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Sample records for action study cas

  1. CRISPR-Cas adaptation: insights into the mechanism of action.

    PubMed

    Amitai, Gil; Sorek, Rotem

    2016-02-01

    Since the first demonstration that CRISPR-Cas systems provide bacteria and archaea with adaptive immunity against phages and plasmids, numerous studies have yielded key insights into the molecular mechanisms governing how these systems attack and degrade foreign DNA. However, the molecular mechanisms underlying the adaptation stage, in which new immunological memory is formed, have until recently represented a major unresolved question. In this Progress article, we discuss recent discoveries that have shown both how foreign DNA is identified by the CRISPR-Cas adaptation machinery and the molecular basis for its integration into the chromosome to form an immunological memory. Furthermore, we describe the roles of each of the specific CRISPR-Cas components that are involved in memory formation, and consider current models for their evolutionary origin.

  2. actions immunoallergiques graves aux antibacillaires: à propos de 10 cas

    PubMed Central

    Alami, Sabah El Machichi; Hammi, Sanae; Bourkadi, Jamal Eddine

    2014-01-01

    L'hypersensibilité aux antituberculeux est l'un des effets secondaires imprévisibles qui apparait chez 4 à 5 % de la population exposée et s’élève à 25% chez les sujets VIH positifs. Dans notre étude parmi 39 patients ayant présenté des réactions immunoallergiques, 10 avaient des formes graves. Le délai moyen d'apparition des signes était de 23 jours. Les réactions immunoallergiques observées étaient 5 cas de toxidermie généralisée fébrile, un cas de Dress syndrome, un cas de neutropénie, un cas de pancitopénie et 2 cas de thrombopénie. Tous nos patients avaient bien évolué cliniquement et bactériologiquement après l'adoption d'un régime thérapeutique excluant le ou les médicaments incriminés. En pratique, si l'effet indésirable imputé à un antituberculeux est grave, il est impératif de l'arrêter, de traiter l'incident et d'associer une autre molécule chez certains cas. Notre étude a montré une fréquence significative des complications graves probablement sous-estimée, surtout dans les pays fortement touchés par l'infection HIV.

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

    ERIC Educational Resources Information Center

    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…

  4. The CAS Classroom

    ERIC Educational Resources Information Center

    Garner, Sue

    2004-01-01

    The Victorian Curriculum and Assessment Authority (VCAA) Computer Algebra System (CAS)Pilot study (2001-2005) is monitoring the use of CAS in senior secondary mathematics. This article explores the author's experiences in the CAS classroom and delineates changes in teaching style, as a result of the introduction of CAS into the senior mathematics…

  5. NASA Controller Acceptability Study 1(CAS-1) Experiment Description and Initial Observations

    NASA Technical Reports Server (NTRS)

    Chamberlain, James P.; Consiglio, Maria C.; Comstock, James R., Jr.; Ghatas, Rania W.; Munoz, Cesar

    2015-01-01

    This paper describes the Controller Acceptability Study 1 (CAS-1) experiment that was conducted by NASA Langley Research Center personnel from January through March 2014 and presents partial CAS-1 results. CAS-1 employed 14 air traffic controller volunteers as research subjects to assess the viability of simulated future unmanned aircraft systems (UAS) operating alongside manned aircraft in moderate-density, moderate-complexity Class E airspace. These simulated UAS were equipped with a prototype pilot-in-the-loop (PITL) Detect and Avoid (DAA) system, specifically the Self-Separation (SS) function of such a system based on Stratway+ software to replace the see-and-avoid capabilities of manned aircraft pilots. A quantitative CAS-1 objective was to determine horizontal miss distance (HMD) values for SS encounters that were most acceptable to air traffic controllers, specifically HMD values that were assessed as neither unsafely small nor disruptively large. HMD values between 0.5 and 3.0 nautical miles (nmi) were assessed for a wide array of encounter geometries between UAS and manned aircraft. The paper includes brief introductory material about DAA systems and their SS functions, followed by descriptions of the CAS-1 simulation environment, prototype PITL SS capability, and experiment design, and concludes with presentation and discussion of partial CAS-1 data and results.

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

    NASA Astrophysics Data System (ADS)

    Bakış, V.; Bakış, H.; Bilir, S.; Eker, Z.

    2016-09-01

    An early-type, massive, short-period (Porb=2d.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 M1=19.04± 0.07 M⊙, M2=13.73± 0.05 M⊙ and R1=7.22± 0.06 R⊙, R2=5.79± 0.06 R⊙, respectively. Both components present synchronous rotation (Vrot1=160 km s-1, Vrot2=130 km s-1) with their orbit. Orbital period analysis yielded a physically bound additional component in the system with a minimum mass of M3=0.88 M⊙ orbiting in an eccentric orbit (e = 0.37 ± 0.2) with an orbital period of P 12 = 42 ± 9 yr. High precision absolute parameters of the system allowed us to derive a distance to DN Cas as 1.7 ± 0.2 kpc which locates the system within the borders of the Cas OB6 association (d = 1.8 kpc). The space velocities and the age of DN Cas are in agreement with those of Cas OB6. The age of DN Cas (τ = 3-5 Myr) is found to be 1-2 Myr older than the embedded clusters (IC 1795, IC 1805, and IC 1848) in the Cas OB6 association, which implies a sequential star formation in the association.

  7. Spectroscopic studies of the small-amplitude Cepheid SU Cas

    NASA Astrophysics Data System (ADS)

    Usenko, I. A.; Klochkova, V. G.; Tavolzhanskaya, N. S.

    2013-09-01

    A new set of 16 high-resolution spectra for the small-amplitude Cepheid SU Cas obtained in 2007-2009 has allowed us to determine its atmospheric parameters ( T eff = 6345 ± 30 K, log g = 2.40, V t = 3.25 km s-1) and to measure its radial velocities. The latter were added to the general list of radial velocities (375 estimates) obtained in the last 90 years. Using a frequency analysis, we have refined the pulsation and orbital periods of the Cepheid. Apart from the well-known fundamental pulsation period [Figure not available: see fulltext.], we have detected a possible secondary period of [Figure not available: see fulltext.]. Their ratio of 0.96 suggests the existence of nonradial pulsations in the Cepheid's atmosphere. Based on photoelectric photometry in the last 60 years, we have shown that the effective temperature undergoes cyclic secular changes of ±200 K with an unknown period. The mean effective temperature T eff = 6395 ± 52 K estimated from photometric data agrees well with our estimate from spectroscopic data. The variations of the mean color index, effective temperature, and γ-velocity (in 90 years of observations) point to a possible orbital motion of the well-known hot companion with the most probable periods of [Figure not available: see fulltext.], [Figure not available: see fulltext.], and [Figure not available: see fulltext.]. The elemental abundances in the atmosphere of SU Cas confirm the conclusion that this Cepheid is a typical yellow supergiant after the first dredge-up. Our T eff estimate gives a radius of 32 R ⊙ and a distance of 455 pc for it, which is inconsistent with its membership in the open cluster Alessi 95. The question about the pulsation mode of SU Cas still remains open.

  8. Spectroscopic studies of three Cepheids with high positive pulsation period increments: SZ Cas, BY Cas, and RU Sct

    NASA Astrophysics Data System (ADS)

    Usenko, I. A.; Klochkova, V. G.

    2015-07-01

    Three high-resolution spectra have been taken at different times with the 6-m SAO RAS telescope (LYNX and PFES spectrographs) for three Cepheids exhibiting high positive period increments: the small-amplitude (DCEPS) SZ Cas and BY Cas and the classical (DCEP) RU Sct. SZ Cas and RU Sct are members of the Galactic open clusters χ and h Per and Trump 35, respectively. Analysis of the spectra has shown that the interstellar Na I D1 and D2 lines in all objects are considerably stronger than the atmospheric ones and are redshifted in SZ Cas and BY Cas and blushifted in RU Sct. The core of the H α absorption line in BY Cas has an asymmetric knifelike shape, while RU Sct exhibits an intense emission in the blue wing of this line. Such phenomena are observed in long-period Cepheids and bright hypergiants with an extended envelope. In this case, the strong Mg Ib 5183.62 Å and Ba II 5853.67, 6141.713, and 6496.90 Å lines with low χlow in SZ Cas and RU Sct also show characteristic knifelike profiles with an asymmetry in the red region, while the Ba II 4934.095 Å line shows similar profiles in the blue one. The absorption lines of neutral atoms and singly ionized metals with different lowerlevel excitation potentials exhibit different degrees of asymmetry: from a pronounced one with secondary components in BY Cas (similar to those in the small-amplitude Cepheid BG Cru pulsating in the first overtone and having an envelope) to its insignificance or virtual absence in SZ Cas and RU Sct. Analysis of the secular changes in mean T eff determined from photometric color indices and spectra over the last 55 years for these stars has revealed periodic fluctuations of 200 K for SZ Cas and BY Cas and 500 K for RU Sct. For SZ Cas and RU Sct, T eff determined in some years from some color indices show much lower values, which together with the temperature fluctuations can be associated with mass loss and dust formation. Based on these facts, we hypothesize the existence of

  9. CRISPR/Cas9 as tool for functional study of genes involved in preimplantation embryo development.

    PubMed

    Kwon, Jeongwoo; Namgoong, Suk; Kim, Nam-Hyung

    2015-01-01

    The CRISPR/Cas9 system has proven to be an efficient gene-editing tool for genome modification of cells and organisms. However, the applicability and efficiency of this system in pig embryos have not been studied in depth. Here, we aimed to remove porcine OCT4 function as a model case using the CRISPR/Cas9 system. Injection of Cas9 and single-guide RNA (sgRNA) against OCT4 decreased the percentages of OCT4-positive embryos to 37-50% of total embryos, while ~100% of control embryos exhibited clear OCT4 immunostaining. We assessed the mutation status near the guide sequence using polymerase chain reaction (PCR) and DNA sequencing, and a portion of blastocysts (20% in exon 2 and 50% in exon 5) had insertions/deletions near protospacer-adjacent motifs (PAMs). Different target sites had frequent deletions, but different concentrations of sgRNA made no impact. OCT4 mRNA levels dramatically decreased at the 8-cell stage, and they were barely detectable in blastocysts, while mRNA levels of other genes, including NANOG, and CDX2 were not affected. In addition, the combination of two sgRNAs led to large-scale deletion (about 1.8 kb) in the same chromosome. Next, we injected an enhanced green fluorescent protein (eGFP) vector targeting the OCT4 exon with Cas9 and sgRNA to create a knockin. We confirmed eGFP fluorescence in blastocysts in the inner cell mass, and also checked the mutation status using PCR and DNA sequencing. A significant portion of blastocysts had eGFP sequence insertions near PAM sites. The CRISPR/CAS9 system provides a good tool for gene functional studies by deleting target genes in the pig. PMID:25775469

  10. CLIM-RUN: Tourism cas study over the French Alps

    NASA Astrophysics Data System (ADS)

    Dubois, C.

    2012-04-01

    Climate information for societal use has becoming a major challenge for tourism management and adaptation in a context of strong climate variability and change. Within the CLIMRUN EU FP7, a case study on summer tourism in the French Alps has been identify. I will introduce the bottom-up approach use in the project where stakeholders and local users meet with climate experts. From those meetings, they thus identify the climate dependence and information which impact their summer activities over this region. All the activities are located in a mountainous region where outdoor leisure is the main economic driver of the region. It has emerged that the climate requirements are as well on past as on future climate information. On one side, the past climate parameters are found to be an invaluable information to evaluate the climate dependence of the different activities. A better knowledge as well as a growing interest in climate variability has been express to quantify the climate dependence on their activities. On the other side, the future climate information requested mainly on seasonal to decadal timescale. A particular interest has been express on the snow cover at the end of the winter season, evolution of heavy precipitations, heatwave, air temperatures and well as the water temperature of the mountainous lakes. Those climate variables are used to create comfort index under climate change. All those targeted climate information are based on on-going projects as well as future model development.

  11. Final corrective action study for the former CCC/USDA facility in Ramona, Kansas.

    SciTech Connect

    LaFreniere, L. M.

    2011-04-20

    Past operations at a grain storage facility formerly leased and operated by the Commodity Credit Corporation of the U.S. Department of Agriculture (CCC/USDA) in Ramona, Kansas, resulted in low concentrations of carbon tetrachloride in groundwater that slightly exceed the regulatory standard in only one location. As requested by the Kansas Department of Health and Environment, the CCC/USDA has prepared a Corrective Action Study (CAS) for the facility. The CAS examines corrective actions to address groundwater impacted by the former CCC/USDA facility but not releases caused by other potential groundwater contamination sources in Ramona. Four remedial alternatives were considered in the CAS. The recommended remedial alternative in the CAS consists of Environmental Use Control to prevent the inadvertent use of groundwater as a water supply source, coupled with groundwater monitoring to verify the continued natural improvement in groundwater quality. The Commodity Credit Corporation of the U.S. Department of Agriculture (CCC/USDA) has directed Argonne National Laboratory to prepare a Corrective Action Study (CAS), consistent with guidance from the Kansas Department of Health and Environment (KDHE 2001a), for the CCC/USDA grain storage facility formerly located in Ramona, Kansas. This effort is pursuant to a KDHE (2007a) request. Although carbon tetrachloride levels at the Ramona site are low, they remain above the Kansas Tier 2 risk-based screening level (RBSL) and the U.S. Environmental Protection Agency (EPA) maximum contaminant level (MCL) of 5 {micro}g/L (Kansas 2003, 2004). In its request for the CAS, the KDHE (2007a) stated that, because of these levels, risk is associated with potential future exposure to contaminated groundwater. The KDHE therefore determined that additional measures are warranted to limit future use of the property and/or exposure to contaminated media as part of site closure. The KDHE further requested comparison of at least two corrective

  12. CRISPR/Cas9 Promotes Functional Study of Testis Specific X-Linked Gene In Vivo

    PubMed Central

    Jiang, Xue; Chen, Yuxi; Zhang, Zhen; Zhang, Xiya; Liang, Puping; Zhan, Shaoquan; Cao, Shanbo; Songyang, Zhou; Huang, Junjiu

    2015-01-01

    Mammalian spermatogenesis is a highly regulated multistage process of sperm generation. It is hard to uncover the real function of a testis specific gene in vitro since the in vitro model is not yet mature. With the development of the CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated 9) system, we can now rapidly generate knockout mouse models of testis specific genes to study the process of spermatogenesis in vivo. SYCP3-like X-linked 2 (SLX2) is a germ cell specific component, which contains a Cor1 domain and belongs to the XLR (X-linked, lymphocyte regulated) family. Previous studies suggested that SLX2 might play an important role in mouse spermatogenesis based on its subcellular localization and interacting proteins. However, the function of SLX2 in vivo is still elusive. Here, to investigate the functions of SLX2 in spermatogenesis, we disrupted the Slx2 gene by using the CRISPR/Cas9 system. Since Slx2 is a testis specific X-linked gene, we obtained knockout male mice in the first generation and accelerated the study process. Compared with wild-type mice, Slx2 knockout mice have normal testis and epididymis. Histological observation of testes sections showed that Slx2 knockout affected none of the three main stages of spermatogenesis: mitosis, meiosis and spermiogenesis. In addition, we further confirmed that disruption of Slx2 did not affect the number of spermatogonial stem cells, meiosis progression or XY body formation by immunofluorescence analysis. As spermatogenesis was normal in Slx2 knockout mice, these mice were fertile. Taken together, we showed that Slx2 itself is not an essential gene for mouse spermatogenesis and CRISPR/Cas9 technique could speed up the functional study of testis specific X-linked gene in vivo. PMID:26599493

  13. Photometric Study of Two Recently Discovered Variable Stars in the Field of BS Cas

    NASA Astrophysics Data System (ADS)

    Virnina, Natalia A.; Kocián, Radek; H´lek, Lubomír; Dubovsky, Pavol; Andronov, Ivan L.; Kudzej, Igor

    Two recently discovered variable stars (CzeV134 = GSC 3682 0018 = USNO-A2.0 1425-1870026 and CzeV135 = GSC 3682 2051 = USNO-A2.0 1425-1825909 = V1094 Cas), which have been identified in the field of the W UMa variable star BS Cas, are studied in the present paper. The phase curves and finding charts for these stars are presented. The ephemeris and other photometric parameters were computed. The phenomenological features indicate that the first star (CzeV134) is probably a low-amplitude RRc Lyrae - type variable star with the period P = 0.419794±0.000029 d and the initial epoch T0 = HJD2453236.50412±0.00056. The amplitude and the shape of the light curve are variable possibly indicating the Blazhko phenomenon. The second star (CzeV135) was classified as an EW-type binary system of subtype A. However, a β Lyrae type may not be excluded, as various classification parameters lie in a range of overlapping values for both classes. The period P=0.51429090±0.00000012 d and the initial epoch T0 = HJD2454543.7920±0.0006. The O'Connell effect is clearly visible. There are slight changes of this effect, noticeable while comparing different seasons of observations. O-C diagrams for these stars were analyzed.

  14. Attitude and CAS Use in Senior Secondary Mathematics: A Case Study of Seven Year 11 Students

    ERIC Educational Resources Information Center

    Cameron, Scott; Ball, Lynda

    2014-01-01

    This paper investigates the possible influence of attitude on seven Year 11 students' use of a Computer Algebra System (CAS) during a class activity where students could choose to use CAS or pen-and-paper in solving a range of problems. Investigation of anxiety, confidence, liking and usefulness through a survey and interview revealed that these…

  15. Using CRISPR/Cas to study gene function and model disease in vivo.

    PubMed

    Tschaharganeh, Darjus F; Lowe, Scott W; Garippa, Ralph J; Livshits, Geulah

    2016-09-01

    The recent discovery of the CRISPR/Cas system and repurposing of this technology to edit a variety of different genomes have revolutionized an array of scientific fields, from genetics and translational research, to agriculture and bioproduction. In particular, the prospect of rapid and precise genome editing in laboratory animals by CRISPR/Cas has generated an immense interest in the scientific community. Here we review current in vivo applications of CRISPR/Cas and how this technology can improve our knowledge of gene function and our understanding of biological processes in animal models. PMID:27149548

  16. Excited-State Photolytic Mechanism of Cyclopentene Containing a Group 14 Element: An MP2-CAS//CASSCF Study.

    PubMed

    Su, Ming-Der

    2015-08-13

    The potential energy surfaces corresponding to the photolytic reactions of 1,2-dimethyl-cyclopentene, 3,4-dimethyl-silacyclopent-3-ene, and 3,4-dimethyl-germacyclopent-3-ene were investigated by employing the CAS(6,6)/6-311G(d) and MP2-CAS-(6,6)/6-311++G(3df,3pd)//CAS(6,6)/6-311G(d) methods. Also, six kinds of substituted germacyclopent-3-ene were used as model reactants by way of the CASSCF and MP2-CAS methods to study their photolytic mechanisms. The theoretical findings indicate that the photolysis of the above reactants all adopt the same reaction path as follows: reactant → Franck-Condon region → conical intersection → germylene and 1,3-butadiene. However, the theoretical results demonstrate that no photolysis ((1)(π →π*)) can be observed in the 1,2-dimethyl-cyclopentene system. Above all, the theoretical investigations strongly suggest that both steric effects, originating from the bulky substituents, and the atomic radius of the group 14 element (C, Si, and Ge) play a crucial role in determining the cis/trans selectivity of the conformation of 1,3-butadiene during their photolytic reactions. PMID:26204240

  17. Application of CRISPR/Cas9 genome editing to the study and treatment of disease.

    PubMed

    Pellagatti, Andrea; Dolatshad, Hamid; Valletta, Simona; Boultwood, Jacqueline

    2015-07-01

    CRISPR/Cas is a microbial adaptive immune system that uses RNA-guided nucleases to cleave foreign genetic elements. The CRISPR/Cas9 method has been engineered from the type II prokaryotic CRISPR system and uses a single-guide RNA to target the Cas9 nuclease to a specific genomic sequence. Cas9 induces double-stranded DNA breaks which are repaired either by imperfect non-homologous end joining to generate insertions or deletions (indels) or, if a repair template is provided, by homology-directed repair. Due to its specificity, simplicity and versatility, the CRISPR/Cas9 system has recently emerged as a powerful tool for genome engineering in various species. This technology can be used to investigate the function of a gene of interest or to correct gene mutations in cells via genome editing, paving the way for future gene therapy approaches. Improvements to the efficiency of CRISPR repair, in particular to increase the rate of gene correction and to reduce undesired off-target effects, and the development of more effective delivery methods will be required for its broad therapeutic application.

  18. Visualization analysis of CRISPR/Cas9 gene editing technology studies*

    PubMed Central

    Du, Quan-sheng; Cui, Jie; Zhang, Chun-jie; He, Ke

    2016-01-01

    The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system is an adaptive immune defense system that resists the invasion of viruses and plasmids heterologous genetic material in bacteria and archaea. Taking the literature related to gene editing technology of CRISPR/Cas9 from the Web of Science database from 2002 to 2015, we use the software CiteSpaceV to analyze co-cited literature in order to establish the research hotspots and fronts recently in this field by knowledge mapping. PMID:27704749

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

    PubMed Central

    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 “Multiplexing of Cas9 at Artificial Loci”—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

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

    PubMed Central

    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

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

    PubMed

    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

  2. Corrective Action Investigation Plan for Corrective Action Unit 550: Smoky Contamination Area Nevada National Security Site, Nevada, Revision 0

    SciTech Connect

    Grant Evenson

    2012-05-01

    Corrective Action Unit (CAU) 550 is located in Areas 7, 8, and 10 of the Nevada National Security Site, which is approximately 65 miles northwest of Las Vegas, Nevada. CAU 550, Smoky Contamination Area, comprises 19 corrective action sites (CASs). Based on process knowledge of the releases associated with the nuclear tests and radiological survey information about the location and shape of the resulting contamination plumes, it was determined that some of the CAS releases are co-located and will be investigated as study groups. This document describes the planned investigation of the following CASs (by study group): (1) Study Group 1, Atmospheric Test - CAS 08-23-04, Atmospheric Test Site T-2C; (2) Study Group 2, Safety Experiments - CAS 08-23-03, Atmospheric Test Site T-8B - CAS 08-23-06, Atmospheric Test Site T-8A - CAS 08-23-07, Atmospheric Test Site T-8C; (3) Study Group 3, Washes - Potential stormwater migration of contaminants from CASs; (4) Study Group 4, Debris - CAS 08-01-01, Storage Tank - CAS 08-22-05, Drum - CAS 08-22-07, Drum - CAS 08-22-08, Drums (3) - CAS 08-22-09, Drum - CAS 08-24-03, Battery - CAS 08-24-04, Battery - CAS 08-24-07, Batteries (3) - CAS 08-24-08, Batteries (3) - CAS 08-26-01, Lead Bricks (200) - CAS 10-22-17, Buckets (3) - CAS 10-22-18, Gas Block/Drum - CAS 10-22-19, Drum; Stains - CAS 10-22-20, Drum - CAS 10-24-10, Battery. These sites are being investigated because existing information on the nature and extent of potential contamination is insufficient to evaluate and recommend corrective action alternatives (CAAs). Additional information will be obtained by conducting a corrective action investigation before evaluating CAAs and selecting the appropriate corrective action for each study group. The results of the field investigation will support a defensible evaluation of viable CAAs that will be presented in the Corrective Action Decision Document. The sites will be investigated based on the data quality objectives (DQOs) developed

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

    PubMed

    Isaac, R Stefan; Jiang, Fuguo; Doudna, Jennifer A; Lim, Wendell A; Narlikar, Geeta J; Almeida, Ricardo

    2016-04-28

    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 Cas9 is variable over several orders of magnitude depending on dynamic properties of the DNA sequence and the distance of the PAM site from the nucleosome dyad. We further find that chromatin remodeling enzymes stimulate Cas9 activity on nucleosomal templates. Our findings imply that the spontaneous breathing of nucleosomal DNA together with the action of chromatin remodelers allow Cas9 to effectively act on chromatin in vivo.

  4. Following the Action in Action Learning: Towards Ethnomethodological Studies of (Critical) Action Learning

    ERIC Educational Resources Information Center

    Fox, Steve

    2009-01-01

    Action learning is a pedagogical practice that helps participants learn by talking about their workplace action with fellow participants ("comrades in adversity") in their action learning set. This paper raises questions about the action in action learning, such as: how do members of an action learning set learn from and through each other? How do…

  5. Genome modification by CRISPR/Cas9.

    PubMed

    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.

  6. Action Learning in ActionAid Nepal: A Case Study

    ERIC Educational Resources Information Center

    Lustig, Patricia; Rai, Deep Ranjani

    2009-01-01

    This article describes an example of how action learning was used as a framework for an organisational intervention to fundamentally change the organisational culture over a period of time. It also identifies our learning over that period of time and what worked well (and not so well) in an International Non-Governmental Organisation in Nepal.

  7. Validation of Subject Areas of CAS Professional Studies Standards for Master's Level Student Affairs Professional Preparation Programs

    ERIC Educational Resources Information Center

    Young, Dallin George; Dean, Laura A.

    2015-01-01

    The standards for Master's Level Student Affairs Professional Preparation Programs, first published in 1986, were among the first standards published by the Council for the Advancement of Standards in Higher Education (CAS; Ebbers & Kruempel, 1992). With the latest revision in 2012, the CAS standards for preparation of student affairs…

  8. Understanding communicative actions: a repetitive TMS study.

    PubMed

    Stolk, Arjen; Noordzij, Matthijs L; Volman, Inge; Verhagen, Lennart; Overeem, Sebastiaan; van Elswijk, Gijs; Bloem, Bas; Hagoort, Peter; Toni, Ivan

    2014-02-01

    Despite the ambiguity inherent in human communication, people are remarkably efficient in establishing mutual understanding. Studying how people communicate in novel settings provides a window into the mechanisms supporting the human competence to rapidly generate and understand novel shared symbols, a fundamental property of human communication. Previous work indicates that the right posterior superior temporal sulcus (pSTS) is involved when people understand the intended meaning of novel communicative actions. Here, we set out to test whether normal functioning of this cerebral structure is required for understanding novel communicative actions using inhibitory low-frequency repetitive transcranial magnetic stimulation (rTMS). A factorial experimental design contrasted two tightly matched stimulation sites (right pSTS vs left MT+, i.e., a contiguous homotopic task-relevant region) and tasks (a communicative task vs a visual tracking task that used the same sequences of stimuli). Overall task performance was not affected by rTMS, whereas changes in task performance over time were disrupted according to TMS site and task combinations. Namely, rTMS over pSTS led to a diminished ability to improve action understanding on the basis of recent communicative history, while rTMS over MT+ perturbed improvement in visual tracking over trials. These findings qualify the contributions of the right pSTS to human communicative abilities, showing that this region might be necessary for incorporating previous knowledge, accumulated during interactions with a communicative partner, to constrain the inferential process that leads to action understanding. PMID:24268321

  9. Understanding communicative actions: a repetitive TMS study.

    PubMed

    Stolk, Arjen; Noordzij, Matthijs L; Volman, Inge; Verhagen, Lennart; Overeem, Sebastiaan; van Elswijk, Gijs; Bloem, Bas; Hagoort, Peter; Toni, Ivan

    2014-02-01

    Despite the ambiguity inherent in human communication, people are remarkably efficient in establishing mutual understanding. Studying how people communicate in novel settings provides a window into the mechanisms supporting the human competence to rapidly generate and understand novel shared symbols, a fundamental property of human communication. Previous work indicates that the right posterior superior temporal sulcus (pSTS) is involved when people understand the intended meaning of novel communicative actions. Here, we set out to test whether normal functioning of this cerebral structure is required for understanding novel communicative actions using inhibitory low-frequency repetitive transcranial magnetic stimulation (rTMS). A factorial experimental design contrasted two tightly matched stimulation sites (right pSTS vs left MT+, i.e., a contiguous homotopic task-relevant region) and tasks (a communicative task vs a visual tracking task that used the same sequences of stimuli). Overall task performance was not affected by rTMS, whereas changes in task performance over time were disrupted according to TMS site and task combinations. Namely, rTMS over pSTS led to a diminished ability to improve action understanding on the basis of recent communicative history, while rTMS over MT+ perturbed improvement in visual tracking over trials. These findings qualify the contributions of the right pSTS to human communicative abilities, showing that this region might be necessary for incorporating previous knowledge, accumulated during interactions with a communicative partner, to constrain the inferential process that leads to action understanding.

  10. Social Action As An Objective of Social Studies Instruction.

    ERIC Educational Resources Information Center

    Curtis, Charles K.

    This paper presents a rationale for making social action a major goal of elementary and secondary school social studies education. In addition, it describes social action models, suggests social action approaches appropriate for students at various grade levels, and reviews literature on social action by public school students. Social action is…

  11. Period and light-curve study of the contact eclipsing binary V523 Cas

    NASA Astrophysics Data System (ADS)

    Mohammadi, Mahya; Abedi, Abbas; Riazi, Nematollah

    2016-04-01

    CCD photometry of the eclipsing W Uma binary system V523 Cas in U, B, V and RC filters was carried out during eight nights in 2012. The physical and geometrical parameters of this system are obtained. A possible pulsation period of one of the components is obtained by analyzing the residuals of the ephemeris light curve. Our observations contain 16 times of minimum light. We combined these with all available published times of minimum. By fitting a quadratic curve to the O-C values, a new ephemeris of the system is calculated. By attributing the period change to mass transfer, we find a mass transfer rate of 4 ×10-12 M⊙/yr. Also, Period (80.58 yr) and the minimum mass (0.3 M⊙) of a possible third body is estimated. In addition, the possible existence of a fourth body with a mass of order 0.15 M⊙ is discussed. These third and fourth bodies could be low-mass main-sequence stars (red dwarfs).

  12. Plaque morphology detected with Duplex ultrasound before carotid angioplasty and stenting (CAS) is not a predictor of carotid artery in-stent restenosis, a case control study

    PubMed Central

    2013-01-01

    Background In-stent restenosis (ISR) is an important factor endangering the long-term safety and efficacy of carotid artery angioplasty and stenting (CAS). It is plausible that soft vulnerable plaques are more likely to be injured during CAS procedure and are therefore more likely to initiate the cascade finally leading to ISR. The aim of this study was to investigate if plaque morphology detected by a simple applicable Duplex ultrasound score before CAS can be used as a predictor for ISR. Methods Within a prospectively collected single-centre CAS database of 281 patients (comprising 300 arteries) with high-grade carotid artery stenosis, who underwent CAS between May 2003 and January 2013, we conducted a nested case–control study. Plaque morphology before CAS was analysed by a blinded investigator and each parameter of the Total Plaque Risk Score (TPRS) as well as the whole score was evaluated with regard to its diagnostic validity for ISR. Results We analysed the data of 10 patients with ISR and 50 patients without ISR. There were no significant differences with respect to baseline characteristics, vascular risk factors, and degree of stenosis between patients with and without ISR. The duration of follow-up was longer in patients with ISR (p = 0.024) and these patients were more likely to show increased PSV (p = 0.012) immediately after CAS than patients without ISR. Neither individual parameters of the TPRS score nor the score as a whole were suitable as a diagnostic test for ISR development. Conclusions In the present study we could demonstrate that the non-contrast enhanced DUS of the pre-interventional plaque formation cannot be used as a predictor for the development of ISR. Evaluating a more sophisticated, but not routinely available approach e.g. by ultrasound based plaque perfusion imaging or CT based plaque analysis could be helpful in the future in order to assess the role of plaque morphology in the context of ISR development. PMID:24191865

  13. Effects of Using a Computer Algebra System (CAS) on Junior College Students' Attitudes towards CAS and Achievement in Mathematics

    ERIC Educational Resources Information Center

    Leng, Ng Wee; Choo, Kwee Tiow; Soon, Lau Hock; Yi-Huak, Koh; Sun, Yap Yew

    2005-01-01

    This study examines the effects of using Texas Instruments' Voyage 200 calculator (V200), a graphing calculator with a built-in computer algebra system (CAS), on attitudes towards CAS and achievement in mathematics of junior college students (17 year olds). Students' attitudes towards CAS were examined using a 40-item Likert-type instrument…

  14. Interacting Parallel Constructions of Knowledge in a CAS Context

    ERIC Educational Resources Information Center

    Kidron, Ivy; Dreyfus, Tommy

    2010-01-01

    We consider the influence of a CAS context on a learner's process of constructing a justification for the bifurcations in a logistic dynamical process. We describe how instrumentation led to cognitive constructions and how the roles of the learner and the CAS intertwine, especially close to the branching and combining of constructing actions. The…

  15. The Impact on Student Achievement of When CAS Technology Is Introduced

    ERIC Educational Resources Information Center

    Driver, David

    2012-01-01

    When a Computer Algebra System (CAS) is used as a pedagogical and functional tool in class and as a functional tool in exams, its effect on student achievement can be quite profound. The timing of when students are first introduced to a CAS has an impact on gains in student achievement. In this action research project, the CAS calculator was…

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

    PubMed

    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

  17. [CAS General Standards 2012

    ERIC Educational Resources Information Center

    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…

  18. Dance Education Action Research: A Twin Study

    ERIC Educational Resources Information Center

    Giguere, Miriam

    2015-01-01

    In this article, the author compares the practices, philosophy, and history of action research, also known as participatory action research, to the purposes and practices of dance education. The comparison yields connections in four categories, enhancing self-reflective teaching and curriculum design, taking responsibility for teaching outcomes,…

  19. A Study of Actions in Operative Notes

    PubMed Central

    Wang, Yan; Pakhomov, Serguei; Burkart, Nora E.; Ryan, James O.; Melton, Genevieve B.

    2012-01-01

    Operative notes contain rich information about techniques, instruments, and materials used in procedures. To assist development of effective information extraction (IE) techniques for operative notes, we investigated the sublanguage used to describe actions within the operative report ‘procedure description’ section. Deep parsing results of 362,310 operative notes with an expanded Stanford parser using the SPECIALIST Lexicon resulted in 200 verbs (92% coverage) including 147 action verbs. Nominal action predicates for each action verb were gathered from WordNet, SPECIALIST Lexicon, New Oxford American Dictionary and Stedman’s Medical Dictionary. Coverage gaps were seen in existing lexical, domain, and semantic resources (Unified Medical Language System (UMLS) Metathesaurus, SPECIALIST Lexicon, WordNet and FrameNet). Our findings demonstrate the need to construct surgical domain-specific semantic resources for IE from operative notes. PMID:23304423

  20. A study of actions in operative notes.

    PubMed

    Wang, Yan; Pakhomov, Serguei; Burkart, Nora E; Ryan, James O; Melton, Genevieve B

    2012-01-01

    Operative notes contain rich information about techniques, instruments, and materials used in procedures. To assist development of effective information extraction (IE) techniques for operative notes, we investigated the sublanguage used to describe actions within the operative report 'procedure description' section. Deep parsing results of 362,310 operative notes with an expanded Stanford parser using the SPECIALIST Lexicon resulted in 200 verbs (92% coverage) including 147 action verbs. Nominal action predicates for each action verb were gathered from WordNet, SPECIALIST Lexicon, New Oxford American Dictionary and Stedman's Medical Dictionary. Coverage gaps were seen in existing lexical, domain, and semantic resources (Unified Medical Language System (UMLS) Metathesaurus, SPECIALIST Lexicon, WordNet and FrameNet). Our findings demonstrate the need to construct surgical domain-specific semantic resources for IE from operative notes. PMID:23304423

  1. A study of actions in operative notes.

    PubMed

    Wang, Yan; Pakhomov, Serguei; Burkart, Nora E; Ryan, James O; Melton, Genevieve B

    2012-01-01

    Operative notes contain rich information about techniques, instruments, and materials used in procedures. To assist development of effective information extraction (IE) techniques for operative notes, we investigated the sublanguage used to describe actions within the operative report 'procedure description' section. Deep parsing results of 362,310 operative notes with an expanded Stanford parser using the SPECIALIST Lexicon resulted in 200 verbs (92% coverage) including 147 action verbs. Nominal action predicates for each action verb were gathered from WordNet, SPECIALIST Lexicon, New Oxford American Dictionary and Stedman's Medical Dictionary. Coverage gaps were seen in existing lexical, domain, and semantic resources (Unified Medical Language System (UMLS) Metathesaurus, SPECIALIST Lexicon, WordNet and FrameNet). Our findings demonstrate the need to construct surgical domain-specific semantic resources for IE from operative notes.

  2. Crystal Structure of Streptococcus pyogenes Cas1 and Its Interaction with Csn2 in the Type II CRISPR-Cas System.

    PubMed

    Ka, Donghyun; Lee, Hasup; Jung, Yi-Deun; Kim, Kyunggon; Seok, Chaok; Suh, Nayoung; Bae, Euiyoung

    2016-01-01

    CRISPRs and Cas proteins constitute an RNA-guided microbial immune system against invading nucleic acids. Cas1 is a universal Cas protein found in all three types of CRISPR-Cas systems, and its role is implicated in new spacer acquisition during CRISPR-mediated adaptive immunity. Here, we report the crystal structure of Streptococcus pyogenes Cas1 (SpCas1) in a type II CRISPR-Cas system and characterize its interaction with S. pyogenes Csn2 (SpCsn2). The SpCas1 structure reveals a unique conformational state distinct from type I Cas1 structures, resulting in a more extensive dimerization interface, a more globular overall structure, and a disruption of potential metal-binding sites for catalysis. We demonstrate that SpCas1 directly interacts with SpCsn2, and identify the binding interface and key residues for Cas complex formation. These results provide structural information for a type II Cas1 protein, and lay a foundation for studying multiprotein Cas complexes functioning in type II CRISPR-Cas systems.

