靶向编辑技术综述%Review about CRISPR/Cas system as a new targeted genome editing tech-nology

    Institute of Scientific and Technical Information of China (English)

    贾良杰

    2014-01-01

    CRISPR (clustered, regularly interspaced, short, palindromic repeats)/Cas (CRISPR-associated) systems are a unique prokaryotic defense against foreign genetic elements. It also be considered as a targeted genome editing tool in molecular biology research. Because of its simplicity, high success rate and high efficiency in genome targeting, Cas system became the best genome targeting tools compared with ZFNs (Zinc-finger nucleases) and TALENs (Transcrip-tion activator-like effector nucleases). According to the recent researches, Cas system could be used as an efficient system for site-specific transcriptional repression or activation. Additionally, a specific Cas9 protein has been observed to target an RNA substrate, suggesting that Cas9 may have the same ability as RNAi technology to be programmed to target RNA as well. Overall, the targeted genome editing technology via CRISPR/Cas system has been Widely promot-ed and applied in many field such as the research on gene functions, the the disease model of gene knock-out animal construction and the gene therapy. So it will have significant impact on future advancements in genome engineering.%Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated(Cas)是细菌特有的一种获得性免疫系统,研究人员将其改造成为靶向基因组编辑的工具,由于操作简单、成功率高且效率高,成为了靶向基因组编辑工具中的佼佼者。同时CRISPR/Cas系统也被改造作为一种极为有效的位点特异性的转录抑制/激活的工具而在研究工作中广泛应用,不仅如此,具有靶向细胞转录过程中产生的RNA底物的利用前景,从而起到与RNAi技术相类似的效果。 CRISPR-Cas技术已经在基因功能研究、动物模型建立、基因治疗等领域得到广泛的推广和应用,有力地推动了相关领域的研究进展。

  10. Immune Deficiency Foundation

    Science.gov (United States)

    ... The IDF Resource for the CGD Community For Healthcare Professionals Consulting Immunologist Program Free physician to physician consults ... Publications Educational materials developed by expert immunologists and healthcare professionals Connect IDF Social Networks IDF Friends Our social ...

  11. CR ISPR-Cas9-based Genome Engineering%CRISPR-Cas9介导的基因组编辑技术的研究进展

    Institute of Scientific and Technical Information of China (English)

    郑小梅; 张晓立; 于建东; 郑平; 孙际宾

    2015-01-01

    CRISPR-Cas ( clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) systems are the adaptive immune system in bacteria and archaea that defends against infectious viruses and plasmids.Immunity is mediated by Cas nucleases, which use small RNA guides ( the crRNAs) to specify a cleavage site within the genome of invading nucleic acids. In typeⅡCRISPR-Cas systems, the DNA-cleaving activity is performed by a single enzyme Cas9 guided by an RNA duplex. Using synthetic single RNA guides, Cas9 can be reprogrammed to create specific double-stranded DNA breaks in the genomes of a variety of organisms, ranging from human cells to bacteria, and thus constitutes a powerful tool for genetic engineering.In this review, we described the development and applications of CRISPR-Cas9 system with highlighting the practical considerations for implementing CRISPR-Cas9 technology.%CRISPR-Cas ( clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins)系统为细菌与古生菌中抵御外源病毒或质粒DNA入侵的获得性免疫系统。该系统在crRNA的指导下,使核酸酶Cas识别并降解外源DNA。其中,Ⅱ型CRISPR-Cas系统最为简单,仅包括一个核酸酶Cas9与tracrRNA:crRNA二聚体便可完成其生物功能。基于CRISPR-Cas9的基因组编辑技术的核心为将tracrRNA:crRNA设计为引导RNA,在引导RNA的指导下Cas9定位于特定DNA序列上,进行DNA双链切割,实现基因组的定向编辑。 CRISPR-Cas9系统以设计操纵简便、编辑高效与通用性广等优势成为新一代基因组编辑技术,为基因组定向改造调控与应用等带来突破性革命。从CRISPRC-as 9介导的基因组编辑技术的发展与应用等方面综述其最新研究进展,并着重介绍该技术的关键影响因素,为相关研究者提供参考。

  12. How Is Alpha-1 Antitrypsin Deficiency Diagnosed?

    Science.gov (United States)

    ... Alpha-1 Antitrypsin Deficiency Diagnosed? Alpha-1 antitrypsin (AAT) deficiency usually is diagnosed after you develop a ... related to the condition. Your doctor may suspect AAT deficiency if you have signs or symptoms of ...

  13. How Is Alpha-1 Antitrypsin Deficiency Treated?

    Science.gov (United States)

    ... Alpha-1 Antitrypsin Deficiency Treated? Alpha-1 antitrypsin (AAT) deficiency has no cure, but its related lung ... pulmonary disease). If you have symptoms related to AAT deficiency, your doctor may recommend: Medicines called inhaled ...

  14. Glucose-6-Phosphate Dehydrogenase Deficiency Overview

    Science.gov (United States)

    ... Drugs GARD Information Navigator FAQs About Rare Diseases Glucose-6-phosphate dehydrogenase deficiency Title Other Names: G6PD ... G6PD deficiency Categories: Newborn Screening Summary Summary Listen Glucose 6 phosphate dehydrogenase (G6PD) deficiency is a hereditary ...

  15. Genetics Home Reference: factor V deficiency

    Science.gov (United States)

    ... Genetics Home Health Conditions factor V deficiency factor V deficiency Enable Javascript to view the expand/collapse ... Print All Open All Close All Description Factor V deficiency is a rare bleeding disorder. The signs ...

  16. Monocular Elevation Deficiency - Double Elevator Palsy

    Science.gov (United States)

    ... Español Condiciones Chinese Conditions Monocular Elevation Deficiency/ Double Elevator Palsy En Español Read in Chinese What is monocular elevation deficiency (Double Elevator Palsy)? Monocular Elevation Deficiency, also known by the ...

  17. Genetics Home Reference: leptin receptor deficiency

    Science.gov (United States)

    ... Understand Genetics Home Health Conditions leptin receptor deficiency leptin receptor deficiency Enable Javascript to view the expand/ ... boxes. Print All Open All Close All Description Leptin receptor deficiency is a condition that causes severe ...

  18. Genetics Home Reference: congenital leptin deficiency

    Science.gov (United States)

    ... Genetics Home Health Conditions congenital leptin deficiency congenital leptin deficiency Enable Javascript to view the expand/collapse ... Print All Open All Close All Description Congenital leptin deficiency is a condition that causes severe obesity ...

  19. Genetics Home Reference: carnitine palmitoyltransferase I deficiency

    Science.gov (United States)

    ... Understand Genetics Home Health Conditions CPT I deficiency carnitine palmitoyltransferase I deficiency Enable Javascript to view the ... boxes. Print All Open All Close All Description Carnitine palmitoyltransferase I (CPT I) deficiency is a condition ...

  20. Genetics Home Reference: factor XIII deficiency

    Science.gov (United States)

    ... InfoSearch: Factor XIII deficiency Factor XIII Registry Database: Introduction to Factor XIII Deficiency MalaCards: factor xiii deficiency ... Library of Medicine Lister Hill National Center for Biomedical Communications 8600 Rockville Pike, Bethesda, MD 20894, USA ...

  1. Genetics Home Reference: combined pituitary hormone deficiency

    Science.gov (United States)

    ... Genetics Home Health Conditions combined pituitary hormone deficiency combined pituitary hormone deficiency Enable Javascript to view the ... boxes. Print All Open All Close All Description Combined pituitary hormone deficiency is a condition that causes ...

  2. Mortality and GH deficiency

    DEFF Research Database (Denmark)

    Stochholm, Kirstine; Gravholt, Claus Højbjerg; Laursen, Torben;

    2007-01-01

    OBJECTIVE: To estimate the mortality in Denmark in patients suffering from GH deficiency (GHD). DESIGN: Mortality was analyzed in 1794 GHD patients and 8014 controls matched on age and gender. All records in GHD patients were studied and additional morbidity noted. Patients were divided into...... childhood onset (CO) and adult onset (AO), discriminated by an age cutoff below or above 18 years at onset of GHD. METHOD: Data on death were identified in national registries. Sex- and cause-specific mortalities were identified in CO and AO GHD when compared with controls. RESULTS: Mortality was increased...... in CO and AO GHD in both genders, when compared with controls. The hazard ratio (HR) for CO males was 8.3 (95% confidence interval (CI) 4.5-15.1) and for females 9.4 (CI 4.6-19.4). For AO males, HR was 1.9 (CI 1.7-2.2) and for females 3.4 (CI 2.9-4.0). We found a significantly higher HR in AO females...