  3. Harnessing a Hurricane: Social Studies in Action.

    ERIC Educational Resources Information Center

    Floyd, Kathleen L.

    1991-01-01

    Describes how a sixth grade class in Findlay, Ohio, became involved in events in McClellanville, South Carolina, where Hurricane Hugo severely damaged a school. After students viewed a videotape of the damage, they planned actions to provide relief that ultimately involved their entire school. Underscores the project's meaningfulness and…

  4. Action!

    ERIC Educational Resources Information Center

    Senese, Joseph

    1998-01-01

    A small group of teachers at one Illinois high school is helping to effect and promote change. Through the Action Research Laboratory (ARL), teams of teachers conduct collaborative action research to improve classroom practices. Data from the first two years of the ARL indicate that teachers are eager to participate in, and have thrived in, their…

  5. A database of whole-body action videos for the study of action, emotion, and untrustworthiness.

    PubMed

    Keefe, Bruce D; Villing, Matthias; Racey, Chris; Strong, Samantha L; Wincenciak, Joanna; Barraclough, Nick E

    2014-12-01

    We present a database of high-definition (HD) videos for the study of traits inferred from whole-body actions. Twenty-nine actors (19 female) were filmed performing different actions-walking, picking up a box, putting down a box, jumping, sitting down, and standing and acting-while conveying different traits, including four emotions (anger, fear, happiness, sadness), untrustworthiness, and neutral, where no specific trait was conveyed. For the actions conveying the four emotions and untrustworthiness, the actions were filmed multiple times, with the actor conveying the traits with different levels of intensity. In total, we made 2,783 action videos (in both two-dimensional and three-dimensional format), each lasting 7 s with a frame rate of 50 fps. All videos were filmed in a green-screen studio in order to isolate the action information from all contextual detail and to provide a flexible stimulus set for future use. In order to validate the traits conveyed by each action, we asked participants to rate each of the actions corresponding to the trait that the actor portrayed in the two-dimensional videos. To provide a useful database of stimuli of multiple actions conveying multiple traits, each video name contains information on the gender of the actor, the action executed, the trait conveyed, and the rating of its perceived intensity. All videos can be downloaded free at the following address: http://www-users.york.ac.uk/~neb506/databases.html. We discuss potential uses for the database in the analysis of the perception of whole-body actions. PMID:24584971

  6. Corrective Action Plan for Corrective Action Unit 271: Areas 25, 26, and 27 Septic Systems, Nevada Test Site, Nevada

    SciTech Connect

    R. B. Jackson

    2003-05-01

    The Areas 25, 26 and 27 Septic Systems are in the Federal Facility Agreement and Consent Order (FFACO) of 1996 as Corrective Action Unit (CAU) 271. This Corrective Action Plan (CAP) provides selected corrective action alternatives and proposes the closure methodology for CAU 271. CAU 271 is located on the Nevada Test Site (NTS) approximately 105 kilometers (65 miles) northwest of Las Vegas, Nevada, and consists of the following 15 Corrective Action Sites (CAS): CAS 25-04-1, Septic System; CAS 25-04-03, Septic System; CAS25-04-04, Septic System; CAS 25-04-08, Septic System; CAS 25-04-09, Septic System; CAS 25-04-10, Septic System; CAS 25-04-11, Septic System; CAS 26-03-01, Contaminated Water Reservoir; CAS 26-04-1, Septic System; CAS 26-04-02, Septic System; CAS 26-05-01, Radioactive Leachfield; CAS-26-05-03, Septic System; CAS 26-05-04, Septic System; CAS 26-05-05, Septic System; and CAS 27-05-02, Leachfield.

  7. CRISPR-Cas: biology, mechanisms and relevance

    PubMed Central

    Hille, Frank

    2016-01-01

    Prokaryotes have evolved several defence mechanisms to protect themselves from viral predators. Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) display a prokaryotic adaptive immune system that memorizes previous infections by integrating short sequences of invading genomes—termed spacers—into the CRISPR locus. The spacers interspaced with repeats are expressed as small guide CRISPR RNAs (crRNAs) that are employed by Cas proteins to target invaders sequence-specifically upon a reoccurring infection. The ability of the minimal CRISPR-Cas9 system to target DNA sequences using programmable RNAs has opened new avenues in genome editing in a broad range of cells and organisms with high potential in therapeutical applications. While numerous scientific studies have shed light on the biochemical processes behind CRISPR-Cas systems, several aspects of the immunity steps, however, still lack sufficient understanding. This review summarizes major discoveries in the CRISPR-Cas field, discusses the role of CRISPR-Cas in prokaryotic immunity and other physiological properties, and describes applications of the system as a DNA editing technology and antimicrobial agent. This article is part of the themed issue ‘The new bacteriology’. PMID:27672148

  8. CRISPR-Cas: biology, mechanisms and relevance.

    PubMed

    Hille, Frank; Charpentier, Emmanuelle

    2016-11-01

    Prokaryotes have evolved several defence mechanisms to protect themselves from viral predators. Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) display a prokaryotic adaptive immune system that memorizes previous infections by integrating short sequences of invading genomes-termed spacers-into the CRISPR locus. The spacers interspaced with repeats are expressed as small guide CRISPR RNAs (crRNAs) that are employed by Cas proteins to target invaders sequence-specifically upon a reoccurring infection. The ability of the minimal CRISPR-Cas9 system to target DNA sequences using programmable RNAs has opened new avenues in genome editing in a broad range of cells and organisms with high potential in therapeutical applications. While numerous scientific studies have shed light on the biochemical processes behind CRISPR-Cas systems, several aspects of the immunity steps, however, still lack sufficient understanding. This review summarizes major discoveries in the CRISPR-Cas field, discusses the role of CRISPR-Cas in prokaryotic immunity and other physiological properties, and describes applications of the system as a DNA editing technology and antimicrobial agent.This article is part of the themed issue 'The new bacteriology'.

  9. CRISPR-Cas: biology, mechanisms and relevance.

    PubMed

    Hille, Frank; Charpentier, Emmanuelle

    2016-11-01

    Prokaryotes have evolved several defence mechanisms to protect themselves from viral predators. Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins (Cas) display a prokaryotic adaptive immune system that memorizes previous infections by integrating short sequences of invading genomes-termed spacers-into the CRISPR locus. The spacers interspaced with repeats are expressed as small guide CRISPR RNAs (crRNAs) that are employed by Cas proteins to target invaders sequence-specifically upon a reoccurring infection. The ability of the minimal CRISPR-Cas9 system to target DNA sequences using programmable RNAs has opened new avenues in genome editing in a broad range of cells and organisms with high potential in therapeutical applications. While numerous scientific studies have shed light on the biochemical processes behind CRISPR-Cas systems, several aspects of the immunity steps, however, still lack sufficient understanding. This review summarizes major discoveries in the CRISPR-Cas field, discusses the role of CRISPR-Cas in prokaryotic immunity and other physiological properties, and describes applications of the system as a DNA editing technology and antimicrobial agent.This article is part of the themed issue 'The new bacteriology'. PMID:27672148

  10. Nucleosome breathing and remodeling constrain CRISPR-Cas9 function

    PubMed Central

    Isaac, R Stefan; Jiang, Fuguo; Doudna, Jennifer A; Lim, Wendell A; Narlikar, Geeta J; Almeida, Ricardo

    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 Cas9 is variable over several orders of magnitude depending on dynamic properties of the DNA sequence and the distance of the PAM site from the nucleosome dyad. We further find that chromatin remodeling enzymes stimulate Cas9 activity on nucleosomal templates. Our findings imply that the spontaneous breathing of nucleosomal DNA together with the action of chromatin remodelers allow Cas9 to effectively act on chromatin in vivo. DOI: http://dx.doi.org/10.7554/eLife.13450.001 PMID:27130520

  11. Advances in therapeutic CRISPR/Cas9 genome editing.

    PubMed

    Savić, Nataša; Schwank, Gerald

    2016-02-01

    Targeted nucleases are widely used as tools for genome editing. Two years ago the clustered regularly interspaced short palindromic repeat (CRISPR)-associated Cas9 nuclease was used for the first time, and since then has largely revolutionized the field. The tremendous success of the CRISPR/Cas9 genome editing tool is powered by the ease design principle of the guide RNA that targets Cas9 to the desired DNA locus, and by the high specificity and efficiency of CRISPR/Cas9-generated DNA breaks. Several studies recently used CRISPR/Cas9 to successfully modulate disease-causing alleles in vivo in animal models and ex vivo in somatic and induced pluripotent stem cells, raising hope for therapeutic genome editing in the clinics. In this review, we will summarize and discuss such preclinical CRISPR/Cas9 gene therapy reports.

  12. Engineering new balancer chromosomes in C. elegans via CRISPR/Cas9.

    PubMed

    Iwata, Satoru; Yoshina, Sawako; Suehiro, Yuji; Hori, Sayaka; Mitani, Shohei

    2016-01-01

    Balancer chromosomes are convenient tools used to maintain lethal mutations in heterozygotes. We established a method for engineering new balancers in C. elegans by using the CRISPR/Cas9 system in a non-homologous end-joining mutant. Our studies will make it easier for researchers to maintain lethal mutations and should provide a path for the development of a system that generates rearrangements at specific sites of interest to model and analyse the mechanisms of action of genes. PMID:27650892

  13. Engineering new balancer chromosomes in C. elegans via CRISPR/Cas9

    PubMed Central

    Iwata, Satoru; Yoshina, Sawako; Suehiro, Yuji; Hori, Sayaka; Mitani, Shohei

    2016-01-01

    Balancer chromosomes are convenient tools used to maintain lethal mutations in heterozygotes. We established a method for engineering new balancers in C. elegans by using the CRISPR/Cas9 system in a non-homologous end-joining mutant. Our studies will make it easier for researchers to maintain lethal mutations and should provide a path for the development of a system that generates rearrangements at specific sites of interest to model and analyse the mechanisms of action of genes. PMID:27650892

  14. Embodied Learning and Creative Writing: An Action Research Study

    ERIC Educational Resources Information Center

    Tobin, Jennifer Ann

    2012-01-01

    This action research study used narrative analysis to explore the role of the body in the writing process of creative writers. Specifically, the purpose of this action research study was threefold: it was first to examine how professional creative writers describe their writing process with particular attention to their perceptions of the role and…

  15. Middle School Responses to Cyberbullying: An Action Research Study

    ERIC Educational Resources Information Center

    Zidack, Astri Marie

    2013-01-01

    This action research study engaged a small public middle school in the northwest United States in a collaborative process to address cyberbullying issues that often lead to academic and behavior problems in schools (Hinduja, 2010; Olweus, 2010). The specific purpose of this action research study was to address the middle school's cyberbullying…

  16. Student Voice in High School: An Action Research Study

    ERIC Educational Resources Information Center

    Termini, Lorraine

    2013-01-01

    This action research study examined the effects of student voice in one high school and the self-reflection of the researcher-administrator involved in the effort. Using three cycles of action research, the researcher-administrator completed a pilot study, implemented a student voice project in one class, and developed a professional development…

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

    PubMed Central

    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

  18. Cas9 Functionally Opens Chromatin.

    PubMed

    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. CRISPR/Cas9 Genome Editing as a Strategy to Study the Tumor Microenvironment in Transgenic Mice.

    PubMed

    Yamanaka, Yojiro

    2016-01-01

    Development of engineered site-specific endonucleases like zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and CRISPR/Cas9 has been revolutionizing genetic approaches in biomedical research fields. These new tools have opened opportunities to carry out targeted genome editing in mouse zygotes without the need for manipulating embryonic stem cells, which have a higher technical burden and many constraints in strain availability. Specific genetic modifications can be directly generated in working genetic backgrounds. This new approach saves time and costs associated with generation and backcrossing of genetically modified animals and will facilitate their use in various cancer research fields. PMID:27581028

  20. Studying Action Representation in Children via Motor Imagery

    ERIC Educational Resources Information Center

    Gabbard, Carl

    2009-01-01

    The use of motor imagery is a widely used experimental paradigm for the study of cognitive aspects of action planning and control in adults. Furthermore, there are indications that motor imagery provides a window into the process of action representation. These notions complement internal model theory suggesting that such representations allow…

  1. Fostering Reading through Intrinsic Motivation: An Action Research Study.

    ERIC Educational Resources Information Center

    Joyce, Marilyn Z.

    2003-01-01

    Discusses a two-year action research study conducted at a high school that transformed reluctant students into lifelong readers by emphasizing intrinsic motivation as opposed to programs that use rewards to motivate students to read. Explains how to design an action research question. (LRW)

  2. Leadership development through action learning sets: an evaluation study.

    PubMed

    Walia, Surinder; Marks-Maran, Di

    2014-11-01

    This article examines the use of action learning sets in a leadership module delivered by a university in south east England. An evaluation research study was undertaking using survey method to evaluate student engagement with action learning sets, and their value, impact and sustainability. Data were collected through a questionnaire with a mix of Likert-style and open-ended questions and qualitative and quantitative data analysis was undertaken. Findings show that engagement in the action learning sets was very high. Action learning sets also had a positive impact on the development of leadership knowledge and skills and are highly valued by participants. It is likely that they would be sustainable as the majority would recommend action learning to colleagues and would consider taking another module that used action learning sets. When compared to existing literature on action learning, this study offers new insights as there is little empirical literature on student engagement with action learning sets and even less on value and sustainability.

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

    PubMed Central

    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. PMID:25775043

  4. An ERP study of motor compatibility effects in action language.

    PubMed

    Santana, Eduardo J; de Vega, Manuel

    2013-08-14

    This ERP study explores the brain's response to the manipulation of motor compatibility in action-related language. In Experiment 1 participants read sentences in which a protagonist performed two different manual actions either simultaneously or consecutively (e.g. While/after cleaning the wound he unrolled the bandage…). The ERPs were measured in the second-clause verb (e.g. unrolled) and noun (e.g. bandage). Notably, only the noun showed compatibility effects, namely a larger N400 in the simultaneous (incompatible) version than in the consecutive (compatible) version, suggesting that readers need to integrate the meaning of the whole sentence to evaluate the feasibility of the actions. In Experiment 2, motor compatibility was manipulated in a different way: all the sentences described the protagonist as performing two simultaneous actions that were both manual (While cleaning the wound he unrolled the bandage), or one action that was perceptual and the other manual (While looking at the wound he unrolled the bandage). The N400 effects for the former incompatible condition were replicated, again in the second-clause noun. The results demonstrated that readers of action language employ their pragmatic world knowledge to test the feasibility of motor actions, taking into account the embodied constraints of such actions. PMID:23796780

  5. An ERP study of motor compatibility effects in action language.

    PubMed

    Santana, Eduardo J; de Vega, Manuel

    2013-08-14

    This ERP study explores the brain's response to the manipulation of motor compatibility in action-related language. In Experiment 1 participants read sentences in which a protagonist performed two different manual actions either simultaneously or consecutively (e.g. While/after cleaning the wound he unrolled the bandage…). The ERPs were measured in the second-clause verb (e.g. unrolled) and noun (e.g. bandage). Notably, only the noun showed compatibility effects, namely a larger N400 in the simultaneous (incompatible) version than in the consecutive (compatible) version, suggesting that readers need to integrate the meaning of the whole sentence to evaluate the feasibility of the actions. In Experiment 2, motor compatibility was manipulated in a different way: all the sentences described the protagonist as performing two simultaneous actions that were both manual (While cleaning the wound he unrolled the bandage), or one action that was perceptual and the other manual (While looking at the wound he unrolled the bandage). The N400 effects for the former incompatible condition were replicated, again in the second-clause noun. The results demonstrated that readers of action language employ their pragmatic world knowledge to test the feasibility of motor actions, taking into account the embodied constraints of such actions.

  6. From Calculus to Dynamical Systems through DGS and CAS

    ERIC Educational Resources Information Center

    García, Jeanett López; Zamudio, Jorge Javier Jiménez

    2015-01-01

    Several factors have motivated the use of CAS or DGS in the teaching-learning process, such as: the development of new technologies, the availability of computers, and the widespread use of the Internet, among others. Even more, the trend to include CAS and DGS in the curricula of some undergraduate studies has resulted in the instruction of the…

  7. Studying Distance Students: Methods, Findings, Actions

    ERIC Educational Resources Information Center

    Wahl, Diane; Avery, Beth; Henry, Lisa

    2013-01-01

    University of North Texas (UNT) Libraries began studying the library needs of distance learners in 2009 using a variety of approaches to explore and confirm these needs as well as obtain input into how to meet them. Approaches used to date include analysis of both quantitative and qualitative responses by online students to the LibQUAL+[R] surveys…

  8. New Case Studies of Citizen Action

    ERIC Educational Resources Information Center

    Tanner, Thomas

    1977-01-01

    Describes a six-unit case study curriculum package designed for secondary and college-level courses relating to environmental education. The units deal with nuclear power, stream channelization, a river dam project, overgrazing of public lands, agribusiness versus the family farm, and swamp preservation. (Author/DB)

  9. Professional Vision in Action: An Exploratory Study

    ERIC Educational Resources Information Center

    Sherin, Miriam Gamoran; Russ, Rosemary S.; Sherin, Bruce L.; Colestock, Adam

    2008-01-01

    The study of teachers' professional vision poses some unique challenges. The application of professional vision happens in a manner that is fleeting, and that is distributed through the moments of instruction. Because of the ongoing nature of instruction, it is not realistic to expect that one could "pause" instruction momentarily, ask a teacher…

  10. [Epilobium parviflorum--in vitro study of biological action].

    PubMed

    Hevesi Tóth, Barbara; Kéry, Agnes

    2009-01-01

    Epilobium parviflorum Schreb. (willow-herb) is used for the treatment of benign prostatic hyperplasia (BPH), but its biological action is not entirely identified. This paper aims to report data on willow-herbs probable biological effect. In vitro studies have been made to investigate different aspects of its antioxidant capacity, anti-inflammatory (COX-inhibitory) action, steroid-receptor -agonistic and -antagonostic effect as well as aromatase-inhibitory effect. Based on our results, willow-herb possess remarkable antioxidant and COX-inhibitory action. PMID:19526676

  11. Action fluency in Parkinson's disease: a follow-up study.

    PubMed

    Signorini, Matteo; Volpato, Chiara

    2006-04-01

    The impairment in action fluency task present in Parkinson's disease (PD) patients has been previously interpreted as an indicator of conversion from PD to PD with dementia or as a grammatical deficit for verbs and ascribed to a frontostriatal loop pathophysiology. In the present study, 20 patients with PD without dementia were longitudinally tested with overall cognitive decline scales and semantic, letter, and action fluency tasks in a 24-month follow-up study. In comparison with healthy age-matched controls, PD patients showed a stable and consistent impairment on action fluency without any sign of cognitive decline. Our findings suggest that action fluency task may be an early sign of impairment of frontostriatal circuits in PD and it cannot be considered an indicator of conversion from PD to PD with dementia.

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

    ERIC Educational Resources Information Center

    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…

  13. A Study of SN Ejecta in the Core-Collapse Supernova Remnant G292.0+1.8: Cas A's Older Cousin

    NASA Astrophysics Data System (ADS)

    Fesen, Robert

    2006-07-01

    Recent studies of the southern oxygen-rich supernova remnant {SNR} G292.0+1.8 have shown it to be the only Galactic SNR to exhibit all the features we expect in young remnants of core-collapse supernovae: an outer shell behind an expanding primary shock, high-velocity fragments of undiluted metal-rich ejecta, and a central pulsar surrounded by a pulsar-wind nebula. G292.0+1.8's optical emission consists of numerous knots and filaments of O- and S-rich ejecta spread throughout much of the remnant shell, many with radially oriented pencil-like geometries that may trace their origins to Rayleigh-Taylor instabilities during the SN event. The evolution and fine-scale structure of SN debris in young remnants is poorly understood and largely uncharted territory. For testing models for the distribution of metal-rich ejecta from core-collapse SNe, how the ejecta evolve and clump, and how SN shocks interact with the local circumstellar medium, the 3000-yr-old G292.0+1.8 remnant rivals the 320-yr-old Cas A remnant in importance. We therefore propose the first HST images of G292.0+1.8 in order to characterize the fine-scale spatial distribution of the ejecta, their sub-arcsecond chemical make-up, and the detailed structure and scale lengths for metal-rich SN ejecta clumps. The proposed HST images of G292.0+1.8 will be used in conjunction with existing Spitzer Cycle 1 infrared data and an upcoming 0.5 Msec Chandra X-ray image. We expect to achieve the same kind of results for G292 that have already been obtained for Cas A. High-resolution HST images of this remnant, combined with Spitzer and Chandra data and contrasted with a similar data set on Cas A, will provide superb multiwavelength benchmarks for both very young and older core-collapse SNRs.

  14. NTP Toxicology and Carcinogenesis Studies of Oxazepam (CAS No. 604-75-1) in F344/N Rats (Feed Studies).

    PubMed

    1998-10-01

    Oxazepam and related benzodiazepine drugs are used in the treatment of anxiety. All benzodiazepines currently in use share a number of effects, including sedation, hypnosis, decreased anxiety, muscle relaxation, amnesia, and anticonvulsant activity. Oxazepam and four other benzodiazepines (chlordiazepoxide, chlorazepate, diazepam, and flurazepam) were nominated for study by the Food and Drug Administration (FDA) and by the NIEHS based on their widespread use, use by pregnant women, and the lack of adequate rodent carcinogenicity studies. Oxazepam was evaluated in 14-week and 2-year studies by the NTP, and Technical Report No. 443 contains the results of the studies performed with the Swiss-Webster and B6C3F1 strains of mice. Studies with rats were not initiated at the same time as the mouse studies because adequate carcinogenicity studies of oxazepam with the Sprague-Dawley rat strain had been submitted to the FDA. Subsequently, because of the marked neoplastic responses found in the two mouse strains, the NTP initiated 2-year studies of oxazepam with the F344/N rat. Groups of male and female F344/N rats were exposed to oxazepam (greater than 99% pure) in feed for 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and cultured Chinese hamster ovary cells, and mouse peripheral blood samples were analyzed for the frequency of micronucleated normochromatic erythrocytes. 2-YEAR STUDY: Groups of 50 male and 50 female F344/N rats were fed diets containing 0, 625, 2,500, or 5,000 ppm oxazepam for up to 105 weeks. A stop-exposure group of 50 males and 50 females received 10,000 ppm oxazepam in feed for 26 weeks, after which animals received undosed feed for the remainder of the 2-year study. The continuous-exposure concentrations resulted in average daily doses of 25, 100, or 250 mg oxazepam/kg body weight to males and 25, 110, or 220 mg/kg to females. Stop- exposure males and females received an average daily dose of 630 mg/kg during the exposure

  15. Toxicology and Carcinogenesis Studies of Hexachloroethane (CAS No. 67-72-1) in F344/N Rats (Gavage Studies).

    PubMed

    1989-08-01

    Hexachloroethane is used in organic synthesis as a retarding agent in fermentation, as a camphor substitute in nitrocellulose, in pyrotechnics and smoke devices, in explosives, and as a solvent. In previous long-term gavage studies with B6C3F1 mice and Osbourne-Mendel rats (78 weeks of exposure followed by 12-34 weeks of observation), hexachloroethane caused increased incidences of hepatocellular carcinomas in mice. However, survival of low and high dose rats was reduced compared with that of vehicle controls, and the effects on rats were inconclusive. Therefore, additional toxicology and carcinogenesis studies were conducted in F344/N rats by administering hexachloroethane (approximately 99% pure) in corn oil by gavage to groups of males and females for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and in Chinese hamster ovary (CHO) cells. Urinalysis was performed in conjunction with the 13-week studies. Sixteen-Day Studies: In the 16-day studies (dose range, 187-3,000 mg/kg), all rats that received 1,500 or 3,000 mg/kg and 1/5 males and 2/5 females that received 750 mg/kg died before the end of the studies. Final mean body weights of rats that received 750 mg/kg were 25% lower than that of vehicle controls for males and 37% lower for females. Compound-related clinical signs seen at 750 mg/kg or more included dyspnea, ataxia, prostration, and excessive lacrimation. Other compound-related effects included hyaline droplet formation in the tubular epithelial cells in all dosed males and tubular cell regeneration and granular casts in the tubules at the corticomedullary junction in the kidney in males receiving 187 and 375 mg/kg. Thirteen-Week Studies: In the 13-week studies (dose range, 47-750 mg/kg), 5/10 male rats and 2/10 female rats that received 750 mg/kg died before the end of the studies. The final mean body weight of male rats that received 750 mg/kg was 19% lower than that of vehicle controls. Compound

  16. Historic light curve of V890 Cas

    NASA Astrophysics Data System (ADS)

    Nesci, R.

    2016-05-01

    The variability of V890 Cas is studied with 87 -band plates of the Asiago archive. The star shows variations of about 5 mag with an average magnitude =13 and a period of 486 days. An 5.0 color index is also derived near the maximum luminosity.

  17. Using the CAS Standards in Assessment Projects

    ERIC Educational Resources Information Center

    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.

  18. NTP Toxicology and Carcinogenesis Studies of Pentachlorophenol (CAS NO. 87-86-5) in F344/N Rats (Feed Studies).

    PubMed

    1999-04-01

    Pentachlorophenol has been used as an herbicide, algicide, defoliant, wood preservative, germicide, fungicide, and molluscicide. Pentachlorophenol was nominated by the National Cancer Institute for carcinogenicity testing based on its widespread use as a wood preservative, potential for entering the environment (pentachlorophenol residues have been found worldwide in soil, water, and air samples; in food products; and in human and animal tissues and body fluids), and likelihood of bioaccumulation in the environment (pentachlorophenol is persistent in soil, having a half-life of up to 5 years). Technical Report No. 349 contains the results of the 2-year studies of pentachlorophenol performed by the NTP with B6C3F1 mice. Male and female F344/N rats were exposed to pentachlorophenol (approximately 99% pure) in feed for 28 days or 2 years. Genetic toxicology studies were conducted in vitro in Salmonella typhimurium and cultured Chinese hamster ovary cells and in vivo in rat and mouse bone marrow cells. 28-DAY STUDY IN RATS: Groups of 10 male and 10 female F344/N rats were given 0, 200, 400, 800, 1,600, or 3,200 ppm pentachlorophenol, equivalent to average daily doses of approximately 20, 40, 75, 150, or 270 mg pentachlorophenol/kg body weight to males and females in feed for 28 days. With the exception of one male and two females exposed to 3,200 ppm, all rats survived until the end of the study. The final mean body weights and body weight gains of male rats exposed to 1,600 or 3,200 ppm and female rats exposed to 400, 800, 1,600, or 3,200 ppm were significantly less than those of the controls; rats exposed to 3,200 ppm lost weight during the study. Feed consumption by 3,200 ppm males was less than that by the control group throughout the study. The absolute and relative liver weights of 400, 800, and 1,600 ppm males and all exposed groups of females were significantly greater than those of the controls. Compared to the control groups, the incidences of minimal to mild

  19. Toxicology and Carcinogenesis Studies of Ochratoxin A (CAS No. 303-47-9) in F344/N Rats (Gavage Studies).

    PubMed

    1989-05-01

    Ochratoxin A is a naturally occurring fungal toxin that is a contaminant in corn, peanuts, storage grains, cottonseed, meats, dried fish, and nuts. Toxicology and carcinogenesis studies were conducted by administering ochratoxin A (98% pure) in corn oil by gavage to groups of F344/N rats of each sex for 16 days, 13 weeks, 9 months, 15 months, or 2 years. Only rats were studied because ochratoxin A has been shown to be carcinogenic in mice. Genetic toxicology tests were performed with bacterial and mammalian cells. Urinalysis, hematologic and serum chemical analyses, and bone marrow cellularity determinations were conducted at 9, 15, and 24 months in the 2-year studies. Sixteen-Day and Thirteen-Week Studies: Rats were administered 0, 1, 4, or 16 mg/kg ochratoxin A in corn oil by gavage 5 days per week for a total of 12 doses over 16 days. All rats that received 16 mg/kg ochratoxin A died within 6 days. Rats that received 4 mg/kg lost weight. Compound-related lesions in rats included bone marrow hyperplasia, thymic atrophy, necrosis and hyperplasia of the forestomach epithelium, renal tubular cell degenerative and regenerative changes (nephropathy), and adrenal gland hemorrhage. Renal tubular changes were most severe in animals that received 4 mg/kg. Rats that received 16 mg/kg had less severe renal lesions than those at 4 mg/kg, perhaps because the acute toxicity and early death did not allow sufficient time for full development of lesions. No compound-related deaths occurred in the 13-week studies (doses were 0 and 0.0625 to 1 mg/kg). The final mean body weight of rats that received 0.25, 0.5, or 1 mg/kg was 7%, 11%, or 19% lower than that of vehicle controls for males and 3%, 4%, or 9% lower for females. Compound-related lesions in the kidney were characterized as degeneration and regeneration of the epithelium of the proximal convoluted tubules with individual cell necrosis of moderate severity (see page 3 of the Technical Report). Karyomegaly of tubular

  20. NTP Toxicology and Carcinogenesis Studies of Pentachlorophenol (CAS NO. 87-86-5) in F344/N Rats (Feed Studies).

    PubMed

    1999-04-01

    Pentachlorophenol has been used as an herbicide, algicide, defoliant, wood preservative, germicide, fungicide, and molluscicide. Pentachlorophenol was nominated by the National Cancer Institute for carcinogenicity testing based on its widespread use as a wood preservative, potential for entering the environment (pentachlorophenol residues have been found worldwide in soil, water, and air samples; in food products; and in human and animal tissues and body fluids), and likelihood of bioaccumulation in the environment (pentachlorophenol is persistent in soil, having a half-life of up to 5 years). Technical Report No. 349 contains the results of the 2-year studies of pentachlorophenol performed by the NTP with B6C3F1 mice. Male and female F344/N rats were exposed to pentachlorophenol (approximately 99% pure) in feed for 28 days or 2 years. Genetic toxicology studies were conducted in vitro in Salmonella typhimurium and cultured Chinese hamster ovary cells and in vivo in rat and mouse bone marrow cells. 28-DAY STUDY IN RATS: Groups of 10 male and 10 female F344/N rats were given 0, 200, 400, 800, 1,600, or 3,200 ppm pentachlorophenol, equivalent to average daily doses of approximately 20, 40, 75, 150, or 270 mg pentachlorophenol/kg body weight to males and females in feed for 28 days. With the exception of one male and two females exposed to 3,200 ppm, all rats survived until the end of the study. The final mean body weights and body weight gains of male rats exposed to 1,600 or 3,200 ppm and female rats exposed to 400, 800, 1,600, or 3,200 ppm were significantly less than those of the controls; rats exposed to 3,200 ppm lost weight during the study. Feed consumption by 3,200 ppm males was less than that by the control group throughout the study. The absolute and relative liver weights of 400, 800, and 1,600 ppm males and all exposed groups of females were significantly greater than those of the controls. Compared to the control groups, the incidences of minimal to mild

  1. Validity in Action: Lessons from Studies of Data Use

    ERIC Educational Resources Information Center

    Moss, Pamela A.

    2013-01-01

    Studies of data use illuminate ways in which education professionals have used test scores and other evidence relevant to students' learning--in action in their own contexts of work--to make decisions about their practice. These studies raise instructive challenges for a validity theory that focuses on intended interpretations and uses of test…

  2. Insulin action in the human brain: evidence from neuroimaging studies.

    PubMed

    Kullmann, S; Heni, M; Fritsche, A; Preissl, H

    2015-06-01

    Thus far, little is known about the action of insulin in the human brain. Nonetheless, recent advances in modern neuroimaging techniques, such as functional magnetic resonance imaging (fMRI) or magnetoencephalography (MEG), have made it possible to investigate the action of insulin in the brain in humans, providing new insights into the pathogenesis of brain insulin resistance and obesity. Using MEG, the clinical relevance of the action of insulin in the brain was first identified, linking cerebral insulin resistance with peripheral insulin resistance, genetic predisposition and weight loss success in obese adults. Although MEG is a suitable tool for measuring brain activity mainly in cortical areas, fMRI provides high spatial resolution for cortical as well as subcortical regions. Thus, the action of insulin can be detected within all eating behaviour relevant regions, which include regions deeply located within the brain, such as the hypothalamus, midbrain and brainstem, as well as regions within the striatum. In this review, we outline recent advances in the field of neuroimaging aiming to investigate the action of insulin in the human brain using different routes of insulin administration. fMRI studies have shown a significant insulin-induced attenuation predominantly in the occipital and prefrontal cortical regions and the hypothalamus, successfully localising insulin-sensitive brain regions in healthy, mostly normal-weight individuals. However, further studies are needed to localise brain areas affected by insulin resistance in obese individuals, which is an important prerequisite for selectively targeting brain insulin resistance in obesity.

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

    PubMed

    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.

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

    PubMed

    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.

  5. Case Studies of Action Research in Various Adult Education Settings.

    ERIC Educational Resources Information Center

    Kuhne, Gary W.; Weirauch, Drucie; Fetterman, David J.; Mearns, Raiana M.; Kalinosky, Kathy; Cegles, Kathleen A.; Ritchey, Linda

    1997-01-01

    Six case studies illustrate action research in adult education: faculty development in a museum, participation in a church congregation, retention of literacy volunteers in a corrections center, learner participation in a homeless shelter, technology innovation in a university, and infection control in a hospital. (SK)

  6. Espousing Democratic Leadership Practices: A Study of Values in Action

    ERIC Educational Resources Information Center

    Devereaux, Lorraine

    2003-01-01

    This article examines principals' espoused values and their values in action. It provides a reanalysis of previously collected data through a values lens. The original research study was an international quantitative and qualitative investigation of principals' leadership approaches that was based in 15 schools. This particular excerpt of the…

  7. A Longitudinal Study of the Impact of an Environmental Action

    ERIC Educational Resources Information Center

    Rioux, Liliane; Pasquier, Daniel

    2013-01-01

    In a previous study, we investigated the impact of an awareness-raising campaign on the behaviour of secondary school children in the Centre Region of France, regarding the recycling of used batteries. But, was it a question of pro-environmental behaviour or simply an environmental action? To answer this question, a three-year longitudinal study…

  8. Study Challenges Assumptions about Affirmative-Action Bans

    ERIC Educational Resources Information Center

    Schmidt, Peter

    2008-01-01

    This article reports the results of a new study on the impact of bans on race-conscious admissions policies which seem to confirm what many critics of affirmative action have long suspected: It is Asian-Americans, rather than whites, who are most disadvantaged by elite universities' consideration of ethnicity and race. Left unanswered are the…

  9. Minesweeper and Hypothetical Thinking Action Research & Pilot Study

    ERIC Educational Resources Information Center

    Walker, Jacob J.

    2010-01-01

    This Action Research project and Pilot Study was designed and implemented to improve students' hypothetical thinking abilities by exploring the possibility that learning and playing the computer game Minesweeper may inherently help improve hypothetical thinking. One objective was to use educational tools to make it easier for students to learn the…

  10. A diary study of action slips in healthy individuals.

    PubMed

    Jónsdóttir, María K; Adólfsdóttir, Steinunn; Cortez, Rúna Dögg; Gunnarsdóttir, María; Gústafsdóttir, Agústa Hlín

    2007-12-01

    Memory complaints following minor head injury or whiplash are common and often bear similarity to absent mindedness or action slips (Reason, 1979). We replicated Reason's study by asking 189 healthy volunteers to keep diaries of their action slips for a week. The mean number of slips was 6.4 (SD = 4.9). Perceived stress did not correlate with number of slips but there was a weak positive correlation between action slips and scores on a memory failures questionnaire. Memory diaries may be clinically useful when assessing individuals who worry about cognitive sequelae of minor injuries. Diaries clarify the nature of the complaints and may have therapeutic value by demonstrating that the memory slips are less frequent than estimated by the patients. PMID:17853144

  11. Spectra of Cas A's Highest Velocity Ejecta

    NASA Astrophysics Data System (ADS)

    Fesen, Robert A.; Milisavljevic, Dan

    2010-08-01

    The young age and close distance of the Galactic supernova remnant Cassiopeia A (Cas A) make it perhaps our best case study and clearest look at the explosion dynamics of a core-collapse supernova (CCSN). Interestingly, Cas A exhibits two nearly opposing streams of high velocity ejecta or `jets' in its NE and SW regions racing outward at speeds more than twice that of the main shell. The nature of these jets, however, and their possible association with an aspherical supernova explosion mechanism is controversial. A handful of existing low-resolution spectra of outer knots in the NE jet display chemical abundances hinting at an origin from the S-Si-Ca- Ar rich layer deep inside the progenitor. If these abundances could be firmly established in both the NE and SW jets, it would be very strong evidence in support of a highly asymmetrical explosion engine for Cas A's progenitor and, in turn, for CCSNe in general. We request KPNO 4m telescope + MARS time to obtain high quality multi-object spectroscopy of Cas A's highest velocity ejecta to measure their nitrogen, sulfur, oxygen, calcium, and argon abundances. These spectra will be analyzed with the metal-rich shock models of J. Raymond and then compared to current sets of CCSN models paying particular attention to knot composition vs. ejection velocity and ejecta mixing.