  3. Iodine deficiency disorders.

    Science.gov (United States)

    Elliott, T C

    1987-01-01

    Iodine deficiency disorder (IDD) affects 800 million people in the world, yet iodine supplementation is one of the most cost-effective nutritional interventions known. Iodine is incorporated into thyroid hormones, necessary for regulating metabolic rate, growth, and development of the brain and nervous system. IDD may appear as goiter in adults, usually not a serious problem, or in cretinism in children, which is marked by severe mental and physical retardation, with irreversible hearing and speech defects and either deaf-mutism, squint and paralysis, or stunting and edema. Children supplemented by age 1 or 2 can sometimes be helped. Foods contain variable amounts of iodine dependent on the soil where they are grown, hence mountainous and some inland regions have high goiter and IDD incidence. There are also goitrogenic foods, typically those of the cabbage family. Diagnosis is clinical or by blood tests for thyroid hormone levels and ratios. Finger-stick methods are available. Prevention of IDD is simple with either iodized salt or flour, iodinated central water supplies, injectable or oral iodine-containing oil. All cost about $.04 per person per year, except injections, which cost about $1 per person, but have the advantage that they could be combined with immunizations. Local problems with supplements are loss of iodine in salt with storage in tropics, and local production of cheaper uniodinated salt. Emphasis should be given to pregnant women and young children. There is no harm in giving pregnant women iodine injections in 2nd or 3rd trimester. PMID:12343033

  4. Utilization of the CRISPR/Cas9 system for the efficient production of mutant mice using crRNA/tracrRNA with Cas9 nickase and FokI-dCas9

    Science.gov (United States)

    Terao, Miho; Tamano, Moe; Hara, Satoshi; Kato, Tomoko; Kinoshita, Masato; Takada, Shuji

    2016-01-01

    The CRISPR/Cas9 system is a powerful genome editing tool for the production of genetically modified animals. To produce mutant mice, chimeric single-guide RNA (sgRNA) is cloned in a plasmid vector and a mixture of sgRNA and Cas9 are microinjected into the fertilized eggs. An issue associated with gene manipulation using the CRISPR/Cas9 system is that there can be off-target effects. To simplify the production of mutant mice with low risks of off-target effects caused by the CRISPR/Cas9 system, we demonstrated that genetically modified mice can be efficiently obtained using chemically synthesized CRISPR RNA (crRNA), trans-activating crRNA (tracrRNA), and modified Cas9s, such as the nickase version and FokI-fused catalytically inactive Cas9, by microinjection into fertilized eggs. Using this method, it is no longer necessary to clone sgRNA into a plasmid vector, and this enables high-throughput production of mutant mice. PMID:26972821

  5. SSFinder: High Throughput CRISPR-Cas Target Sites Prediction Tool

    Directory of Open Access Journals (Sweden)

    Santosh Kumar Upadhyay

    2014-01-01

    Full Text Available Clustered regularly interspaced short palindromic repeats (CRISPR and CRISPR-associated protein (Cas system facilitates targeted genome editing in organisms. Despite high demand of this system, finding a reliable tool for the determination of specific target sites in large genomic data remained challenging. Here, we report SSFinder, a python script to perform high throughput detection of specific target sites in large nucleotide datasets. The SSFinder is a user-friendly tool, compatible with Windows, Mac OS, and Linux operating systems, and freely available online.

  6. CAS - CERN Accelerator School: Course on Superconductivity for Accelerators

    CERN Document Server

    2014-01-01

    These proceedings collate lectures given at the twenty-seventh specialized course organised by the CERN Accelerator School (CAS). The course was held at the Ettore Majorana Foundation and Centre for Scientific Culture (EMFCSC) in Erice, Italy, from 24 April to 4 May 2013. Following recapitulation lectures on basic accelerator physics and superconductivity, the course covered topics related to the design, production and operation of superconducting RF systems and superconducting magnets for accelerators. The participants pursued one of six case studies in order to get ’hands-on’ experience of the issues connected with the design of superconducting systems. A series of topical seminars completed the programme.

  7. Meeting report: GARNet/OpenPlant CRISPR-Cas workshop.

    Science.gov (United States)

    Parry, Geraint; Patron, Nicola; Bastow, Ruth; Matthewman, Colette

    2016-01-01

    Targeted genome engineering has been described as a "game-changing technology" for fields as diverse as human genetics and plant biotechnology. One technique used for precise gene editing utilises the CRISPR-Cas system and is an effective method for genetic engineering in a wide variety of plants. However, many researchers remain unaware of both the technical challenges that emerge when using this technique or of its potential benefits. Therefore in September 2015, GARNet and OpenPlant organized a two-day workshop at the John Innes Centre that provided both background information and hands-on training for this important technology. PMID:26823675

  8. CRISPR/Cas9 technology for targeted genome editing

    Directory of Open Access Journals (Sweden)

    Lomov N. A.

    2015-08-01

    Full Text Available CRISPRs (Clustered Regularly Interspaced Short Palindromic Repeats are the segments of prokaryotic DNA containing short repeats in its nucleotide sequence. Today we know that this is a bacterial protection system against viral DNA. The molecular components of CRISPR/Cas9 system have been used for a gene editing in eukaryotes since 2013. But as any other method it also has the limitations and drawbacks. Here we are going to review the history of CRISPR biology and to discuss the possibilities that this new technology provides to researchers as well as the prospects for its use in the medical research and treatment.

  9. Tuberculose orbitaire: à propos d'un cas

    OpenAIRE

    Khrifi, Zineb; Abdellaoui, Meriem; Alaoui, Abdellah; Benatiya, Idriss Andaloussi; Tahri, Hicham

    2014-01-01

    La tuberculose sévit à l’état endémique au Maroc, l'atteinte orbitaire est rare et peut se faire par voie hématogène ou par contigüité à partir d'un foyer de voisinage. Nous rapportons le cas d'une patiente de 42 ans qui présente une exophtalmie droite dont l'enquête étiologique révèle une tuberculose orbitaire.

  10. Clinical manifestations of zinc deficiency.

    Science.gov (United States)

    Prasad, A S

    1985-01-01

    The essentiality of zinc for humans was recognized in the early 1960s. The causes of zinc deficiency include malnutrition, alcoholism, malabsorption, extensive burns, chronic debilitating disorders, chronic renal diseases, following uses of certain drugs such as penicillamine for Wilson's disease and diuretics in some cases, and genetic disorders such as acrodermatitis enteropathica and sickle cell disease. In pregnancy and during periods of growth the requirement of zinc is increased. The clinical manifestations in severe cases of zinc deficiency include bullous-pustular dermatitis, alopecia, diarrhea, emotional disorder, weight loss, intercurrent infections, hypogonadism in males; it is fatal if unrecognized and untreated. A moderate deficiency of zinc is characterized by growth retardation and delayed puberty in adolescents, hypogonadism in males, rough skin, poor appetite, mental lethargy, delayed wound healing, taste abnormalities, and abnormal dark adaptation. In mild cases of zinc deficiency in human subjects, we have observed oligospermia, slight weight loss, and hyperammonemia. Zinc is a growth factor. Its deficiency adversely affects growth in many animal species and humans. Inasmuch as zinc is needed for protein and DNA synthesis and for cell division, it is believed that the growth effect of zinc is related to its effect on protein synthesis. Whether or not zinc is required for the metabolism of somatomedin needs to be investigated in the future. Testicular functions are affected adversely as a result of zinc deficiency in both humans and experimental animals. This effect of zinc is at the end organ level; the hypothalamic-pituitary axis is intact in zinc-deficient subjects. Inasmuch as zinc is intimately involved in cell division, its deficiency may adversely affect testicular size and thus affect its functions. Zinc is required for the functions of several enzymes and whether or not it has an enzymatic role in steroidogenesis is not known at present

  11. [Immune deficiencies in nutritional anemias].

    Science.gov (United States)

    Bonnet Gajdos, M; Navarro, J; Belas, F; Traineau, R

    1982-12-16

    A transient cellular immunologic defect caused by folic acid deficiency was seen in a goat-milk-fed infant with severe enterocolitis. Data on the immunologic consequences of folic acid, protein and iron deficiencies were reviewed in the medical literature. Investigations are difficult because of the patients' poor general condition. Results are difficult to interpret as many etiologic factors are often combined and mechanisms of immunologic responses are complex. Attention is drawn to the danger of iron therapy in patients with transferrin deficiency. PMID:6297076

  12. Iron deficiency - a global problem

    International Nuclear Information System (INIS)

    Iron deficiency is an important nutritional global problem. This paper contains summery of information gathered from a dietary survey as iron deficiency anaemia is major public health problem in many developing countries including Pakistan. Comparison of anaemia in different age group and sex versus various regions in the world are given. In Pakistan also anaemia is widespread. According to the report of Micro-Nutrient survey of Pakistan 40% of the population are found to have low level of haemoglobin, more than half of pregnant women suffered from marginal or deficient haemoglobin. (A.B.)

  13. Genetics Home Reference: adenosine monophosphate deaminase deficiency

    Science.gov (United States)

    ... links) CLIMB: Children Living with Inherited Metabolic Diseases Muscular Dystrophy Association: Myoadenylate Deaminase Deficiency Genetic Testing Registry (1 link) Muscle AMP deaminase deficiency ...

  14. Campylobacter jejuni acquire new host-derived CRISPR spacers when in association with bacteriophages harboring a CRISPR-like Cas4 protein

    OpenAIRE

    Hooton, Steven P. T.; Connerton, I.F.

    2015-01-01

    Campylobacter jejuni is a worldwide cause of human diarrhoeal disease. Clustered Repetitively Interspaced Palindromic Repeats (CRISPRs) and associated proteins allow Bacteria and Archaea to evade bacteriophage and plasmid infection. Type II CRISPR systems are found in association with combinations of genes encoding the CRISPR-associated Cas1, Cas2, Cas4 or Csn2, and Cas9 proteins. C. jejuni possesses a minimal subtype II-C CRISPR system containing cas1, cas2, and cas9 genes whilst cas4 is not...

  15. Occurrence and Diversity of CRISPR-Cas Systems in the Genus Bifidobacterium.

    Directory of Open Access Journals (Sweden)

    Alexandra E Briner

    Full Text Available CRISPR-Cas systems constitute adaptive immune systems for antiviral defense in bacteria. We investigated the occurrence and diversity of CRISPR-Cas systems in 48 Bifidobacterium genomes to gain insights into the diversity and co-evolution of CRISPR-Cas systems within the genus and investigate CRISPR spacer content. We identified the elements necessary for the successful targeting and inference of foreign DNA in select Type II CRISPR-Cas systems, including the tracrRNA and target PAM sequence. Bifidobacterium species have a very high frequency of CRISPR-Cas occurrence (77%, 37 of 48. We found that many Bifidobacterium species have unusually large and diverse CRISPR-Cas systems that contain spacer sequences showing homology to foreign genetic elements like prophages. A large number of CRISPR spacers in bifidobacteria show perfect homology to prophage sequences harbored in the chromosomes of other species of Bifidobacterium, including some spacers that self-target the chromosome. A correlation was observed between strains that lacked CRISPR-Cas systems and the number of times prophages in that chromosome were targeted by other CRISPR spacers. The presence of prophage-targeting CRISPR spacers and prophage content may shed light on evolutionary processes and strain divergence. Finally, elements of Type II CRISPR-Cas systems, including the tracrRNA and crRNAs, set the stage for the development of genome editing and genetic engineering tools.