  12. Students of Action? A Comparative Investigation of Secondary Science and Social Studies Students' Action Repertoires in a Land Use Context

    ERIC Educational Resources Information Center

    Kumler, Lori M.

    2011-01-01

    Environmental education (EE) and social studies education share an interest in behavioral outcomes. This study compares behavioral outcomes--including both self-reported knowledge of actions and reported actions taken--in the context of a land use curriculum enacted in secondary science versus social studies classes with 500 students and nine…

  13. Closure Report for Corrective Action Unit 107: Low Impact Soil Sites, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2009-06-01

    Corrective Action Unit (CAU) 107 is identified in the Federal Facility Agreement and Consent Order (FFACO) as 'Low Impact Soil Sites' and consists of the following 15 Corrective Action Sites (CASs), located in Areas 1, 2, 3, 4, 5, 9, 10, and 18 of the Nevada Test Site: CAS 01-23-02, Atmospheric Test Site - High Alt; CAS 02-23-02, Contaminated Areas (2); CAS 02-23-03, Contaminated Berm; CAS 02-23-10, Gourd-Amber Contamination Area; CAS 02-23-11, Sappho Contamination Area; CAS 02-23-12, Scuttle Contamination Area; CAS 03-23-24, Seaweed B Contamination Area; CAS 03-23-27, Adze Contamination Area; CAS 03-23-28, Manzanas Contamination Area; CAS 03-23-29, Truchas-Chamisal Contamination Area; CAS 04-23-02, Atmospheric Test Site T4-a; CAS 05-23-06, Atmospheric Test Site; CAS 09-23-06, Mound of Contaminated Soil; CAS 10-23-04, Atmospheric Test Site M-10; and CAS 18-23-02, U-18d Crater (Sulky). Closure activities were conducted from February through April 2009 according to the FFACO (1996; as amended February 2008) and Revision 1 of the Streamlined Approach for Environmental Restoration Plan for CAU 107 (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office, 2009). The corrective action alternatives included No Further Action and Closure in Place with Administrative Controls. Closure activities are summarized.

  14. Performance of the Cas9 nickase system in Drosophila melanogaster.

    PubMed

    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

    2014-10-01

    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 Cas9(D10A) nickase flies efficiently produces heritable indel mutants. We then determined the effective distance between the two sgRNA targets and their orientations that affected the ability of the sgRNA pairs to generate mutations when expressed in the transgenic nickase flies. Interestingly, Cas9 nickase greatly reduces the ability to generate mutants with one sgRNA, suggesting that the application of Cas9 nickase and sgRNA pairs can almost avoid off-target effects when generating indel mutants. Finally, a defined piwi mutant allele is generated with this system through homology-directed repair. However, Cas9(D10A) is not as effective as Cas9 in replacing the entire coding sequence of piwi with two sgRNAs. PMID:25128437

  15. CRISPR/Cas9 in Genome Editing and Beyond.

    PubMed

    Wang, Haifeng; La Russa, Marie; Qi, Lei S

    2016-06-01

    The Cas9 protein (CRISPR-associated protein 9), derived from type II CRISPR (clustered regularly interspaced short palindromic repeats) bacterial immune systems, is emerging as a powerful tool for engineering the genome in diverse organisms. As an RNA-guided DNA endonuclease, Cas9 can be easily programmed to target new sites by altering its guide RNA sequence, and its development as a tool has made sequence-specific gene editing several magnitudes easier. The nuclease-deactivated form of Cas9 further provides a versatile RNA-guided DNA-targeting platform for regulating and imaging the genome, as well as for rewriting the epigenetic status, all in a sequence-specific manner. With all of these advances, we have just begun to explore the possible applications of Cas9 in biomedical research and therapeutics. In this review, we describe the current models of Cas9 function and the structural and biochemical studies that support it. We focus on the applications of Cas9 for genome editing, regulation, and imaging, discuss other possible applications and some technical considerations, and highlight the many advantages that CRISPR/Cas9 technology offers.

  16. Primary processing of CRISPR RNA by the endonuclease Cas6 in Staphylococcus epidermidis.

    PubMed

    Wakefield, Noelle; Rajan, Rakhi; Sontheimer, Erik J

    2015-10-01

    In many bacteria and archaea, an adaptive immune system (CRISPR-Cas) provides immunity against foreign genetic elements. This system uses CRISPR RNAs (crRNAs) derived from the CRISPR array, along with CRISPR-associated (Cas) proteins, to target foreign nucleic acids. In most CRISPR systems, endonucleolytic processing of crRNA precursors (pre-crRNAs) is essential for the pathway. Here we study the Cas6 endonuclease responsible for crRNA processing in the Type III-A CRISPR-Cas system from Staphylococcus epidermidis RP62a, a model for Type III-A CRISPR-Cas systems, and define substrate requirements for SeCas6 activity. We find that SeCas6 is necessary and sufficient for full-length crRNA biogenesis in vitro, and that it relies on both sequence and stem-loop structure in the 3' half of the CRISPR repeat for recognition and processing.

  17. Controlling UCAVs by JTACs in CAS missions

    NASA Astrophysics Data System (ADS)

    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.

  18. An action research study of secondary science assessment praxes

    NASA Astrophysics Data System (ADS)

    Ryan, Thomas Gerald

    This practical participatory action research study illuminates the assessment praxes of four Ontario secondary level science teachers at one school using a facilitative approach. Participants were joined by a thematic concern, that is, a commitment to inform and improve assessment. Hence, two distinct sets of research questions emerged. The first involves the nature of assessment as we asked, what was the current state of assessment practice in secondary science? What were participants' initial understandings of assessment and actual practices at the onset of this research? To what extent did these initial understandings and actual practices change due to the illumination of assessment praxes through action research involvement? What was their level of awareness of current Ontario government pronouncements and in what ways did they implement this knowledge? The second theme, concerning the nature of action research, was realised by asking what did participants learn about action research? What other learning and professional gains were realised during this study? And, what did I learn about action research and assessment through my involvement in this study? Data were collected via supportive discussion groups, individual interviews, classroom visitations, journals and documentation. This professional development experience facilitated 'interactive professionalism' as teachers worked in a small group and interacted frequently in the course of planning, testing new ideas, attempting to solve different problems, and assess the effectiveness of those ideas. In addition, this action research effort was strategic and systematic, to attain a high degree of specific interactions, (personal interviews, group meetings, classroom observations, evidence collection). This series of deliberate and planned intentions helped participants solve assessment dilemmas. We developed an awareness and understanding of the need for more preservice and inservice assessment training

  19. Multiple mechanisms for CRISPR-Cas inhibition by anti-CRISPR proteins.

    PubMed

    Bondy-Denomy, Joseph; Garcia, Bianca; Strum, Scott; Du, Mingjian; Rollins, MaryClare F; Hidalgo-Reyes, Yurima; Wiedenheft, Blake; Maxwell, Karen L; Davidson, Alan R

    2015-10-01

    The battle for survival between bacteria and the viruses that infect them (phages) has led to the evolution of many bacterial defence systems and phage-encoded antagonists of these systems. Clustered regularly interspaced short palindromic repeats (CRISPR) and the CRISPR-associated (cas) genes comprise an adaptive immune system that is one of the most widespread means by which bacteria defend themselves against phages. We identified the first examples of proteins produced by phages that inhibit a CRISPR-Cas system. Here we performed biochemical and in vivo investigations of three of these anti-CRISPR proteins, and show that each inhibits CRISPR-Cas activity through a distinct mechanism. Two block the DNA-binding activity of the CRISPR-Cas complex, yet do this by interacting with different protein subunits, and using steric or non-steric modes of inhibition. The third anti-CRISPR protein operates by binding to the Cas3 helicase-nuclease and preventing its recruitment to the DNA-bound CRISPR-Cas complex. In vivo, this anti-CRISPR can convert the CRISPR-Cas system into a transcriptional repressor, providing the first example-to our knowledge-of modulation of CRISPR-Cas activity by a protein interactor. The diverse sequences and mechanisms of action of these anti-CRISPR proteins imply an independent evolution, and foreshadow the existence of other means by which proteins may alter CRISPR-Cas function.

  20. Multiple mechanisms for CRISPR-Cas inhibition by anti-CRISPR proteins.

    PubMed

    Bondy-Denomy, Joseph; Garcia, Bianca; Strum, Scott; Du, Mingjian; Rollins, MaryClare F; Hidalgo-Reyes, Yurima; Wiedenheft, Blake; Maxwell, Karen L; Davidson, Alan R

    2015-10-01

    The battle for survival between bacteria and the viruses that infect them (phages) has led to the evolution of many bacterial defence systems and phage-encoded antagonists of these systems. Clustered regularly interspaced short palindromic repeats (CRISPR) and the CRISPR-associated (cas) genes comprise an adaptive immune system that is one of the most widespread means by which bacteria defend themselves against phages. We identified the first examples of proteins produced by phages that inhibit a CRISPR-Cas system. Here we performed biochemical and in vivo investigations of three of these anti-CRISPR proteins, and show that each inhibits CRISPR-Cas activity through a distinct mechanism. Two block the DNA-binding activity of the CRISPR-Cas complex, yet do this by interacting with different protein subunits, and using steric or non-steric modes of inhibition. The third anti-CRISPR protein operates by binding to the Cas3 helicase-nuclease and preventing its recruitment to the DNA-bound CRISPR-Cas complex. In vivo, this anti-CRISPR can convert the CRISPR-Cas system into a transcriptional repressor, providing the first example-to our knowledge-of modulation of CRISPR-Cas activity by a protein interactor. The diverse sequences and mechanisms of action of these anti-CRISPR proteins imply an independent evolution, and foreshadow the existence of other means by which proteins may alter CRISPR-Cas function. PMID:26416740

  1. Action Researchers' Perspectives about the Distinguishing Characteristics of Action Research: A Delphi and Learning Circles Mixed-Methods Study

    ERIC Educational Resources Information Center

    Rowell, Lonnie L.; Polush, Elena Yu; Riel, Margaret; Bruewer, Aaron

    2015-01-01

    The purpose of this study was to identify distinguishing characteristics of action research within the Action Research Special Interest Group of the American Educational Research Association. The authors sought to delineate the foundational framework endorsed by this community. The study was conducted during January-April 2012 and employed an…

  2. Verb gapping: an action-gap compatibility study.

    PubMed

    Claus, Berry

    2015-03-01

    This study addresses the processing of verb-gapping sentences, e.g., John closes a juice bottle and Jim [ ] a lemonade bottle. The goal was to explore if there would be an interaction between language comprehension and motor action not only for overt action verbs but also for gapped verbs. Participants read gapping sentences that either described clockwise or counter-clockwise manual rotations (e.g., closes vs. opens a juice bottle). Adopting a paradigm developed by Zwaan and Taylor (2006), sentence presentation was frame-by-frame. Participants proceeded from frame to frame by turning a knob either clockwise or counter clockwise. Analyses of the frame reading-times yielded a significant effect of compatibility between the linguistically conveyed action and the knob turning for the overt-verb (e.g., closes/opens a juice bottle) as well as for the gapped-verb frame (e.g., a lemonade bottle) - with longer reading times in the match condition than in the mismatch condition - but not for any of the other frames (e.g., and Jim). The results are promising in providing novel evidence for the real-time reactivation of gapped verbs and in suggesting that action simulation is not bound to the processing of overt verbs. PMID:25103783

  3. Corrective Action Investigation Plan for Corrective Action Unit 106: Areas 5, 11 Frenchman Flat Atmospheric Sites, Nevada National Security Site, Nevada

    SciTech Connect

    Patrick Matthews

    2011-07-01

    Corrective Action Unit 106 comprises the four corrective action sites (CASs) listed below: • 05-20-02, Evaporation Pond • 05-23-05, Atmospheric Test Site - Able • 05-45-04, 306 GZ Rad Contaminated Area • 05-45-05, 307 GZ Rad Contaminated Area These sites are being investigated because existing information on the nature and extent of potential contamination is insufficient to evaluate and recommend corrective action alternatives (CAAs). Additional information will be obtained by conducting a corrective action investigation before evaluating CAAs and selecting the appropriate corrective action for each CAS. The results of the field investigation will support a defensible evaluation of viable CAAs that will be presented in the Corrective Action Decision Document. The sites will be investigated based on the data quality objectives (DQOs) developed on January 19, 2010, by representatives of the Nevada Division of Environmental Protection and the U.S. Department of Energy (DOE), National Nuclear Security Administration Nevada Site Office. The DQO process was used to identify and define the type, amount, and quality of data needed to develop and evaluate appropriate corrective actions for CAU 106. The presence and nature of contamination at CAU 106 will be evaluated based on information collected from a field investigation. The CAU includes land areas impacted by the release of radionuclides from groundwater pumping during the Radionuclide Migration study program (CAS 05-20-02), a weapons-related airdrop test (CAS 05-23-05), and unknown support activities at two sites (CAS 05-45-04 and CAS 05-45-05). The presence and nature of contamination from surface-deposited radiological contamination from CAS 05-23-05, Atmospheric Test Site - Able, and other types of releases (such as migration and excavation as well as any potential releases discovered during the investigation) from the remaining three CASs will be evaluated using soil samples collected from the locations

  4. Cell transfection as a tool to study growth hormone action

    SciTech Connect

    Norstedt, G.; Enberg, B.; Francis, S.

    1994-12-31

    The isolation of growth hormone receptor (GHR) cDNA clones has made possible the transfection of GHRs into cultured cells. Our aim in this minireview is to show how the application of such approaches have benefited GHR research. GH stimulation of cells expressing GHR cDNAs can cause an alteration of cellular function that mimic those of the endogenous GHR. GHR cDNA transfected cells also offer a system where the mechanism of GH action can be studied. Such a system has been used to demonstrate that the GHR itself becomes tyrosine phosphorylated and that further phosphorylation of downstream proteins is important in GH action. The GH signals are transmitted to the nucleus and GH regulated genes have now begun to be characterized. The ability to use cell transfection for mechanistic studies of GH action will be instrumental to define domains within the receptor that are of functional importance and to determined pathways whereby GH signals are conveyed within the cell. 33 refs., 2 tabs.

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

    PubMed

    Kim, Hyun-Min; Colaiácovo, Monica P

    2016-07-01

    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.

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

    PubMed

    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

  7. Adenovirus-Mediated Somatic Genome Editing of Pten by CRISPR/Cas9 in Mouse Liver in Spite of Cas9-Specific Immune Responses.

    PubMed

    Wang, Dan; Mou, Haiwei; Li, Shaoyong; Li, Yingxiang; Hough, Soren; Tran, Karen; Li, Jia; Yin, Hao; Anderson, Daniel G; Sontheimer, Erik J; Weng, Zhiping; Gao, Guangping; Xue, Wen

    2015-07-01

    CRISPR/Cas9 derived from the bacterial adaptive immunity pathway is a powerful tool for genome editing, but the safety profiles of in vivo delivered Cas9 (including host immune responses to the bacterial Cas9 protein) have not been comprehensively investigated in model organisms. Nonalcoholic steatohepatitis (NASH) is a prevalent human liver disease characterized by excessive fat accumulation in the liver. In this study, we used adenovirus (Ad) vector to deliver a Streptococcus pyogenes-derived Cas9 system (SpCas9) targeting Pten, a gene involved in NASH and a negative regulator of the PI3K-AKT pathway, in mouse liver. We found that the Ad vector mediated efficient Pten gene editing even in the presence of typical Ad vector-associated immunotoxicity in the liver. Four months after vector infusion, mice receiving the Pten gene-editing Ad vector showed massive hepatomegaly and features of NASH, consistent with the phenotypes following Cre-loxP-induced Pten deficiency in mouse liver. We also detected induction of humoral immunity against SpCas9 and the potential presence of an SpCas9-specific cellular immune response. Our findings provide a strategy to model human liver diseases in mice and highlight the importance considering Cas9-specific immune responses in future translational studies involving in vivo delivery of CRISPR/Cas9. PMID:26086867

  8. Priorities for action in a rural older adults study.

    PubMed

    Averill, Jennifer B

    2012-01-01

    This article reports the findings from a recent study of older adults in the rural southwestern United States and discusses practice and research implications. The aim of the study was to analyze health disparities and strengths in the contexts of rurality, aging, a depressed economy, and limited health resources. Identified themes needing action included sustained access to prescriptions, transportation solutions for older adults in isolated communities, inadequate access to care, poor infrastructure and coordination of services, scarce assisted living and in-home care for frail older adults, and barriers related to culture, language, and economics. PMID:22929381

  9. Priorities for Action in a Rural Older Adults Study

    PubMed Central

    Averill, Jennifer B.

    2013-01-01

    This article reports the findings from a recent study of older adults in the rural southwestern United States and discusses practice and research implications. The aim of the study was to analyze health disparities and strengths in the contexts of rurality, aging, a depressed economy, and limited health resources. Identified themes needing action included sustained access to prescriptions, transportation solutions for older adults in isolated communities, inadequate access to care, poor infrastructure and coordination of services, scarce assisted living and in-home care for frail older adults, and barriers related to culture, language, and economics. PMID:22929381

  10. Passive reading and motor imagery about hand actions and tool-use actions: an fMRI study.

    PubMed

    Yang, Jie; Shu, Hua

    2014-02-01

    Recent studies have shown that motor activations in action verb comprehension can be modulated by task demands (e.g., motor imagery vs. passive reading) and the specificity of action verb meaning. However, how the two factors work together to influence the involvement of the motor system during action verb comprehension is still unclear. To address the issue, the current study investigated the brain activations in motor imagery and passive reading of verbs about hand actions and tool-use actions. Three types of Chinese verbs were used, including hand-action verbs and two types of tool-use verbs emphasizing either the hand or tools information. Results indicated that all three types of verbs elicited common activations in hand motor areas during passive reading and motor imagery. Contrast analyses showed that in the hand verbs and the tool verbs where the hand information was emphasized, motor imagery elicited stronger effects than passive reading in the superior frontal gyrus, supplemental motor area and cingulate cortex that are related to motor control and regulation. For tool-use verbs emphasizing tools information, the motor imagery task elicited stronger activity than passive reading in occipital regions related to visual imagery. These results suggest that motor activations during action verb comprehension can be modulated by task demands and semantic features of action verbs. The sensorimotor simulation during language comprehension is flexible and determined by the interactions between linguistic and extralinguistic contexts.

  11. Goal anticipation during action observation is influenced by synonymous action capabilities, a puzzling developmental study.

    PubMed

    Gredebäck, Gustaf; Kochukhova, Olga

    2010-04-01

    Eighteen- and 25-month-old human toddlers' ability to manually solve a puzzle and their ability to anticipate the goal during observation of similar actions were investigated. Results demonstrate that goal anticipation during action observation is dependent on manual ability, both on a group level (only 25-month-olds solved the manual task and anticipated the goal during observation) and individually within the older age group (r (xy) = 0.53). These findings suggests a connection between manual ability and the ability to anticipate the goal of others' actions in toddlers, in accordance with the direct matching hypothesis. PMID:20041233

  12. Corrective Action Plan for Corrective Action Unit 490: Station 44 Burn Area, Tonopah Test Range, Nevada

    SciTech Connect

    K. B. Campbell

    2002-04-01

    Corrective Action Unit (CAU) 490, Station 44 Burn Area is located on the Tonopah Test Range (TTR). CAU 490 is listed in the Federal Facility Agreement and Consent Order (FFACO, 1996) and includes for Corrective Action Sites (CASs): (1) Fire Training Area (CAS 03-56-001-03BA); (2) Station 44 Burn Area (CAS RG-56-001-RGBA); (3) Sandia Service Yard (CAS 03-58-001-03FN); and (4) Gun Propellant Burn Area (CAS 09-54-001-09L2).

  13. Closure Report for Corrective Action Unit 134: Aboveground Storage Tanks, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2009-06-30

    Corrective Action Unit (CAU) 134 is identified in the Federal Facility Agreement and Consent Order (FFACO) as “Aboveground Storage Tanks” and consists of the following four Corrective Action Sites (CASs), located in Areas 3, 15, and 29 of the Nevada Test Site: · CAS 03-01-03, Aboveground Storage Tank · CAS 03-01-04, Tank · CAS 15-01-05, Aboveground Storage Tank · CAS 29-01-01, Hydrocarbon Stain

  14. CRISPR-Cas: evolution of an RNA-based adaptive immunity system in prokaryotes.

    PubMed

    Koonin, Eugene V; Makarova, Kira S

    2013-05-01

    The CRISPR-Cas (clustered regularly interspaced short palindromic repeats, CRISPR-associated genes) is an adaptive immunity system in bacteria and archaea that functions via a distinct self-non-self recognition mechanism that is partially analogous to the mechanism of eukaryotic RNA interference (RNAi). The CRISPR-Cas system incorporates fragments of virus or plasmid DNA into the CRISPR repeat cassettes and employs the processed transcripts of these spacers as guide RNAs to cleave the cognate foreign DNA or RNA. The Cas proteins, however, are not homologous to the proteins involved in RNAi and comprise numerous, highly diverged families. The majority of the Cas proteins contain diverse variants of the RNA recognition motif (RRM), a widespread RNA-binding domain. Despite the fast evolution that is typical of the cas genes, the presence of diverse versions of the RRM in most Cas proteins provides for a simple scenario for the evolution of the three distinct types of CRISPR-cas systems. In addition to several proteins that are directly implicated in the immune response, the cas genes encode a variety of proteins that are homologous to prokaryotic toxins that typically possess nuclease activity. The predicted toxins associated with CRISPR-Cas systems include the essential Cas2 protein, proteins of COG1517 that, in addition to a ligand-binding domain and a helix-turn-helix domain, typically contain different nuclease domains and several other predicted nucleases. The tight association of the CRISPR-Cas immunity systems with predicted toxins that, upon activation, would induce dormancy or cell death suggests that adaptive immunity and dormancy/suicide response are functionally coupled. Such coupling could manifest in the persistence state being induced and potentially providing conditions for more effective action of the immune system or in cell death being triggered when immunity fails.

  15. An Approach to the Study of Systems of Equations with Geogebra: Learning Opportunities Provided by the Integration of CAS View: Story of a Workshop Experience with Teachers

    ERIC Educational Resources Information Center

    Alejandra, Almirón; Fernando, Bifano; Leonardo, Lupinacci

    2015-01-01

    Solving systems of equations at school, at least in Argentina, is usually a task that students are given as a series of techniques that "allow" them to find a solution. How to overcome educational obstacles that are generated from a fragmented approach of knowledge? What can DGS do, in particular the CAS environment? What epistemic and…

  16. Closure Report for Corrective Action Unit 350: Miscellaneous Housekeeping Sites, Nevada Test Site, Nevada

    SciTech Connect

    R. B. Jackson

    2003-05-01

    This Closure Report documents the closure activities conducted for Corrective Action Unit (CAU) 350: Miscellaneous Housekeeping sites. CAU 350 is listed in Appendix III of the Federal Facility Agreement and Consent Order (FFACO) (FFACO, 1996) and consists of the following eight Corrective Action Sites (CASs) located in Areas 12 and 15 of the Nevada Test Site (NTS): CAS 12-26-01, Lead Shot; CAS 15-22-04, Drums(2); CAS 15-22-06, Drums(10); CAS 15-22-16, Drums(3); CAS 15-22-22, Hydrocarbon Impacted Soil; CAS 15-22-29, Drums(2); CAS 15-24-07, Batteries; and CAS 15-99-02, Gas Cylinder. Closure activities consisted of closing each CAS by removing debris and/or material, disposing of the generated waste, and verifying that each site was clean-closed by visual inspection and/or laboratory analysis of soil samples.

  17. Corrective Action Plan for Corrective Action Unit 562: Waste Systems, Nevada National Security Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2011-04-30

    This Corrective Action Plan has been prepared for Corrective Action Unit (CAU) 562, Waste Systems, in accordance with the Federal Facility Agreement and Consent Order (1996; as amended March 2010). CAU 562 consists of 13 Corrective Action Sites (CASs) located in Areas 2, 23, and 25 of the Nevada National Security Site. Site characterization activities were performed in 2009 and 2010, and the results are presented in Appendix A of the Corrective Action Decision Document for CAU 562. The scope of work required to implement the recommended closure alternatives is summarized. (1) CAS 02-26-11, Lead Shot, will be clean closed by removing shot. (2) CAS 02-44-02, Paint Spills and French Drain, will be clean closed by removing paint and contaminated soil. As a best management practice (BMP), asbestos tile will be removed. (3) CAS 02-59-01, Septic System, will be clean closed by removing septic tank contents. As a BMP, the septic tank will be removed. (4) CAS 02-60-01, Concrete Drain, contains no contaminants of concern (COCs) above action levels. No further action is required; however, as a BMP, the concrete drain will be removed. (5) CAS 02-60-02, French Drain, was clean closed. Corrective actions were completed during corrective action investigation activities. As a BMP, the drain grates and drain pipe will be removed. (6) CAS 02-60-03, Steam Cleaning Drain, will be clean closed by removing contaminated soil. As a BMP, the steam cleaning sump grate and outfall pipe will be removed. (7) CAS 02-60-04, French Drain, was clean closed. Corrective actions were completed during corrective action investigation activities. (8) CAS 02-60-05, French Drain, will be clean closed by removing contaminated soil. (9) CAS 02-60-06, French Drain, contains no COCs above action levels. No further action is required. (10) CAS 02-60-07, French Drain, requires no further action. The french drain identified in historical documentation was not located during corrective action investigation

  18. Closure Report for Corrective Action Unit 398: Area 25 Spill Sites, Nevada Test Site, Nevada

    SciTech Connect

    K. B. Campbell

    2003-04-01

    This Closure Report (CR) documents the activities performed to close Corrective Action Unit (CAU) 398: Area 25 Spill Sites, in accordance with the Federal Facility Agreement and Consent Order (FFACO) of 1996, and the Nevada Division of Environmental Protection (NDEP)-approved Streamlined Approach for Environmental Restoration (SA4FER) Plan for CAU 398: Area 25 Spill Sites, Nevada Test Site, Nevada (U.S. Department of Energy, Nevada Operations Office [DOEN], 2001). CAU 398 consists of the following thirteen Corrective Action Sites (CASs) all located in Area 25 of the Nevada Test Site (NTS) (Figure 1): CAS 25-25-02, Oil Spills, CAS 25-25-03, Oil Spills, CAS 25-25-04, Oil Spills, CAS 25-25-05, Oil Spills, CAS 25-25-06, Oil Spills, CAS 25-25-07, Hydraulic Oil Spill(s), CAS 25-25-08, Hydraulic Oil Spill(s), CAS 25-25-16, Diesel Spill (from CAS 25-01-02), CAS 25-25-17, Subsurface Hydraulic Oil Spill, CAS 25-44-0 1, Fuel Spill, CAS 25-44-04, Acid Spill (from CAS 25-01-01), CAS 25-44-02, Spill, and CAS 25-44-03, Spill. Copies of the analytical results for the site verification samples are included in Appendix B. Copies of the CAU Use Restriction Information forms are included in Appendix C.

  19. Closure Report for Corrective Action Unit 346: Areas 8, 10 Housekeeping Sites, Nevada Test Site, Nevada

    SciTech Connect

    K. B. Campbell

    2003-08-01

    This Closure Report documents the closure activities conducted for Corrective Action Unit (CAU) 346: Areas 8, 10 Housekeeping Sites. CAU 346 is listed in Appendix III of the Federal Facility Agreement and Consent Order (FFACO, 1996) and consists of the following 14 Corrective Action Sites (CASs) located in Areas 8 and 10 of the Nevada Test Site (NTS): (1) CAS 08-22-04: Drums (2); (2) CAS 08-22-11: Drums; Bucket; (3) CAS 08-24-02: Battery; (4) CAS 10-14-01: Transformer; (5) CAS 10-22-06: Drum (Gas Block); (6) CAS 10-22-10: Drum (Gas Block); (7) CAS 10-22-12: Drum (Gas Block); (8) CAS 10-22-13: Drum (Gas Block); (9) CAS 10-22-16: Drum (Gas Block); (10) CAS 10-22-22: Drum; (11) CAS 10-22-25: Drum; (12) CAS 10-22-36: Paint Can; (13) CAS 10-22-37: Gas Block; and (14) CAS 10-24-11: Battery. Closure activities consisted of closing each CAS by removing debris and/or material, disposing of the generated waste, and verifying that each site was clean-closed by visual inspection and/or laboratory analysis of soil verification samples.

  20. A Study of Hypergiant Mass Loss in the Near-To-Mid Infrared: VY CMa, IRC +10420, mu Cep and rho Cas

    NASA Astrophysics Data System (ADS)

    Shenoy, Dinesh Prabhakar

    2016-01-01

    Stars of initial mass greater than 9 M_sun become red supergiants (RSGs), a short-lived stage during which they experience mass-loss that strongly influences their post-RSG evolution and end state. The highest luminosity RSGs, referred to here as hypergiants, experience episodic mass-loss whose mechanism remains poorly understood and motivates observations to help constrain it. This thesis studies mass loss from hypergiant stars with near-to-mid infrared imaging over a range of angular scales. The recent mass-loss history of the extreme red supergiant VY Canis Majoris and the warm hypergiant star IRC +10420 are studied at the sub-arcsecond scale with adaptive optics imaging and imaging polarimetry from 1 - 5 micron using LMIRCam on the Large Binocular Telescope (LBT) and MMT-Pol at the MMT Observatory. The nebular features of VY CMa are found to be highly polarized at 1.3 and 3.1 micron, with optically thick scattering required to reproduce the observed surface brightness. The flux of VY CMa's peculiar ``Southwest Clump'' is demonstrated to be due almost entirely to optically thick scattering, with little thermal emission, and with a lower limit mass of 5E-03 M_sun in this single feature. The imaging polarimetry of IRC +10420 at 2.2 micron resolves nebular emission with intrinsic polarization of 30%, with a high surface brightness indicating optically thick scattering largely in the plane of the sky. Using the polarimetry to constrain the scattered light emission, it is shown that the nebula's the emission is mostly thermal with a color temperature well above that for typical astrophysical dust. To probe further into hypergiants' history of mass-loss, mid-IR imaging with MMT/ MIRAC and SOFIA/FORCAST is used to study VY CMa, IRC +10420 and two additional hypergiants: the RSG mu Cep and the warm hypergiant rho Cas. Using DUSTY 1-D radiative transfer models, mu Cep's mass-loss rate is found to have declined by about a factor of 5 over a 13,000 history, ranging from 5E

  1. Neuronal and immunological basis of action of antidepressants in chronic pain - clinical and experimental studies.

    PubMed

    Mika, Joanna; Zychowska, Magdalena; Makuch, Wioletta; Rojewska, Ewelina; Przewlocka, Barbara

    2013-01-01

    The current knowledge of the pharmacological actions of the tricyclic antidepressants (TCAs) has slowly evolved through their over 40-year history. Chronic pain represents one of the most important public health problems, and antidepressants are an essential part of the therapeutic strategy in addition to classical analgesics. This article reviews the available evidence on the efficacy and safety of antidepressants in chronic pain conditions; namely, headaches, low back pain, fibromyalgia, cancer pain and especially neuropathic pain. TCAs are traditionally the main type of depression medication used to treat chronic pain. Recently, new antidepressants were introduced into clinical use, with a significant reduction in side effects and equivalent efficacy on mood disorders. These new drugs that are effective for chronic pain belong to the tetracyclic antidepressants (TeCAs) group (amoxapine, maprotiline), the serotonin and noradrenaline reuptake inhibitors (SNRIs) group (duloxetine, venlafaxine, milnacipran) and the atypical antidepressants group (bupropion, trazodone, mirtazapine, nefazodone). In this review, we present the available publications on TCAs (amitriptyline, doxepin, imipramine, desipramine, nortriptyline), TeCAs (amoxapine, maprotiline), selective serotonin reuptake inhibitors (SSRIs) (citalopram, fluoxetine, paroxetine), SNRIs (duloxetine, venlafaxine, milnacipran) and atypical antidepressants (bupropion) for the treatment of neuropathic pain. We also review analgesics acting as both opioid receptor agonists and also acting as aminergic reuptake inhibitors. Existing data are insufficient to conclude which of these new classes of antidepressants has the best clinical profile and will be the most effective in the treatment of neuropathic pain; in addition, a lower incidence of side effects should be considered. Increased experimental and translational research is a key for further improvement of the treatment of chronic pain with antidepressants. However

  2. A Self-Study of the Teaching of Action Research in a University Context

    ERIC Educational Resources Information Center

    Choi, Jung-ah

    2011-01-01

    Despite the potential benefits of action research, teaching action research in a university setting can present challenges. Analyzing my own experiences of teaching a university-based course on action research, this self-study investigates what my students (all classroom teachers) did and did not understand about action research and what hindered…

  3. [Application of CRISPR/Cas9 mediated genome editing in farm animals].

    PubMed

    Yuyun, Xing; Qiang, Yang; Jun, Ren

    2016-03-01

    CRISPR (Clustered regularly interspaced short palindromic repeats)/Cas (CRISPR associated proteins) is an acquired immune system found in bacteria and archaea that fight against invasion of viruses or plasmids. CRISPR/Cas systems are currently classified into three main types: I, II and III, of which type II has relatively simple components. The CRISPR/Cas9 technology modified from type II CRISPR/Cas system has been developed as an efficient genome editing tool. Since the initial application of the CRISPR/Cas9 technology in mammals in 2013, the reports of this system for genomic editing has skyrocketed. Farm animals are not only economically important animals, but also ideal animal models for human diseases and biomedical studies. In this review, we summarize the applications of CRISPR/Cas9 in farm animals, briefly describe the off-target effects and the main solutions, and finally highlight the future perspectives of this technology.

  4. Study of Synthetic Vision Systems (SVS) and Velocity-vector Based Command Augmentation System (V-CAS) on Pilot Performance

    NASA Technical Reports Server (NTRS)

    Liu, Dahai; Goodrich, Ken; Peak, Bob

    2006-01-01

    This study investigated the effects of synthetic vision system (SVS) concepts and advanced flight controls on single pilot performance (SPP). Specifically, we evaluated the benefits and interactions of two levels of terrain portrayal, guidance symbology, and control-system response type on SPP in the context of lower-landing minima (LLM) approaches. Performance measures consisted of flight technical error (FTE) and pilot perceived workload. In this study, pilot rating, control type, and guidance symbology were not found to significantly affect FTE or workload. It is likely that transfer from prior experience, limited scope of the evaluation task, specific implementation limitations, and limited sample size were major factors in obtaining these results.

  5. BCAR3/AND-34 can signal independent of complex formation with CAS family members or the presence of p130Cas

    PubMed Central

    Borre, Pierre Vanden; Near, Richard I.; Makkinje, Anthony; Mostoslavsky, Gustavo; Lerner, Adam

    2011-01-01

    BCAR3 binds to the carboxy-terminus of p130Cas, a focal adhesion adapter protein. Both BCAR3 and p130Cas have been linked to resistance to anti-estrogens in breast cancer, Rac activation and cell motility. Using R743A BCAR3, a point mutant that has lost the ability to bind p130Cas, we find that BCAR3-p130Cas complex formation is not required for BCAR3-mediated anti-estrogen resistance, Rac activation or discohesion of epithelial breast cancer cells. Complex formation was also not required for BCAR3-induced lamellipodia formation in BALB/c-3T3 fibroblasts but was required for optimal BCAR3-induced motility. Although both wildtype and R743A BCAR3 induced phosphorylation of p130Cas and the related adapter protein HEF1/NEDD9, chimeric NSP3:BCAR3 experiments demonstrate that such phosphorylation does not correlate with BCAR3-induced anti-estrogen resistance or lamellipodia formation. Wildtype but not R743A BCAR3 induced lamellipodia formation and augmented cell motility in p130Cas-/- murine embryonic fibroblasts (MEFs), suggesting that while p130Cas itself is not strictly required for these endpoints, complex formation with other CAS family members is, at least in cells lacking p130Cas. Overall, our work suggests that many, but not all, BCAR3-mediated signaling events in epithelial and mesenchymal cells are independent of p130Cas association. These studies also indicate that disruption of the BCAR3-p130Cas complex is unlikely to reverse BCAR3-mediated anti-estrogen resistance. PMID:21262352

  6. Observational Study Of The Pacific Western Boundary Currents And The Indonesian Throughflow by the CAS Strategic Priority Project

    NASA Astrophysics Data System (ADS)

    Yuan, D.; Wang, J.

    2014-12-01

    The warm pool in the western Pacific Ocean has significant impact on the evolution of ENSO and the East Asian monsoon. Ocean circulation in the western Pacific Ocean and in Indonesian seas plays an important role in the interannual climate variations and predictability of the tropical Indo-Pacific Ocean. A major observational program of the Chinese Academy of Sciences is recently launched to study the western Pacific Ocean circulation and the warm pool to test these scientific hypotheses. The physical oceanography project called the "Western Pacific Ocean Circulation and the Warm Pool Variability" is by far the largest and the most intensive observational program in history in the western Pacific ocean study. In this talk, the background and scientific hypotheses of the project, the observational design in the western Pacific Ocean, Indonesian seas, and the eastern Indian Ocean region, and some preliminary results of the program will be presented. The talk serves to encourage more scientists to collaborate in the studies of the ocean circulation and climate in the western Pacific and eastern Indian Oceans.

  7. A Cloud-Resolving Modeling Intercomparison Study on Properties of Cloud Microphysics, Convection, and Precipitation for a Squall Line Cas

    NASA Astrophysics Data System (ADS)

    Fan, J.; Han, B.; Morrison, H.; Varble, A.; Mansell, E.; Milbrandt, J.; Wang, Y.; Lin, Y.; Dong, X.; Giangrande, S. E.; Jensen, M. P.; Collis, S. M.; North, K.; Kollias, P.

    2015-12-01

    The large spread in CRM model simulations of deep convection and aerosol effects on deep convective clouds (DCCs) makes it difficult (1) to further our understanding of deep convection and (2) to define "benchmarks" and recommendations for their use in parameterization developments. Past model intercomparison studies used different models with different complexities of dynamic-microphysics interactions, making it hard to isolate the causes of differences between simulations. In this intercomparison study, we employed a much more constrained approach - with the same model and same experiment setups for simulations with different cloud microphysics schemes (one-moment, two-moment, and bin models). Both the piggybacking and interactive approaches are employed to explore the major microphysical processes that control the model differences and the significance of their feedback to dynamics through latent heating/cooling and cold pool characteristics. Real-case simulations are conducted for the squall line case 20 May 2011 from the MC3E field campaign. Results from the piggybacking approach show substantially different responses of the microphysics schemes to the same dynamical fields. Although the interactive microphysics-dynamics simulations buffer some differences compared with those from the piggyback runs, large differences still exist and are mainly contributed by ice microphysical processes parameterizations. The presentation will include in-depth analyses of the major microphysical processes for the squall line case, the significance of the feedback of the processes to dynamics, and how those results differ in different cloud microphysics schemes.

  8. CAS-Induced Difficulties in Learning Mathematics?

    ERIC Educational Resources Information Center

    Jankvist, Uffe Thomas; Misfeldt, Morten

    2015-01-01

    In recent years computer algebra systems (CAS) have become an integrated part of the upper secondary school mathematics program. Despite the many positive possibilities of CAS, there also seems to be a flip side of the coin in relation to actual difficulties in learning mathematics, not least because a strong dependence on CAS for mathematical…

  9. Prospects and challenges of CRISPR/Cas genome editing for the study and control of neglected vector-borne nematode diseases.

    PubMed

    Zamanian, Mostafa; Andersen, Erik C

    2016-09-01

    Neglected tropical diseases caused by parasitic nematodes inflict an immense health and socioeconomic burden throughout much of the developing world. Current estimates indicate that more than two billion people are infected with nematodes, resulting in the loss of 14 million disability-adjusted life years per annum. Although these parasites cause significant mortality, they primarily cause chronic morbidity through a wide range of severe clinical ailments. Treatment options for nematode infections are restricted to a small number of anthelmintic drugs, and the rapid expansion of anthelmintic mass drug administration raises concerns of drug resistance. Preservation of existing drugs is necessary, as well as the development of new treatment options and methods of control. We focus this review on how the democratization of CRISPR/Cas9 genome editing technology can be enlisted to improve our understanding of the biology of nematode parasites and our ability to treat the infections they cause. We will first explore how this robust method of genome manipulation can be used to newly exploit the powerful model nematode Caenorhabditis elegans for parasitology research. We will then discuss potential avenues to develop CRISPR/Cas9 editing protocols in filarial nematodes. Lastly, we will propose potential ways in which CRISPR/Cas9 can be used to engineer gene drives that target the transmission of mosquito-borne filarial nematodes. PMID:27300487

  10. Action anticipation beyond the action observation network: a functional magnetic resonance imaging study in expert basketball players.

    PubMed

    Abreu, A M; Macaluso, E; Azevedo, R T; Cesari, P; Urgesi, C; Aglioti, S M

    2012-05-01

    The ability to predict the actions of others is quintessential for effective social interactions, particularly in competitive contexts (e.g. in sport) when knowledge about upcoming movements allows anticipating rather than reacting to opponents. Studies suggest that we predict what others are doing by using our own motor system as an internal forward model and that the fronto-parietal action observation network (AON) is fundamental for this ability. However, multiple-duty cells dealing with action perception and execution have been found in a variety of cortical regions. Here we used functional magnetic resonance imaging to explore, in expert basketball athletes and novices, whether the ability to make early predictions about the fate of sport-specific actions (i.e. free throws) is underpinned by neural regions beyond the classical AON. We found that, although involved in action prediction, the fronto-parietal AON was similarly activated in novices and experts. Importantly, athletes exhibited relatively greater activity in the extrastriate body area during the prediction task, probably due to their expert reading of the observed action kinematics. Moreover, experts exhibited higher activation in the bilateral inferior frontal gyrus and in the right anterior insular cortex when producing errors, suggesting that they might become aware of their own errors. Correct action prediction induced higher posterior insular cortex activity in experts and higher orbito-frontal activity in novices, suggesting that body awareness is important for performance monitoring in experts, whereas novices rely more on higher-order decision-making strategies. This functional reorganization highlights the tight relationship between action anticipation, error awareness and motor expertise leading to body-related processing and differences in decision-making processes.

  11. How Action and Context Priming Influence Categorization: A Developmental Study

    ERIC Educational Resources Information Center

    Kalenine, Solene; Bonthoux, Francoise; Borghi, Anna M.

    2009-01-01

    Embodied views of cognition propose that concepts are grounded in sensorimotor experience. Diverse aspects of sensorimotor experience, like action and context information, could play a key role in the formation and processing of manipulable object concepts. Specifically, contextual information could help to link specific actions experienced with…

  12. Insight and Action Analytics: Three Case Studies to Consider

    ERIC Educational Resources Information Center

    Milliron, Mark David; Malcolm, Laura; Kil, David

    2014-01-01

    Civitas Learning was conceived as a community of practice, bringing together forward-thinking leaders from diverse higher education institutions to leverage insight and action analytics in their ongoing efforts to help students learn well and finish strong. We define insight and action analytics as drawing, federating, and analyzing data from…

  13. Phonological Awareness and Early Reading Development in Childhood Apraxia of Speech (CAS)

    ERIC Educational Resources Information Center

    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…

  14. Computational Neural Modeling of Speech Motor Control in Childhood Apraxia of Speech (CAS)

    ERIC Educational Resources Information Center

    Terband, Hayo; Maassen, Ben; Guenther, Frank H.; Brumberg, Jonathan

    2009-01-01

    Purpose: Childhood apraxia of speech (CAS) has been associated with a wide variety of diagnostic descriptions and has been shown to involve different symptoms during successive stages of development. In the present study, the authors attempted to associate the symptoms of CAS in a particular developmental stage with particular…

  15. Nitazoxanide: Nematicidal Mode of Action and Drug Combination Studies

    PubMed Central

    Somvanshi, Vishal S.; Ellis, Brian L.; Hu, Yan; Aroian, Raffi V.

    2014-01-01

    Intestinal nematodes or roundworms (aka soil-transmitted helminths or STHs) cause great disease. They infect upwards of two billion people, leading to high morbidity and a range of health problems, especially in infected children and pregnant women. Development of resistance to the two main classes of drugs used to treat intestinal nematode infections of humans has been reported. To fight STH infections, we need new and more effective drugs and ways to improve the efficacy of the old drugs. One promising alternative drug is nitazoxanide (NTZ). NTZ, approved for treating human protozoan infections, was serendipitously shown to have therapeutic activity against STHs. However, its mechanism of action against nematodes is not known. Using the laboratory nematode Caenorhabditis elegans, we show that NTZ acts on the nematodes through avr-14, an alpha-type subunit of a glutamate-gated chloride ion channel known for its role in ivermectin susceptibility. In addition, a forward genetic screen to select C. elegans mutants resistant to NTZ resulted in isolation of two NTZ resistant mutants that are not in avr-14, suggesting that additional mechanisms are involved in resistance to NTZ. We found that NTZ combines synergistically with other classes of anthelmintic drugs, i.e. albendazole and pyrantel, making it a good candidate for further studies on its use in drug combination therapy of STH infections. Given NTZ acts against a wide range of nematode parasites, our findings also validate avr-14 as an excellent target for pan-STH therapy. PMID:24412397

  16. Acceptable Risk? The Nuclear Age in the United States. Study/Action Guide [and] Companion to Study/Action Guide for Congregations and Religious Groups.

    ERIC Educational Resources Information Center

    American Friends Service Committee, Philadelphia, PA. National Action/Research on the Military Industrial Complex.

    A study-action guide and a companion guide are intended to help citizens explore some of the challenging dilemmas of U.S. nuclear policy. The two guides place strong emphasis on group discussion and participation as well as action citizens might want to take to bring about a non-nuclear world. The companion guide is intended for congregations and…

  17. The RNA- and DNA-targeting CRISPR–Cas immune systems of Pyrococcus furiosus

    PubMed Central

    Terns, Rebecca M.; Terns, Michael P.

    2014-01-01

    Using the hyperthermophile Pyrococcus furiosus, we have delineated several key steps in CRISPR (clustered regularly interspaced short palindromic repeats)–Cas (CRISPR-associated) invader defence pathways. P. furiosus has seven transcriptionally active CRISPR loci that together encode a total of 200 crRNAs (CRISPR RNAs). The 27 Cas proteins in this organism represent three distinct pathways and are primarily encoded in two large gene clusters. The Cas6 protein dices CRISPR locus transcripts to generate individual invader-targeting crRNAs. The mature crRNAs include a signature sequence element (the 5′ tag) derived from the CRISPR locus repeat sequence that is important for function. crRNAs are tailored into distinct species and integrated into three distinct crRNA–Cas protein complexes that are all candidate effector complexes. The complex formed by the Cmr [Cas module RAMP (repeat-associated mysterious proteins)] (subtype III-B) proteins cleaves complementary target RNAs and can be programmed to cleave novel target RNAs in a prokaryotic RNAi-like manner. Evidence suggests that the other two CRISPR–Cas systems in P. furiosus, Csa (Cas subtype Apern) (subtype I-A) and Cst (Cas subtype Tneap) (subtype I-B), target invaders at the DNA level. Studies of the CRISPR–Cas systems from P. furiosus are yielding fundamental knowledge of mechanisms of crRNA biogenesis and silencing for three of the diverse CRISPR–Cas pathways, and reveal that organisms such as P. furiosus possess an arsenal of multiple RNA-guided mechanisms to resist diverse invaders. Our knowledge of the fascinating CRISPR–Cas pathways is leading in turn to our ability to co-opt these systems for exciting new biomedical and biotechnological applications. PMID:24256230

  18. Students' use of CAS calculators - effects on the trustworthiness and fairness of mathematics assessments

    NASA Astrophysics Data System (ADS)

    Lind Pantzare, Anna

    2012-10-01

    Calculators with computer algebra systems (CAS) are powerful tools when working with equations and algebraic expressions in mathematics. When calculators are allowed to be used during assessments but are not available or provided to every student, they may cause bias. The CAS calculators may also have an impact on the trustworthiness of results. In this study students' use of the CAS calculator in their work with released assessment items from TIMSS Advanced 2008 is studied using two approaches. Eight students familiar with CAS, from two mathematics classes in the 12th form, were video filmed when encouraged to think aloud during their work with the items. In addition, a questionnaire was distributed to all 33 students in the two classes who had been working with a CAS. The main finding is that even if the students are used to working with the CAS calculator, they are not using the calculator to a large extent. The analysis indicates that the difference in performance between the high- and low-achieving students has slightly increased due to the use of the calculator. From a validity perspective one could therefore argue that the CAS calculator is no major threat to the trustworthiness of the assessment. Nevertheless, the result indicates that those students in the study, mainly high achieving, who know how to use the CAS calculator, get an additional advantage. The advantage brings an amount of unfairness into the assessment and could be a threat to the trustworthiness and fairness.

  19. Exploring Coaching Actions Based on Developed Values: A Case Study of a Female Hockey Coach

    ERIC Educational Resources Information Center

    Callary, Bettina; Werthner, Penny; Trudel, Pierre

    2013-01-01

    There are few empirical studies that demonstrate how values are developed and how they are linked to coaching actions. There can be a discrepancy between the statement of coaches' values and their actual coaching actions. In order to examine how coaching actions are influenced by values that are developed over a lifetime, the purpose of this…

  20. Mode of action and clinical studies with alemtuzumab.

    PubMed

    Jones, Joanne L; Coles, Alasdair J

    2014-12-01

    The lymphocyte depleting anti-CD52 monoclonal antibody alemtuzumab has been used in Cambridge, UK, as an experimental treatment of multiple sclerosis since 1991. One phase-2 trial (CAMMS-223) and two phase-3 studies (CARE-MS1 and CARE-MS2) have confirmed its efficacy in treatment-naive patients, and have established superiority over interferon beta-1a in patients who continue to relapse in spite of first-line therapy (Cohen et al., 2012; Coles et al., 2008; Coles et al., 2012a; Coles et al., 2012b). Despite causing a prolonged T cell lymphopenia, significant infections have not been an issue following treatment; rather alemtuzumab's primary safety concern is secondary autoimmunity, occurring up to five years after treatment and maximally at two years: 30% of patients develops thyroid autoimmunity, and 1% develops idiopathic thrombocytopenic purpura (ITP). In addition, 4 out of 1486 patients (<0.3%) treated on the commercially sponsored studies developed glomerulonephritis. Two of these patients developed anti-glomerular basement membrane disease, a condition which may result in renal failure unless treated aggressively. In September 2013, the European Medicine Agency (EMA) ruled that the benefit-to-risk balance for alemtuzumab was favourable, approving it as a first-line therapy for adults with active relapsing remitting multiple sclerosis (under the trade name Lemtrada). Lemtrada is now also approved as a treatment of multiple sclerosis in Canada, Australia, Switzerland, Israel, Mexico and Brazil. However, in December 2013, Lemtrada failed to gain approval from the U.S. Food and Drug Administration (FDA), with concerns over trial design and safety stated as the main reasons. In this review we describe our local experience and explain the rationale behind its initial use as a treatment of multiple sclerosis and behind the design of the commercially sponsored trials, summarising their key findings. We also sum up our understanding of its mechanism of action.

  1. Light Curves and Analyses of the Eclipsing Binaries EG Cas and EP Cas

    NASA Astrophysics Data System (ADS)

    Bradstreet, David H.; Sanders, S. J.; McClain, T. R.

    2010-01-01

    New precision V & Rc light curves of the eclipsing binaries EG Cas and EP Cas have been obtained using the 41-cm telescope at the Eastern University Observatory equipped with an SBIG ST-10XME CCD. EG Cas (P = 0.6115 days, Vmax = 12.9) has no published light curves and only a few dozen (mostly visual) timings of minimum light. The system is being observed throughout the fall of 2009 and the current light curves distinctly show that the system is a totally eclipsing overcontact binary. The light curves are also significantly asymmetric (strong O'Connell effect) indicating the presence of large, cool starspots, most likely on both stars. Preliminary analysis indicates that the binary is an A-type (the larger, more massive star is the hotter component), has a mass ratio of 0.32, very large temperature difference between the stars greater than 1600 K, and a fillout of 0.26. The modern timings of minimum light combined with those in the literature indicate that the binary's period is decreasing. The large temperature difference coupled to a significant fillout factor seems contradictory, and further data acquisition and analysis will hopefully resolve this seeming enigma. EP Cas (P = 0.8134 days, Vmax = 11.2) is a partially eclipsing detached system with a relatively deep primary eclipse of 1.0 mag in Rc. No published light curves exist for this system although many timings of minimum light have been published. The O-C curve indicates that the period for the binary has remained relatively constant since observations were first published in 1936. Preliminary light curve models indicate a partially eclipsing system consisting of detached but tidally distorted stars. The complete light curve analyses as well as a period study of all published times of minimum light will be presented for both systems.

  2. CRISPR/Cas9 mediates efficient conditional mutagenesis in Drosophila.

    PubMed

    Xue, Zhaoyu; Wu, Menghua; Wen, Kejia; Ren, Menda; Long, Li; Zhang, Xuedi; Gao, Guanjun

    2014-11-01

    Existing transgenic RNA interference (RNAi) methods greatly facilitate functional genome studies via controlled silencing of targeted mRNA in Drosophila. Although the RNAi approach is extremely powerful, concerns still linger about its low efficiency. Here, we developed a CRISPR/Cas9-mediated conditional mutagenesis system by combining tissue-specific expression of Cas9 driven by the Gal4/upstream activating site system with various ubiquitously expressed guide RNA transgenes to effectively inactivate gene expression in a temporally and spatially controlled manner. Furthermore, by including multiple guide RNAs in a transgenic vector to target a single gene, we achieved a high degree of gene mutagenesis in specific tissues. The CRISPR/Cas9-mediated conditional mutagenesis system provides a simple and effective tool for gene function analysis, and complements the existing RNAi approach. PMID:25193494

  3. CRISPR/Cas9 Genome Editing in Embryonic Stem Cells.

    PubMed

    Andrey, Guillaume; Spielmann, Malte

    2017-01-01

    Targeted mutagenesis is required to evaluate the function of DNA segments across the genome. In recent years the CRISPR/Cas9 technology has been widely used for functional genome studies and is partially replacing classical homologous recombination methods in different aspects. CRISPR/Cas9-derived tools indeed allow the production of a wide-range of engineered mutations: from point mutations to large chromosomal rearrangements such as deletions, duplications and inversions. Here we present a protocol to engineer Embryonic Stem Cells (ESC) with desired mutations using transfection of custom-made CRISPR/Cas9 vectors. These methods allow the in vivo modeling of congenital mutations and the functional interrogation of DNA sequences. PMID:27662879

  4. Harnessing CRISPR-Cas systems for bacterial genome editing.

    PubMed

    Selle, Kurt; Barrangou, Rodolphe

    2015-04-01

    Manipulation of genomic sequences facilitates the identification and characterization of key genetic determinants in the investigation of biological processes. Genome editing via clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) constitutes a next-generation method for programmable and high-throughput functional genomics. CRISPR-Cas systems are readily reprogrammed to induce sequence-specific DNA breaks at target loci, resulting in fixed mutations via host-dependent DNA repair mechanisms. Although bacterial genome editing is a relatively unexplored and underrepresented application of CRISPR-Cas systems, recent studies provide valuable insights for the widespread future implementation of this technology. This review summarizes recent progress in bacterial genome editing and identifies fundamental genetic and phenotypic outcomes of CRISPR targeting in bacteria, in the context of tool development, genome homeostasis, and DNA repair.

  5. Corrective Action Plan for Corrective Action Unit 139: Waste Disposal Sites, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2007-07-01

    Corrective Action Unit (CAU) 139, Waste Disposal Sites, is listed in the Federal Facility Agreement and Consent Order (FFACO) of 1996 (FFACO, 1996). CAU 139 consists of seven Corrective Action Sites (CASs) located in Areas 3, 4, 6, and 9 of the Nevada Test Site (NTS), which is located approximately 65 miles (mi) northwest of Las Vegas, Nevada (Figure 1). CAU 139 consists of the following CASs: CAS 03-35-01, Burn Pit; CAS 04-08-02, Waste Disposal Site; CAS 04-99-01, Contaminated Surface Debris; CAS 06-19-02, Waste Disposal Site/Burn Pit; CAS 06-19-03, Waste Disposal Trenches; CAS 09-23-01, Area 9 Gravel Gertie; and CAS 09-34-01, Underground Detection Station. Details of the site history and site characterization results for CAU 139 are provided in the approved Corrective Action Investigation Plan (CAIP) (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2006) and in the approved Corrective Action Decision Document (CADD) (NNSA/NSO, 2007). The purpose of this Corrective Action Plan (CAP) is to present the detailed scope of work required to implement the recommended corrective actions as specified in Section 4.0 of the approved CADD (NNSA/NSO, 2007). The approved closure activities for CAU 139 include removal of soil and debris contaminated with plutonium (Pu)-239, excavation of geophysical anomalies, removal of surface debris, construction of an engineered soil cover, and implementation of use restrictions (URs). Table 1 presents a summary of CAS-specific closure activities and contaminants of concern (COCs). Specific details of the corrective actions to be performed at each CAS are presented in Section 2.0 of this report.

  6. Spectroscopic Observations of UU Cas

    NASA Astrophysics Data System (ADS)

    Markov, H.; Vince, I.; Markova, N.; Djurasevic, G.

    2010-09-01

    High dispersion (R = 30000) spectra of the eclipsing binary system UU Cas are presented for the first time. Spectra covering 15 different phases were taken in two spectral regions - the vicinity of H_α and another centered on 5800Å. Seven spectral lines were identified in common. Four of them belonging to the brighter component were used to construct radial velocity curve. Some constrains on the spectral classification of the components are presented.

  7. Expanding the Biologist's Toolkit with CRISPR-Cas9.

    PubMed

    Sternberg, Samuel H; Doudna, Jennifer A

    2015-05-21

    Few discoveries transform a discipline overnight, but biologists today can manipulate cells in ways never possible before, thanks to a peculiar form of prokaryotic adaptive immunity mediated by clustered regularly interspaced short palindromic repeats (CRISPR). From elegant studies that deciphered how these immune systems function in bacteria, researchers quickly uncovered the technological potential of Cas9, an RNA-guided DNA cleaving enzyme, for genome engineering. Here we highlight the recent explosion in visionary applications of CRISPR-Cas9 that promises to usher in a new era of biological understanding and control.

  8. Applications of CRISPR-Cas systems in neuroscience.

    PubMed

    Heidenreich, Matthias; Zhang, Feng

    2016-01-01

    Genome-editing tools, and in particular those based on CRISPR-Cas (clustered regularly interspaced short palindromic repeat (CRISPR)-CRISPR-associated protein) systems, are accelerating the pace of biological research and enabling targeted genetic interrogation in almost any organism and cell type. These tools have opened the door to the development of new model systems for studying the complexity of the nervous system, including animal models and stem cell-derived in vitro models. Precise and efficient gene editing using CRISPR-Cas systems has the potential to advance both basic and translational neuroscience research.

  9. Applications of CRISPR-Cas9 mediated genome engineering.

    PubMed

    Yang, Xiao

    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.

  10. Corrective Action Plan for Corrective Action Unit 543: Liquid Disposal Units, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2006-09-01

    Corrective Action Unit (CAU) 543, Liquid Disposal Units, is listed in Appendix III of the Federal Facility Agreement and Consent Order of 1996. CAU 543 consists of seven Corrective Action Sites (CASs) located in Areas 6 and 15 of the Nevada Test Site, which is approximately 65 miles northwest of Las Vegas, Nevada. CAU 543 consists of the following seven CASs: {sm_bullet} CAS 06-07-01, Decon Pad {sm_bullet} CAS 15-01-03, Aboveground Storage Tank {sm_bullet} CAS 15-04-01, Septic Tank {sm_bullet} CAS 15-05-01, Leachfield {sm_bullet} CAS 15-08-01, Liquid Manure Tank {sm_bullet} CAS 15-23-01, Underground Radioactive Material Area {sm_bullet} CAS 15-23-03, Contaminated Sump, Piping From January 24, 2005 through April 14, 2005, CAU 543 site characterization activities were conducted, and are reported in Appendix A of the CAU 543 Corrective Action Decision Document (CADD) (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2005). The recommended corrective action as stated in the approved CADD is No Further Action for five of the CAU 543 CASs, and Closure In Place for the remaining two CASs.

  11. Breast Cancer Anti-estrogen Resistance 3 (BCAR3) Protein Augments Binding of the c-Src SH3 Domain to Crk-associated Substrate (p130cas)*

    PubMed Central

    Makkinje, Anthony; Vanden Borre, Pierre; Near, Richard I.; Patel, Prayag S.; Lerner, Adam

    2012-01-01

    The focal adhesion adapter protein p130cas regulates adhesion and growth factor-related signaling, in part through Src-mediated tyrosine phosphorylation of p130cas. AND-34/BCAR3, one of three NSP family members, binds the p130cas carboxyl terminus, adjacent to a bipartite p130cas Src-binding domain (SBD) and induces anti-estrogen resistance in breast cancer cell lines as well as phosphorylation of p130cas. Only a subset of the signaling properties of BCAR3, specifically augmented motility, are dependent upon formation of the BCAR3-p130cas complex. Using GST pull-down and immunoprecipitation studies, we show that among NSP family members, only BCAR3 augments the ability of p130cas to bind the Src SH3 domain through an RPLPSPP motif in the p130cas SBD. Although our prior work identified phosphorylation of the serine within the p130cas RPLPSPP motif, mutation of this residue to alanine or glutamic acid did not alter BCAR3-induced Src SH3 domain binding to p130cas. The ability of BCAR3 to augment Src SH3 binding requires formation of a BCAR3-p130cas complex because mutations that reduce association between these two proteins block augmentation of Src SH3 domain binding. Similarly, in MCF-7 cells, BCAR3-induced tyrosine phosphorylation of the p130cas substrate domain, previously shown to be Src-dependent, was reduced by an R743A mutation that blocks BCAR3 association with p130cas. Immunofluorescence studies demonstrate that BCAR3 expression alters the intracellular location of both p130cas and Src and that all three proteins co-localize. Our work suggests that BCAR3 expression may regulate Src signaling in a BCAR3-p130cas complex-dependent fashion by altering the ability of the Src SH3 domain to bind the p130cas SBD. PMID:22711540

  12. Comparative Actions of Barbiturates Studied by Pollen Grain Germination.

    ERIC Educational Resources Information Center

    Kordan, Herbert A.; Mumford, Pauline M.

    1979-01-01

    Describes a simple experimental system whereby the comparative actions of long, medium, and short-acting barbiturates can be demonstrated in a relatively short period of time under optical microscopy using pollen grains as the biological test or assay system. (Author/HM)

  13. Action Research. Case Studies in TESOL Practice Series.

    ERIC Educational Resources Information Center

    Edge, Julian, Ed.

    Chapter titles in this book include the following: "Attitude and Access: Building a New Teaching/Learning Community in TESOL" (Julian Edge); "Here It Is, Rough Though It May Be: Basic Computer for ESL" (Alison Perkins); "An 'It's Not Action Research Yet, but I'm Getting There' Approach to Teaching Writing" (Neil Cowie); "Early Reflections:…

  14. Action Learning with Second Life--A Pilot Study

    ERIC Educational Resources Information Center

    Wagner, Christian; Ip, Rachael K. F.

    2009-01-01

    Virtual worlds, computer-based simulated environments in which users interact via avatars, provide an opportunity for the highly realistic enactment of real life activities online. Unlike computer games, which have a pre-defined purpose, pay-off structure, and action patterns, virtual worlds can leave many of these elements for users to determine.…

  15. Motivating Struggling Adolescent Readers: An Action Research Study

    ERIC Educational Resources Information Center

    Richardson, Eileen M.

    2016-01-01

    In a high-school reading class, the author used Reader's Theater as an instructional and motivational strategy for underachieving students. This action research focused on the extent to which implementing Reader's Theater motivated students to read and improve their reading skills. Consistent increases in scores for all students occurred over the…

  16. Targeting cellular mRNAs translation by CRISPR-Cas9

    PubMed Central

    Liu, Yuchen; Chen, Zhicong; He, Anbang; Zhan, Yonghao; Li, Jianfa; Liu, Li; Wu, Hanwei; Zhuang, Chengle; Lin, Junhao; Zhang, Qiaoxia; Huang, Weiren

    2016-01-01

    Recently CRISPR-Cas9 system has been reported to be capable of targeting a viral RNA, and this phenomenon thus raises an interesting question of whether Cas9 can also influence translation of cellular mRNAs. Here, we show that both natural and catalytically dead Cas9 can repress mRNA translation of cellular genes, and that only the first 14 nt in the 5′ end of sgRNA is essential for this process. CRISPR-Cas9 can suppress the protein expression of an unintended target gene without affecting its DNA sequence and causes unexpected phenotypic changes. Using the designed RNA aptamer-ligand complexes which physically obstruct translation machinery, we indicate that roadblock mechanism is responsible for this phenomenon. Our work suggests that studies on Cas9 should avoid the potential off-target effects by detecting the alteration of genes at both the DNA and protein levels. PMID:27405721

  17. Pseudorabies virus can escape from CRISPR-Cas9-mediated inhibition.

    PubMed

    Peng, Zhiyuan; Ouyang, Ting; Pang, Daxin; Ma, Teng; Chen, Xinrong; Guo, Ning; Chen, Fuwang; Yuan, Lin; Ouyang, Hongsheng; Ren, Linzhu

    2016-09-01

    The CRISPR-Cas9 system is a newly developed genome-engineering tool used to inhibit virus infection by targeting the conserved regions of the viral genomic DNA. In the present study, we constructed a cell line stably expressing Cas9 endonuclease and sgRNA targeting the conserved UL30 gene of pseudorabies virus (PRV). During the PRV infection, the CRISPR-Cas9 system was efficient in cleaving the UL30 gene in each passage. However, deletions and insertions occurred at low passages, while substitutions were frequently observed at high passages. Furthermore, copy numbers and virus titers of PRV were significantly increased in a passage-dependent manner, indicating that viral genomic replication and assembly were more effective at the high passages than at low passages. These results demonstrated that PRV could escape from CRISPR-Cas9-mediated inhibition. Therefore, whether the CRISPR-Cas9 system is suitable for antiviral application should be considered and carefully verified.

  18. Targeting cellular mRNAs translation by CRISPR-Cas9.

    PubMed

    Liu, Yuchen; Chen, Zhicong; He, Anbang; Zhan, Yonghao; Li, Jianfa; Liu, Li; Wu, Hanwei; Zhuang, Chengle; Lin, Junhao; Zhang, Qiaoxia; Huang, Weiren

    2016-07-13

    Recently CRISPR-Cas9 system has been reported to be capable of targeting a viral RNA, and this phenomenon thus raises an interesting question of whether Cas9 can also influence translation of cellular mRNAs. Here, we show that both natural and catalytically dead Cas9 can repress mRNA translation of cellular genes, and that only the first 14 nt in the 5' end of sgRNA is essential for this process. CRISPR-Cas9 can suppress the protein expression of an unintended target gene without affecting its DNA sequence and causes unexpected phenotypic changes. Using the designed RNA aptamer-ligand complexes which physically obstruct translation machinery, we indicate that roadblock mechanism is responsible for this phenomenon. Our work suggests that studies on Cas9 should avoid the potential off-target effects by detecting the alteration of genes at both the DNA and protein levels.

  19. Temporal prediction restores the evaluative processing of delayed action feedback: an electrophysiological study.

    PubMed

    Kimura, Kenta; Kimura, Motohiro

    2016-09-28

    The evaluative processing of the valence of action feedback is reflected by an event-related brain potential component called feedback-related negativity (FRN) or reward positivity (RewP). Recent studies have shown that FRN/RewP is markedly reduced when the action-feedback interval is long (e.g. 6000 ms), indicating that an increase in the action-feedback interval can undermine the evaluative processing of the valence of action feedback. The aim of the present study was to investigate whether or not such undermined evaluative processing of delayed action feedback could be restored by improving the accuracy of the prediction in terms of the timing of action feedback. With a typical gambling task in which the participant chose one of two cards and received an action feedback indicating monetary gain or loss, the present study showed that FRN/RewP was significantly elicited even when the action-feedback interval was 6000 ms, when an auditory stimulus sequence was additionally presented during the action-feedback interval as a temporal cue. This result suggests that the undermined evaluative processing of delayed action feedback can be restored by increasing the accuracy of the prediction on the timing of the action feedback.

  20. Temporal prediction restores the evaluative processing of delayed action feedback: an electrophysiological study.

    PubMed

    Kimura, Kenta; Kimura, Motohiro

    2016-09-28

    The evaluative processing of the valence of action feedback is reflected by an event-related brain potential component called feedback-related negativity (FRN) or reward positivity (RewP). Recent studies have shown that FRN/RewP is markedly reduced when the action-feedback interval is long (e.g. 6000 ms), indicating that an increase in the action-feedback interval can undermine the evaluative processing of the valence of action feedback. The aim of the present study was to investigate whether or not such undermined evaluative processing of delayed action feedback could be restored by improving the accuracy of the prediction in terms of the timing of action feedback. With a typical gambling task in which the participant chose one of two cards and received an action feedback indicating monetary gain or loss, the present study showed that FRN/RewP was significantly elicited even when the action-feedback interval was 6000 ms, when an auditory stimulus sequence was additionally presented during the action-feedback interval as a temporal cue. This result suggests that the undermined evaluative processing of delayed action feedback can be restored by increasing the accuracy of the prediction on the timing of the action feedback. PMID:27513200

  1. Prospective Mathematics Teachers' Interactions with CAS-Based Textbook Elements

    ERIC Educational Resources Information Center

    Davis, Jon D.

    2015-01-01

    This study investigated how a group of 10 prospective secondary mathematics teachers (PST) read, evaluated, and adapted a textbook lesson involving the symbolic manipulation capabilities of computer algebra systems (CASS). PST read the entire lesson and tended to focus on the organizing question at the beginning of the student lesson and the CAS-S…

  2. Learning from Action Evaluation of the Use of Multimedia Case Studies in Management Information Systems Courses

    ERIC Educational Resources Information Center

    Kawulich, Barbara B.

    2011-01-01

    This manuscript shares lessons learned from conducting an action evaluation of the use of multimedia case studies in Management Information Systems (MIS) courses. Three undergraduate MIS classes took part in the study. The purpose for using case studies in these classes was to teach students about the role of MIS in business. An action evaluation…

  3. CRISPR/Cas9-mediated targeted mutagenesis in the liverwort Marchantia polymorpha L.

    PubMed

    Sugano, Shigeo S; Shirakawa, Makoto; Takagi, Junpei; Matsuda, Yoriko; Shimada, Tomoo; Hara-Nishimura, Ikuko; Kohchi, Takayuki

    2014-03-01

    Targeted genome modification technologies are key tools for functional genomics. The clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease Cas9 system (CRISPR/Cas9) is an emerging technology for targeted genome modification. The CRISPR/Cas9 system consists of a short guide RNA (gRNA), which specifies the target genome sequence, and the Cas9 protein, which has endonuclease activity. The CRISPR/Cas9 system has been applied to model animals and flowering plants, including rice, sorghum, wheat, tobacco and Arabidopsis. Here, we report the application of CRISPR/Cas9 to targeted mutagenesis in the liverwort Marchantia polymorpha L., which has emerged as a model species for studying land plant evolution. The U6 promoter of M. polymorpha was identified and cloned to express the gRNA. The target sequence of the gRNA was designed to disrupt the gene encoding auxin response factor 1 (ARF1) in M. polymorpha. Using Agrobacterium-mediated transformation, we isolated stable mutants in the gametophyte generation of M. polymorpha. CRISPR/Cas9-based site-directed mutagenesis in vivo was achieved using either the Cauliflower mosaic virus 35S or M. polymorpha EF1α promoter to express Cas9. Isolated mutant individuals showing an auxin-resistant phenotype were not chimeric. Moreover, stable mutants were produced by asexual reproduction of T1 plants. Multiple arf1 alleles were easily established using CRIPSR/Cas9-based targeted mutagenesis. Our results provide a rapid and simple approach for molecular genetics in M. polymorpha, and raise the possibility that CRISPR/Cas9 may be applied to a wide variety of plant species.

  4. Responses of the human motor system to observing actions across species: A transcranial magnetic stimulation study.

    PubMed

    White, Nicole C; Reid, Connor; Welsh, Timothy N

    2014-10-22

    Ample evidence suggests that the role of the mirror neuron system (MNS) in monkeys is to represent the meaning of actions. The MNS becomes active in monkeys during execution, observation, and auditory experience of meaningful, object-oriented actions, suggesting that these cells represent the same action based on a variety of cues. The present study sought to determine whether the human motor system, part of the putative human MNS, similarly represents and reflects the meaning of actions rather than simply the mechanics of the actions. To this end, transcranial magnetic stimulation (TMS) of primary motor cortex was used to generate motor-evoked potentials (MEPs) from muscles involved in grasping while participants viewed object-oriented grasping actions performed by either a human, an elephant, a rat, or a body-less robotic arm. The analysis of MEP amplitudes suggested that activity in primary motor cortex during action observation was greatest during observation of the grasping actions of the rat and elephant, and smallest for the human and robotic arm. Based on these data, we conclude that the human action observation system can represent actions executed by non-human animals and shows sensitivity to species-specific differences in action mechanics. PMID:25463135

  5. Structure and activity of the Cas3 HD nuclease MJ0384, an effector enzyme of the CRISPR interference

    SciTech Connect

    Beloglazova, Natalia; Petit, Pierre; Flick, Robert; Brown, Greg; Savchenko, Alexei; Yakunin, Alexander F.