  16. Lentivirus pre-packed with Cas9 protein for safer gene editing.

    Science.gov (United States)

    Choi, J G; Dang, Y; Abraham, S; Ma, H; Zhang, J; Guo, H; Cai, Y; Mikkelsen, J G; Wu, H; Shankar, P; Manjunath, N

    2016-07-01

    The CRISPR/Cas9 system provides an easy way to edit specific site/s in the genome and thus offers tremendous opportunity for human gene therapy for a wide range of diseases. However, one major concern is off-target effects, particularly with long-term expression of Cas9 nuclease when traditional expression methods such as via plasmid/viral vectors are used. To overcome this limitation, we pre-packaged Cas9 protein (Cas9P LV) in lentiviral particles for transient exposure and showed its effectiveness for gene disruption in cells, including primary T cells expressing specific single guide RNAs (sgRNAs). We then constructed an 'all in one virus' to express sgRNAs in association with pre-packaged Cas9 protein (sgRNA/Cas9P LV). We successfully edited CCR5 in TZM-bl cells by this approach. Using an sgRNA-targeting HIV long terminal repeat, we also were able to disrupt HIV provirus in the J-LAT model of viral latency. Moreover, we also found that pre-packaging Cas9 protein in LV particle reduced off-target editing of chromosome 4:-29134166 locus by CCR5 sgRNA, compared with continued expression from the vector. These results show that sgRNA/Cas9P LV can be used as a safer approach for human gene therapy applications. PMID:27052803

  17. Diverse evolutionary roots and mechanistic variations of the CRISPR-Cas systems

    NARCIS (Netherlands)

    Mohanraju, Prarthana; Makarova, Kira S.; Zetsche, Bernd; Zhang, Feng; Koonin, Eugene V.; Oost, van der John

    2016-01-01

    Adaptive immunity had been long thought of as an exclusive feature of animals. However, the discovery of the CRISPR-Cas defense system, present in almost half of prokaryotic genomes, proves otherwise. Because of the everlasting parasite-host arms race, CRISPR-Cas has rapidly evolved through horiz

  18. Optimization of genome engineering approaches with the CRISPR/Cas9 system

    DEFF Research Database (Denmark)

    Li, Kai; Wang, Gang; Andersen, Troels;

    2014-01-01

    Designer nucleases such as TALENS and Cas9 have opened new opportunities to scarlessly edit the mammalian genome. Here we explored several parameters that influence Cas9-mediated scarless genome editing efficiency in murine embryonic stem cells. Optimization of transfection conditions and enrichi...

  19. Phonological Awareness and Early Reading Development in Childhood Apraxia of Speech (CAS)

    Science.gov (United States)

    McNeill, B. C.; Gillon, G. T.; Dodd, B.

    2009-01-01

    Background: Childhood apraxia of speech (CAS) is associated with phonological awareness, reading, and spelling deficits. Comparing literacy skills in CAS with other developmental speech disorders is critical for understanding the complexity of the disorder. Aims: This study compared the phonological awareness and reading development of children…

  20. Efficient fdCas9 Synthetic Endonuclease with Improved Specificity for Precise Genome Engineering

    KAUST Repository

    Aouida, Mustapha

    2015-07-30

    The Cas9 endonuclease is used for genome editing applications in diverse eukaryotic species. A high frequency of off-target activity has been reported in many cell types, limiting its applications to genome engineering, especially in genomic medicine. Here, we generated a synthetic chimeric protein between the catalytic domain of the FokI endonuclease and the catalytically inactive Cas9 protein (fdCas9). A pair of guide RNAs (gRNAs) that bind to sense and antisense strands with a defined spacer sequence range can be used to form a catalytically active dimeric fdCas9 protein and generate double-strand breaks (DSBs) within the spacer sequence. Our data demonstrate an improved catalytic activity of the fdCas9 endonuclease, with a spacer range of 15–39 nucleotides, on surrogate reporters and genomic targets. Furthermore, we observed no detectable fdCas9 activity at known Cas9 off-target sites. Taken together, our data suggest that the fdCas9 endonuclease variant is a superior platform for genome editing applications in eukaryotic systems including mammalian cells.

  1. Structure of a CRISPR-associated protein Cas2 from Desulfovibrio vulgaris

    International Nuclear Information System (INIS)

    A 1.35 Å resolution crystal structure of Cas2 from the bacterium Desulfovibrio vulgaris (DvuCas2) is reported. CRISPRs (clustered regularly interspaced short palindromic repeats) provide bacteria and archaea with RNA-guided acquired immunity to invasive DNAs. CRISPR-associated (Cas) proteins carry out the immune effector functions. Cas2 is a universal component of the CRISPR system. Here, a 1.35 Å resolution crystal structure of Cas2 from the bacterium Desulfovibrio vulgaris (DvuCas2) is reported. DvuCas2 is a homodimer, with each protomer consisting of an N-terminal βαββαβ ferredoxin fold (amino acids 1–78) to which is appended a C-terminal segment (amino acids 79–102) that includes a short 310-helix and a fifth β-strand. The β5 strands align with the β4 strands of the opposite protomers, resulting in two five-stranded antiparallel β-sheets that form a sandwich at the dimer interface. The DvuCas2 dimer is stabilized by a distinctive network of hydrophilic cross-protomer side-chain interactions

  2. Transgene-Free Genome Editing in Caenorhabditis elegans Using CRISPR-Cas

    OpenAIRE

    Chiu, Hui; Schwartz, Hillel T.; Antoshechkin, Igor; Paul W Sternberg

    2013-01-01

    CRISPR-Cas is an efficient method for genome editing in organisms from bacteria to human cells. We describe a transgene-free method for CRISPR-Cas-mediated cleavage in nematodes, enabling RNA-homology-targeted deletions that cause loss of gene function; analysis of whole-genome sequencing indicates that the nuclease activity is highly specific.

  3. Promoting a Research Hub of Water Resources by Pooling Forccs from CAS Institutes

    Institute of Scientific and Technical Information of China (English)

    LIU Wenhua; ZHAN Chesheng; XIA Jun; UU Xiaojie

    2012-01-01

    CAS inaugurated its Center for Water Resources Research in 2006 with an objective to bring together R&D resources housed in different CAS institutes so as to play a major role in the comprehensive, strategic, interdisciplinary and foresighted studies of water resources at the national level and in key regions. In light of China's strategic demands, the Center carries out macroscopic,

  4. 76 FR 79545 - Cost Accounting Standards: Change to the CAS Applicability Threshold for the Inflation Adjustment...

    Science.gov (United States)

    2011-12-22

    ... request for comment (76 FR 40817) for the purpose of revising the Cost Accounting Standards (CAS... Acquisition Regulations Council on August 30, 2010 (at 75 FR 53129). By revising the CAS applicability... Council (Councils) published a final rule in the Federal Register on August 30, 2010 (75 FR...

  5. An Undergraduate Laboratory Class Using CRISPR/Cas9 Technology to Mutate Drosophila Genes

    Science.gov (United States)

    Adame, Vanesa; Chapapas, Holly; Cisneros, Marilyn; Deaton, Carol; Deichmann, Sophia; Gadek, Chauncey; Lovato, TyAnna L.; Chechenova, Maria B.; Guerin, Paul; Cripps, Richard M.

    2016-01-01

    CRISPR/Cas9 genome editing technology is used in the manipulation of genome sequences and gene expression. Because of the ease and rapidity with which genes can be mutated using CRISPR/Cas9, we sought to determine if a single-semester undergraduate class could be successfully taught, wherein students isolate mutants for specific genes using…

  6. Evolutionary Processes and Mental Deficiency

    Science.gov (United States)

    Spitz, Herman H.

    1973-01-01

    The author hypothesizes that central nervous system damage of deficiency associated with mental retardation affects primarily those cortical processes which developed at a late stage in man's evolutionary history. (Author)

  7. [Niacin deficiency and cutaneous immunity].

    Science.gov (United States)

    Ikenouchi-Sugita, Atsuko; Sugita, Kazunari

    2015-01-01

    Niacin, also known as vitamin B3, is required for the synthesis of coenzymes, nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP). Niacin binds with G protein-coupled receptor (GPR) 109A on cutaneous Langerhans cells and causes vasodilation with flushing in head and neck area. Niacin deficiency due to excessive alcohol consumption, certain drugs or inadequate uptake in diet causes pellagra, a photosensitivity dermatitis. Recently several studies have revealed the mechanism of photosensitivity in niacin deficiency, which may pave a way for new therapeutic approaches. The expression level of prostaglandin E synthase (PTGES) is up-regulated in the skin of both pellagra patients and niacin deficient pellagra mouse models. In addition, pellagra is mediated through prostaglandin E₂-EP4 (PGE₂-EP4) signaling via reactive oxygen species (ROS) production in keratinocytes. In this article, we have reviewed the role of niacin in immunity and the mechanism of niacin deficiency-induced photosensitivity. PMID:25765687

  8. Cutaneous findings of nutritional deficiencies in children.

    Science.gov (United States)

    Goskowicz, M; Eichenfield, L F

    1993-08-01

    Nutritional deficiencies may be associated with a variety of cutaneous findings in children. This review emphasizes new developments relating to cutaneous findings of nutritional deficiencies. Zinc deficiency, acrodermatitis enteropathica, and acrodermatitis enteropathica-like eruptions are seen with a variety of conditions including cystic fibrosis, anorexia nervosa, and breastfeeding. Similar cutaneous findings not related to zinc deficiency may also occur with such metabolic disorders as methylmalonic aciduria, multiple carboxylase deficiency, essential fatty acid deficiency and other amino acid deficiencies. Vitamin K deficiency is associated with hemorrhagic disease of the newborn and coagulopathy. Vitamin A deficiency presents with a variety of systemic findings and distinctive dermatologic findings. Acute vitamin A deficiency may be seen in children infected with measles and is associated with more severe disease. The systemic and cutaneous findings of vitamin C deficiency, scurvy, are discussed. PMID:8374671

  9. CRISPR-Cas Systems in the Cyanobacterium Synechocystis sp. PCC6803 Exhibit Distinct Processing Pathways Involving at Least Two Cas6 and a Cmr2 Protein

    OpenAIRE

    Scholz, Ingeborg; Lange, Sita J.; Hein, Stephanie; Wolfgang R Hess; Backofen, Rolf

    2013-01-01

    The CRISPR-Cas (Clustered Regularly Interspaced Short Palindrome Repeats – CRISPR associated proteins) system provides adaptive immunity in archaea and bacteria. A hallmark of CRISPR-Cas is the involvement of short crRNAs that guide associated proteins in the destruction of invading DNA or RNA. We present three fundamentally distinct processing pathways in the cyanobacterium Synechocystis sp. PCC6803 for a subtype I-D (CRISPR1), and two type III systems (CRISPR2 and CRISPR3), which are locate...

  10. Health Consequences of Iodine Deficiency

    OpenAIRE

    Kapil, Umesh

    2007-01-01

    Iodine Deficiency Disorders (IDD) are one of the biggest worldwide public health problem of today. Their effect is hidden and profoundly affects the quality of human life. Iodine deficiency occurs when the soil is poor in iodine, causing a low concentration in food products and insufficient iodine intake in the population. When iodine requirements are not met, the thyroid may no longer be able to synthesize sufficient amounts of thyroid hormone. The resulting low-level of thyroid hormones in ...

  11. Organophosphates and monocyte esterase deficiency.

    OpenAIRE

    1995-01-01

    AIMS--To examine the possibility that monocyte esterase deficiency (MED) could be caused by exposure to organophosphates. METHODS--Pseudocholinesterase, paraoxonase and arylesterase activities were measured in the serum and acetylcholinesterase activity was measured in the red cells of a group of monocyte esterase deficient subjects and compared with the enzyme activities of a control group of monocyte esterase positive subjects. RESULTS--No significant difference was found between the enzyme...

  12. The linear collider alignment and survey (LiCAS) project

    International Nuclear Information System (INIS)

    For the next generation of Linear Colliders (LC) the precision alignment of accelerator components will be critical. The DESY applied geodesy group has developed the concept of an automated 'survey train'. The train runs along the accelerator wall measuring the 3D position of a set of equispaced reference markers. This reference structure is then used to align the accelerator components. The LiCAS group is developing a measurement system for the survey train. It will use a combination of Laser Straightness Monitors (SM) and Frequency Scanning Interferometry (FSI). FSI is an interferometric length measurement technique originally developed for the online alignment of the ATLAS Inner Detector. This novel combination of optical techniques is expected to overcome the limitations of traditional open air survey. The authors describe the LiCAS project, the measurement systems and their integration into the survey train. The technical parameters and constraints will be mentioned. There will also be brief discussion of the second phase of the project to allow on-line monitoring of the LC alignment. (author)

  13. Hernie de Spiegel: a propos d’un cas

    Directory of Open Access Journals (Sweden)

    Karim Ibn Majdoub Hassani

    2010-02-01

    Full Text Available La hernie de Spiegel ou hernie ventrale latérale est une déhiscence inhabituelle apparaissant sur la ligne ou fascia semi-lunaire de Spiegel. C’est une entité clinique rare, représente 0.10 à 1 pourcent des hernies. Aussi, nous a-t-il paru opportun de rapporter ce cas colligé dans le service de chirurgie B du CHU Hassan II de Fès. Nous rapportons l’observation d’une patiente âgée de 60 ans, sans antécédent particulier qui présentais une tuméfaction para ombilicale gauche augmentant progressivement de volume, Une hernie de Spiegel a été suspectée à l’examen clinique, et le diagnostic d’éventration antérolatérale gauche a été retenu à la tomodensitométrie abdominale. Une cure de la hernie par plaque de prolène a été réalisée et les suites opératoires étaient simples. La hernie de Spiegel est une affection rare, son diagnostic clinique peut être difficile. Elle est asymptomatique dans 90 pourcent des cas et Son diagnostic positif est radiologique. Le risque d’étranglement non négligeable impose un traitement chirurgical une fois le diagnostic est confirmé.

  14. Maladie de Haglund: à propos de trois cas

    Science.gov (United States)

    Adigo, Amégninou Mawuko Yao; Gnakadja, Néille Gbèssi; Dellanh, Yaovi Yanick; Adambounou, Kokou; Djagnikpo, Oni; Agoda-Kousséma, Lama Kegdigoma; Adoko, Abikou Léon; Adjénou, Komlanvi Victor

    2015-01-01

    La maladie de Haglund est une pathologie relativement sous évaluée. Elle est liée à un conflit calcanéo-achilléen. Nous rapportons les cas de patients âgés de 40, 42 et 37 ans, révélés par des œdèmes douloureux de la cheville. Le diagnostic a été confirmé à la radiographie standard de la cheville en charge et à l’échographie chez tous les patients. Un seul patient avait bénéficié d'une exploration IRM. Le traitement, initialement médical dans tous les cas, s'est soldé par une chirurgie de résection de l'angle postéro-supérieur du calcanéum chez un patient. L’évolution a été favorable chez tous les patients. PMID:26664538

  15. An X-ray flare from 47 Cas

    Energy Technology Data Exchange (ETDEWEB)

    Pandey, Jeewan C.; Karmakar, Subhajeet, E-mail: jeewan@aries.res.in [Aryabhatta Research Institute of Observational Sciences (ARIES), Nainital-263 001 (India)

    2015-02-01

    Using XMM-Newton observations, we investigate properties of a flare from the very active but poorly known stellar system 47 Cas. The luminosity at the peak of the flare is found to be 3.54 × 10{sup 30} erg s{sup −1}, which is ∼2 times higher than that at a quiescent state. The quiescent state corona of 47 Cas can be represented by two temperature plasma: 3.7 and 11.0 MK. The time-resolved X-ray spectroscopy of the flare show the variable nature of the temperature, the emission measure, and the abundance. The maximum temperature during the flare is derived as 72.8 MK. We infer the length of a flaring loop to be 3.3 × 10{sup 10} cm using a hydrodynamic loop model. Using the RGS spectra, the density during the flare is estimated as 4.0 × 10{sup 10} cm{sup −3}. The loop scaling laws are also applied when deriving physical parameters of the flaring plasma.

  16. Photometric analysis of the overcontact binary CW Cas

    International Nuclear Information System (INIS)

    New CCD photometric observations of overcontact binary CW Cas were carried out in 2004 and 2011. In particular, the light curve obtained in 2004 shows a remarkable O'Connell effect. Compared with light curves in different observing seasons, variations were found. These variations can be explained by dark spot activities on the surface of at least one component. Using the Wilson-Devinney code with a spot model, we find that the photometric solutions confirm CW Cas is a shallow W-subtype overcontact binary with a spotted massive component. Our new determined times of minimum light together with the others published in the literature were analyzed to find a change of orbital period. From the O – C curves, the period of the system shows a cyclic period change (P 3 = 69.9 yr, A 3 = 0.03196 days) superposed on the linear increase. The cyclic variation, if explained as the light-travel time effect, reveals the presence of a tertiary companion.

  17. Photometric Analysis of the Overcontact Binary CW Cas

    Science.gov (United States)

    Wang, J. J.; Qian, S. B.; He, J. J.; Li, L. J.; Zhao, E. G.

    2014-11-01

    New CCD photometric observations of overcontact binary CW Cas were carried out in 2004 and 2011. In particular, the light curve obtained in 2004 shows a remarkable O'Connell effect. Compared with light curves in different observing seasons, variations were found. These variations can be explained by dark spot activities on the surface of at least one component. Using the Wilson-Devinney code with a spot model, we find that the photometric solutions confirm CW Cas is a shallow W-subtype overcontact binary with a spotted massive component. Our new determined times of minimum light together with the others published in the literature were analyzed to find a change of orbital period. From the O - C curves, the period of the system shows a cyclic period change (P 3 = 69.9 yr, A 3 = 0.03196 days) superposed on the linear increase. The cyclic variation, if explained as the light-travel time effect, reveals the presence of a tertiary companion.

  18. Characterization of CRISPR RNA Biogenesis and Cas6 Cleavage-Mediated Inhibition of a Provirus in the Haloarchaeon Haloferax mediterranei

    OpenAIRE

    Li, Ming; Liu, Hailong; Han, Jing; Liu, Jingfang; Wang, Rui; Zhao, Dahe; Zhou, Jian; Xiang, Hua

    2013-01-01

    The adaptive immune system comprising CRISPR (clustered regularly interspaced short palindromic repeats) arrays and cas (CRISPR-associated) genes has been discovered in a wide range of bacteria and archaea and has recently attracted comprehensive investigations. However, the subtype I-B CRISPR-Cas system in haloarchaea has been less characterized. Here, we investigated Cas6-mediated RNA processing in Haloferax mediterranei. The Cas6 cleavage site, as well as the CRISPR transcription start sit...