    2012-03-15

    Clustered regularly interspaced short palindromic repeats (CRISPRs) and Cas proteins represent an adaptive microbial immunity system against viruses and plasmids. Cas3 proteins have been proposed to play a key role in the CRISPR mechanism through the direct cleavage of invasive DNA. Here, we show that the Cas3 HD domain protein MJ0384 from Methanocaldococcus jannaschii cleaves endonucleolytically and exonucleolytically (3'-5') single-stranded DNAs and RNAs, as well as 3'-flaps, splayed arms, and R-loops. The degradation of branched DNA substrates by MJ0384 is stimulated by the Cas3 helicase MJ0383 and ATP. The crystal structure of MJ0384 revealed the active site with two bound metal cations and together with site-directed mutagenesis suggested a catalytic mechanism. Our studies suggest that the Cas3 HD nucleases working together with the Cas3 helicases can completely degrade invasive DNAs through the combination of endo- and exonuclease activities.

  6. Atrazine and reproductive function: mode and mechanism of action studies.

    PubMed

    Cooper, Ralph L; Laws, Susan C; Das, Parikshit C; Narotsky, Michael G; Goldman, Jerome M; Lee Tyrey, E; Stoker, Tammy E

    2007-04-01

    Atrazine, a chlorotriazine herbicide, is used to control annual grasses and broadleaf weeds. In this review, we summarize our laboratory's work evaluating the neuroendocrine toxicity of atrazine (and related chlorotriazines) from an historic perspective. We provide the rationale for our work as we have endeavored to determine: 1) the underlying reproductive changes leading to the development of mammary gland tumors in the atrazine-exposed female rat; 2) the cascade of physiological events that are responsible for these changes (i.e., the mode of action for mammary tumors); 3) the potential cellular mechanisms involving adverse effects of atrazine; and 4) the range of reproductive alterations associated with this pesticide.

  7. Experimental study of cyclic action of plasma on tungsten

    NASA Astrophysics Data System (ADS)

    Voronin, A. V.; Aleksandrov, A. E.; Ber, B. Ya.; Brunkov, P. N.; Bormatov, A. A.; Gusev, V. K.; Demina, E. V.; Novokhatskii, A. N.; Pavlov, S. I.; Prusakova, M. D.; Sotnikova, G. Yu.; Yagovkina, M. A.

    2016-03-01

    We report on experimental results on multiple action of hydrogen, deuterium, and helium plasmas produced by a plasma gun and the Globus-M tokamak on tungsten. The surface temperature in the course of irradiation is measured with a bichromatic pyrometer with a time resolution of ⩾1 μs. The morphology of the surface layer is investigated and X-ray structure analysis of tungsten exposed to multiple radiations by the plasma under various conditions is carried out. A slight decrease in the lattice parameter in the sample subjected to the maximal number of irradiation cycles is detected. It is shown that the morphology of the tungsten surface irradiated by the hydrogen plasma from the gun and by the deuterium plasma from the Globus-M tokamak changes (the structure becomes smoother). The characteristic depth of the layer in which impurities have been accumulated exceeds 0.5 μm. This depth was the largest for the sample exposed to 1000 shots from the gun and 2370 shots from the tokamak. It is shown that the helium jet from the plasma gun makes it possible to simulate the action of helium ions on the International Thermonuclear Experimental Reactor (ITER) diverter, producing a layer of submicrometer particles (bubbles).

  8. Action semantics modulate action prediction.

    PubMed

    Springer, Anne; Prinz, Wolfgang

    2010-11-01

    Previous studies have demonstrated that action prediction involves an internal action simulation that runs time-locked to the real action. The present study replicates and extends these findings by indicating a real-time simulation process (Graf et al., 2007), which can be differentiated from a similarity-based evaluation of internal action representations. Moreover, results showed that action semantics modulate action prediction accuracy. The semantic effect was specified by the processing of action verbs and concrete nouns (Experiment 1) and, more specifically, by the dynamics described by action verbs (Experiment 2) and the speed described by the verbs (e.g., "to catch" vs. "to grasp" vs. "to stretch"; Experiment 3). These results propose a linkage between action simulation and action semantics as two yet unrelated domains, a view that coincides with a recent notion of a close link between motor processes and the understanding of action language.

  9. The Impact of Special Education on Self-Perception: An Autoethnographic-Action Research Study

    ERIC Educational Resources Information Center

    Zabonick, Lisa A.

    2013-01-01

    This qualitative autoethnographic-action research study examined how lack of voice as a special education student in the mid-1970s influenced my self-perception. This study also examined, through the use of action research, what influence storytelling had on teacher perceptions of students with disabilities. Autoethnographic data results were used…

  10. Creative Expression as a Way of Knowing in Diabetes Adult Health Education: An Action Research Study

    ERIC Educational Resources Information Center

    Stuckey, Heather L.

    2009-01-01

    This action research study explores the meaning-making process using forms of creative expression for eight women with insulin-dependent diabetes. The study is theoretically informed by arts-based ways of knowing and aspects of feminist poststructuralism, and explains the process of creativity used in the action research process. The findings…

  11. Transforming Language Ideologies through Action Research: A Case Study of Bilingual Science Learning

    ERIC Educational Resources Information Center

    Yang, Eunah

    2012-01-01

    This qualitative case study explored a third grade bilingual teacher's transformative language ideologies through participating in a collaborative action research project. By merging language ideologies theory, Cultural Historical Activity Theory (CHAT), and action research, I was able to identify the analytic focus of this study. I analyzed…

  12. CAS as Environments for Implementing Mathematical Microworlds.

    ERIC Educational Resources Information Center

    Alpers, Burkhard

    2002-01-01

    Investigates whether computer algebra systems (CAS) are suitable environments for implementing mathematical microworlds. Recalls what constitutes a microworld and explores how CAS can be used for implementation, stating potentials as well as limitations. Provides as an example the microworld "Formula 1", implemented in Maple Software. (Author/KHR)

  13. Streamlined Approach for Environmental Restoration Plan for Corrective Action Unit 107: Low Impact Soil Sites, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2008-09-30

    This Streamlined Approach for Environmental Restoration Plan covers activities associated with Corrective Action Unit (CAU) 107 of the Federal Facility Agreement and Consent Order (FFACO, 1996 [as amended February 2008]). CAU 107 consists of the following Corrective Action Sites (CASs) located in Areas 1, 2, 3, 4, 5, 9, 10, and 18 of the Nevada Test Site. (1) CAS 01-23-02, Atmospheric Test Site - High Alt; (2) CAS 02-23-02, Contaminated Areas (2); (3) CAS 02-23-03, Contaminated Berm; (4) CAS 02-23-10, Gourd-Amber Contamination Area; (5) CAS 02-23-11, Sappho Contamination Area; (6) CAS 02-23-12, Scuttle Contamination Area; (7) CAS 03-23-24, Seaweed B Contamination Area; (8) CAS 03-23-27, Adze Contamination Area; (9) CAS 03-23-28, Manzanas Contamination Area; (10) CAS 03-23-29, Truchas-Chamisal Contamination Area; (11) CAS 04-23-02, Atmospheric Test Site T4-a; (12) CAS 05-23-06, Atmospheric Test Site; (13) CAS 09-23-06, Mound of Contaminated Soil; (14) CAS 10-23-04, Atmospheric Test Site M-10; and (15) CAS 18-23-02, U-18d Crater (Sulky). Based on historical documentation, personnel interviews, site process knowledge, site visits, photographs, engineering drawings, field screening, analytical results, and the results of data quality objectives process (Section 3.0), closure in place with administrative controls or no further action will be implemented for CAU 107. CAU 107 closure activities will consist of verifying that the current postings required under Title 10 Code of Federal Regulations (CFR) Part 835 are in place and implementing use restrictions (URs) at two sites, CAS 03-23-29 and CAS 18-23-02. The current radiological postings combined with the URs are adequate administrative controls to limit site access and worker dose.

  14. Applying CRISPR-Cas9 tools to identify and characterize transcriptional enhancers.

    PubMed

    Lopes, Rui; Korkmaz, Gozde; Agami, Reuven

    2016-09-01

    The development of the CRISPR-Cas9 system triggered a revolution in the field of genome engineering. Initially, the use of this system was focused on the study of protein-coding genes but, recently, a number of CRISPR-Cas9-based tools have been developed to study non-coding transcriptional regulatory elements. These technological advances offer unprecedented opportunities for elucidating the functions of enhancers in their endogenous context. Here, we discuss the application, current limitations and future development of CRISPR-Cas9 systems to identify and characterize enhancer elements in a high-throughput manner.

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

    PubMed Central

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

    2014-01-01

    In bacteria, the clustered regularly interspaced short palindromic repeats (CRISPR)–associated (Cas) DNA-targeting complex Cascade (CRISPR-associated complex for antiviral defense) uses CRISPR RNA (crRNA) guides to bind complementary DNA targets at sites adjacent to a trinucleotide signature sequence called the protospacer adjacent motif (PAM). The Cascade complex then recruits Cas3, a nuclease-helicase that catalyzes unwinding and cleavage of foreign double-stranded DNA (dsDNA) bearing a sequence matching that of the crRNA. Cascade comprises the CasA–E proteins and one crRNA, forming a structure that binds and unwinds dsDNA to form an R loop in which the target strand of the DNA base pairs with the 32-nt RNA guide sequence. Single-particle electron microscopy reconstructions of dsDNA-bound Cascade with and without Cas3 reveal that Cascade positions the PAM-proximal end of the DNA duplex at the CasA subunit and near the site of Cas3 association. The finding that the DNA target and Cas3 colocalize with CasA implicates this subunit in a key target-validation step during DNA interference. We show biochemically that base pairing of the PAM region is unnecessary for target binding but critical for Cas3-mediated degradation. In addition, the L1 loop of CasA, previously implicated in PAM recognition, is essential for Cas3 activation following target binding by Cascade. Together, these data show that the CasA subunit of Cascade functions as an essential partner of Cas3 by recognizing DNA target sites and positioning Cas3 adjacent to the PAM to ensure cleavage. PMID:24748111

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

    PubMed

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

    2014-05-01

    In bacteria, the clustered regularly interspaced short palindromic repeats (CRISPR)-associated (Cas) DNA-targeting complex Cascade (CRISPR-associated complex for antiviral defense) uses CRISPR RNA (crRNA) guides to bind complementary DNA targets at sites adjacent to a trinucleotide signature sequence called the protospacer adjacent motif (PAM). The Cascade complex then recruits Cas3, a nuclease-helicase that catalyzes unwinding and cleavage of foreign double-stranded DNA (dsDNA) bearing a sequence matching that of the crRNA. Cascade comprises the CasA-E proteins and one crRNA, forming a structure that binds and unwinds dsDNA to form an R loop in which the target strand of the DNA base pairs with the 32-nt RNA guide sequence. Single-particle electron microscopy reconstructions of dsDNA-bound Cascade with and without Cas3 reveal that Cascade positions the PAM-proximal end of the DNA duplex at the CasA subunit and near the site of Cas3 association. The finding that the DNA target and Cas3 colocalize with CasA implicates this subunit in a key target-validation step during DNA interference. We show biochemically that base pairing of the PAM region is unnecessary for target binding but critical for Cas3-mediated degradation. In addition, the L1 loop of CasA, previously implicated in PAM recognition, is essential for Cas3 activation following target binding by Cascade. Together, these data show that the CasA subunit of Cascade functions as an essential partner of Cas3 by recognizing DNA target sites and positioning Cas3 adjacent to the PAM to ensure cleavage.

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

    PubMed

    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.

  18. An Implementation of the Action Space Concept in Behavioral Analysis.

    ERIC Educational Resources Information Center

    Higgs, Gary K.

    The Contact Action Space (CAS) of an individual, or group of individuals, has a significant impact on the location of activities and the organization of the use of space. Beginning with the most basic components of a CAS, the individual behavior pattern element is developed, and operational variations affecting alignment and configuration are…

  19. Spermatogenic Cell-Specific Gene Mutation in Mice via CRISPR-Cas9.

    PubMed

    Bai, Meizhu; Liang, Dan; Wang, Yinghua; Li, Qing; Wu, Yuxuan; Li, Jinsong

    2016-05-20

    Tissue-specific knockout technology enables the analysis of the gene function in specific tissues in adult mammals. However, conventional strategy for producing tissue-specific knockout mice is a time- and labor-consuming process, restricting rapid study of the gene function in vivo. CRISPR-Cas9 system from bacteria is a simple and efficient gene-editing technique, which has enabled rapid generation of gene knockout lines in mouse by direct injection of CRISPR-Cas9 into zygotes. Here, we demonstrate CRISPR-Cas9-mediated spermatogenic cell-specific disruption of Scp3 gene in testes in one step. We first generated transgenic mice by pronuclear injection of a plasmid containing Hspa2 promoter driving Cas9 expression and showed Cas9 specific expression in spermatogenic cells. We then produced transgenic mice carrying Hspa2 promoter driven Cas9 and constitutive expressed sgRNA targeting Scp3 gene. Male founders were infertile due to developmental arrest of spermatogenic cells while female founders could produce progeny normally. Consistently, male progeny from female founders were infertile and females could transmit the transgenes to the next generation. Our study establishes a CRISPR-Cas9-based one-step strategy to analyze the gene function in adult tissues by a temporal-spatial pattern.

  20. CAS

    SciTech Connect

    Martinez, B.; Pomeroy, G. )

    1989-12-02

    The Security Alarm System is a data acquisition and control system which collects data from intrusion sensors and displays the information in a real-time environment for operators. The Access Control System monitors and controls the movement of personnel with the use of card readers and biometrics hand readers.

  1. Choosing Actions

    PubMed Central

    Rosenbaum, David A.; Chapman, Kate M.; Coelho, Chase J.; Gong, Lanyun; Studenka, Breanna E.

    2013-01-01

    Actions that are chosen have properties that distinguish them from actions that are not. Of the nearly infinite possible actions that can achieve any given task, many of the unchosen actions are irrelevant, incorrect, or inappropriate. Others are relevant, correct, or appropriate but are disfavored for other reasons. Our research focuses on the question of what distinguishes actions that are chosen from actions that are possible but are not. We review studies that use simple preference methods to identify factors that contribute to action choices, especially for object-manipulation tasks. We can determine which factors are especially important through simple behavioral experiments. PMID:23761769

  2. Knowledge-based discovery for designing CRISPR-CAS systems against invading mobilomes in thermophiles.

    PubMed

    Chellapandi, P; Ranjani, J

    2015-09-01

    Clustered regularly interspaced short palindromic repeats (CRISPRs) are direct features of the prokaryotic genomes involved in resistance to their bacterial viruses and phages. Herein, we have identified CRISPR loci together with CRISPR-associated sequences (CAS) genes to reveal their immunity against genome invaders in the thermophilic archaea and bacteria. Genomic survey of this study implied that genomic distribution of CRISPR-CAS systems was varied from strain to strain, which was determined by the degree of invading mobiloms. Direct repeats found to be equal in some extent in many thermopiles, but their spacers were differed in each strain. Phylogenetic analyses of CAS superfamily revealed that genes cmr, csh, csx11, HD domain, devR were belonged to the subtypes of cas gene family. The members in cas gene family of thermophiles were functionally diverged within closely related genomes and may contribute to develop several defense strategies. Nevertheless, genome dynamics, geological variation and host defense mechanism were contributed to share their molecular functions across the thermophiles. A thermophilic archaean, Thermococcus gammotolerans and thermophilic bacteria, Petrotoga mobilis and Thermotoga lettingae have shown superoperons-like appearance to cluster cas genes, which were typically evolved for their defense pathways. A cmr operon was identified with a specific promoter in a thermophilic archaean, Caldivirga maquilingensis. Overall, we concluded that knowledge-based genomic survey and phylogeny-based functional assignment have suggested for designing a reliable genetic regulatory circuit naturally from CRISPR-CAS systems, acquired defense pathways, to thermophiles in future synthetic biology.

  3. Occurrence and activity of a type II CRISPR-Cas system in Lactobacillus gasseri.

    PubMed

    Sanozky-Dawes, Rosemary; Selle, Kurt; O'Flaherty, Sarah; Klaenhammer, Todd; Barrangou, Rodolphe

    2015-09-01

    Bacteria encode clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated genes (cas), which collectively form an RNA-guided adaptive immune system against invasive genetic elements. In silico surveys have revealed that lactic acid bacteria harbour a prolific and diverse set of CRISPR-Cas systems. Thus, the natural evolutionary role of CRISPR-Cas systems may be investigated in these ecologically, industrially, scientifically and medically important microbes. In this study, 17 Lactobacillus gasseri strains were investigated and 6 harboured a type II-A CRISPR-Cas system, with considerable diversity in array size and spacer content. Several of the spacers showed similarity to phage and plasmid sequences, which are typical targets of CRISPR-Cas immune systems. Aligning the protospacers facilitated inference of the protospacer adjacent motif sequence, determined to be 5'-NTAA-3' flanking the 3' end of the protospacer. The system in L. gasseri JV-V03 and NCK 1342 interfered with transforming plasmids containing sequences matching the most recently acquired CRISPR spacers in each strain. We report the distribution and function of a native type II-A CRISPR-Cas system in the commensal species L. gasseri. Collectively, these results open avenues for applications for bacteriophage protection and genome modification in L. gasseri, and contribute to the fundamental understanding of CRISPR-Cas systems in bacteria.

  4. Genome editing with Cas9 in adult mice corrects a disease mutation and phenotype

    PubMed Central

    Yin, Hao; Xue, Wen; Chen, Sidi; Bogorad, Roman L; Benedetti, Eric; Grompe, Markus; Koteliansky, Victor; Sharp, Phillip A; Jacks, Tyler; Anderson, Daniel G

    2014-01-01

    We demonstrate CRISPR-Cas9–mediated correction of a Fah mutation in hepatocytes in a mouse model of the human disease hereditary tyrosinemia. Delivery of components of the CRISPR-Cas9 system by hydrodynamic injection resulted in initial expression of the wild-type Fah protein in ~1/250 liver cells. Expansion of Fah-positive hepatocytes rescued the body weight loss phenotype. Our study indicates that CRISPR-Cas9–mediated genome editing is possible in adult animals and has potential for correction of human genetic diseases. PMID:24681508

  5. Genome editing with Cas9 in adult mice corrects a disease mutation and phenotype.

    PubMed

    Yin, Hao; Xue, Wen; Chen, Sidi; Bogorad, Roman L; Benedetti, Eric; Grompe, Markus; Koteliansky, Victor; Sharp, Phillip A; Jacks, Tyler; Anderson, Daniel G

    2014-06-01

    We demonstrate CRISPR-Cas9-mediated correction of a Fah mutation in hepatocytes in a mouse model of the human disease hereditary tyrosinemia. Delivery of components of the CRISPR-Cas9 system by hydrodynamic injection resulted in initial expression of the wild-type Fah protein in ∼1/250 liver cells. Expansion of Fah-positive hepatocytes rescued the body weight loss phenotype. Our study indicates that CRISPR-Cas9-mediated genome editing is possible in adult animals and has potential for correction of human genetic diseases.

  6. The Early Development of Object Knowledge: A Study of Infants' Visual Anticipations during Action Observation

    ERIC Educational Resources Information Center

    Hunnius, Sabine; Bekkering, Harold

    2010-01-01

    This study examined the developing object knowledge of infants through their visual anticipation of action targets during action observation. Infants (6, 8, 12, 14, and 16 months) and adults watched short movies of a person using 3 different everyday objects. Participants were presented with objects being brought either to a correct or to an…

  7. Immune response of apoptosis-related cysteine peptidases from the red abalone Haliotis rufescens (HrCas8 and HrCas3): molecular characterization and transcription expression.

    PubMed

    Chávez-Mardones, Jacqueline; Gallardo-Escárate, Cristian

    2014-07-01

    Caspases play an important role in the different stages of programmed cell death, or apoptosis, which has been related to the immune response in multicellular organisms. The present study characterized an initiator caspase (HrCas8) and an effector caspase (HrCas3) from the red abalone Haliotis rufescens using the RACE method and qPCR analysis. HrCas8 showed a complete sequence of 2529 base pairs (bp) with an open-reading frame (ORF) of 1911 bp, a 5'UTR of 201 bp, and a 3'UTR of 417 bp. The estimated molecular mass for the 636 amino acids from HrCas8 was 71.5 kDa with an isoelectric point of 6.2. The HrCas8 sequence had two death-effector domains (DEDs) and the subunits p20 and p10, in addition to an active site characteristic of cysteine proteins. Meanwhile, the effector caspase HrCas3 showed a complete sequence of 1404 bp, a 5'UTR of 82 bp, and a 3'UTR of 574 bp. The ORF of this caspase had 747 bp that coded for 248 residues. Moreover, the predicted molecular mass of HrCas3 was 29.4 kDa; the theoretical isoelectric point was 5.70, and the sequence evidenced a conserved caspase recruitment domain (CARD). The distribution of the caspases in distinct tissues revealed that HrCas8 was principally expressed in the hemolymph, while HrCas3 had a higher expression in the gills. A basal level of expression was found for both caspases in muscle tissue. The immune response of caspases in H. rufescens was evaluated through an injection of Vibrio anguillarum. The results showed an increase in the transcription of HrCas8 post-challenge, as well as an activation of HrCas3, which together suggest the initiation of apoptosis as a response to bacterial infection in H. rufescens. PMID:24821426

  8. Campylobacter jejuni acquire new host-derived CRISPR spacers when in association with bacteriophages harboring a CRISPR-like Cas4 protein

    PubMed Central

    Hooton, Steven P. T.; Connerton, Ian 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 notably absent. Cas4 proteins possess 5′-3′ exonuclease activity to create recombinogenic-ends for spacer acquisition. Here we report a conserved Cas4-like protein in Campylobacter bacteriophages that creates a novel split arrangement between the bacteriophage and host that represents a new twist in the bacteriophage/host co-evolutionary arms race. The continuous association of bacteriophage and host in the carrier state life cycle of C. jejuni provided an opportunity to study spacer acquisition in this species. Remarkably all the spacer sequences observed were of host origin. We hypothesize that Campylobacter bacteriophages can use Cas4-like protein to activate spacer acquisition to use host DNA as an effective decoy to bacteriophage DNA. Bacteria that acquire self-spacers and escape phage infection must overcome CRISPR-mediated autoimmunity either by loss of the interference functions leaving them susceptible to foreign DNA incursion or tolerate changes in gene regulation. PMID:25601859

  9. NTP Toxicology and Carcinogenesis Studies of Diethylphthalate (CAS No. 84-66-2) in F344/N Rats and B6C3F1 Mice (Dermal Studies) with Dermal Initiation/ Promotion Study of Diethylphthalate and Dimethylphthalate (CAS No. 131-11-3) in Male Swiss (CD-1(R)) Mice.

    PubMed

    1995-05-01

    Diethylphthalate and dimethylphthalate are used as phthalate plasticizers, in an extensive array of products. The chronic dermal toxicity of diethylphthalate was evaluated in male and female F344/N rats and B6C3F1 mice in 2-year studies. In a series of special studies, the tumor initiation or promotion potential of diethylphthalate or dimethylphthalate was evaluated in male Swiss (CD-1(R)) mice by an initiation/promotion model of skin carcinogenesis. The genetic toxicity of diethylphthalate and dimethylphthalate in Salmonella typhimurium and cultured Chinese hamster ovary cells was also evaluated. 4-WEEK STUDY IN F344/N RATS: Groups of 10 male and 10 female rats were dermally administered diethylphthalate at volumes of 0, 37.5, 75, 150, or 300 &mgr;L (0, 46, 92, 184, or 369 &mgr;g) applied neat, 5 days per week for 4 weeks. All male and female rats survived to the end of the study. No evidence of dermatotoxicity was observed, with no adverse clinical signs observed and no effects on weight gain or feed consumption. Relative liver weights of 300 &mgr;L males and females and 150 &mgr;L females were greater than those of controls. Relative kidney weights of 150 and 300 &mgr;L males and 150 &mgr;L females were greater than those of controls. No other adverse effects were observed in this study. 4-WEEK STUDY IN B6C3F1 MICE: Groups of 10 male and 10 female mice were dermally administered diethylphthalate at volumes of 0, 12.5, 25, 50, or 100 &mgr;L (0, 15, 31, 62, or 123 &mgr;) applied neat, five days per week for 4 weeks. One control female died before the end of the study; all other mice survived. No evidence of dermatotoxicity or other adverse clinical signs were observed, and no clear adverse effects on weight gain or feed consumption were seen. Absolute and relative liver weights of 25 and 100 &mgr;L females were greater than those of the controls. Based on these 4-week study results, doses of 0, 35, and 100 &mgr;L diethylphthalate were recommended for the 2-year

  10. Assisting Students' Cognitive Strategies with the Use of CAS

    ERIC Educational Resources Information Center

    Sarvari, Csaba; Lavicza, Zsolt; Klincsik, Mihaly

    2010-01-01

    This paper examines various cognitive strategies applied while CAS (Computer Algebra System) are used in undergraduate-level engineering mathematics teaching and learning. We posed some questions in relation to such CAS use: What kind of tools can CAS offer to enhance different cognitive strategies of students? How can the use of CAS widen the…

  11. Data Communications Requirements for CAS Applications

    NASA Technical Reports Server (NTRS)

    McCabe, James D.; Blaylock, Bruce T. (Technical Monitor)

    1994-01-01

    This paper discusses the long-haul data communications requirements needed to support visualization of Computational Aeroscience (CAS) simulations. A scientific computing model is presented, from which current communications requirements are derived. This model is then expanded to describe the CAS Collaborative Visualization Environment, in which scientists from multiple NASA centers will collaborate on multidisciplinary, time-accurate simulations. An analysis of this model, along with data from the TFCC Workload Model, shows that CAS data communications requirements will increase to 10-100 megabytes/second by 1996.

  12. Nucleosomes Inhibit Cas9 Endonuclease Activity in Vitro.

    PubMed

    Hinz, John M; Laughery, Marian F; Wyrick, John J

    2015-12-01

    During Cas9 genome editing in eukaryotic cells, the bacterial Cas9 enzyme cleaves DNA targets within chromatin. To understand how chromatin affects Cas9 targeting, we characterized Cas9 activity on nucleosome substrates in vitro. We find that Cas9 endonuclease activity is strongly inhibited when its target site is located within the nucleosome core. In contrast, the nucleosome structure does not affect Cas9 activity at a target site within the adjacent linker DNA. Analysis of target sites that partially overlap with the nucleosome edge indicates that the accessibility of the protospacer-adjacent motif (PAM) is the critical determinant of Cas9 activity on a nucleosome.

  13. NTP Toxicology and Carcinogenesis Studies of Trichloroethylene (CAS No. 79-01-6) in Four Strains of Rats (ACI, August, Marshall, Osborne-Mendel) (Gavage Studies).

    PubMed

    1988-04-01

    Trichloroethylene is an industrial solvent used primarily for vapor degreasing and cold cleaning. It was selected for study because of its industrial use and for potential for human exposure. (An estimated 3.5 million workers are exposed to trichloroethylene.) In an earlier study trichloroethylene (stabilized with epichlorohydrin and 1,2-epoxybutane) administered by gavage caused hepatocellular carcinomas in male and female B6C3F1 mice. Trichloroethylene administration did not increase the incidence of tumors in male or female Osborne-Mendel rats. However, the survival of dosed rats was reduced, thereby compromising the sensitivity of the study to detect a carcinogenic effect. The studies described in this report were conducted to compare the sensitivities of four strains of rats (ACI, August, Marshall, and Osborne-Mendel) to diisopropylamine-stabilized trichloroethylene. The results of the present studies demonstrate that long-term administration of trichloroethylene produces nephrotoxicity in four strains of rats and that the susceptibilities of these strains to the nephrotoxic effects of the chemical are similar. Because of chemically induced toxicity, reduced survival, and incomplete documentation of the experimental data, the studies are considered inadequate for either comparing or assessing trichloroethylene-induced carcinogenesis in these strains of rats. Toxicology and carcinogenesis studies of trichloroethylene (more than 99% pure, stabilized with 8 ppm diisopropylamine) were conducted by administering the chemical in corn oil gavage at doses of 0, 500, or 1,000 mg/kg per day, 5 day per week, for 103 weeks to groups of 50 male and 50 female ACI, August, Marshall, and Osborne-Mendel rats. The doses were selected on the basis of results from 13-week gavage studies in which groups of 10 male and 10 female ACI, August, and Marshall rats received daily doses or trichloroethylene (male: 125-2,000 mg/kg; female: 63-1,000 mg/kg). Doses for Osborne-Mendel rats

  14. NTP Toxicology and Carcinogenesis Studies of 3,3'-Dimethoxybenzidine Dihydrochloride (CAS No. 20325-40-0) in F344/N Rats (Drinking Water Studies).

    PubMed

    1990-01-01

    3,3'-Dimethoxybenzidine dihydrochloride is an off-white powder with a melting point of 274 degrees C. 3,3'-Dimethoxybenzidine is used principally as an intermediate in the production of commercial bisazobiphenyl dyes for coloring textiles, paper, plastic, rubber, and leather. In the synthesis of the bisazobiphenyl dyes, the amine groups of 3,3'-dimethoxybenzidine are chemically linked with other aromatic amines. A small quantity of 3,3'-dimethoxybenzidine is also used as an intermediate in the production of o-dianisidine diisocyanate, which is used in isocyanate-based adhesive systems and as a component of polyurethane elastomers. 3,3'-Dimethoxybenzidine dihydrochloride was evaluated in toxicity and carcinogenicity studies as part of the National Toxicology Program's Benzidine Dye Initiative. This Initiative was designed to evaluate the representative benzidine congeners and benzidine congener-derived and benzidine-derived dyes. 3,3'-Dimethoxybenzidine dihydrochloride was nominated for study because of the potential for human exposure during production of bisazobiphenyl dyes and because benzidine, a structurally related chemical, is a known human carcinogen. NTP Toxicology and Carcinogenesis studies were conducted by administering 3,3'-dimethoxybenzidine dihydrochloride (greater than 97.5% pure) in drinking water to groups of F344/N rats of each sex for 14 days, 13 weeks, 9 months, or 21-months. The 21-month studies were intended to last 24 months but were terminated early because of rapidly declining survival due to neoplasia. Studies were performed only in rats because similar studies are being performed in mice at the National Center for Toxicology Research. Genetic toxicology studies were conducted with Salmonella typhimurium, Chinese hamster over (CHO) cells, and Drosophila melanogaster. Fourteen-Day Studies: All rats receiving drinking water concentrations up to 4,500 ppm lived to the end of the studies. Rats that received water containing 4,500 ppm 3

  15. NTP Toxicology and Carcinogenesis Studies of Salicylazosulfapyridine (CAS No. 599-79-1) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

    PubMed

    1997-05-01

    Salicylazosulfapyridine is widely used for the treatment of ulcerative colitis and Crohn's disease. It has been beneficial in the treatment of psoriasis and rheumatoid arthritis, and it has been used in veterinary medicine for the treatment of granulomatous colitis. Salicylazosulfapyridine was nominated for toxicity and carcinogenicity testing by the National Cancer Institute on the basis of its widespread use in humans and because it is a representative chemical from a class of aryl sulfonamides. Salicylazosulfapyridine is a suspect carcinogen because reductive cleavage of the azo linkage yields a p-amino aryl sulfonamide (sulfapyridine), and a related p-amino aryl sulfonamide (sulfamethoxazole) has been shown to produce thyroid neoplasms in rats. Toxicology and carcinogenicity studies were conducted in F344/N rats and B6C3F1 mice. Rats and mice were administered salicylazosulfapyridine (96% to 98% pure) in corn oil by gavage for 16 days, 13 weeks, or 2 years. The gavage route of administration was selected for these studies because it approximates the typical route of human exposure to the chemical. Genetic toxicology studies were conducted in vitro in Salmonella typhimurium and cultured Chinese hamster ovary cells and in vivo in rat and mouse bone marrow and mouse peripheral blood cells. 16-DAY STUDY IN RATS: Groups of five male and five female rats were administered 0, 675, 1,350, or 2,700 mg salicylazosulfapyridine/kg body weight in corn oil by gavage for 16 days excluding weekends. All rats survived to the end of the study. With the exception of the 675 mg/kg male group, the final mean body weights of all dosed groups of males and females were significantly lower than those of controls. Mean body weight gains of all dosed groups were less than those of controls. Clinical findings included ruffled fur and distended abdomens in male and female rats receiving 2,700 mg/kg. Hypothyroidism, evidenced by decreased serum triiodothyronine and thyroxine concentrations

  16. NTP Toxicology and Carcinogenesis Studies of Benzaldehyde (CAS No. 100-52-7) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

    PubMed

    1990-03-01

    Benzaldehyde is an aromatic aldehyde used in the food, beverage, pharmaceutical, perfume, soap, and dyestuff industries. NTP Toxicology and Carcinogenesis studies were conducted by administering benzaldehyde (99% pure) in corn oil by gavage to groups of F344/N rats and B6C3F1 mice of each sex for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, mouse lymphoma cells, Chinese hamster ovary (CHO) cells, and Drosophila melanogaster. Sixteen-Day Studies: All rats that received 1,600 mg/kg died by day 2, and 2/5 males and 2/5 females that received 800 mg/kg died before the end of the studies. Final mean body weights of dosed and vehicle control rats were similar, with the exception of the 800 mg/kg groups, in which males were 14% lighter and females were 11% lighter than vehicle controls. All mice that received 1,600 or 3,200 mg/kg died by day 3. Final mean body weights of dosed and vehicle control mice were similar. No gross lesions attributable to benzaldehyde were detected upon necropsy. Thirteen-Week Studies: Six of 10 male rats and 3/10 female rats that received 800 mg/kg and 1/10 female rats that received 400 mg/kg died near the end of the studies. Final mean body weights of dosed and vehicle control rats were similar, with the exception of male rats receiving 800 mg/kg, which were 26% lighter than vehicle controls. Compound-related lesions seen in rats receiving 800 mg/kg, but not in those receiving 400 mg/kg, included degeneration and necrosis in the cerebellum, necrosis in the hippocampus, hyperplasia and/or hyperkeratosis in the forestomach, and degeneration or necrosis of the liver and of the tubular epithelium in the kidney. Nine of 10 male mice and 1/10 female mice that received 1,200 mg/kg benzaldehyde died by the end of the first week. Compound-related renal tubule degeneration and/or necrosis and reduction in final body weight were observed in the 600 mg/kg group of male mice. No reductions in body weight

  17. Lifetime carcinogenesis studies of chrysotile asbestos (CAS No. 12001-29-5) in syrian golden hamsters (feed studies). Technical report series

    SciTech Connect

    Not Available

    1990-07-01

    Carcinogenesis studies of short range (SR), intermediate range (IR) or intermediate range chrysotile asbestos in combination with the intestinal carcinogen 1,2-dimethylhydrazine dihydrochloride (DMH) were conducted with male and female Syrian golden hamsters. Both forms of chrysotile asbestos were administered at a concentration of 1% in pelleted diet for the entire lifetime of the hamsters, starting with mothers of the test animals. Group sizes varied from 125 to 253. Starting at 6 weeks of age, male and female hamsters in the intermediate range chrysotile/DMH study were given oral doses of DMH (4 mg/kg) every other week for a total of 5 doses. There was no adverse effect on body weight gain or survival by either form of asbestos or by asbestos in combination with DMH. Under the conditions of these studies, neither short range chrysotile nor intermediate range chrysotile asbestos was carcinogenic when ingested at 1% levels in the diet by male and female Syrian golden hamsters. While there were increases in the rates of adrenal cortical adenomas in male and female hamsters exposed to intermediate range chrysotile asbestos compared with pooled control groups, these incidence rates were not different when compared with the concurrent control groups. Additionally, the biologic importance of adrenal tumors in the absence of target organ (gastrointestinal tract) neoplasia is questionable.

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  19. NTP Technical Report on the comparative toxicity studies of allyl acetate (CAS No. 591-87-7), allyl alcohol (CAS No. 107-18-6) and acrolein (CAS No. 107-02-8) administered by gavage to F344/N rats and B6C3F1 mice.

    PubMed

    Irwin, Rick D

    2006-07-01

    Allyl acetate, allyl alcohol, and acrolein are used in the manufacture of detergents, plastics, pharmaceuticals, and chemicals and as agricultural agents and food additives. Male and female F344/N rats and B6C3F(1) mice received allyl acetate, allyl alcohol, or acrolein by gavage for 14 weeks. Genetic toxicology studies were conducted in Salmonella typhimurium, Drosophila melanogaster, cultured Chinese hamster ovary cells, rat bone marrow erythrocytes, and mouse peripheral blood erythrocytes. Groups of 10 male and 10 female rats were administered 0, 6, 12, 25, 50, or 100 mg allyl acetate/kg body weight, 0, 1.5, 3, 6, 12, or 25 mg/kg allyl alcohol, or 0, 0.75, 1.25, 2.5, 5, or 10 mg/kg acrolein in 0.5% methylcellulose by gavage, 5 days per week for 14 weeks. Groups of 10 male and 10 female mice were administered 0, 8, 16, 32, 62.5, or 125 mg/kg allyl acetate, 0, 3, 6, 12, 25, or 50 mg/kg allyl alcohol, or 0, 1.25, 2.5, 5, 10, or 20 mg/kg acrolein in 0.5% methylcellulose by gavage, 5 days per week for 14 weeks. In the allyl acetate rat study, all males and females in the 100 mg/kg groups died or were killed moribund by day 8; there were no other deaths. In the allyl alcohol study, all rats survived to the end of the study except one 6 mg/kg female. In the acrolein rat study, eight males and eight females in the 10 mg/kg groups died by week 9 of the study. Two males in the 2.5 and 5 mg/kg groups and one or two females in the 1.25, 2.5, and 5 mg/kg groups also died early; two of these deaths were gavage accidents. In the allyl acetate mouse study, all males and females in the 125 mg/kg group died during the first week of the study. All other early deaths, except five 62.5 mg/kg males and one 32 mg/kg female, were gavage accidents. In the allyl alcohol mouse study, one 50 mg/kg female died due to a gavage accident; all other animals survived to the end of the study. In the acrolein mouse study, all males and females administered 20 mg/kg died during the first week of

  20. Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system

    PubMed Central

    Bikard, David; Jiang, Wenyan; Samai, Poulami; Hochschild, Ann; Zhang, Feng; Marraffini, Luciano A.

    2013-01-01

    The ability to artificially control transcription is essential both to the study of gene function and to the construction of synthetic gene networks with desired properties. Cas9 is an RNA-guided double-stranded DNA nuclease that participates in the CRISPR-Cas immune defense against prokaryotic viruses. We describe the use of a Cas9 nuclease mutant that retains DNA-binding activity and can be engineered as a programmable transcription repressor by preventing the binding of the RNA polymerase (RNAP) to promoter sequences or as a transcription terminator by blocking the running RNAP. In addition, a fusion between the omega subunit of the RNAP and a Cas9 nuclease mutant directed to bind upstream promoter regions can achieve programmable transcription activation. The simple and efficient modulation of gene expression achieved by this technology is a useful asset for the study of gene networks and for the development of synthetic biology and biotechnological applications. PMID:23761437

  1. NTP Toxicology and Carcinogenesis Studies of Tetrafluoroethylene (CAS No. 116-14-3) in F344 Rats and B6C3F1 Mice (Inhalation Studies).

    PubMed

    1997-04-01

    Tetrafluoroethylene is used in the production of polytetrafluoroethylene (Teflon(R)) and other polymers. Tetrafluoroethylene was nominated by the National Cancer Institute for toxicity and carcinogenicity studies based on the potential for human exposure to the chemical due to the large production volume and on the lack of adequate data for tetrafluoroethylene in the literature. Male and female F344/N rats and B6C3F1 mice were exposed to tetrafluoroethylene (98% to 99% pure) by whole body inhalation exposure for 16 days, 13 weeks, or 2 years. Genetic toxicity studies were conducted in mouse peripheral blood erythrocytes. 16-DAY STUDY IN RATS: Groups of five male and five female F344/N rats were exposed to 0, 312, 625, 1,250, 2,500, or 5,000 ppm tetrafluoroethylene by inhalation for 6 hours per day, 5 days per week for a total of 12 exposures during a 16-day period. All rats survived to the end of the study. The final mean body weights and body weight gains of males and females exposed to 5,000 ppm were significantly less than those of the controls. The mean body weight gain of females exposed to 2,500 ppm was also significantly less than that of the controls. There were no exposure-related clinical findings in male or female rats. There were no significant differences in hematology parameters that were considered to be related to tetrafluoroethylene exposure. Absolute and relative kidney weights of all exposed groups of males were significantly greater than those of the controls, as were those of females in the 2,500 and 5,000 ppm groups. The absolute kidney weight of females exposed to 1,250 ppm was also significantly greater than that of the controls. The relative liver weights of all exposed groups of males and the absolute liver weights of males in the 625 and 2,500 ppm groups were significantly greater than those of the controls. Increased incidences of renal tubule degeneration occurred in males and females exposed to 625 ppm or greater; this lesion was

  2. Closure Report for Corrective Action Unit 540: Spill Sites, Nevada Test Site, Nevada, Rev. No.: 0

    SciTech Connect

    McClure, Lloyd

    2006-10-01

    This Closure Report (CR) presents information supporting the closure of Corrective Action Unit (CAU) 540: Spill Sites, Nevada Test Site, Nevada. This CR complies with the requirements of the 'Federal Facility Agreement and Consent Order' (1996) that was agreed to by the State of Nevada, the U.S. Department of Energy, and the U.S. Department of Defense. Corrective Action Unit 540 is located within Areas 12 and 19 of the Nevada Test Site and is comprised of the following Corrective Action Sites (CASs): CAS 12-44-01, ER 12-1 Well Site Release; CAS 12-99-01, Oil Stained Dirt; CAS 19-25-02, Oil Spill; CAS 19-25-04, Oil Spill; CAS 19-25-05, Oil Spill; CAS 19-25-06, Oil Spill; CAS 19-25-07, Oil Spill; CAS 19-25-08, Oil Spills (3); and CAS 19-44-03, U-19bf Drill Site Release. The purpose of this CR is to provide documentation supporting recommendations of no further action for the CASs within CAU 540. To achieve this, the following actions were performed: (1) Reviewed the current site conditions, including the concentration and extent of contamination; (2) Performed closure activities to address the presence of substances regulated by 'Nevada Administrative Code' 445A.2272 (NAC, 2002); and (3) Documented Notice of Completion and closure of CAU 540 issued by the Nevada Division of Environmental Protection.

  3. Organisational Competence: The Study of a School Council in Action

    ERIC Educational Resources Information Center

    Cotmore, Richard

    2004-01-01

    Despite the growth in the numbers of school councils, there has been little analytical and conceptual research about this form of participation. A case study of a school council in a junior school was adopted in this study. Complementary theories of "competent participation" and "place in time and space" contributed to the central organising…

  4. Mathematics in Action: Two New Zealand Case Studies.

    ERIC Educational Resources Information Center

    Ryan, David M.

    Mathematics is playing an increasingly important role in business and industry. In this paper we present two case studies to illustrate the power and impact of mathematics in two important practical applications in New Zealand. The first case study describes the development of a mathematical optimization model to maximize the value of aluminum…

  5. Educational Cost Analysis in Action: Case Studies for Planners -- III.

    ERIC Educational Resources Information Center

    Coombs, Philip H.; Hallak, Jacques

    This document is the third in a series of three documents, which together contain 27 case studies on the uses of cost analysis in educational planning. The case studies have been presented to help planners and administrators see how cost analysis can be used to improve the efficiency of their educational systems, and to get the best value from…

  6. Educational Cost Analysis in Action: Case Studies for Planners -- II.

    ERIC Educational Resources Information Center

    Coombs, Philip H.; Hallak, Jacques

    This document is the second in a series of three documents, which together contain 27 case studies on the uses of cost analysis in educational planning. The case studies are presented to help planners and administrators see how cost analysis can be used to improve the efficiency of their educational systems, or to get the best value existing…

  7. The Girl Child and the Family: An Action Research Study.

    ERIC Educational Resources Information Center

    Anandalakshmy, S., Comp.

    This report describes a nationwide study of female children and the family in rural India. The objectives of the study were to generate data on the situation of female children; to identify the major problems related to their status; to start a series of programs to help remedy those shortcomings; and to assist communities, and women in…

  8. NTP Toxicology and Carcinogenesis Studies of Nitromethane (CAS No. 75-52-5) in F344/N Rats and B6C3F1 Mice (Inhalation Studies).

    PubMed

    1997-02-01

    Nitromethane is used as a rocket and engine fuel; as a synthesis intermediate for agricultural fumigants, biocides, and other products; as a solvent; and as an explosive in mining, oil-well drilling, and seismic exploration. It has been detected in air, in surface and drinking water, and in cigarette smoke. Nitromethane was studied because of the potential for widespread human exposure and because it is structurally related to the carcinogens 2-nitropropane and tetranitromethane. Male and female F344/N rats and B6C3F1 mice received nitromethane (purity 98% or greater) by inhalation for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, and peripheral blood erythrocytes of mice. 16-DAY STUDY IN RATS: Groups of five male and five female rats were exposed to 0, 94, 188, 375, 750, or 1,500 ppm nitromethane by inhalation, 6 hours per day, 5 days per week, for 16 days. All rats survived until the end of the study. The mean body weight gain of male rats in the 1,500 ppm group was slightly but significantly less than that of the controls; the final mean body weights and mean body weight gains of exposed females were similar to those of the controls. Clinical findings in all male and female rats in the 1,500 ppm groups included increased preening, rapid breathing, hyperactivity early in the study, and hypoactivity and loss of coordination in the hindlimbs near the end of the study. The relative liver weights of all exposed groups of male rats and the absolute and relative liver weights of females exposed to 375 ppm or greater were significantly greater than those of the controls. Minimal to mild degeneration of the olfactory epithelium was observed in the nose of males and females exposed to 375 ppm or greater. Sciatic nerve degeneration was present in all male and female rats exposed to 375 ppm or greater; rats exposed to 750 or 1,500 ppm also had reduced myelin around sciatic axons. 16

  9. Toxicology and Carcinogenesis Studies of Furfuryl Alcohol (CAS No. 98-00-0) in F344/N Rats and B6C3F1 Mice (Inhalation Studies).

    PubMed

    1999-02-01

    Furfuryl alcohol-based resins are used as binding agents in foundry sand and as corrosion inhibitors in mortar, grout, and cement. Because of their heat resistance, furan resins are used in the manufacture of fiberglass-reinforced plastic equipment. Furfuryl alcohol was selected for evaluation because of the absence of data on its carcinogenic potential and its large production volume, widespread use in manufacturing, and ubiquitous presence in consumer goods. Male and female F344/N rats and B6C3F1 mice were exposed to furfuryl alcohol (greater than 98% pure) by inhalation for 16 days, 14 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, and mouse bone marrow cells. 16-DAY STUDY IN RATS: Groups of five male and five female rats were exposed to concentrations of 0, 16, 31, 63, 125, or 250 ppm furfuryl alcohol by inhalation, 6 hours per day, 5 days per week for 16 days. All male and female rats exposed to 250 ppm died by day 2 of the study, and one male rat exposed to 125 ppm died on day 5. Final mean body weights of male and female rats exposed to 125 ppm were significantly less than those of the chamber control groups. Male rats exposed to 31, 63, or 125 ppm and female rats exposed to 125 ppm gained less weight than the chamber control groups. Clinical findings included dyspnea, hypoactivity, and nasal and ocular discharge in males and females exposed to 63, 125, or 250 ppm. All exposed animals developed lesions in the nasal respiratory epithelium and olfactory epithelium, and the severities of these lesions generally increased with increasing exposure concentration. 16-DAY STUDY IN MICE: Groups of five male and five female mice were exposed to concentrations of 0, 16, 31, 63, 125, or 250 ppm furfuryl alcohol by inhalation, 6 hours per day, 5 days per week for 16 days. All male and female mice exposed to 250 ppm died by day 4 of the study, and one female mouse exposed to 125 ppm died on day

  10. NTP Toxicology and Carcinogenesis Studies of Two Pentachlorophenol Technical-Grade Mixtures (CAS No. 87-86-5) in B6C3F1 Mice (Feed Studies).

    PubMed

    1989-03-01

    Toxicology studies of pentachlorophenol, a biocide used primarily as a wood preservative, were conducted by feeding diets containing a technical-grade composite, Dowicide EC-7 (a technical grade formulation), or pure pentachlorophenol to groups of B6C3F1 mice for 30 days. These three grades plus another commercial grade of pentachlorophenol (DP-2) were used in 6-month studies. These studies were followed by 2-year carcinogenicity studies of technical-grade pentachlorophenol and of Dowicide EC-7 in feed. Genetic toxicology studies were conducted in Salmonella typhimurium and in Chinese hamster ovary (CHO) cells. Thirty-Day and Sixteen-Month Studies: Groups of 19 male mice and 5-15 female mice were fed diets containing 0, 20, 100, 500, 2,500, or 12,500 ppm technical-grade pentachlorophenol, Dowicide EC-7, or pure pentachlorophenol for 30 consecutive days. Necropsies and histopathologic examinations were performed on all animals. Selected organs were weighed. Supplemental analyses included hematology, serum chemistry, urinalysis, immunology, and hepatic enzyme induction. Compound-related deaths were observed at the highest dose (12,500 ppm) with all three materials and at 2,500 ppm with EC-7 and pure pentachlorophenol (males only). Decreases in body weight gain were also observed in the groups in which deaths occurred. Diffuse centrilobular cytomegaly, karyomegaly, nuclear atypia, degeneration, or necrosis of the liver were compound-related lesions observed in all groups that received pure pentachlorophenol, technical-grade pentachlorophenol, or EC-7 at 500 ppm and above. Serum enzymes associated with liver injury were increased. In the 6-month studies, groups of 10 male and 10 female mice were given diets containing the various grades of pentachlorophenol at the following dietary concentrations: 200, 600, or 1,800 ppm technical-grade pentachlorophenol; 200, 600, or 1,200 ppm DP-2 (not used in the 30-day studies); 200, 600, or 1,200 ppm EC-7; or 200, 500, or 1,500 ppm

  11. NTP Toxicology and Carcinogenesis Studies of Polyvinyl Alcohol (CAS No.9002-89-5) in Female B6C3F1 Mice (Intravaginal Studies).

    PubMed

    1998-05-01

    Polyvinyl alcohol is produced primarily for use in textile sizing, adhesives, polymerization aids, and paper coatings. It is also used in surgical drapes, towels, and gauze sponges; protective gloves; cosmetic formulations; topical ophthalmic preparations; plastic sponge implants for reconstructive surgery; and intravaginal contraceptive foam and film. In addition, polyvinyl alcohol is used with magnesium sulfate to dilate the cervix of women prior to induction of labor. It is estimated that hundreds of thousands of women in the United States use an intravaginal product containing polyvinyl alcohol each year. The Food and Drug Administration nominated low-viscosity polyvinyl alcohol for a 2-year study because of concern about the lack of information about the long-term toxic and carcinogenic effects by the intravaginal route. Female B6C3F1 mice received polyvinyl alcohol (approximately 99% pure) in deionized water by intravaginal administration for 30 days or 2 years. 30-DAY STUDY IN MICE: Three groups of 50 female B6C3F1 mice were used in this intravaginal study. The vehicle control group received only 20 &mgr;L of a deionized water vehicle. The other two groups each received 20 &mgr;L of 25% polyvinyl alcohol in deionized water. Animals in one dose group were returned to their cages after dosing; animals in the other dose group were restrained in a vertical nose-down position in restraint bags for several minutes after dosing. Animals were dosed daily for 30 consecutive days. All mice survived to the end of the study. The final mean body weights and body weight gains of dosed mice were similar to those of the vehicle control group. Abnormalities noted in the vaginal area after dosing included vaginal plugs, secretions, and swelling. These vaginal changes were minimal to mild and occurred in vehicle controls as well as in dosed mice. Restraint of mice after dosing appeared to eliminate vaginal secretions but increased both the incidence of vaginal irritation and

  12. NTP Toxicology and Carcinogenesis Studies of Glycidol (CAS No. 556-52-5) In F344/N Rats and B6C3F1 Mice (Gavage Studies).

    PubMed

    1990-03-01

    Glycidol is a viscous liquid that is used as a stabilizer in the manufacture of vinyl polymers, as an additive for oil and synthetic hydraulic fluids, and as a diluent in some epoxy resins. NTP Toxicology and Carcinogenesis studies were conducted by administering glycidol (94% pure, containing 1.2% 3-methoxy-1,2-propanediol, 0.4% 3-chloro-1,2-propanediol, 2.8% diglycidyl ether, and 1.1% 2,6-dimethanol-1,4-dioxane) in water by gavage to groups of F344/N rats and B6C3F1 mice of each sex for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Chinese hamster ovary (CHO) cells, Drosophila melanogaster, and the bone marrow of male B6C3F1 mice. Sixteen-Day Studies: Glycidol doses for groups of five rats or five mice of each sex ranged from 37.5 to 600 mg/kg; vehicle controls received distilled water. All rats that received 600 mg/kg died between days 3 and 13. Edema and degeneration of the epididymal stroma, atrophy of the testis, and granulomatous inflammation of the epididymis occurred in males that received 300 mg/kg. All mice that received 600 mg/kg and two males and two females that received 300 mg/kg died by day 4 of the studies. Focal demyelination in the medulla and thalamus of the brain occurred in all female mice that received 300 mg/kg. Thirteen-Week Studies: Doses for groups of 10 rats ranged from 25 to 400 mg/kg, and doses for groups of 10 mice ranged from 19 to 300 mg/kg; vehicle controls received distilled water. All rats that received 400 mg/kg died by week 2; three males and one female that received 200 mg/kg died during weeks 11-12. Final mean body weights of male rats that received 50, 100, or 200 mg/kg were 96%-85% that of vehicle controls; final mean body weights of female rats receiving the same doses were 95%-89% that of vehicle controls. Sperm count and sperm motility were reduced in male rats that received 100 or 200 mg/kg. Necrosis of the cerebellum, demyelineation in the medulla of the brain

  13. NTP Toxicology and Carcinogenesis Studies of Pentachloroanisole (CAS No. 1825-21-4) in F344 Rats and B6C3F1 Mice (Feed Studies).

    PubMed

    1993-04-01

    Pentachloroanisole is a chlorinated aromatic compound which is widely distributed at low levels in the environment and in food products. Formation of pentachloroanisole in the environment may result from the degradation of structurally related, commercially important, ubiquitous chlorinated aromatic compounds such as pentachlorophenol and pentachloronitrobenzene which are known rodent toxins or carcinogens. Toxicology and carcinogenesis studies were conducted by administering pentachloroanisole (>99% pure) in corn oil by gavage to groups of male and female F344/N rats and B6C3F1 mice for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium strains, mouse lymphoma cells, and Chinese hamster ovary cells. 16-DAY STUDIES IN RATS: Groups of five male and five female F344/N rats were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 16 days at doses of 0, 100, 125, 150, 175, or 200 mg/kg body weight. Deaths occurred during days 2 and 3 in rats receiving doses of 125 mg/kg or greater; these deaths were considered directly related to pentachloroanisole administration. No biologically significant changes in mean body weight gains or final body weights were noted in the 100 mg/kg groups of rats. Because of the high early mortality rate, valid comparisons of body weight differences in other dose groups could not be made. Inactivity was noted in all dose groups. Rats administered doses of 125 mg/kg or greater also exhibited dyspnea. 16-DAY STUDIES IN MICE: Groups of five male and five female B6C3F1 mice were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 16 days at doses of 0, 100, 175, 250, 325, or 400 mg/kg. Deaths occurred during days 2 and 3 in mice receiving doses of 175 mg/kg or greater; these deaths were considered directly related to chemical administration. No biologically significant changes in mean body weight gains or final body weights

  14. NTP Technical Report on the Toxicology and Carcinogenesis Studies of Chloral Hydrate (Cas No. 302-17-0) in B6C3F1 mice (Gavage Studies).

    PubMed

    2002-02-01

    Chloral hydrate is used medically as a sedative or hypnotic and as a rubefacient in topical preparations, and it is often given to children as a sedative during dental and other medical procedures. Chloral hydrate is used as a central nervous system depressant and sedative in veterinary medicine and as a general anesthetic in cattle and horses. It is a byproduct of the chlorination of water and has been detected in plant effluent after the bleaching of softwood pulp. Chloral, the anhydrous form of chloral hydrate, is used as a synthetic intermediate in the production of insecticides and herbicides. Chloral hydrate was nominated for study by the Food and Drug Administration based upon widespread human exposure and its potential hepatotoxicity and the toxicity of related chemicals. One goal of the study was to assess the effect of the animal’s age and the duration of dosing on the tumorigenicity of chloral hydrate. Beginning on postnatal day 28, female B6C3F1 mice received chloral hydrate (99.5% pure) in water by gavage for 3, 6, or 12 months, 2 years, or as a single dose; on postnatal day 15, male and female B6C3F1 mice received a single dose by gavage. Tumorigenicity was assessed for 2 years after the initial dose. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, Drosophila melanogaster, and mouse bone marrow cells. 2-YEAR STUDY Groups of female mice (regimens A, B, C, and D) and groups of male mice (regimen E) received chloral hydrate in distilled water by gavage; control groups received distilled water only. In regimen A, groups of 48 female mice received 0, 25, 50, or 100 mg chloral hydrate/kg body weight 5 days per week for 104 weeks beginning when they were 28 days old. In regimen B, 24 female mice received 0 mg/kg and three groups of 48 female mice received 100 mg/kg 5 days per week beginning when they were 28 days old. Eight mice from the 0 and 100 mg/kg groups were killed at 3, 6, or 12 months. The

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

    PubMed

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

    2015-03-12

    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 molecules that guide the Cas9 nuclease to the viral targets (protospacers). Streptococcus pyogenes Cas9 cleavage of the viral genome requires the presence of a 5'-NGG-3' protospacer adjacent motif (PAM) sequence immediately downstream of the viral target. It is not known whether and how viral sequences flanked by the correct PAM are chosen as new spacers. Here we show that Cas9 selects functional spacers by recognizing their PAM during spacer acquisition. The replacement of cas9 with alleles that lack the PAM recognition motif or recognize an NGGNG PAM eliminated or changed PAM specificity during spacer acquisition, respectively. Cas9 associates with other proteins of the acquisition machinery (Cas1, Cas2 and Csn2), presumably to provide PAM-specificity to this process. These results establish a new function for Cas9 in the genesis of prokaryotic immunological memory.

  16. Carcinogenesis Studies of Allyl Isovalerate (CAS No. 2835-39-4) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

    PubMed

    1983-05-01

    Allyl isovalerate, a synthetic fragrance and flavoring ingredient in use since the 1950's, may be found in various products at the following concentrations: soap, 30 ppm; detergent, 3 ppm; creams, 15 ppm; perfume, 50 ppm; nonalcoholic beverages, 9 ppm; ice cream, 18 ppm; candy, 22 ppm; baked goods, 15-48 ppm; and gelatins and puddings, 1 ppm. A colorless liquid with an apple-like odor and taste, allyl isovalerate is approved by the U.S. Food and Drug Administration for use in foods. Specific production figures are not available, but U.S. production in 1980 exceeded 1,000 pounds. Carcinogenesis studies of allyl isovalerate (96% pure) were conducted by administering the test chemical in corn oil gavage to groups of 50 male and 50 female F344/N rats and to groups of 50 male and 50 female B6C3F1 mice at doses of 31 or 62 mg/kg. The doses selected were based on the chemically-induced toxic effects and depressed weight gains obtained from the 13-week studies. Doses were administered five times per week for 103 weeks. Groups of 50 rats and 50 mice of each sex received corn oil by gavage on the same dosing schedule and served as vehicle controls. Survival and mean body weight gain of rats of each sex and male mice were not adversely affected by the administration of allyl isovalerate. The significantly lower survival (P=0.001) and the lower mean body weight of low-dose female as compared with controls are likely consequences of the high incidence of a genital tract infection in the low-dose females. This infection was probably responsible for the deaths of 11/19 control, 22/33 low-dose, and 13/25 high-dose female mice that died before the end of the study. Squamous cell papillomas and epithelial hyperplasia of the nonglandular stomach were observed in dosed male mice in the 2-year studies (squamous cell papillomas: 0/50, 1/50, 2%, 3/48, 6%; epithelial hyperplasia: 1/50, 2%, 1/50, 2%, 7/48, 15%). The papillomas occurred with a significant positive trend (P<0.05). The

  17. Corrective Action Investigation Plan for Corrective Action Unit 137: Waste Disposal Sites, Nevada Test Site, Nevada, Rev. No.:0

    SciTech Connect

    Wickline, Alfred

    2005-12-01

    This Corrective Action Investigation Plan (CAIP) contains project-specific information including facility descriptions, environmental sample collection objectives, and criteria for conducting site investigation activities at Corrective Action Unit (CAU) 137: Waste Disposal Sites. This CAIP has been developed in accordance with the ''Federal Facility Agreement and Consent Order'' (FFACO) (1996) that was agreed to by the State of Nevada, the U.S. Department of Energy (DOE), and the U.S. Department of Defense. Corrective Action Unit 137 contains sites that are located in Areas 1, 3, 7, 9, and 12 of the Nevada Test Site (NTS), which is approximately 65 miles (mi) northwest of Las Vegas, Nevada (Figure 1-1). Corrective Action Unit 137 is comprised of the eight corrective action sites (CASs) shown on Figure 1-1 and listed below: (1) CAS 01-08-01, Waste Disposal Site; (2) CAS 03-23-01, Waste Disposal Site; (3) CAS 03-23-07, Radioactive Waste Disposal Site; (4) CAS 03-99-15, Waste Disposal Site; (5) CAS 07-23-02, Radioactive Waste Disposal Site; (6) CAS 09-23-07, Radioactive Waste Disposal Site; (7) CAS 12-08-01, Waste Disposal Site; and (8) CAS 12-23-07, Waste Disposal Site. The Corrective Action Investigation (CAI) will include field inspections, radiological surveys, geophysical surveys, sampling of environmental media, analysis of samples, and assessment of investigation results, where appropriate. Data will be obtained to support corrective action alternative evaluations and waste management decisions. The CASs in CAU 137 are being investigated because hazardous and/or radioactive constituents may be present in concentrations that could potentially pose a threat to human health and the environment. Existing information on the nature and extent of potential contamination is insufficient to evaluate and recommend corrective action alternatives for the CASs. Additional information will be generated by conducting a CAI before evaluating and selecting corrective action

  18. Toxicology and carcinogenesis studies of pulegone (CAS No. 89-82-7) in F344/N rats and B6C3F1 mice (gavage studies).

    PubMed

    2011-08-01

    Several essential oils contain pulegone and are used for flavoring foods, drinks, and dental products, as fragrance agents, and in herbal medicines. Pulegone was nominated for study by the National Institute of Environmental Health Sciences based on the potential for human exposure and the absence of carcinogenicity data. Male and female F344/N rats and B6C3F1 mice received pulegone (approximately 96% pure) by gavage for 2 weeks, 3 months, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Escherichia coli, and mouse peripheral blood erythrocytes. 2-WEEK STUDY IN RATS: Groups of five male and five female rats were administered 0, 37.5, 75, 150, 300, or 600 mg pulegone/kg body weight in corn oil by gavage, 5 days per week for 16 days. All male rats and nearly all female rats in the 300 and 600 mg/kg groups died prior to the end of the study. All moribund sacrifices and early deaths were attributed to liver toxicity. Mean body weight gains of males administered 37.5 or 150 mg/kg were significantly less than that of the vehicle controls. Clinical findings in 300 and 600 mg/kg rats included nasal/eye discharge, thinness, lethargy, and ruffled fur. Liver and kidney weights of dosed groups of females were generally significantly greater than those of the vehicle control group. The incidences of necrosis and cytoplasmic vacuolization of the liver in 300 and 600 mg/kg males and females were significantly greater than those in the vehicle control groups. 2-WEEK STUDY IN MICE: Groups of five male and five female mice were administered 0, 18.75, 37.5, 75, 150, or 300 mg pulegone/kg body weight in corn oil by gavage, 5 days per week for 16 days. Four females and one male in the 300 mg/kg groups died by study day 5. All early deaths were attributed to liver toxicity. Mean body weights of the dosed groups were similar to those of the vehicle controls. Clinical findings were observed only in 300 mg/kg mice and included thinness, lethargy, and ruffled

  19. NTP Toxicology and Carcinogenesis Studies of Nickel Oxide (CAS No. 1313-99-1) in F344 Rats and B6C3F1 Mice (Inhalation Studies).

    PubMed

    1996-07-01

    Nickel oxide (NiO) "sinters" are used in stainless steel and alloy steel production. Nickel oxide was nominated by the National Cancer Institute to the NTP for testing because exposure to this form of nickel is prevalent in the nickel industry. Increased incidences of lung and nasal sinus cancers have occurred among workers in certain nickel refining facilities, and nickel oxide was studied as part of a class study of nickel compounds. Male and female F344/N rats and B6C3F1 mice were exposed to nickel oxide (high temperature, green nickel oxide; mass median diameter 2.2 +/- 2.6 &mgr;m; at least 99% pure) by inhalation for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in peripheral blood of B6C3F1 mice exposed to nickel oxide for 13 weeks. 16-DAY STUDY IN RATS: Groups of five male and five female F344/N rats were exposed to 0, 1.2, 2.5, 5, 10, or 30 mg nickel oxide/m(3)(equivalent to 0, 0.9, 2.0, 3.9, 7.9, or 23.6 mg nickel/m(3)) by inhalation for 6 hours per day, 5 days per week for a total of 12 exposure days during a 16-day period. Additional groups of five male and five female rats were exposed to 0, 1.2, 5, or 10 mg/m(3) for tissue burden studies. All core study rats survived until the end of the study, final mean body weights of exposed male and female rats were similar to those of the controls, and there were no clinical findings related to nickel oxide exposure. Absolute and relative lung weights of male and female rats exposed to 10 or 30 mg/m(3) were significantly greater than those of the controls. Pigment particles in alveolar macrophages or within the alveolar spaces were observed in the lungs of exposed groups of males and females. Chronic-active inflammation and accumulation of macrophages in alveolar spaces of the lungs and hyperplasia in the respiratory tract lymph nodes were most severe in 10 and 30 mg/m(3) males and females. Hyperplasia of bronchial lymph nodes occurred in 30 mg/m(3) rats. Atrophy of the olfactory

  20. NTP Toxicology and Carcinogenesis Studies of Nickel Subsulfide (CAS No. 12035-72-2) in F344 Rats and B6C3F1 Mice (Inhalation Studies).

    PubMed

    1996-07-01

    Nickel subsulfide is used in the manufacture of lithium batteries and is a major component in the refining of certain nickel ores. Nickel subsulfide was nominated as part of a class study of nickel compounds, for which there was little information on the toxic and carcinogenic effects of inhalation exposure. Male and female F334/N rats and B6C3F1 mice were exposed to nickel subsulfide (at least 97% pure; the mean value for the mass median aerodynamic diameter at each exposure concentration ranged from 2.0 to 2.2 mm by inhalation 6 hours per day, 5 days per week, for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, and mouse peripheral blood samples were analyzed for frequency of micronucleated normochromatic erythrocytes. 16-DAY STUDY IN RATS: Groups of five male and five female F344/N rats were exposed to atmospheres containing 0, 0.6, 1.2, 2.5, 5, or 10 mg nickel subsulfide/m(3) (equivalent to 0, 0.44, 0.88, 1.83, 3.65, and 7.33 mg nickel/m(3)) 6 hours per day, 5 days per week for a total of 12 exposure days during a 16-day period. Additionalmgroups of three male and three female rats were exposed to 0, 0.6, 2.5, or 10 mg/m(3) for tissue burden studies. One male exposed to 10 mg nickel subsulfide/m(3) in the core study died on day 14; all other rats survived until the end of the study. Final mean body weights and mean body weight gains of males exposed to 5 or 10 mg nickel subsulfide/m(3) and females exposed to 2.5, 5, or 10 mg/m(3) were significantly lower than those of the controls. Clinical findings of toxicity on day 5 of the study included labored respiration in 10 mg/m(3) males and 5 and 10 mg/m(3) females and dehydration in 5 and 10 mg/m(3) females. Absolute and relative lung weights of 2.5, 5, and 10 mg/m(3) males and all exposed groups of females were significantlymgreater than those of the controls, as was the absolute lung weight of 1.2 mg/m(3) males. Inflammation of the lung and atrophy of the nasal

  1. An fMRI study of perception and action in deaf signers.

    PubMed

    Okada, Kayoko; Rogalsky, Corianne; O'Grady, Lucinda; Hanaumi, Leila; Bellugi, Ursula; Corina, David; Hickok, Gregory

    2016-02-01

    Since the discovery of mirror neurons, there has been a great deal of interest in understanding the relationship between perception and action, and the role of the human mirror system in language comprehension and production. Two questions have dominated research. One concerns the role of Broca's area in speech perception. The other concerns the role of the motor system more broadly in understanding action-related language. The current study investigates both of these questions in a way that bridges research on language with research on manual actions. We studied the neural basis of observing and executing American Sign Language (ASL) object and action signs. In an fMRI experiment, deaf signers produced signs depicting actions and objects as well as observed/comprehended signs of actions and objects. Different patterns of activation were found for observation and execution although with overlap in Broca's area, providing prima facie support for the claim that the motor system participates in language perception. In contrast, we found no evidence that action related signs differentially involved the motor system compared to object related signs. These findings are discussed in the context of lesion studies of sign language execution and observation. In this broader context, we conclude that the activation in Broca's area during ASL observation is not causally related to sign language understanding. PMID:26796716

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

    PubMed Central

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

    2014-01-01

    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 produced remarkable sequence diversity. We selected eight representatives of phylogenetically defined type II CRISPR-Cas groups to analyze possible coevolution of Cas9 and dual-RNA. We demonstrate that these two components are interchangeable only between closely related type II systems when the PAM sequence is adjusted to the investigated Cas9 protein. Comparison of the taxonomy of bacterial species that harbor type II CRISPR-Cas systems with the Cas9 phylogeny corroborates horizontal transfer of the CRISPR-Cas loci. The reported collection of dual-RNA:Cas9 with associated PAMs expands the possibilities for multiplex genome editing and could provide means to improve the specificity of the RNA-programmable Cas9 tool. PMID:24270795

  3. Identifying and Visualizing Functional PAM Diversity across CRISPR-Cas Systems.

    PubMed

    Leenay, Ryan T; Maksimchuk, Kenneth R; Slotkowski, Rebecca A; Agrawal, Roma N; Gomaa, Ahmed A; Briner, Alexandra E; Barrangou, Rodolphe; Beisel, Chase L

    2016-04-01

    CRISPR-Cas adaptive immune systems in prokaryotes boast a diversity of protein families and mechanisms of action, where most systems rely on protospacer-adjacent motifs (PAMs) for DNA target recognition. Here, we developed an in vivo, positive, and tunable screen termed PAM-SCANR (PAM screen achieved by NOT-gate repression) to elucidate functional PAMs as well as an interactive visualization scheme termed the PAM wheel to convey individual PAM sequences and their activities. PAM-SCANR and the PAM wheel identified known functional PAMs while revealing complex sequence-activity landscapes for the Bacillus halodurans I-C (Cascade), Escherichia coli I-E (Cascade), Streptococcus thermophilus II-A CRISPR1 (Cas9), and Francisella novicida V-A (Cpf1) systems. The PAM wheel was also readily applicable to existing high-throughput screens and garnered insights into SpyCas9 and SauCas9 PAM diversity. These tools offer powerful means of elucidating and visualizing functional PAMs toward accelerating our ability to understand and exploit the multitude of CRISPR-Cas systems in nature. PMID:27041224

  4. 75 FR 66822 - Study on Extraterritorial Private Rights of Action

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-29

    ... of the following methods: Electronic Comments Use the Commission's Internet comment form ( http://www... Commission will post all comments on the Commission's Internet Web site ( http://www.sec.gov ). Comments are...) directs that the study shall consider and analyze, among other things-- \\2\\ Section 929Y(a) of the...

  5. From Numbers to Action: A Preliminary Study of Retention.

    ERIC Educational Resources Information Center

    Greenlee, Shelia Parker; Greenlee, Harry

    This study examined retention at Christopher Newport University (CNU) in Newport News, Virginia, focusing on the demographic characteristics of those students who left the university as well as the reasons why they left. A total of 159 students who had attended CNU during the 1994-95 academic year but who did not re-enroll in the fall of 1995 were…

  6. Academies in Action: Case Studies from Camden and Pimlico, 2007

    ERIC Educational Resources Information Center

    Benn, Melissa

    2008-01-01

    Both main UK political parties lend enthusiastic public support to academies, in the name of supporting the nation's poorest pupils. But Gordon Brown's Labour is, in reality, unsure about this undemocratic model while the Tories may well in the future exploit academy "independence" for retrograde ends. Two contemporary case studies from London, in…

  7. Reading Comprehension across Different Genres: An Action Research Study

    ERIC Educational Resources Information Center

    Lubawski, Michael; Sheehan, Caitlyn

    2010-01-01

    Reading comprehension is often a forgotten skill in secondary classrooms. Teachers of older students assign readings of different styles and levels, but don't teach the skills necessary to process different kinds of texts. This study examined which strategies are effective in giving students the skills necessary to read for understanding by…

  8. A Multidisciplinary Osteoporosis Service-Based Action Research Study

    ERIC Educational Resources Information Center

    Whitehead, Dean; Keast, John; Montgomery, Val; Hayman, Sue

    2004-01-01

    Objective: To investigate an existing Trust-based osteoporosis service's preventative activity, determine any issues and problems and use this data to reorganise the service, as part of a National Health Service Executive/Regional Office-commissioned and funded study. Setting: A UK Hospital Trust's Osteoporosis Service. Design & Method: A…

  9. Distributed Curriculum Leadership in Action: A Hong Kong Case Study

    ERIC Educational Resources Information Center

    Law, Edmond; Galton, Maurice; Wan, Sally

    2010-01-01

    This study was designed primarily to investigate the impact of school-based curriculum development teams on teacher development within the tradition of school-based curriculum development. The results are expected to provide valuable insights for teachers, school management and policy making. Teacher interviews in a primary school in Hong Kong…

  10. CRISPR-Cas9 Can Inhibit HIV-1 Replication but NHEJ Repair Facilitates Virus Escape

    PubMed Central

    Wang, Gang; Zhao, Na; Berkhout, Ben; Das, Atze T

    2016-01-01

    Several recent studies demonstrated that the clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease Cas9 can be used for guide RNA (gRNA)-directed, sequence-specific cleavage of HIV proviral DNA in infected cells. We here demonstrate profound inhibition of HIV-1 replication by harnessing T cells with Cas9 and antiviral gRNAs. However, the virus rapidly and consistently escaped from this inhibition. Sequencing of the HIV-1 escape variants revealed nucleotide insertions, deletions, and substitutions around the Cas9/gRNA cleavage site that are typical for DNA repair by the nonhomologous end-joining pathway. We thus demonstrate the potency of CRISPR-Cas9 as an antiviral approach, but any therapeutic strategy should consider the viral escape implications. PMID:26796669

  11. [CRISPR/Cas system for genome editing in pluripotent stem cells].