  19. Diverse evolutionary roots and mechanistic variations of the CRISPR-Cas systems.

    Science.gov (United States)

    Mohanraju, Prarthana; Makarova, Kira S; Zetsche, Bernd; Zhang, Feng; Koonin, Eugene V; van der Oost, John

    2016-08-01

    Adaptive immunity had been long thought of as an exclusive feature of animals. However, the discovery of the CRISPR-Cas defense system, present in almost half of prokaryotic genomes, proves otherwise. Because of the everlasting parasite-host arms race, CRISPR-Cas has rapidly evolved through horizontal transfer of complete loci or individual modules, resulting in extreme structural and functional diversity. CRISPR-Cas systems are divided into two distinct classes that each consist of three types and multiple subtypes. We discuss recent advances in CRISPR-Cas research that reveal elaborate molecular mechanisms and provide for a plausible scenario of CRISPR-Cas evolution. We also briefly describe the latest developments of a wide range of CRISPR-based applications. PMID:27493190

  20. Mouse genome engineering via CRISPR-Cas9 for study of immune function.

    Science.gov (United States)

    Pelletier, Stephane; Gingras, Sebastien; Green, Douglas R

    2015-01-20

    Clustered regularly interspaced palindromic repeats (CRISPR)-associated (Cas9) technology has proven a formidable addition to our armory of approaches for genomic editing. Derived from pathways in archaea and bacteria that mediate the resistance to exogenous genomic material, the CRISPR-Cas9 system utilizes a short single guide RNA (sgRNA) to direct the endonuclease Cas9 to virtually anywhere in the genome. Upon targeting, Cas9 generates DNA double-strand breaks (DSBs) and facilitates the repair or insertion of mutations, insertion of recombinase recognition sites, or large DNA elements. Here, we discuss the practical advantages of the CRISPR-Cas9 system over conventional and other nuclease-based targeting technologies and provide suggestions for the use of this technology to address immunological questions. PMID:25607456

  1. CRISPR/Cas9-mediated target validation of the Splicing Inhibitor Pladienolide B

    KAUST Repository

    Aouida, Mustapha

    2016-02-24

    CRISPR/Cas9 system confers molecular immunity in archeal and bacterial species against invading foreign nucleic acids. CRISPR/Cas9 system is used for genome engineering applications across diverse eukaryotic species. In this study, we demonstrate the utility of the CRISPR/Cas9 genome engineering system for drug target validation in human cells. Pladienolide B is a natural macrolide with antitumor activities mediated through the inhibition of pre-mRNA splicing. To validate the spliceosomal target of Pladienolide B, we employed the CRSIPR/Cas9 system to introduce targeted mutations in the subunits of the SF3B complex in the HEK293T cells. Our data reveal that targeted mutagenesis of the SF3b1 subunit exhibited higher levels of resistance to Pladienolide B. Therefore, our data validate the spliceosomal target of Pladienolide B and provide a proof of concept on using the CRISPR/Cas9 system for drug target identification and validation.

  2. Zinc and its deficiency diseases.

    Science.gov (United States)

    Evans, G W

    1986-01-01

    The pervasive role of zinc in the metabolic function of the body results from its function as a cofactor of a multitude of enzymes. Zinc is found in every tissue in the body, and because zinc metalloenzymes are found in every known class of enzymes, the metal has a function in every conceivable type of biochemical pathway. Symptoms resulting from zinc deficiency are as diverse as the enzymes with which the metal is associated. If chronic, severe, and untreated, zinc deficiency can be fatal. Less drastic symptoms include infections, hypogonadism, weight loss, emotional disturbance, dermatitis, alopecia, impaired taste acuity, night blindness, poor appetite, delayed wound healing, and elevated blood ammonia levels. Many symptoms of zinc deficiency result from poor diet consumption, but often the most severe symptoms result from other factors including excessive alcohol use, liver diseases, malabsorption syndromes, renal disease, enteral or parenteral alimentation, administration of sulfhydryl-containing drugs, and sickle cell disease. The most severe symptoms of zinc deficiency occur in young children affected with the autosomal-recessive trait, acrodermatitis enteropathica. This disease results in decreased synthesis of picolinic acid which causes an impaired ability to utilize zinc from common food. Because simple laboratory analyses are often not reliable in determining zinc nutriture of a patient, those symptoms caused by suspected zinc deficiency are best verified by the oral administration of zinc dipicolinate. This zinc compound is efficacious and safe and would provide an accurate means of identifying symptoms that do result from zinc deficiency. PMID:3514057

  3. CRISPR/Cas9系统在基因表达调控中的应用%Application of CRISPR/Cas9 System for Gene Regulation

    Institute of Scientific and Technical Information of China (English)

    魏泽辉; 贾存灵; 张智英

    2014-01-01

    CRISPR/Cas9系统是2012年发展起来由RNA导向的基因组编辑技术.相比ZFN和TALENs技术,CRISPR/Cas9系统具有类似的打靶效率,且更易于操作.通过将核酸酶Cas9蛋白D10A和H840A位点突变,可以获得失去切割活性的dCas9蛋白.借助导向RNA将dCas9与DNA特定位点结合的作用,可以实现抑制或激活基因转录,从而达到对特定基因表达进行调控的目的.本文主要从作用原理、影响因素等方面综述了CRISPR/Cas9系统在基因调控中的应用,以期为相关研究提供参考.

  4. International Scientists Programs:A New Gateway to Cooperation

    Institute of Scientific and Technical Information of China (English)

    XIN Ling

    2010-01-01

    @@ The Chinese Academy of Sciences(CAS)launched in 2009 a major effort to promote international cooperation and scientific innovation: the Visiting Professorship Program for Senior International Scientists and the Fellowship Program for Young International Scientists.As part of the Academy's long endeavor to attract foreign researchers,both programs received hundreds of applications from abroad.

  5. CRISPR Primer Designer:Design primers for knockout and chromosome imaging CRISPR-Cas system

    Institute of Scientific and Technical Information of China (English)

    Meng Yan; Shi-Rong Zhou; Hong-Wei Xue

    2015-01-01

    The clustered regularly interspaced short palin-dromic repeats (CRISPR)-associated system enables biologists to edit genomes precisely and provides a powerful tool for perturbing endogenous gene regulation, modulation of epigenetic markers, and genome architecture. However, there are concerns about the specificity of the system, especial y the usages of knocking out a gene. Previous designing tools either were mostly built-in websites or ran as command-line programs, and none of them ran local y and acquired a user-friendly interface. In addition, with the development of CRISPR-derived systems, such as chromosome imaging, there were stil no tools helping users to generate specific end-user spacers. We herein present CRISPR Primer Designer for researchers to design primers for CRISPR applications. The program has a user-friendly interface, can analyze the BLAST results by using multiple parameters, score for each candidate spacer, and generate the primers when using a certain plasmid. In addition, CRISPR Primer Designer runs local y and can be used to search spacer clusters, and exports primers for the CRISPR-Cas system-based chromosome imaging system.

  6. CRISPRseek: a bioconductor package to identify target-specific guide RNAs for CRISPR-Cas9 genome-editing systems.

    Directory of Open Access Journals (Sweden)

    Lihua J Zhu

    Full Text Available CRISPR-Cas systems are a diverse family of RNA-protein complexes in bacteria that target foreign DNA sequences for cleavage. Derivatives of these complexes have been engineered to cleave specific target sequences depending on the sequence of a CRISPR-derived guide RNA (gRNA and the source of the Cas9 protein. Important considerations for the design of gRNAs are to maximize aimed activity at the desired target site while minimizing off-target cleavage. Because of the rapid advances in the understanding of existing CRISPR-Cas9-derived RNA-guided nucleases and the development of novel RNA-guided nuclease systems, it is critical to have computational tools that can accommodate a wide range of different parameters for the design of target-specific RNA-guided nuclease systems. We have developed CRISPRseek, a highly flexible, open source software package to identify gRNAs that target a given input sequence while minimizing off-target cleavage at other sites within any selected genome. CRISPRseek will identify potential gRNAs that target a sequence of interest for CRISPR-Cas9 systems from different bacterial species and generate a cleavage score for potential off-target sequences utilizing published or user-supplied weight matrices with position-specific mismatch penalty scores. Identified gRNAs may be further filtered to only include those that occur in paired orientations for increased specificity and/or those that overlap restriction enzyme sites. For applications where gRNAs are desired to discriminate between two related sequences, CRISPRseek can rank gRNAs based on the difference between predicted cleavage scores in each input sequence. CRISPRseek is implemented as a Bioconductor package within the R statistical programming environment, allowing it to be incorporated into computational pipelines to automate the design of gRNAs for target sequences identified in a wide variety of genome-wide analyses. CRISPRseek is available under the GNU General

  7. Molecular basis of complement C3 deficiency in guinea pigs.

    OpenAIRE

    Auerbach, H S; de Burger, R; Dodds, A.; Colten, H R

    1990-01-01

    In experiments to ascertain the biochemical basis of a genetically determined deficiency of the third component of complement (C3) in guinea pigs, we found that C3-deficient liver and peritoneal macrophages contain C3 messenger RNA of normal size (approximately 5 kb) and amounts, that this mRNA programs synthesis of pro-C3 in oocytes primed with liver RNA and in primary macrophage cultures. In each instance, heterodimeric native C3 protein was secreted with normal kinetics but the C3 protein ...

  8. Targeted and genome-wide sequencing reveal single nucleotide variations impacting specificity of Cas9 in human stem cells

    OpenAIRE

    Yang, Luhan; Grishin, Dennis; Wang, Gang; Aach, John; Zhang, Cheng-Zhong; Chari, Raj; Homsy, Jason; Cai, Xuyu; ZHAO, YUE; Fan, Jian-Bing; Seidman, Christine; Seidman, Jonathan; Pu, William; Church, George

    2014-01-01

    CRISPR/Cas9 has demonstrated a high-efficiency in site-specific gene targeting. However, potential off-target effects of the Cas9 nuclease represent a major safety concern for any therapeutic application. Here, we knock out the Tafazzin gene by CRISPR/Cas9 in human-induced pluripotent stem cells with 54% efficiency. We combine whole-genome sequencing and deep-targeted sequencing to characterise the off-target effects of Cas9 editing. Whole-genome sequencing of Cas9-modified hiPSC clones detec...