    PubMed

    Vasil'eva, E A; Melino, D; Barlev, N A

    2015-01-01

    Genome editing systems based on site-specific nucleases became very popular for genome editing in modern bioengineering. Human pluripotent stem cells provide a unique platform for genes function study, disease modeling, and drugs testing. Consequently, technology for fast, accurate and well controlled genome manipulation is required. CRISPR/Cas (clustered regularly interspaced short palindromic repeat/CRISPR-associated) system could be employed for these purposes. This system is based on site-specific programmable nuclease Cas9. Numerous advantages of the CRISPR/Cas system and its successful application to human stem cells provide wide opportunities for genome therapy and regeneration medicine. In this publication, we describe and compare the main genome editing systems based on site-specific programmable nucleases and discuss opportunities and perspectives of the CRISPR/Cas system for application to pluripotent stem cells.

  12. CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling

    PubMed Central

    Platt, Randall J.; Chen, Sidi; Zhou, Yang; Yim, Michael J.; Swiech, Lukasz; Kempton, Hannah R.; Dahlman, James E.; Parnas, Oren; Eisenhaure, Thomas M.; Jovanovic, Marko; Graham, Daniel B.; Jhunjhunwala, Siddharth; Xavier, Ramnik J.; Langer, Robert; Anderson, Daniel G.; Hacohen, Nir; Regev, Aviv; Feng, Guoping; Sharp, Phillip A.; Zhang, Feng

    2014-01-01

    SUMMARY CRISPR-Cas9 is a versatile genome editing technology for studying the function of genetic elements. To broadly enable the application of Cas9 in vivo, we established a Cre-dependent Cas9 knockin mouse. We demonstrated in vivo as well as ex vivo genome editing using adeno-associated virus (AAV)-, lenti-virus-, or particle-mediated delivery of guide RNA in neurons, immune cells, and endothelial cells. Using these mice, we simultaneously modeled the dynamics of KRAS, p53, and LKB1, the top three significantly mutated genes in lung adenocarcinoma. Delivery of a single AAV vector in the lung generated loss-of-function mutations in p53 and LKB1, as well as homology-directed repair-mediated KRASG12D mutations, leading to macroscopic tumors of adeno-carcinoma pathology. Together, these results suggest that Cas9 mice empower a wide range of biological and disease modeling applications. PMID:25263330

  13. In vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9

    PubMed Central

    Swiech, Lukasz; Heidenreich, Matthias; Banerjee, Abhishek; Habib, Naomi; Li, Yinqing; Trombetta, John; Sur, Mriganka; Zhang, Feng

    2015-01-01

    Probing gene function in the mammalian brain can be greatly assisted with methods to manipulate the genome of neurons in vivo. The clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated endonuclease (Cas)9 from Streptococcus pyogenes (SpCas9)1 can be used to edit single or multiple genes in replicating eukaryotic cells, resulting in frame-shifting insertion/deletion (indel) mutations and subsequent protein depletion. Here, we delivered SpCas9 and guide RNAs using adeno-associated viral (AAV) vectors to target single (Mecp2) as well as multiple genes (Dnmt1, Dnmt3a and Dnmt3b) in the adult mouse brain in vivo. We characterized the effects of genome modifications in postmitotic neurons using biochemical, genetic, electrophysiological and behavioral readouts. Our results demonstrate that AAV-mediated SpCas9 genome editing can enable reverse genetic studies of gene function in the brain. PMID:25326897

  14. A multifunctional AAV-CRISPR-Cas9 and its host response.

    PubMed

    Chew, Wei Leong; Tabebordbar, Mohammadsharif; Cheng, Jason K W; Mali, Prashant; Wu, Elizabeth Y; Ng, Alex H M; Zhu, Kexian; Wagers, Amy J; Church, George M

    2016-10-01

    CRISPR-Cas9 delivery by adeno-associated virus (AAV) holds promise for gene therapy but faces critical barriers on account of its potential immunogenicity and limited payload capacity. Here, we demonstrate genome engineering in postnatal mice using AAV-split-Cas9, a multifunctional platform customizable for genome editing, transcriptional regulation, and other previously impracticable applications of AAV-CRISPR-Cas9. We identify crucial parameters that impact efficacy and clinical translation of our platform, including viral biodistribution, editing efficiencies in various organs, antigenicity, immunological reactions, and physiological outcomes. These results reveal that AAV-CRISPR-Cas9 evokes host responses with distinct cellular and molecular signatures, but unlike alternative delivery methods, does not induce extensive cellular damage in vivo. Our study provides a foundation for developing effective genome therapeutics.

  15. CRISPR/Cas9-mediated genome editing and gene replacement in plants: Transitioning from lab to field.

    PubMed

    Schaeffer, Scott M; Nakata, Paul A

    2015-11-01

    The CRISPR/Cas9 genome engineering system has ignited and swept through the scientific community like wildfire. Owing largely to its efficiency, specificity, and flexibility, the CRISPR/Cas9 system has quickly become the preferred genome-editing tool of plant scientists. In plants, much of the early CRISPR/Cas9 work has been limited to proof of concept and functional studies in model systems. These studies, along with those in other fields of biology, have led to the development of several utilities of CRISPR/Cas9 beyond single gene editing. Such utilities include multiplexing for inducing multiple cleavage events, controlling gene expression, and site specific transgene insertion. With much of the conceptual CRISPR/Cas9 work nearly complete, plant researchers are beginning to apply this gene editing technology for crop trait improvement. Before rational strategies can be designed to implement this technology to engineer a wide array of crops there is a need to expand the availability of crop-specific vectors, genome resources, and transformation protocols. We anticipate that these challenges will be met along with the continued evolution of the CRISPR/Cas9 system particularly in the areas of manipulation of large genomic regions, transgene-free genetic modification, development of breeding resources, discovery of gene function, and improvements upon CRISPR/Cas9 components. The CRISPR/Cas9 editing system appears poised to transform crop trait improvement.

  16. Photometry of stars in the Cas OB5 Associations

    NASA Astrophysics Data System (ADS)

    Tanriver, Mehmet; Keskin, Ahmet

    2016-07-01

    OB associations are a grouping of very young associations, contain 10-100 very hot massive stars, spectral types O and B. Also, the OB associations contain low and intermediate mass stars, too. Association members are believed to form within the same small volume inside a giant molecular cloud. Once the surrounding dust and gas is blown away, the remaining stars become not tied up and begin to drift separately. It is believed that the majority of all stars in the Milky Way were formed in OB associations. O type stars are short-lived, and will be at an end as supernovae after roundly a million years. OB associations are generally only a few million years in age or less. In this study, the photometry of UU Cas and field star which been Cas OB5 association member was carried out. Light curves and color diagrams are given in the study.

  17. CLOSURE REPORT FOR CORRECTIVE ACTION UNIT 204: STORAGE BUNKERS, NEVADA TEST SITE, NEVADA

    SciTech Connect

    2006-04-01

    Corrective Action Unit (CAU) 330 consists of four Corrective Action Sites (CASs) located in Areas 6, 22, and 23 of the Nevada Test Site (NTS). The unit is listed in the Federal Facility Agreement and Consent Order (FFACO, 1996) as CAU 330: Areas 6, 22, and 23 Tanks and Spill Sites. CAU 330 consists of the following CASs: CAS 06-02-04, Underground Storage Tank (UST) and Piping CAS 22-99-06, Fuel Spill CAS 23-01-02, Large Aboveground Storage Tank (AST) Farm CAS 23-25-05, Asphalt Oil Spill/Tar Release

  18. Streamlined Approach for Environmental Restoration Plan for Corrective Action Unit 330: Areas 6, 22, and 23 Tanks and Spill Sites, Nevada Test Site, Nevada

    SciTech Connect

    T. M. Fitzmaurice

    2001-08-01

    This Streamlined Approach for Environmental restoration (SAFER) plan addresses the action necessary for the closure of Corrective Action Unit (CAU) 330, Areas 6,22, and 23 Tanks and Spill Sites. The CAUs are currently listed in Appendix III of the Federal Facility Agreement and Consent Order (FFACO). This CAU is located at the Nevada Test Site (NTS) (Figure 1). CAU 330 consists of the following Corrective Action Sites (CASs): (1) CAS 06-02-04 - Consists of an underground tank and piping. This CAS is close to an area that was part of the Animal Investigation Program (AIP), conducted under the U.S. Public Health Service. Its purpose was to study and perform tests on the cattle and wild animals in and around the NTS that were exposed to radionuclides. It is unknown if this tank was part of these operations. (2) CAS 22-99-06 - Is a fuel spill that is believed to be a waste oil release which occurred when Camp Desert Rock was an active facility. This CAS was originally identified as being a small depression where liquids were poured onto the ground, located on the west side of Building T-1001. This building has been identified as housing a fire station, radio station, and radio net remote and telephone switchboard. (3) CAS 23-01-02 - Is a large aboveground storage tank (AST) farm that was constructed to provide gasoline and diesel storage in Area 23. The site consists of two ASTs, a concrete foundation, a surrounding earthen berm, associated piping, and unloading stations. (4) CAS 23-25-05 - Consists of an asphalt oil spill/tar release that contains a wash covered with asphalt oil/tar material, a half buried 208-liter (L) (55-gallon [gal]) drum, rebar, and concrete located in the vicinity.

  19. Rhetoric to action: a study of stakeholder perceptions of aging well in two local communities.

    PubMed

    Everingham, Jo-Anne; Lui, Chi-Wai; Bartlett, Helen; Warburton, Jeni; Cuthill, Michael

    2010-11-01

    This qualitative study of local perceptions of policy goals and action in relation to aging reports 31 stakeholder interviews within 2 Australian communities exploring (a) the meaning of aging well; and (b) preferred policy actions to achieve positive aging outcomes. Findings suggest that community perceptions of aging well are broadly consistent with the goals of national and international policy frameworks in focusing on 3 dimensions--health, social engagement, and security. Further, participants believe that achievement of positive aging outcomes requires a mix of self-help, community action, and government intervention--particularly government support and encouragement for aging well initiatives. PMID:20972930

  20. Sustained action tetracycline preparation--tetrabid-organon blood level study.

    PubMed

    Lucas, C R; Mugglestone, C J; Thomas, D R

    1977-01-01

    A comparison of the plasma levels of tetracycline obtained whilst taking standard therapeutic doses of Tetrabid-Organon was made in twelve healthy volunteers. Two standard production batches were used in the study which was conducted under double-blind conditions. Sampling 12 hours and 8 hours after dosing showed no significant differences in plasma levels, with each batch. Even at these lowest levels satisfactory concentrations were rapidly obtained following initial administration, and were maintained when the drug was given at 12-hourly intervals. No side-effects of the drug were noted.

  1. The cerebellum in action: a simulation and robotics study.

    PubMed

    Hofstötter, Constanze; Mintz, Matti; Verschure, Paul F M J

    2002-10-01

    The control or prediction of the precise timing of events are central aspects of the many tasks assigned to the cerebellum. Despite much detailed knowledge of its physiology and anatomy, it remains unclear how the cerebellar circuitry can achieve such an adaptive timing function. We present a computational model pursuing this question for one extensively studied type of cerebellar-mediated learning: the classical conditioning of discrete motor responses. This model combines multiple current assumptions on the function of the cerebellar circuitry and was used to investigate whether plasticity in the cerebellar cortex alone can mediate adaptive conditioned response timing. In particular, we studied the effect of changes in the strength of the synapses formed between parallel fibres and Purkinje cells under the control of a negative feedback loop formed between inferior olive, cerebellar cortex and cerebellar deep nuclei. The learning performance of the model was evaluated at the circuit level in simulated conditioning experiments as well as at the behavioural level using a mobile robot. We demonstrate that the model supports adaptively timed responses under real-world conditions. Thus, in contrast to many other models that have focused on cerebellar-mediated conditioning, we investigated whether and how the suggested underlying mechanisms could give rise to behavioural phenomena.

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

    PubMed Central

    2012-01-01

    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 present study is the first

  3. Corrective Action Plan for Corrective Action Unit 563: Septic Systems, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2009-03-31

    This Corrective Action Plan (CAP) has been prepared for Corrective Action Unit (CAU) 563, Septic Systems, in accordance with the Federal Facility Agreement and Consent Order. CAU 563 consists of four Corrective Action Sites (CASs) located in Areas 3 and 12 of the Nevada Test Site. CAU 563 consists of the following CASs: CAS 03-04-02, Area 3 Subdock Septic Tank CAS 03-59-05, Area 3 Subdock Cesspool CAS 12-59-01, Drilling/Welding Shop Septic Tanks CAS 12-60-01, Drilling/Welding Shop Outfalls Site characterization activities were performed in 2007, and the results are presented in Appendix A of the CAU 563 Corrective Action Decision Document. The scope of work required to implement the recommended closure alternatives is summarized below. CAS 03-04-02, Area 3 Subdock Septic Tank, contains no contaminants of concern (COCs) above action levels. No further action is required for this site; however, as a best management practice (BMP), all aboveground features (e.g., riser pipes and bumper posts) will be removed, the septic tank will be removed, and all open pipe ends will be sealed with grout. CAS 03-59-05, Area 3 Subdock Cesspool, contains no COCs above action levels. No further action is required for this site; however, as a BMP, all aboveground features (e.g., riser pipes and bumper posts) will be removed, the cesspool will be abandoned by filling it with sand or native soil, and all open pipe ends will be sealed with grout. CAS 12-59-01, Drilling/Welding Shop Septic Tanks, will be clean closed by excavating approximately 4 cubic yards (yd3) of arsenic- and chromium-impacted soil. In addition, as a BMP, the liquid in the South Tank will be removed, the North Tank will be removed or filled with grout and left in place, the South Tank will be filled with grout and left in place, all open pipe ends will be sealed with grout or similar material, approximately 10 yd3 of chlordane-impacted soil will be excavated, and debris within the CAS boundary will be removed. CAS 12

  4. Genetically modified mouse models in studies of luteinising hormone action.

    PubMed

    Huhtaniemi, Ilpo; Ahtiainen, Petteri; Pakarainen, Tomi; Rulli, Susana B; Zhang, Fu-Ping; Poutanen, Matti

    2006-06-27

    Numerous genetically modified mouse models have recently been developed for the study of the pituitary-gonadal interactions. They include spontaneous or engineered knockouts (KO) of the gonadotrophin-releasing hormone (GnRH) and its receptor, the gonadotrophin common-alpha(Calpha), luteinising hormone (LH) beta and follicle-stimulating hormone (FSH) beta subunits, and the two gonadotrophin receptors (R), LHR and FSHR. In addition, there are also transgenic (TG) mice overexpressing gonadotrophin subunits and producing supraphysiological levels of these hormones. These models have offered relevant phenocopies for similar mutations in humans and to a great extent expanded our knowledge on normal and pathological functions of the hypothalamic-pituitary-gonadal (HPG) axis. The purpose of this article is to review some of our recent findings on two such mouse models, the LHR KO mouse (LuRKO), and the hCG overexpressing TG mouse (hCG+).

  5. Microcalorimetry studies of the antimicrobial actions of Aconitum alkaloids.

    PubMed

    Shi, Yan-bin; Liu, Lian; Shao, Wei; Wei, Ting; Lin, Gui-mei

    2015-08-01

    The metabolic activity of organisms can be measured by recording the heat output using microcalorimetry. In this paper, the total alkaloids in the traditional Chinese medicine Radix Aconiti Lateralis were extracted and applied to Escherichia coli and Staphylococcus aureus. The effect of alkaloids on bacteria growth was studied by microcalorimetry. The power-time curves were plotted with a thermal activity monitor (TAM) air isothermal microcalorimeter and parameters such as growth rate constant (μ), peak-time (Tm), inhibitory ratio (I), and enhancement ratio (E) were calculated. The relationships between the concentration of Aconitum alkaloids and μ of E. coli or S. aureus were discussed. The results showed that Aconitum alkaloids had little effect on E. coli and had a potentially inhibitory effect on the growth of S. aureus.

  6. Canceling planned action: an FMRI study of countermanding saccades.

    PubMed

    Curtis, Clayton E; Cole, Michael W; Rao, Vikas Y; D'Esposito, Mark

    2005-09-01

    We investigated the voluntary control of motor behavior by studying the process of deciding whether or not to execute a movement. We imaged the human dorsal cortex while subjects performed a countermanding task that allowed us to manipulate the probability that subjects would be able to cancel a planned saccade in response to an imperative stop signal. We modeled the behavioral data as a race between gaze-shifting mechanisms and gaze-holding mechanisms towards a finish line where a saccade is generated or canceled, and estimated that saccade cancelation took approximately 160 ms. The frontal eye fields showed greater activation on stop signal trials regardless of successful cancelation, suggesting coactivation of saccade and fixation mechanisms. The supplementary eye fields, however, distinguished between successful and unsuccessful cancelation, suggesting a role in monitoring performance. These oculomotor regions play distinct roles in the decision processes mediating saccadic choice. PMID:15616130

  7. In Vivo Transcriptional Activation Using CRISPR/Cas9 in Drosophila.

    PubMed

    Lin, Shuailiang; Ewen-Campen, Ben; Ni, Xiaochun; Housden, Benjamin E; Perrimon, Norbert

    2015-10-01

    A number of approaches for Cas9-mediated transcriptional activation have recently been developed, allowing target genes to be overexpressed from their endogenous genomic loci. However, these approaches have thus far been limited to cell culture, and this technique has not been demonstrated in vivo in any animal. The technique involving the fewest separate components, and therefore the most amenable to in vivo applications, is the dCas9-VPR system, where a nuclease-dead Cas9 is fused to a highly active chimeric activator domain. In this study, we characterize the dCas9-VPR system in Drosophila cells and in vivo. We show that this system can be used in cell culture to upregulate a range of target genes, singly and in multiplex, and that a single guide RNA upstream of the transcription start site can activate high levels of target transcription. We observe marked heterogeneity in guide RNA efficacy for any given gene, and we confirm that transcription is inhibited by guide RNAs binding downstream of the transcription start site. To demonstrate one application of this technique in cells, we used dCas9-VPR to identify target genes for Twist and Snail, two highly conserved transcription factors that cooperate during Drosophila mesoderm development. In addition, we simultaneously activated both Twist and Snail to identify synergistic responses to this physiologically relevant combination. Finally, we show that dCas9-VPR can activate target genes and cause dominant phenotypes in vivo, providing the first demonstration of dCas9 activation in a multicellular animal. Transcriptional activation using dCas9-VPR thus offers a simple and broadly applicable technique for a variety of overexpression studies. PMID:26245833

  8. In Vivo Transcriptional Activation Using CRISPR/Cas9 in Drosophila

    PubMed Central

    Lin, Shuailiang; Ewen-Campen, Ben; Ni, Xiaochun; Housden, Benjamin E.; Perrimon, Norbert

    2015-01-01

    A number of approaches for Cas9-mediated transcriptional activation have recently been developed, allowing target genes to be overexpressed from their endogenous genomic loci. However, these approaches have thus far been limited to cell culture, and this technique has not been demonstrated in vivo in any animal. The technique involving the fewest separate components, and therefore the most amenable to in vivo applications, is the dCas9-VPR system, where a nuclease-dead Cas9 is fused to a highly active chimeric activator domain. In this study, we characterize the dCas9-VPR system in Drosophila cells and in vivo. We show that this system can be used in cell culture to upregulate a range of target genes, singly and in multiplex, and that a single guide RNA upstream of the transcription start site can activate high levels of target transcription. We observe marked heterogeneity in guide RNA efficacy for any given gene, and we confirm that transcription is inhibited by guide RNAs binding downstream of the transcription start site. To demonstrate one application of this technique in cells, we used dCas9-VPR to identify target genes for Twist and Snail, two highly conserved transcription factors that cooperate during Drosophila mesoderm development. In addition, we simultaneously activated both Twist and Snail to identify synergistic responses to this physiologically relevant combination. Finally, we show that dCas9-VPR can activate target genes and cause dominant phenotypes in vivo, providing the first demonstration of dCas9 activation in a multicellular animal. Transcriptional activation using dCas9-VPR thus offers a simple and broadly applicable technique for a variety of overexpression studies. PMID:26245833

  9. In Vivo Transcriptional Activation Using CRISPR/Cas9 in Drosophila.

    PubMed

    Lin, Shuailiang; Ewen-Campen, Ben; Ni, Xiaochun; Housden, Benjamin E; Perrimon, Norbert

    2015-10-01

    A number of approaches for Cas9-mediated transcriptional activation have recently been developed, allowing target genes to be overexpressed from their endogenous genomic loci. However, these approaches have thus far been limited to cell culture, and this technique has not been demonstrated in vivo in any animal. The technique involving the fewest separate components, and therefore the most amenable to in vivo applications, is the dCas9-VPR system, where a nuclease-dead Cas9 is fused to a highly active chimeric activator domain. In this study, we characterize the dCas9-VPR system in Drosophila cells and in vivo. We show that this system can be used in cell culture to upregulate a range of target genes, singly and in multiplex, and that a single guide RNA upstream of the transcription start site can activate high levels of target transcription. We observe marked heterogeneity in guide RNA efficacy for any given gene, and we confirm that transcription is inhibited by guide RNAs binding downstream of the transcription start site. To demonstrate one application of this technique in cells, we used dCas9-VPR to identify target genes for Twist and Snail, two highly conserved transcription factors that cooperate during Drosophila mesoderm development. In addition, we simultaneously activated both Twist and Snail to identify synergistic responses to this physiologically relevant combination. Finally, we show that dCas9-VPR can activate target genes and cause dominant phenotypes in vivo, providing the first demonstration of dCas9 activation in a multicellular animal. Transcriptional activation using dCas9-VPR thus offers a simple and broadly applicable technique for a variety of overexpression studies.

  10. Studies on the mode of action of the essential oil of lavender (Lavandula angustifolia P. Miller).

    PubMed

    Lis-Balchin, M; Hart, S

    1999-09-01

    Lavender (Lavandula angustifolia, P. Miller) is used in aromatherapy as a holistic relaxant and is said to have carminative, antiflatulence and anticolic properties. Its sedative nature, on inhalation, has been shown both in animals and man. Lavender has a spasmolytic activity on guineapig ileum and rat uterus in vitro and it also decreases the tone in the skeletal muscle preparation of the phrenic nerve-diaphragm of rats. As the mechanism of action has not been studied previously, the spasmolytic activity was studied in vitro using a guinea-pig ileum smooth muscle preparation. The mechanism of action was postsynaptic and not atropine-like. The spasmolytic effect of lavender oil was most likely to be mediated through cAMP, and not through cGMP. The mode of action of linalool, one of lavender's major components, reflected that of the whole oil. The mode of action of lavender oil resembled that of geranium and peppermint oils.

  11. Activity-Based Teaching in Social Studies Education: An Action Research

    ERIC Educational Resources Information Center

    Akkus, Zekerya

    2015-01-01

    The aim of this study was to determine pre-service social studies teachers' skills to plan and apply the activity-based teaching and contribute to their development of these skills. In the study, the action research design of qualitative research was used. The sample of the study consisted of 6 pre-service teachers who were 4th year students at…

  12. Biosphere reserves in action: Case studies of the American experience

    SciTech Connect

    1995-06-26

    For nearly 20 years, biosphere reserves have offered a unique framework for building the knowledge, skills, and attitudes required for conservation and sustainable use of ecosystems. The 12 case studies in this volume chronicle many of the cooperative efforts to implement the biosphere reserve concept in the United States. Considered together, these efforts involve more than 20 types of protected areas, and the participation of all levels of government, and many private organizations, academic institutions, citizens groups, and individuals. Biosphere reserves are multi-purpose areas that are nominated by the national committee of the Man and the Biosphere Program (MAB) and designated by the United Nations Educational, Scientific and Cultural Organization (UNESCO) to serve as demonstration areas for cooperation in building harmonious relationships between human activities and the conservation of ecosystems and biological diversity. Each biosphere reserve exemplifies the characteristic ecosystems of one of the worlds biogeographical regions. It is a land or coas%arine area involving human communities as integral components and including resources managed for objectives ranging from complete protection to intensive, yet sustainable development. A biosphere reserve is envisioned as a regional ''landscape for learning'' in which monitoring, research, education, and training are encouraged to support sustainable conservation of natural and managed ecosystems. It is a framework for regional cooperation involving government decisionmakers, scientists, resource managers, private organizations and local people (i.e., the biosphere reserve ''stakeholders''). Finally, each biosphere reserve is part of a global network for sharing information and experience to help address complex problems of conservation and development. The 12 case studies presented in this report represent only a few of the possible evolutions of a biosphere reserve in its efforts to reach out to the local

  13. Dopaminergic action beyond its effects on motor function: imaging studies.

    PubMed

    Brooks, David J

    2006-08-01

    Along with motor programming, it is now thought that tonic release of dopamine in the striatum acts to focus and filter non-motor activities such as working memory, implicit learning, decision making, and planning. Additionally, thresholds to painful stimuli may well be dopamine dependant. Phasic (burst) release of dopamine in the basal ganglia and frontal areas is thought to play a role in alerting organisms to novel and potentially rewarding stimuli and in mediating contextual learning. Dopamine release also drives a craving for stimuli and facilitates their enjoyment. Functional imaging can help elucidate the role of dopamine in mediating non-motor activities. The integrity of dopamine terminal function can be measured with PET and SPECT in vivo in health and Parkinson's disease (PD) and this can be correlated with performance of executive tasks. In addition, these imaging modalities allow dopamine release in response to stimuli (both rewarding and unrewarding) to be detected, as reflected by changes in D2 receptor availability to radioligands. Finally, the functional effects of dopamine deficiency and its replacement can be monitored by studying patterns of brain activation, as evidenced by regional blood flow changes. In this review, some of the insights that imaging has given us concerning the role of dopamine in non-motor functions is presented.

  14. A 'suicide' CRISPR-Cas9 system to promote gene deletion and restoration by electroporation in Cryptococcus neoformans.

    PubMed

    Wang, Yu; Wei, Dongsheng; Zhu, Xiangyang; Pan, Jiao; Zhang, Ping; Huo, Liang; Zhu, Xudong

    2016-01-01

    Loss-of-function mutagenesis is an important tool used to characterize gene functions, and the CRISPR-Cas9 system is a powerful method for performing targeted mutagenesis in organisms that present low recombination frequencies, such as the serotype D strains of Cryptococcus neoformans. However, when the CRISPR-Cas9 system persists in the host cells, off-target effects and Cas9 cytotoxicity may occur, which might block subsequent genetic manipulation. Here, we report a method of spontaneously eliminating the CRISPR-Cas9 system without impairing its robust editing function. We successfully expressed single guide RNA under the driver of an endogenous U6 promoter and the human codon-optimized Cas9 endonuclease with an ACT1 promoter. This system can effectively generate an indel mutation and efficiently perform targeted gene disruption via homology-directed repair by electroporation in yeast. We then demonstrated the spontaneous elimination of the system via a cis arrangement of the CRISPR-Cas9 expression cassettes to the recombination construct. After a system-mediated double crossover, the CRISPR-Cas9 cassettes were cleaved and degraded, which was validated by Southern blotting. This 'suicide' CRISPR-Cas9 system enables the validation of gene functions by subsequent complementation and has the potential to minimize off-target effects. Thus, this technique has the potential for use in functional genomics studies of C. neoformans. PMID:27503169

  15. Foreign DNA acquisition by the I-F CRISPR-Cas system requires all components of the interference machinery.

    PubMed

    Vorontsova, Daria; Datsenko, Kirill A; Medvedeva, Sofia; Bondy-Denomy, Joseph; Savitskaya, Ekaterina E; Pougach, Ksenia; Logacheva, Maria; Wiedenheft, Blake; Davidson, Alan R; Severinov, Konstantin; Semenova, Ekaterina

    2015-12-15

    CRISPR immunity depends on acquisition of fragments of foreign DNA into CRISPR arrays. For type I-E CRISPR-Cas systems two modes of spacer acquisition, naïve and primed adaptation, were described. Naïve adaptation requires just two most conserved Cas1 and Cas2 proteins; it leads to spacer acquisition from both foreign and bacterial DNA and results in multiple spacers incapable of immune response. Primed adaptation requires all Cas proteins and a CRISPR RNA recognizing a partially matching target. It leads to selective acquisition of spacers from DNA molecules recognized by priming CRISPR RNA, with most spacers capable of protecting the host. Here, we studied spacer acquisition by a type I-F CRISPR-Cas system. We observe both naïve and primed adaptation. Both processes require not just Cas1 and Cas2, but also intact Csy complex and CRISPR RNA. Primed adaptation shows a gradient of acquisition efficiency as a function of distance from the priming site and a strand bias that is consistent with existence of single-stranded adaption intermediates. The results provide new insights into the mechanism of spacer acquisition and illustrate surprising mechanistic diversity of related CRISPR-Cas systems.

  16. A 'suicide' CRISPR-Cas9 system to promote gene deletion and restoration by electroporation in Cryptococcus neoformans.

    PubMed

    Wang, Yu; Wei, Dongsheng; Zhu, Xiangyang; Pan, Jiao; Zhang, Ping; Huo, Liang; Zhu, Xudong

    2016-08-09

    Loss-of-function mutagenesis is an important tool used to characterize gene functions, and the CRISPR-Cas9 system is a powerful method for performing targeted mutagenesis in organisms that present low recombination frequencies, such as the serotype D strains of Cryptococcus neoformans. However, when the CRISPR-Cas9 system persists in the host cells, off-target effects and Cas9 cytotoxicity may occur, which might block subsequent genetic manipulation. Here, we report a method of spontaneously eliminating the CRISPR-Cas9 system without impairing its robust editing function. We successfully expressed single guide RNA under the driver of an endogenous U6 promoter and the human codon-optimized Cas9 endonuclease with an ACT1 promoter. This system can effectively generate an indel mutation and efficiently perform targeted gene disruption via homology-directed repair by electroporation in yeast. We then demonstrated the spontaneous elimination of the system via a cis arrangement of the CRISPR-Cas9 expression cassettes to the recombination construct. After a system-mediated double crossover, the CRISPR-Cas9 cassettes were cleaved and degraded, which was validated by Southern blotting. This 'suicide' CRISPR-Cas9 system enables the validation of gene functions by subsequent complementation and has the potential to minimize off-target effects. Thus, this technique has the potential for use in functional genomics studies of C. neoformans.

  17. A ‘suicide’ CRISPR-Cas9 system to promote gene deletion and restoration by electroporation in Cryptococcus neoformans

    PubMed Central

    Wang, Yu; Wei, Dongsheng; Zhu, Xiangyang; Pan, Jiao; Zhang, Ping; Huo, Liang; Zhu, Xudong

    2016-01-01

    Loss-of-function mutagenesis is an important tool used to characterize gene functions, and the CRISPR-Cas9 system is a powerful method for performing targeted mutagenesis in organisms that present low recombination frequencies, such as the serotype D strains of Cryptococcus neoformans. However, when the CRISPR-Cas9 system persists in the host cells, off-target effects and Cas9 cytotoxicity may occur, which might block subsequent genetic manipulation. Here, we report a method of spontaneously eliminating the CRISPR-Cas9 system without impairing its robust editing function. We successfully expressed single guide RNA under the driver of an endogenous U6 promoter and the human codon-optimized Cas9 endonuclease with an ACT1 promoter. This system can effectively generate an indel mutation and efficiently perform targeted gene disruption via homology-directed repair by electroporation in yeast. We then demonstrated the spontaneous elimination of the system via a cis arrangement of the CRISPR-Cas9 expression cassettes to the recombination construct. After a system-mediated double crossover, the CRISPR-Cas9 cassettes were cleaved and degraded, which was validated by Southern blotting. This ‘suicide’ CRISPR-Cas9 system enables the validation of gene functions by subsequent complementation and has the potential to minimize off-target effects. Thus, this technique has the potential for use in functional genomics studies of C. neoformans. PMID:27503169

  18. Seeing biological actions in 3D: An fMRI study

    PubMed Central

    Jastorff, Jan; Abdollahi, Rouhollah O.; Fasano, Fabrizio

    2015-01-01

    Abstract Precise kinematics or body configuration cannot be recovered from visual input without disparity information. Yet, no imaging study has investigated the role of disparity on action observation. Here, we investigated the interaction between disparity and the main cues of biological motion, kinematics and configuration, in two fMRI experiments. Stimuli were presented as point‐light figures, depicting complex action sequences lasting 21 s. We hypothesized that interactions could occur at any of the three levels of the action observation network, comprising occipitotemporal, parietal and premotor cortex, with premotor cortex being the most likely location. The main effects of kinematics and configuration confirmed that the biological motion sequences activated all three levels of the action observation network, validating our approach. The interaction between configuration and disparity activated only premotor cortex, whereas interactions between kinematics and disparity occurred at all levels of the action observation network but were strongest at the premotor level. Control experiments demonstrated that these interactions could not be accounted for by low level motion in depth, task effects, spatial attention, or eye movements, including vergence. These results underscore the role of premotor cortex in action observation, and in imitating others or responding to their actions. Hum Brain Mapp 37:203–219, 2016. © 2015 The Authors Human Brain Mapping Published by Wiley Periodicals, Inc. PMID:26510637

  19. Infants' perception of actions and situational constraints: an eye-tracking study.

    PubMed

    Elsner, Birgit; Pfeifer, Caroline; Parker, Charlene; Hauf, Petra

    2013-10-01

    Rational action understanding requires that infants evaluate the efficiency of a movement in achieving a goal with respect to situational constraints. In contrast, recent accounts have highlighted the impact of perceptual characteristics of the demonstrated movement or constraints to explain infants' behavior in so-called rational imitation tasks. The current study employed eye tracking to investigate how 13- to 15-month-old infants distribute their visual attention to different aspects of an action demonstration. In three tasks (touchlight, house, and obstacle), infants watched videos in which a model performed an unusual action while she was or was not restricted by situational constraints. Infants' overall looking to the demonstration as well as looking to four segments of the video (initial segment, constraint demonstration, action performance, and final segment) and to specific areas (constraint area of interest [AOI] and action AOI) was analyzed. Overall, infants looked longer at the demonstration in the constraint condition compared with the no-constraint condition. The condition differences occurred in the two video segments where the constraint or action was displayed and were especially profound for the constraint AOI. These findings indicate that infants processed the situational constraints. However, the pattern of condition differences varied slightly in the three tasks. In sum, the data imply that infants process perceptual characteristics of the movement or constraints and that low-level perceptual processes interact with higher level cognitive processes in infants' action perception. PMID:23410481

  20. Targeting ABCB1-mediated tumor multidrug resistance by CRISPR/Cas9-based genome editing

    PubMed Central

    Yang, Yang; Qiu, Jian-Ge; Li, Yong; Di, Jin-Ming; Zhang, Wen-Ji; Jiang, Qi-Wei; Zheng, Di-Wei; Chen, Yao; Wei, Meng-Ning; Huang, Jia-Rong; Wang, Kun; Shi, Zhi

    2016-01-01

    The RNA-guided clustered regularly interspaced short palindromic (CRISPR) in combination with a CRISPR-associated nuclease 9 (Cas9) nuclease system is a new rapid and precise technology for genome editing. In the present study, we applied the CRISPR/Cas9 system to target ABCB1 (also named MDR1) gene which encodes a 170 kDa transmembrane glycoprotein (P-glycoprotein/P-gp) transporting multiple types of chemotherapeutic drugs including taxanes, epipodophyllotoxins, vinca alkaloids and anthracyclines out of cells to contribute multidrug resistance (MDR) in cancer cells. Our data showed that knockout of ABCB1 by CRISPR/Cas9 system was succesfully archieved with two target sgRNAs in two MDR cancer cells due to the alteration of genome sequences. Knockout of ABCB1 by CRISPR/Cas9 system significantly enhances the sensitivity of ABCB1 substrate chemotherapeutic agents and the intracellular accumulation of rhodamine 123 and doxorubicin in MDR cancer cells. Although now there are lots of limitations to the application of CRISPR/Cas9 for editing cancer genes in human patients, our study provides valuable clues for the use of the CRISPR/Cas9 technology in the investigation and conquest of cancer MDR. PMID:27725879

  1. The impact of CRISPR repeat sequence on structures of a Cas6 protein-RNA complex

    SciTech Connect

    Wang, Ruiying; Zheng, Han; Preamplume, Gan; Shao, Yaming; Li, Hong

    2012-03-15

    The repeat-associated mysterious proteins (RAMPs) comprise the most abundant family of proteins involved in prokaryotic immunity against invading genetic elements conferred by the clustered regularly interspaced short palindromic repeat (CRISPR) system. Cas6 is one of the first characterized RAMP proteins and is a key enzyme required for CRISPR RNA maturation. Despite a strong structural homology with other RAMP proteins that bind hairpin RNA, Cas6 distinctly recognizes single-stranded RNA. Previous structural and biochemical studies show that Cas6 captures the 5' end while cleaving the 3' end of the CRISPR RNA. Here, we describe three structures and complementary biochemical analysis of a noncatalytic Cas6 homolog from Pyrococcus horikoshii bound to CRISPR repeat RNA of different sequences. Our study confirms the specificity of the Cas6 protein for single-stranded RNA and further reveals the importance of the bases at Positions 5-7 in Cas6-RNA interactions. Substitutions of these bases result in structural changes in the protein-RNA complex including its oligomerization state.