  9. Expanding the CRISPR imaging toolset with Staphylococcus aureus Cas9 for simultaneous imaging of multiple genomic loci

    OpenAIRE

    Chen, Baohui; Hu, Jeffrey; Almeida, Ricardo; Liu, Harrison; Balakrishnan, Sanjeev; Covill-Cooke, Christian; Lim, Wendell A.; Huang, Bo

    2016-01-01

    In order to elucidate the functional organization of the genome, it is vital to directly visualize the interactions between genomic elements in living cells. For this purpose, we engineered the Cas9 protein from Staphylococcus aureus (SaCas9) for the imaging of endogenous genomic loci, which showed a similar robustness and efficiency as previously reported for Streptococcus pyogenes Cas9 (SpCas9). Imaging readouts allowed us to characterize the DNA-binding activity of SaCas9 and to optimize i...

  10. Structure and specificity of the RNA-guided endonuclease Cas9 during DNA interrogation, target binding and cleavage

    OpenAIRE

    Josephs, Eric A.; Kocak, D. Dewran; Fitzgibbon, Christopher J.; McMenemy, Joshua; Gersbach, Charles A; Marszalek, Piotr E.

    2015-01-01

    CRISPR-associated endonuclease Cas9 cuts DNA at variable target sites designated by a Cas9-bound RNA molecule. Cas9's ability to be directed by single ‘guide RNA’ molecules to target nearly any sequence has been recently exploited for a number of emerging biological and medical applications. Therefore, understanding the nature of Cas9's off-target activity is of paramount importance for its practical use. Using atomic force microscopy (AFM), we directly resolve individual Cas9 and nuclease-in...

  11. Conditional targeting of Ispd using paired Cas9 nickase and a single DNA template in mice

    Directory of Open Access Journals (Sweden)

    Angus Yiu-fai Lee

    2014-01-01

    Full Text Available CRISPR/Cas9 technology is a highly promising genome editing tool in the mouse, potentially overcoming the costs and time required for more traditional gene targeting methods in embryonic stem (ES cells. Recently, compared to the wildtype nuclease, paired Cas9 nickase (Cas9n combined with single guide RNA (sgRNA molecules has been found to enhance the specificity of genome editing while reducing off-target effects. Paired Cas9n has been shown to be as efficient as Cas9 for generating insertion and deletion (indel mutations by non-homologous end joining and targeted deletion in the genome. However, an efficient and reliable approach to the insertion of loxP sites flanking critical exon(s to create a conditional allele of a target gene remains an elusive goal. In this study, we microinjected Cas9n RNA with sgRNAs together with a single DNA template encoding two loxP sites flanking (floxing exon 2 of the isoprenoid synthase containing domain (Ispd into the pronucleus and cytoplasm of C57BL/6NCr one-cell stage zygotes. After surgical transfer, one F0 mouse expressing a conditional allele was produced (at a frequency of ∼8% of live pups born. The floxed allele was transmitted through the germline to F1 progeny, and could be successfully recombined using Cre recombinase. This study indicates that conditional targeting can be accomplished effectively using paired Cas9n and a single DNA template.

  12. The Neisseria meningitidis CRISPR-Cas9 System Enables Specific Genome Editing in Mammalian Cells

    Science.gov (United States)

    Lee, Ciaran M; Cradick, Thomas J; Bao, Gang

    2016-01-01

    The clustered regularly-interspaced short palindromic repeats (CRISPR)—CRISPR-associated (Cas) system from Streptococcus pyogenes (Spy) has been successfully adapted for RNA-guided genome editing in a wide range of organisms. However, numerous reports have indicated that Spy CRISPR-Cas9 systems may have significant off-target cleavage of genomic DNA sequences differing from the intended on-target site. Here, we report the performance of the Neisseria meningitidis (Nme) CRISPR-Cas9 system that requires a longer protospacer-adjacent motif for site-specific cleavage, and present a comparison between the Spy and Nme CRISPR-Cas9 systems targeting the same protospacer sequence. The results with the native crRNA and tracrRNA as well as a chimeric single guide RNA for the Nme CRISPR-Cas9 system were also compared. Our results suggest that, compared with the Spy system, the Nme CRISPR-Cas9 system has similar or lower on-target cleavage activity but a reduced overall off-target effect on a genomic level when sites containing three or fewer mismatches are considered. Thus, the Nme CRISPR-Cas9 system may represent a safer alternative for precision genome engineering applications. PMID:26782639

  13. Optimization of CRISPR/Cas9 genome editing to modify abiotic stress responses in plants

    Science.gov (United States)

    Osakabe, Yuriko; Watanabe, Takahito; Sugano, Shigeo S; Ueta, Risa; Ishihara, Ryosuke; Shinozaki, Kazuo; Osakabe, Keishi

    2016-01-01

    Genome editing using the CRISPR/Cas9 system can be used to modify plant genomes, however, improvements in specificity and applicability are still needed in order for the editing technique to be useful in various plant species. Here, using genome editing mediated by a truncated gRNA (tru-gRNA)/Cas9 combination, we generated new alleles for OST2, a proton pump in Arabidopsis, with no off-target effects. By following expression of Cas9 and the tru-gRNAs, newly generated mutations in CRIPSR/Cas9 transgenic plants were detected with high average mutation rates of up to 32.8% and no off-target effects using constitutive promoter. Reducing nuclear localization signals in Cas9 decreased the mutation rate. In contrast, tru-gRNA Cas9 cassettes driven by meristematic- and reproductive-tissue-specific promoters increased the heritable mutation rate in Arabidopsis, showing that high expression in the germ line can produce bi-allelic mutations. Finally, the new mutant alleles obtained for OST2 exhibited altered stomatal closing in response to environmental conditions. These results suggest further applications in molecular breeding to improve plant function using optimized plant CRISPR/Cas9 systems. PMID:27226176

  14. DNA interrogation by the CRISPR RNA-guided endonuclease Cas9

    Science.gov (United States)

    Sternberg, Samuel H.; Redding, Sy; Jinek, Martin; Greene, Eric C.; Doudna, Jennifer A.

    2014-03-01

    The clustered regularly interspaced short palindromic repeats (CRISPR)-associated enzyme Cas9 is an RNA-guided endonuclease that uses RNA-DNA base-pairing to target foreign DNA in bacteria. Cas9-guide RNA complexes are also effective genome engineering agents in animals and plants. Here we use single-molecule and bulk biochemical experiments to determine how Cas9-RNA interrogates DNA to find specific cleavage sites. We show that both binding and cleavage of DNA by Cas9-RNA require recognition of a short trinucleotide protospacer adjacent motif (PAM). Non-target DNA binding affinity scales with PAM density, and sequences fully complementary to the guide RNA but lacking a nearby PAM are ignored by Cas9-RNA. Competition assays provide evidence that DNA strand separation and RNA-DNA heteroduplex formation initiate at the PAM and proceed directionally towards the distal end of the target sequence. Furthermore, PAM interactions trigger Cas9 catalytic activity. These results reveal how Cas9 uses PAM recognition to quickly identify potential target sites while scanning large DNA molecules, and to regulate scission of double-stranded DNA.

  15. Optimization of CRISPR/Cas9 genome editing to modify abiotic stress responses in plants.

    Science.gov (United States)

    Osakabe, Yuriko; Watanabe, Takahito; Sugano, Shigeo S; Ueta, Risa; Ishihara, Ryosuke; Shinozaki, Kazuo; Osakabe, Keishi

    2016-01-01

    Genome editing using the CRISPR/Cas9 system can be used to modify plant genomes, however, improvements in specificity and applicability are still needed in order for the editing technique to be useful in various plant species. Here, using genome editing mediated by a truncated gRNA (tru-gRNA)/Cas9 combination, we generated new alleles for OST2, a proton pump in Arabidopsis, with no off-target effects. By following expression of Cas9 and the tru-gRNAs, newly generated mutations in CRIPSR/Cas9 transgenic plants were detected with high average mutation rates of up to 32.8% and no off-target effects using constitutive promoter. Reducing nuclear localization signals in Cas9 decreased the mutation rate. In contrast, tru-gRNA Cas9 cassettes driven by meristematic- and reproductive-tissue-specific promoters increased the heritable mutation rate in Arabidopsis, showing that high expression in the germ line can produce bi-allelic mutations. Finally, the new mutant alleles obtained for OST2 exhibited altered stomatal closing in response to environmental conditions. These results suggest further applications in molecular breeding to improve plant function using optimized plant CRISPR/Cas9 systems. PMID:27226176

  16. CAS Accelerators for Medical Applications in Vösendorf, Austria

    CERN Multimedia

    CERN Accelerator School

    2015-01-01

    The CERN Accelerator School (CAS) and MedAustron jointly organised a course on Accelerators for Medical Applications in Vösendorf, Austria between 26 May and 5 June 2015. The course was held at the Eventhotel Pyramide on the outskirts of Vienna, and was attended by 76 participants from 29 countries, coming from as far away as Canada, China, Lithuania, Thailand, Ukraine and Russia.       The intensive programme comprised 37 lectures. The emphasis was on using charged particle beams for cancer therapy and the programme began by covering the way in which particles interact with biological material, how this translates into the dose needed for treatment and how this dose is best delivered. The different accelerator options for providing the particles needed were then presented in some detail. The production of radioisotopes and how these are used for diagnostics and therapy was also covered, together with a look at novel acceleration techniques that may play a role i...

  17. Cas9-Mediated Genome Engineering in Drosophila melanogaster.

    Science.gov (United States)

    Housden, Benjamin E; Perrimon, Norbert

    2016-01-01

    The recent development of the CRISPR-Cas9 system for genome engineering has revolutionized our ability to modify the endogenous DNA sequence of many organisms, including Drosophila This system allows alteration of DNA sequences in situ with single base-pair precision and is now being used for a wide variety of applications. To use the CRISPR system effectively, various design parameters must be considered, including single guide RNA target site selection and identification of successful editing events. Here, we review recent advances in CRISPR methodology in Drosophila and introduce protocols for some of the more difficult aspects of CRISPR implementation: designing and generating CRISPR reagents and detecting indel mutations by high-resolution melt analysis. PMID:27587786

  18. CAS Accelerator Physics (High-Power Hadron Machines) in Spain

    CERN Multimedia

    CAS

    2011-01-01

    The CERN Accelerator School (CAS) and ESS-Bilbao jointly organised a specialised course on High-Power Hadron Machines, held at the Hotel Barceló Nervión in Bilbao, Spain, from 24 May to 2 June, 2011.   CERN Accelerator School students. After recapitulation lectures on the essentials of accelerator physics and review lectures on the different types of accelerators, the programme focussed on the challenges of designing and operating high-power facilities. The particular problems for RF systems, beam instrumentation, vacuum, cryogenics, collimators and beam dumps were examined. Activation of equipment, radioprotection and remote handling issues were also addressed. The school was very successful, with 69 participants of 22 nationalities. Feedback from the participants was extremely positive, praising the expertise and enthusiasm of the lecturers, as well as the high standard and excellent quality of their lectures. In addition to the academic programme, the participants w...

  19. CAS CERN Accelerator School: Cyclotrons, linacs and their applications. Proceedings

    International Nuclear Information System (INIS)

    These proceedings present the lectures given at the eighth specialized course organized by the CERN Accelerator School (CAS), the topic this time being 'Cyclotrons, Linacs and Their Applications'. Following an introductory lecture on linacs, the fundamental features of electron, ion and induction linacs are described together with their RF systems and particle sources. Cyclotrons are then introduced followed by details of their different types, their magnet and RF design, and their injection and extraction systems, with a glance towards exotic and possible future machines. Chapters are then presented on the use of linacs and cyclotrons for medical, fission, fusion and material applications, as well as for isotope production. Finally, descriptions of the design of a radioisotope facility, the matching of accelerators to their task and the computational tools used in their design are included. (orig.)

  20. CAS course on Power Converters in Baden, Switzerland

    CERN Multimedia

    CERN Accelerator School

    2014-01-01

    The CERN Accelerator School (CAS) and the Paul Scherrer Institute (PSI) recently organised a specialised course on Power Converters, which was held at the Hotel du Parc in Baden, Switzerland from 7 to 14 May 2014.   Photo courtesy of Markus Fischer, Paul Scherrer Institut. Following some recapitulation lectures on accelerators and the requirements on power converters, the course covered a wide range of topics related to the different types of power converters needed for particle accelerators. Topical seminars completed the programme. The course was very successful, attended by 84 students representing 21 nationalities, mostly from European countries but also from America, Brazil, Canada, China, Iran, Jordan and Thailand. Feedback from the participants was very positive, reflecting the high standard of the lectures and teaching. In addition to the academic programme, the participants also had an opportunity to take part in a full-day site visit to ABB and PSI and an excursion to the Rhine Fall...

  1. CAS Accelerator Physics (RF for Accelerators) in Denmark

    CERN Multimedia

    Barbara Strasser

    2010-01-01

    The CERN Accelerator School (CAS) and Aarhus University jointly organised a specialised course on RF for Accelerators, at the Ebeltoft Strand Hotel, Denmark from 8 to 17 June 2010.   Caption The challenging programme focused on the introduction of the underlying theory, the study and the performance of the different components involved in RF systems, the RF gymnastics and RF measurements and diagnostics. This academic part was supplemented with three afternoons dedicated to practical hands-on exercises. The school was very successful, with 100 participants representing 25 nationalities. Feedback from the participants was extremely positive, praising the expertise and enthusiasm of the lecturers, as well as the high standard and excellent quality of their lectures. In addition to the academic programme, the participants were able to visit a small industrial exhibition organised by Aarhus University and take part in a one-day excursion consisting of a visit of the accelerators operated ...

  2. CAS course on advanced accelerator physics in Trondheim, Norway

    CERN Multimedia

    CERN Accelerator School

    2013-01-01

    The CERN Accelerator School (CAS) and the Norwegian University of Science and Technology (NTNU) recently organised a course on advanced accelerator physics. The course was held in Trondheim, Norway, from 18 to 29 August 2013. Accommodation and lectures were at the Hotel Britannia and practical courses were held at the university.   The course's format included lectures in the mornings and practical courses in the afternoons. The lecture programme consisted of 32 lectures supplemented by discussion sessions, private study and tutorials. The practical courses provided "hands-on" experience in three topics: RF measurement techniques, beam instrumentation and diagnostics, and optics design and corrections. Participants selected one of the three courses and followed the chosen topic throughout the course. The programme concluded with seminars and a poster session.  70 students representing 21 nationalities were selected from over 90 applicants, with most participa...

  3. CAS CERN Accelerator School. Third advanced accelerator physics course

    International Nuclear Information System (INIS)

    The third version of the CERN Accelerator School's (CAS) advanced course on General Accelerator Physics was given at Uppsala University from 18-29 September, 1989. Its syllabus was based on the previous courses held in Oxford, 1985 and Berlin, 1987 whose proceedings were published as CERN Yellow Reports 87-03 and 89-01 respectively. However, the opportunity was taken to emphasize the physics of small accelerators and storage rings, to present some topics in new ways, and to introduce new seminars. Thus the lectures contained in the present volume include chromaticity, dynamic aperture, kinetic theory, Landau damping, ion-trapping, Schottky noise, laser cooling and small ring lattice problems while the seminars include interpretation of numerical tracking, internal targets and living with radiation. (orig.)

  4. CAS course on Advanced Accelerator Physics in Warsaw

    CERN Multimedia

    CERN Accelerator School

    2015-01-01

    The CERN Accelerator School (CAS) and the National Centre for Nuclear Research (NCBJ) recently organised a course on Advanced Accelerator Physics. The course was held in Warsaw, Poland from 27 September to 9 October 2015.    The course followed an established format with lectures in the mornings and practical courses in the afternoons. The lecture programme consisted of 34 lectures, supplemented by private study, tutorials and seminars. The practical courses provided ‘hands-on’ experience of three topics: ‘Beam Instrumentation and Diagnostics’, ‘RF Measurement Techniques’ and ‘Optics Design and Corrections’. Participants selected one of the three courses and followed their chosen topic throughout the duration of the school. Sixty-six students representing 18 nationalities attended this course, with most participants coming from European counties, but also from South Korea, Taiwan and Russia. Feedback from th...

  5. Hernie de Spiegel: a propos d’un cas

    OpenAIRE

    Majdoub Hassani, Karim Ibn; Zahid, Fatimzohra; Anoune, Hicham; Toughrai, Imane; Laalim, Said Ait; Mazaz, Khalid

    2010-01-01

    Introduction: La hernie de Spiegel ou hernie ventrale latérale est une déhiscence inhabituelle apparaissant sur la ligne ou fascia semi-lunaire de Spiegel. C’est une entité clinique rare, représente 0.10 à 1% des hernies. Aussi, nous a-t-il paru opportun de rapporter ce cas colligé dans le service de chirurgie B du CHU Hassan II de Fès. Patient et observation: Nous rapportons l’observation d’une patiente âgée de 60ans, sans antécédent particulier qui présentais une tuméfaction para ombilicale...

  6. CAS CERN Accelerator School: Fourth general accelerator physics course

    International Nuclear Information System (INIS)

    The fourth CERN Accelerator School (CAS) basic course on General Accelerator Physics was given at KFA, Juelich, from 17 to 28 September 1990. Its syllabus was based on the previous similar courses held at Gif-sur-Yvette in 1984, Aarhus 1986, and Salamanca 1988, and whose proceedings were published as CERN Reports 85-19, 87-10, and 89-05, respectively. However, certain topics were treated in a different way, improved or extended, while new subjects were introduced. All of these appear in the present proceedings, which include lectures or seminars on the history and applications of accelerators, phase space and emittance, chromaticity, beam-beam effects, synchrotron radiation, radiation damping, tune measurement, transition, electron cooling, the designs of superconducting magnets, ring lattices, conventional RF cavities and ring RF systems, and an introduction to cyclotrons. (orig.)

  7. La tuberculose sternale: à propos de 2 cas

    OpenAIRE

    Ouarssani, Aziz; Atoini, Fouad; Ait Lhou, Fatima; Idrissi Rguibi, Mustapha

    2012-01-01

    La tuberculose ostéoarticulaire touche surtout le rachis. L’atteinte sternale est rare, elle représente moins de 1% des ostéomyélites tuberculeuses. L’abcès froid compliqué par une fracture pathologique constitue le mode de révélation le plus fréquent. Le traitement chirurgical assure l’évacuation des abcès et permet le diagnostique histologique et bactériologique. On rapporte deux cas de tuberculose sternale âgés respectivement de 45 ans et de 10ans, colligés au service de pneumologie, l’exa...

  8. Un cas isolé de tuberculose appendiculaire

    OpenAIRE

    Mountassir, Moujahid; Ennafae, Issam; Kechna, Hicham; Berrada, Samir; El Kandry, Siffedine

    2011-01-01

    La tuberculose est une affection qui sévit à l’état endémique dans notre pays, elle demeure un problème majeur de santé publique .la tuberculose appendiculaire primitive est une affection très rare. Nous rapportons le cas d'un patient de sexe masculin âgé de 17 ans, admis au service pour une prise en charge d'une douleur de la fosse iliaque droite évoluant dans un contexte fébrile avec conservation de l’état général posant le diagnostic d'une appendicite aigue. Une appendicectomie a été réali...

  9. CAS course on Intensity Limitations in Particle Beams at CERN

    CERN Multimedia

    CERN Accelerator School

    2015-01-01

    The CERN Accelerator School (CAS) recently organised a specialised course on Intensity Limitations in Particle Beams, at CERN from 2 to 11 November, 2015.     Many accelerators and storage rings, whether intended for particle physics experiments, synchrotron light sources or industrial applications, require beams of high brightness and the highest possible intensities. A good understanding of the possible limitations is required to achieve the desired performance. This course covered the interaction of beams with their surroundings and with other beams, as well as further collective effects. The lectures on the effects and possible mitigations were complemented by tutorials. The course was very successful, with 66 students representing 14 nationalities attending. Most participants came from European counties, but also from Armenia, China and Russia. Feedback from the participants was positive, reflecting the standard of the lectures and teaching. In addition to the academic pro...

  10. Interstellar and ejecta dust in the cas a supernova remnant

    International Nuclear Information System (INIS)

    Infrared continuum observations provide a means of investigating the physical composition of the dust in the ejecta and swept up medium of the Cas A supernova remnant (SNR). Using low-resolution Spitzer IRS spectra (5-35 μm), and broad-band Herschel PACS imaging (70, 100, and 160 μm), we identify characteristic dust spectra, associated with ejecta layers that underwent distinct nuclear burning histories. The most luminous spectrum exhibits strong emission features at ∼9 and 21 μm and is closely associated with ejecta knots with strong Ar emission lines. The dust features can be reproduced by magnesium silicate grains with relatively low Mg to Si ratios. Another dust spectrum is associated with ejecta having strong Ne emission lines. It has no indication of any silicate features and is best fit by Al2O3 dust. A third characteristic dust spectrum shows features that are best matched by magnesium silicates with a relatively high Mg to Si ratio. This dust is primarily associated with the X-ray-emitting shocked ejecta, but it is also evident in regions where shocked interstellar or circumstellar material is expected. However, the identification of dust composition is not unique, and each spectrum includes an additional featureless dust component of unknown composition. Colder dust of indeterminate composition is associated with emission from the interior of the SNR, where the reverse shock has not yet swept up and heated the ejecta. Most of the dust mass in Cas A is associated with this unidentified cold component, which is ≲ 0.1 M ☉. The mass of warmer dust is only ∼0.04 M ☉.

  11. Folate Deficiency in Chronic Pancreatitis

    Directory of Open Access Journals (Sweden)

    Gopalakrishna Rajesh

    2010-07-01

    Full Text Available Dear Sir, While there has been a spurt of interest in genetic alterations associated with pancreatitis in the past few years, interest in the role of environmental factors has largely focused on alcoholism and smoking with insufficient attention being paid to the contributions of nutritional deficiency, and the role of environmental toxins in the pathogenesis of pancreatitis. Braganza and Dormandy [1] argue convincingly about the role played by cytochrome P450 monooxygenases (especially CYP1A enzyme induction by xenobiotics and the resultant oxidative stress, as also the now increasingly recognized reductive stress posed by the metabolites in initiating pancreatic injury. Their article underlines the important part played by the deficiency of methyl and thiol molecules in different stages of the progression of pancreatic damage. Furthermore, they attempt to establish a link between environmental and genetic factors and bring in a holistic view on the etiopathogenesis of chronic pancreatitis. We have recently demonstrated lower plasma methionine levels in two cohorts of chronic pancreatitis patients; one of tropical chronic pancreatitis and the other, of alcoholic chronic pancreatitis as compared to healthy controls [2] which suggests that deficiency of methyl groups may be a factor in various forms of pancreatitis. Similarly, we have shown lower red cell glutathione levels in chronic pancreatitis patients with tropical chronic pancreatitis and alcoholic chronic pancreatitis, indicating deficiency of thiol molecules. In addition, we have demonstrated significantly higher levels of plasma total homocysteine in chronic pancreatitis patients than in healthy controls. Moreover, our study has shown that there is a deficiency of red cell folate in the majority of chronic pancreatitis patients, more so in tropical chronic pancreatitis; and that folate deficiency appeared to be the key factor in hyperhomocysteinemia in chronic pancreatitis patients

  12. Structural and dynamic views of the CRISPR-Cas system at the single-molecule level.

    Science.gov (United States)

    Lee, Seung Hwan; Bae, Sangsu

    2016-04-01

    The CRISPR-Cas system has emerged as a fascinating and important genome editing tool. It is now widely used in biology, biotechnology, and biomedical research in both academic and industrial settings. To improve the specificity and efficiency of Cas nucleases and to extend the applications of these systems for other areas of research, an understanding of their precise working mechanisms is crucial. In this review, we summarize current studies on the molecular structures and dynamic functions of type I and type II Cas nucleases, with a focus on target DNA searching and cleavage processes as revealed by single-molecule observations. [BMB Reports 2016; 49(4): 201-207]. PMID:26923305

  13. Overcoming doxorubicin resistance of cancer cells by Cas9-mediated gene disruption

    OpenAIRE

    Jong Seong Ha; Juyoung Byun; Dae-Ro Ahn

    2016-01-01

    In this study, Cas9 system was employed to down-regulate mdr1 gene for overcoming multidrug resistance of cancer cells. Disruption of the MDR1 gene was achieved by delivery of the Cas9-sgRNA plasmid or the Cas9-sgRNA ribonucleoprotein complex using a conventional gene transfection agent and protein transduction domain (PTD). Doxorubicin showed considerable cytotoxicity to the drug-resistant breast cancer cells pre-treated with the RNA-guided endonuclease (RGEN) systems, whereas virtually non-...

  14. Asymmetric parental genome engineering by Cas9 during mouse meiotic exit

    OpenAIRE

    Toru Suzuki; Maki Asami; Perry, Anthony C. F.

    2014-01-01

    Mammalian genomes can be edited by injecting pronuclear embryos with Cas9 cRNA and guide RNA (gRNA) but it is unknown whether editing can also occur during the onset of embryonic development, prior to pronuclear embryogenesis. We here report Cas9-mediated editing during sperm-induced meiotic exit and the initiation of development. Injection of unfertilized, mouse metaphase II (mII) oocytes with Cas9 cRNA, gRNA and sperm enabled efficient editing of transgenic and native alleles. Pre-loading o...

  15. DNA-binding domain fusions enhance the targeting range and precision of Cas9

    OpenAIRE

    Bolukbasi, Mehmet Fatih; Gupta, Ankit; Oikemus, Sarah; Derr, Alan G.; Garber, Manuel; Brodsky, Michael H.; Zhu, Lihua Julie; Scot A Wolfe

    2015-01-01

    The CRISPR-Cas9 system is commonly employed in biomedical research; however, the precision of Cas9 is sub-optimal for gene therapy applications that involve editing a large population of cells. Variations on the standard Cas9 system have yielded improvements in the precision of targeted DNA cleavage, but often restrict the range of targetable sequences. It remains unclear whether these variants can limit lesions to a single site within the human genome over a large cohort of treated cells. He...

  16. Comparison of CRISPR/Cas9 expression constructs for efficient targeted mutagenesis in rice

    OpenAIRE

    Mikami, Masafumi; Toki, Seiichi; Endo, Masaki

    2015-01-01

    The CRISPR/Cas9 system is an efficient tool used for genome editing in a variety of organisms. Despite several recent reports of successful targeted mutagenesis using the CRISPR/Cas9 system in plants, in each case the target gene of interest, the Cas9 expression system and guide-RNA (gRNA) used, and the tissues used for transformation and subsequent mutagenesis differed, hence the reported frequencies of targeted mutagenesis cannot be compared directly. Here, we evaluated mutation frequency i...

  17. Editing of mouse and human immunoglobulin genes by CRISPR-Cas9 system

    OpenAIRE

    Cheong, Taek-Chin; Compagno, Mara; Chiarle, Roberto

    2016-01-01

    Applications of the CRISPR-Cas9 system to edit the genome have widely expanded to include DNA gene knock-out, deletions, chromosomal rearrangements, RNA editing and genome-wide screenings. Here we show the application of CRISPR-Cas9 technology to edit the mouse and human immunoglobulin (Ig) genes. By delivering Cas9 and guide-RNA (gRNA) with retro- or lenti-virus to IgM+ mouse B cells and hybridomas, we induce class-switch recombination (CSR) of the IgH chain to the desired subclass. Similarl...

  18. Efficient fdCas9 Synthetic Endonuclease with Improved Specificity for Precise Genome Engineering

    OpenAIRE

    Aouida, Mustapha; Eid, Ayman; Ali, Zahir; Cradick, Thomas; Lee, Ciaran; Deshmukh, Harshavardhan; Atef, Ahmed; AbuSamra, Dina; Gadhoum, Samah Zeineb; Merzaban, Jasmeen; Bao, Gang; Mahfouz, Magdy

    2015-01-01

    The Cas9 endonuclease is used for genome editing applications in diverse eukaryotic species. A high frequency of off-target activity has been reported in many cell types, limiting its applications to genome engineering, especially in genomic medicine. Here, we generated a synthetic chimeric protein between the catalytic domain of the FokI endonuclease and the catalytically inactive Cas9 protein (fdCas9). A pair of guide RNAs (gRNAs) that bind to sense and antisense strands with a defined spac...

  19. Multiplex metabolic pathway engineering using CRISPR/Cas9 in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Jakociunas, Tadas; Bonde, Ida; Herrgard, Markus;

    2015-01-01

    CRISPR/Cas9 is a simple and efficient tool for targeted and marker-free genome engineering. Here, we report the development and successful application of a multiplex CRISPR/Cas9 system for genome engineering of up to 5 different genomic loci in one transformation step in baker's yeast Saccharomyces...... cerevisiae. To assess the specificity of the tool we employed genome re-sequencing to screen for off-target sites in all single knock-out strains targeted by different gRNAs. This extensive analysis identified no more genome variants in CRISPR/Cas9 engineered strains compared to wild-type reference strains...

  20. CRISPR/Cas9-Mediated Genome Editing in Soybean Hairy Roots

    OpenAIRE

    Yupeng Cai; Li Chen; Xiujie Liu; Shi Sun; Cunxiang Wu; Bingjun Jiang; Tianfu Han; Wensheng Hou

    2015-01-01

    As a new technology for gene editing, the CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) system has been rapidly and widely used for genome engineering in various organisms. In the present study, we successfully applied type II CRISPR/Cas9 system to generate and estimate genome editing in the desired target genes in soybean (Glycine max (L.) Merrill.). The single-guide RNA (sgRNA) and Cas9 cassettes were assembled on one vector to improve transformat...