  2. CRISPR-Cas9 for medical genetic screens: applications and future perspectives.

    PubMed

    Xue, Hui-Ying; Ji, Li-Juan; Gao, Ai-Mei; Liu, Ping; He, Jing-Dong; Lu, Xiao-Jie

    2016-02-01

    CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR associated nuclease 9) systems have emerged as versatile and convenient (epi)genome editing tools and have become an important player in medical genetic research. CRISPR-Cas9 and its variants such as catalytically inactivated Cas9 (dead Cas9, dCas9) and scaffold-incorporating single guide sgRNA (scRNA) have been applied in various genomic screen studies. CRISPR screens enable high-throughput interrogation of gene functions in health and diseases. Compared with conventional RNAi screens, CRISPR screens incur less off-target effects and are more versatile in that they can be used in multiple formats such as knockout, knockdown and activation screens, and can target coding and non-coding regions throughout the genome. This powerful screen platform holds the potential of revolutionising functional genomic studies in the near future. Herein, we introduce the mechanisms of (epi)genome editing mediated by CRISPR-Cas9 and its variants, introduce the procedures and applications of CRISPR screen in functional genomics, compare it with conventional screen tools and at last discuss current challenges and opportunities and propose future directions.

  3. Efficiency and Inheritance of Targeted Mutagenesis in Maize Using CRISPR-Cas9.

    PubMed

    Zhu, Jinjie; Song, Ning; Sun, Silong; Yang, Weilong; Zhao, Haiming; Song, Weibin; Lai, Jinsheng

    2016-01-20

    CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins) is an adaptive immune system in bacteria and archaea to defend against invasion from foreign DNA fragments. Recently, it has been developed as a powerful targeted genome editing tool for a wide variety of species. However, its application in maize has only been tested with transiently expressed somatic cells or with a limited number of stable transgenic T0 plants. The exact efficiency and specificity of the CRISPR/Cas system in the highly complex maize genome has not been documented yet. Here we report an extensive study of the well-studied type II CRISPR-Cas9 system for targeted genome editing in maize, with the codon-optimized Cas9 protein and the short non-coding guide RNA generated through a functional maize U6 snRNA promoter. Targeted gene mutagenesis was detected for 90 loci by maize protoplast assay, with an average cleavage efficiency of 10.67%. Stable knockout transformants for maize phytoene synthase gene (PSY1) were obtained. Mutations occurred in germ cells can be stably inherited to the next generation. Moreover, no off-target effect was detected at the computationally predicted putative off-target loci. No significant difference between the transcriptomes of the Cas9 expressed and non-expressed lines was detected. Our results confirmed that the CRISPR-Cas9 could be successfully applied as a robust targeted genome editing system in maize.

  4. Streamlined Approach for Environmental Restoration Plan for Corrective Action Unit 107: Low Impact Soil Sites, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2009-03-31

    This Streamlined Approach for Environmental Restoration Plan covers activities associated with Corrective Action Unit (CAU) 107 of the Federal Facility Agreement and Consent Order (1996 [as amended February 2008]). CAU 107 consists of the following Corrective Action Sites (CASs) located in Areas 1, 2, 3, 4, 5, 9, 10, and 18 of the Nevada Test Site. {sm_bullet} CAS 01-23-02, Atmospheric Test Site - High Alt{sm_bullet} CAS 02-23-02, Contaminated Areas (2){sm_bullet} CAS 02-23-03, Contaminated Berm{sm_bullet} CAS 02-23-10, Gourd-Amber Contamination Area{sm_bullet} CAS 02-23-11, Sappho Contamination Area{sm_bullet} CAS 02-23-12, Scuttle Contamination Area{sm_bullet} CAS 03-23-24, Seaweed B Contamination Area{sm_bullet} CAS 03-23-27, Adze Contamination Area{sm_bullet} CAS 03-23-28, Manzanas Contamination Area{sm_bullet} CAS 03-23-29, Truchas-Chamisal Contamination Area{sm_bullet} CAS 04-23-02, Atmospheric Test Site T4-a{sm_bullet} CAS 05-23-06, Atmospheric Test Site{sm_bullet} CAS 09-23-06, Mound of Contaminated Soil{sm_bullet} CAS 10-23-04, Atmospheric Test Site M-10{sm_bullet} CAS 18-23-02, U-18d Crater (Sulky) Based on historical documentation, personnel interviews, site process knowledge, site visits, photographs, engineering drawings, field screening, analytical results, and the results of data quality objectives process (Section 3.0), closure in place with administrative controls or no further action will be implemented for CAU 107.

  5. Corrective Action Plan for Corrective Action Unit 166: Storage Yards and Contaminated Materials, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2007-10-01

    Corrective Action Unit (CAU) 166, Storage Yards and Contaminated Materials, is listed in the Federal Facility Agreement and Consent Order (FFACO) of 1996 (FFACO, 1996). CAU 166 consists of seven Corrective Action Sites (CASs) located in Areas 2, 3, 5, and 18 of the Nevada Test Site (NTS), which is located approximately 65 miles northwest of Las Vegas, Nevada (Figure 1). CAU 166 consists of the following CASs: (1) CAS 02-42-01, Cond. Release Storage Yd - North; (2) CAS 02-42-02, Cond. Release Storage Yd - South; (3) CAS 02-99-10, D-38 Storage Area; (4) CAS 03-42-01, Conditional Release Storage Yard; (5) CAS 05-19-02, Contaminated Soil and Drum; (6) CAS 18-01-01, Aboveground Storage Tank; and (7) CAS 18-99-03, Wax Piles/Oil Stain. Details of the site history and site characterization results for CAU 166 are provided in the approved Corrective Action Investigation Plan (CAIP) (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2006) and in the approved Corrective Action Decision Document (CADD) (NNSA/NSO, 2007).

  6. Regulation of the Type I-F CRISPR-Cas system by CRP-cAMP and GalM controls spacer acquisition and interference

    PubMed Central

    Patterson, Adrian G.; Chang, James T.; Taylor, Corinda; Fineran, Peter C.

    2015-01-01

    The CRISPR-Cas prokaryotic ‘adaptive immune systems’ represent a sophisticated defence strategy providing bacteria and archaea with protection from invading genetic elements, such as bacteriophages or plasmids. Despite intensive research into their mechanism and application, how CRISPR-Cas systems are regulated is less clear, and nothing is known about the regulation of Type I-F systems. We used Pectobacterium atrosepticum, a Gram-negative phytopathogen, to study CRISPR-Cas regulation, since it contains a single Type I-F system. The CRP-cAMP complex activated the cas operon, increasing the expression of the adaptation genes cas1 and cas2–3 in addition to the genes encoding the Csy surveillance complex. Mutation of crp or cyaA (encoding adenylate cyclase) resulted in reductions in both primed spacer acquisition and interference. Furthermore, we identified a galactose mutarotase, GalM, which reduced cas operon expression in a CRP- and CyaA-dependent manner. We propose that the Type I-F system senses metabolic changes, such as sugar availability, and regulates cas genes to initiate an appropriate defence response. Indeed, elevated glucose levels reduced cas expression in a CRP- and CyaA-dependent manner. Taken together, these findings highlight that a metabolite-sensing regulatory pathway controls expression of the Type I-F CRISPR-Cas system to modulate levels of adaptation and interference. PMID:26007654

  7. Temporal prediction modulates the evaluative processing of "good" action feedback: An electrophysiological study.

    PubMed

    Kimura, Kenta; Kimura, Motohiro; Iwaki, Sunao

    2016-10-01

    The present study aimed to investigate whether or not the evaluative processing of action feedback can be modulated by temporal prediction. For this purpose, we examined the effects of the predictability of the timing of action feedback on an ERP effect that indexed the evaluative processing of action feedback, that is, an ERP effect that has been interpreted as a feedback-related negativity (FRN) elicited by "bad" action feedback or a reward positivity (RewP) elicited by "good" action feedback. In two types of experimental blocks, the participants performed a gambling task in which they chose one of two cards and received an action feedback that indicated monetary gain or loss. In fixed blocks, the time interval between the participant's choice and the onset of the action feedback was fixed at 0, 500, or 1,000 ms in separate blocks; thus, the timing of action feedback was predictable. In mixed blocks, the time interval was randomly chosen from the same three intervals with equal probability; thus, the timing was less predictable. The results showed that the FRN/RewP was smaller in mixed than fixed blocks for the 0-ms interval trial, whereas there was no difference between the two block types for the 500-ms and 1,000-ms interval trials. Interestingly, the smaller FRN/RewP was due to the modulation of gain ERPs rather than loss ERPs. These results suggest that temporal prediction can modulate the evaluative processing of action feedback, and particularly good feedback, such as that which indicates monetary gain.

  8. Temporal prediction modulates the evaluative processing of "good" action feedback: An electrophysiological study.

    PubMed

    Kimura, Kenta; Kimura, Motohiro; Iwaki, Sunao

    2016-10-01

    The present study aimed to investigate whether or not the evaluative processing of action feedback can be modulated by temporal prediction. For this purpose, we examined the effects of the predictability of the timing of action feedback on an ERP effect that indexed the evaluative processing of action feedback, that is, an ERP effect that has been interpreted as a feedback-related negativity (FRN) elicited by "bad" action feedback or a reward positivity (RewP) elicited by "good" action feedback. In two types of experimental blocks, the participants performed a gambling task in which they chose one of two cards and received an action feedback that indicated monetary gain or loss. In fixed blocks, the time interval between the participant's choice and the onset of the action feedback was fixed at 0, 500, or 1,000 ms in separate blocks; thus, the timing of action feedback was predictable. In mixed blocks, the time interval was randomly chosen from the same three intervals with equal probability; thus, the timing was less predictable. The results showed that the FRN/RewP was smaller in mixed than fixed blocks for the 0-ms interval trial, whereas there was no difference between the two block types for the 500-ms and 1,000-ms interval trials. Interestingly, the smaller FRN/RewP was due to the modulation of gain ERPs rather than loss ERPs. These results suggest that temporal prediction can modulate the evaluative processing of action feedback, and particularly good feedback, such as that which indicates monetary gain. PMID:27412662

  9. Expression of OsCAS (Calcium-Sensing Receptor) in an Arabidopsis Mutant Increases Drought Tolerance

    PubMed Central

    Wei, Rongrong; Liu, Yang

    2015-01-01

    The calcium-sensing receptor (CaS), which is localized in the chloroplasts, is a crucial regulator of extracellular calcium-induced stomatal closure in Arabidopsis. It has homologs in Oryza sativa and other plants. These sequences all have a rhodanese-like protein domain, which has been demonstrated to be associated with specific stress conditions. In this study, we cloned the Oryza sativa calcium-sensing receptor gene (OsCAS) and demonstrated that OsCAS could sense an increase of extracellular Ca2+ concentration and mediate an increase in cytosolic Ca2+ concentration. The OsCAS gene was transformed into an Arabidopsis CaS knockout mutant (Salk) and overexpressed in the transgenic plants. OsCAS promoted stomatal closure. We screened homozygous transgenic Arabidopsis plants and determined physiological indices such as the oxidative damage biomarker malondialdehyde (MDA), relative membrane permeability (RMP), proline content, and chlorophyll fluorescence parameters, after 21 days of drought treatment. Our results revealed lower RMP and MDA contents and a higher Proline content in transgenic Arabidopsis plants after drought stress, whereas the opposite was observed in Salk plants. With respect to chlorophyll fluorescence, the electron transport rate and effective PSII quantum yield decreased in all lines under drought stress; however, in the transgenic plants these two parameters changed fewer and were higher than those in wild-type and Salk plants. The quantum yield of regulated energy dissipation and nonregulated energy dissipation in PSII were higher in Salk plants, whereas these values were lower in the transgenic plants than in the wild type under drought stress. The above results suggest that the transgenic plants showed better resistance to drought stress by decreasing damage to the cell membrane, increasing the amount of osmoprotectants, and maintaining a relatively high photosynthetic capacity. In conclusion, OsCAS is an extracellular calcium-sensing receptor

  10. New vectors for simple and streamlined CRISPR-Cas9 genome editing in Saccharomyces cerevisiae.

    PubMed

    Laughery, Marian F; Hunter, Tierra; Brown, Alexander; Hoopes, James; Ostbye, Travis; Shumaker, Taven; Wyrick, John J

    2015-12-01

    Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 technology is an important tool for genome editing because the Cas9 endonuclease can induce targeted DNA double-strand breaks. Targeting of the DNA break is typically controlled by a single-guide RNA (sgRNA), a chimeric RNA containing a structural segment important for Cas9 binding and a 20mer guide sequence that hybridizes to the genomic DNA target. Previous studies have demonstrated that CRISPR-Cas9 technology can be used for efficient, marker-free genome editing in Saccharomyces cerevisiae. However, introducing the 20mer guide sequence into yeast sgRNA expression vectors often requires cloning procedures that are complex, time-consuming and/or expensive. To simplify this process, we have developed a new sgRNA expression cassette with internal restriction enzyme sites that permit rapid, directional cloning of 20mer guide sequences. Here we describe a flexible set of vectors based on this design for cloning and expressing sgRNAs (and Cas9) in yeast using different selectable markers. We anticipate that the Cas9-sgRNA expression vector with the URA3 selectable marker (pML104) will be particularly useful for genome editing in yeast, since the Cas9 machinery can be easily removed by counter-selection using 5-fluoro-orotic acid (5-FOA) following successful genome editing. The availability of new vectors that simplify and streamline the technical steps required for guide sequence cloning should help accelerate the use of CRISPR-Cas9 technology in yeast genome editing.

  11. CRISPR-Cas9 for in vivo Gene Therapy: Promise and Hurdles

    PubMed Central

    Dai, Wei-Jing; Zhu, Li-Yao; Yan, Zhong-Yi; Xu, Yong; Wang, Qi-Long; Lu, Xiao-Jie

    2016-01-01

    Owing to its easy-to-use and multiplexing nature, the genome editing tool CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats (CRISPR) associated nuclease 9) is revolutionizing many areas of medical research and one of the most amazing areas is its gene therapy potentials. Previous explorations into the therapeutic potentials of CRISPR-Cas9 were mainly conducted in vitro or in animal germlines, the translatability of which, however, is either limited (to tissues with adult stem cells amenable to culture and manipulation) or currently impermissible (due to ethic concerns). Recently, important progresses have been made on this regard. Several studies have demonstrated the ability of CRISPR-Cas9 for in vivo gene therapy in adult rodent models of human genetic diseases delivered by methods that are potentially translatable to human use. Although these recent advances represent a significant step forward to the eventual application of CRISPR-Cas9 to the clinic, there are still many hurdles to overcome, such as the off-target effects of CRISPR-Cas9, efficacy of homology-directed repair, fitness of edited cells, immunogenicity of therapeutic CRISPR-Cas9 components, as well as efficiency, specificity, and translatability of in vivo delivery methods. In this article, we introduce the mechanisms and merits of CRISPR-Cas9 in genome editing, briefly retrospect the applications of CRISPR-Cas9 in gene therapy explorations and highlight recent advances, later we discuss in detail the challenges lying ahead in the way of its translatability, propose possible solutions, and future research directions.

  12. CRISPR-Cas Adaptive Immune Systems of the Sulfolobales: Unravelling Their Complexity and Diversity

    PubMed Central

    Garrett, Roger A.; Shah, Shiraz A.; Erdmann, Susanne; Liu, Guannan; Mousaei, Marzieh; León-Sobrino, Carlos; Peng, Wenfang; Gudbergsdottir, Soley; Deng, Ling; Vestergaard, Gisle; Peng, Xu; She, Qunxin

    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-Cas systems of bacteria. Despite this, there is evidence for a complex and differential regulation of expression of the diverse functional modules in response to viral infection. Recent work also supports critical roles for non-core Cas proteins, especially during Type III-directed interference, and this is consistent with these proteins tending to coevolve with core Cas proteins. Various novel aspects of CRISPR-Cas systems of the Sulfolobales are considered including an alternative spacer acquisition mechanism, reversible spacer acquisition, the formation and significance of antisense CRISPR RNAs, and a novel mechanism for avoidance of CRISPR-Cas defense. Finally, questions regarding the basis for the complexity, diversity, and apparent redundancy, of the intracellular CRISPR-Cas systems are discussed. PMID:25764276

  13. How specifically are action verbs represented in the neural motor system: an fMRI study.

    PubMed

    van Dam, Wessel O; Rueschemeyer, Shirley-Ann; Bekkering, Harold

    2010-12-01

    Embodied accounts of language processing suggest that sensorimotor areas, generally dedicated to perception and action, are also involved in the processing and representation of word meaning. Support for such accounts comes from studies showing that language about actions selectively modulates the execution of congruent and incongruent motor responses (e.g., Glenberg & Kaschak, 2002), and from functional neuroimaging studies showing that understanding action-related language recruits sensorimotor brain areas (e.g. Hauk, Johnsrude, & Pulvermueller, 2004). In the current experiment we explored the basis of the neural motor system's involvement in representing words denoting actions. Specifically, we investigated whether the motor system's involvement is modulated by the specificity of the kinematics associated with a word. Previous research in the visual domain indicates that words denoting basic level category members lacking a specific form (e.g., bird) are less richly encoded within visual areas than words denoting subordinate level members (e.g., pelican), for which the visual form is better specified (Gauthier, Anderson, Tarr, Skudlarski, & Gore, 1997). In the present study we extend these findings to the motor domain. Modulation of the BOLD response elicited by verbs denoting a general motor program (e.g., to clean) was compared to modulation elicited by verbs denoting a more specific motor program (e.g., to wipe). Conform with our hypothesis, a region within the bilateral inferior parietal lobule, typically serving the representation of action plans and goals, was sensitive to the specificity of motor programs associated with the action verbs. These findings contribute to the growing body of research on embodied language representations by showing that the concreteness of an action-semantic feature is reflected in the neural response to action verbs.

  14. CRISPR-Cas9: a new and promising player in gene therapy.

    PubMed

    Xiao-Jie, Lu; Hui-Ying, Xue; Zun-Ping, Ke; Jin-Lian, Chen; Li-Juan, Ji

    2015-05-01

    First introduced into mammalian organisms in 2013, the RNA-guided genome editing tool CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease 9) offers several advantages over conventional ones, such as simple-to-design, easy-to-use and multiplexing (capable of editing multiple genes simultaneously). Consequently, it has become a cost-effective and convenient tool for various genome editing purposes including gene therapy studies. In cell lines or animal models, CRISPR-Cas9 can be applied for therapeutic purposes in several ways. It can correct the causal mutations in monogenic disorders and thus rescue the disease phenotypes, which currently represents the most translatable field in CRISPR-Cas9-mediated gene therapy. CRISPR-Cas9 can also engineer pathogen genome such as HIV for therapeutic purposes, or induce protective or therapeutic mutations in host tissues. Moreover, CRISPR-Cas9 has shown potentials in cancer gene therapy such as deactivating oncogenic virus and inducing oncosuppressor expressions. Herein, we review the research on CRISPR-mediated gene therapy, discuss its advantages, limitations and possible solutions, and propose directions for future research, with an emphasis on the opportunities and challenges of CRISPR-Cas9 in cancer gene therapy.

  15. Translational studies of goal-directed action as a framework for classifying deficits across psychiatric disorders

    PubMed Central

    Griffiths, Kristi R.; Morris, Richard W.; Balleine, Bernard W.

    2014-01-01

    The ability to learn contingencies between actions and outcomes in a dynamic environment is critical for flexible, adaptive behavior. Goal-directed actions adapt to changes in action-outcome contingencies as well as to changes in the reward-value of the outcome. When networks involved in reward processing and contingency learning are maladaptive, this fundamental ability can be lost, with detrimental consequences for decision-making. Impaired decision-making is a core feature in a number of psychiatric disorders, ranging from depression to schizophrenia. The argument can be developed, therefore, that seemingly disparate symptoms across psychiatric disorders can be explained by dysfunction within common decision-making circuitry. From this perspective, gaining a better understanding of the neural processes involved in goal-directed action, will allow a comparison of deficits observed across traditional diagnostic boundaries within a unified theoretical framework. This review describes the key processes and neural circuits involved in goal-directed decision-making using evidence from animal studies and human neuroimaging. Select studies are discussed to outline what we currently know about causal judgments regarding actions and their consequences, action-related reward evaluation, and, most importantly, how these processes are integrated in goal-directed learning and performance. Finally, we look at how adaptive decision-making is impaired across a range of psychiatric disorders and how deepening our understanding of this circuitry may offer insights into phenotypes and more targeted interventions. PMID:24904322

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

    PubMed

    Nakanishi, Takashi; Kato, Yasuhiko; Matsuura, Tomoaki; Watanabe, Hajime

    2014-01-01

    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.

  17. AND-34/BCAR3 Regulates Adhesion-Dependent p130Cas Serine Phosphorylation and Breast Cancer Cell Growth Pattern

    PubMed Central

    Makkinje, Anthony; Near, Richard I.; Infusini, Giuseppe; Borre, Pierre Vanden; Bloom, Alexander; Cai, Dongpo; Costello, Catherine E.; Lerner, Adam

    2009-01-01

    NSP protein family members associate with p130Cas, a focal adhesion adapter protein best known as a Src substrate that integrates adhesion-related signaling. Over-expression of AND-34/BCAR3/NSP2 (BCAR3), but not NSP1 or NSP3, induces anti-estrogen resistance in human breast cancer cell lines. BCAR3 over-expression in epithelial MCF-7 cells augments levels of a phosphorylated p130Cas species that migrates more slowly on SDS PAGE while NSP-1 and NSP3 induce modest or no phosphorylation, respectively. Conversely, reduction in BCAR3 expression in mesenchymal MDA-231 cells by inducible shRNA results in loss of such p130Cas phosphorylation. Replacement of NSP3's serine/proline-rich domain with that of AND-34/BCAR3 instills the ability to induce p130Cas phosphorylation. Phospho-amino acid analysis demonstrates that BCAR3 induces p130Cas serine phosphorylation. Mass spectrometry identified phosphorylation at p130Cas serines 139, 437 and 639. p130Cas serine phosphorylation accumulates for several hours after adhesion of MDA-231 cells to fibronectin and is dependent upon BCAR3 expression. BCAR3 knockdown alters p130Cas localization and converts MDA-231 growth to an epithelioid pattern characterized by striking cohesiveness and lack of cellular projections at colony borders. These studies demonstrate that BCAR3 regulates p130Cas serine phosphorylation that is adhesion-dependent, temporally distinct from previously well-characterized rapid Fak and Src kinase-mediated p130Cas tyrosine phosphorylation and that correlates with invasive phenotype. PMID:19454314

  18. Transforming Language Ideologies through Action Research: A Case Study of Bilingual Science Learning

    NASA Astrophysics Data System (ADS)

    Yang, Eunah

    This qualitative case study explored a third grade bilingual teacher's transformative language ideologies through participating in a collaborative action research project. By merging language ideologies theory, Cultural Historical Activity Theory (CHAT), and action research, I was able to identify the analytic focus of this study. I analyzed how one teacher and I, the researcher, collaboratively reflected on classroom language practices during the video analysis meetings and focus groups. Further, I analyzed twelve videos that we coded together to see the changes in the teacher's language practices over time. My unit of analysis was the discourse practice mediated by additive language ideologies. Throughout the collaborative action research process, we both critically reflected on the classroom language use. We also developed a critical consciousness about the participatory shifts and learning of focal English Learner (EL) students. Finally, the teacher made changes to her classroom language practices. The results of this study will contribute to the literacy education research field for theoretical, methodological, and practical insights. The integration of language ideologies, CHAT, and action research can help educational practitioners, researchers, and policy makers understand the importance of transforming teachers' language ideologies in designing additive learning contexts for ELs. From a methodological perspective, the transformative language ideologies through researcher and teacher collaborated video analysis process provide a unique contribution to the language ideologies in education literature, with analytic triangulation. As a practical implication, this study suggests action research can be one of the teacher education tools to help the teachers transform language ideologies for EL education.

  19. The role of immediate and final goals in action planning: an fMRI study.

    PubMed

    Majdandzić, Jasminka; Grol, Meike J; van Schie, Hein T; Verhagen, Lennart; Toni, Ivan; Bekkering, Harold

    2007-08-15

    To interact effectively with our environment, we need to specify the intended outcomes (goals) of our actions. In this process, immediate goals and final goals can be regarded as different levels within a hierarchically organized system for action planning: immediate goals and movement details are selected to accomplish more remote goals. Behavioral studies support this notion of different levels of action planning, but the neurophysiological basis remains unclear. Using fMRI, we examined the neural correlates of preparing object manipulations based on either the desired end-state (the final goal) or the initial movement towards a target (the immediate goal). Subjects had to insert an object (consisting of a large and a small cube) into one of two corresponding large and small slots. The subjects were cued on either which slot to fill (Final Goal trials) or which object part to grasp (Immediate Goal trials). These actions required similar movements, but different planning. During Final Goal trials, there was differential preparatory activity along the superior frontal gyrus (bilaterally) and in left inferior parietal cortex. Immediate Goal trials evoked differential activity in occipito-parietal and occipito-temporal cortex. These findings support the notion that actions can be planned at different levels. We show that different fronto-parietal circuits plan the same action, by a relative emphasis on either selecting a sequence of movements to achieve a desired end-state, or selecting movements spatially compatible with given object properties.

  20. The different neural correlates of action and functional knowledge in semantic memory: an FMRI study.

    PubMed

    Canessa, Nicola; Borgo, Francesca; Cappa, Stefano F; Perani, Daniela; Falini, Andrea; Buccino, Giovanni; Tettamanti, Marco; Shallice, Tim

    2008-04-01

    Previous reports suggest that the internal organization of semantic memory is in terms of different "types of knowledge," including "sensory" (information about perceptual features), "action" (motor-based knowledge of object utilization), and "functional" (abstract properties, as function and context of use). Consistent with this view, a specific loss of action knowledge, with preserved functional knowledge, has been recently observed in patients with left frontoparietal lesions. The opposite pattern (impaired functional knowledge with preserved action knowledge) was reported in association with anterior inferotemporal lesions. In the present study, the cerebral representation of action and functional knowledge was investigated using event-related analysis of functional magnetic resonance imaging data. Fifteen subjects were presented with pictures showing pairs of manipulable objects and asked whether the objects within each pair were used with the same manipulation pattern ("action knowledge" condition) or in the same context ("functional knowledge" condition). Direct comparisons showed action knowledge, relative to functional knowledge, to activate a left frontoparietal network, comprising the intraparietal sulcus, the inferior parietal lobule, and the dorsal premotor cortex. The reverse comparison yielded activations in the retrosplenial and the lateral anterior inferotemporal cortex. These results confirm and extend previous neuropsychological data and support the hypothesis of the existence of different types of information processing in the internal organization of semantic memory.

  1. Memory and action: an experimental study on normal subjects and schizophrenic patients.

    PubMed

    Daprati, Elena; Nico, Daniele; Saimpont, Arnaud; Franck, Nicolas; Sirigu, Angela

    2005-01-01

    Psychologists have shown that recall of sentences describing previously performed actions is enhanced compared to recall of heard-only action-phrases (enactment effect). One interpretation of this effect argues that subjects benefit from a multi-modal encoding where movement plays a major role. In line with this motor account, it is conceivable that the beneficial effect of enactment might rely, at least in part, on procedural learning, thus tapping more directly implicit memory functions. Neuropsychological observations support this hypothesis, as shown by the fact that the enactment effect is quite insensitive to perturbations affecting declarative memories. i.e. Alzheimer disease. Memory for subject performed tasks in patients with Korsakoff syndrome. The present study attempts to evaluate whether pure motor activity is sufficient to guarantee the described memory facilitation or alternatively, whether first-person experience in carrying out the action (i.e. true enactment) would be required. To this purpose, in a first experiment on healthy subjects, we tested whether sentence meaning and content of the executed action should match in order to produce facilitation in recall of enacted action-phrases. In a second experiment, we explored whether the enactment effect is present in patients suffering from psychiatric disorders supposed to spare procedural memory but to alter action awareness (e.g. schizophrenia). We show that better recall for action phrases is found only when the motor component is a true enactment of verbal material. Moreover, this effect is nearly lost in schizophrenia. This latter result, on the one hand, queries the automatic/implicit nature of the enactment effect and supports the role of the experience of having performed the action in the first-person. On the other hand, it questions the nature of the memory impairments detected in schizophrenia.

  2. Corrective Action Plan for Corrective Action Unit 543: Liquid Disposal Units, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2007-04-01

    Corrective Action Unit (CAU) 543: Liquid Disposal Units is listed in Appendix III of the ''Federal Facility Agreement and Consent Order'' (FFACO) which was agreed to by the state of Nevada, the U.S. Department of Energy (DOE), and the U.S. Department of Defense (FFACO, 1996). CAU 543 sites are located in Areas 6 and 15 of the Nevada Test Site (NTS), which is approximately 65 miles northwest of Las Vegas, Nevada. CAU 543 consists of the following seven Corrective Action Sites (CASs) (Figure 1): CAS 06-07-01, Decon Pad; CAS 15-01-03, Aboveground Storage Tank; CAS 15-04-01, Septic Tank; CAS 15-05-01, Leachfield; CAS 15-08-01, Liquid Manure Tank; CAS 15-23-01, Underground Radioactive Material Area; and CAS 15-23-03, Contaminated Sump, Piping. All Area 15 CASs are located at the former U.S. Environmental Protection Agency (EPA) Farm, which operated from 1963 to 1981 and was used to support animal experiments involving the uptake of radionuclides. Each of the Area 15 CASs, except CAS 15-23-01, is associated with the disposal of waste effluent from Building 15-06, which was the primary location of the various tests and experiments conducted onsite. Waste effluent disposal from Building 15-06 involved piping, sumps, outfalls, a septic tank with leachfield, underground storage tanks, and an aboveground storage tank (AST). CAS 15-23-01 was associated with decontamination activities of farm equipment potentially contaminated with radiological constituents, pesticides, and herbicides. While the building structures were removed before the investigation took place, all the original tanks, sumps, piping, and concrete building pads remain in place. The Area 6 CAS is located at the Decontamination Facility in Area 6, a facility which operated from 1971 to 2001 and was used to decontaminate vehicles, equipment, clothing, and other materials that had become contaminated during nuclear testing activities. The CAS includes the effluent collection and distribution systems for Buildings

  3. Diversity of CRISPR-Cas-Mediated Mechanisms of Adaptive Immunity in Prokaryotes and Their Application in Biotechnology.

    PubMed

    Savitskaya, E E; Musharova, O S; Severinov, K V

    2016-07-01

    CRISPR-Cas systems of adaptive immunity in prokaryotes consist of CRISPR arrays (clusters of short repeated genomic DNA fragments separated by unique spacer sequences) and cas (CRISPR-associated) genes that provide cells with resistance against bacteriophages and plasmids containing protospacers, i.e. sequences complementary to CRISPR array spacers. CRISPR-Cas systems are responsible for two different cellular phenomena: CRISPR adaptation and CRISPR interference. CRISPR adaptation is cell genome modification by integration of new spacers that represents a unique case of Lamarckian inheritance. CRISPR interference involves specific recognition of protospacers in foreign DNA followed by introduction of breaks into this DNA and its destruction. According to the mechanisms of action, CRISPR-Cas systems have been subdivided into two classes, five types, and numerous subtypes. The development of techniques based on CRISPR interference mediated by the Type II system Cas9 protein has revolutionized the field of genome editing because it allows selective, efficient, and relatively simple introduction of directed breaks into target DNA loci. However, practical applications of CRISPR-Cas systems are not limited only to genome editing. In this review, we focus on the variety of CRISPR interference and CRISPR adaptation mechanisms and their prospective use in biotechnology. PMID:27449612

  4. [Synthesis of new mandelic acid derivatives with preservative action. Synthesis and acute toxicity study].

    PubMed

    Stan, Cătălina; Năstase, V; Pavelescu, M; Vasile, Cornelia; Dumitrache, M; Gherase, Florenţa; Năstasă, Veronica

    2004-01-01

    Starting from the antiseptic action of DL mandelic acid, there were synthesized a series of esters of the mandelic acid, esters which could have preservative action. This study present the synthesis, structure validation and the acute toxicity study, for the new synthesized compounds. The esters were obtained by acylating 4-hydroxybenzoic acid propyl, ethyl, methyl esters and salicylic acid with the DL mandelic chloride (that was protected initially by the hydroxylic group). The structure of the synthesized compounds was confirmed by quantitative elemental analysis and RMN 1H spectral measurements. The acute toxicity was determined for two of the esters, who proved to had a preservative action (previously studied) and indicated that these esters have a small toxicity.

  5. Manipulating the Biosynthesis of Bioactive Compound Alkaloids for Next-Generation Metabolic Engineering in Opium Poppy Using CRISPR-Cas 9 Genome Editing Technology.

    PubMed

    Alagoz, Yagiz; Gurkok, Tugba; Zhang, Baohong; Unver, Turgay

    2016-01-01

    Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated9 (Cas9) endonuclease system is a powerful RNA-guided genome editing tool. CRISPR/Cas9 has been well studied in model plant species for targeted genome editing. However, few studies have been reported on plant species without whole genome sequence information. Currently, no study has been performed to manipulate metabolic pathways using CRISPR/Cas9. In this study, the type II CRISPR/SpCas9 system was used to knock out, via nonhomologous end-joining genome repair, the 4'OMT2 in opium poppy (Papaver somniferum L.), a gene which regulates the biosythesis of benzylisoquinoline alkaloids (BIAs). For sgRNA transcription, viral-based TRV and synthetic binary plasmids were designed and delivered into plant cells with a Cas9 encoding-synthetic vector by Agrobacterium-mediated transformation. InDels formed by CRISPR/Cas9 were detected by sequence analysis. Our results showed that the biosynthesis of BIAs (e.g. morphine, thebaine) was significantly reduced in the transgenic plants suggesting that 4'OMT2 was efficiently knocked-out by our CRISPR-Cas9 genome editing approach. In addition, a novel uncharacterized alkaloid was observed only in CRISPR/Cas9 edited plants. Thus, the applicabilitiy of the CRISPR/Cas9 system was demonstrated for the first time for medicinal aromatic plants by sgRNAs transcribed from both synthetic and viral vectors to regulate BIA metabolism and biosynthesis. PMID:27483984

  6. Manipulating the Biosynthesis of Bioactive Compound Alkaloids for Next-Generation Metabolic Engineering in Opium Poppy Using CRISPR-Cas 9 Genome Editing Technology.

    PubMed

    Alagoz, Yagiz; Gurkok, Tugba; Zhang, Baohong; Unver, Turgay

    2016-08-03

    Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated9 (Cas9) endonuclease system is a powerful RNA-guided genome editing tool. CRISPR/Cas9 has been well studied in model plant species for targeted genome editing. However, few studies have been reported on plant species without whole genome sequence information. Currently, no study has been performed to manipulate metabolic pathways using CRISPR/Cas9. In this study, the type II CRISPR/SpCas9 system was used to knock out, via nonhomologous end-joining genome repair, the 4'OMT2 in opium poppy (Papaver somniferum L.), a gene which regulates the biosythesis of benzylisoquinoline alkaloids (BIAs). For sgRNA transcription, viral-based TRV and synthetic binary plasmids were designed and delivered into plant cells with a Cas9 encoding-synthetic vector by Agrobacterium-mediated transformation. InDels formed by CRISPR/Cas9 were detected by sequence analysis. Our results showed that the biosynthesis of BIAs (e.g. morphine, thebaine) was significantly reduced in the transgenic plants suggesting that 4'OMT2 was efficiently knocked-out by our CRISPR-Cas9 genome editing approach. In addition, a novel uncharacterized alkaloid was observed only in CRISPR/Cas9 edited plants. Thus, the applicabilitiy of the CRISPR/Cas9 system was demonstrated for the first time for medicinal aromatic plants by sgRNAs transcribed from both synthetic and viral vectors to regulate BIA metabolism and biosynthesis.

  7. Significant Life Experiences Affect Environmental Action: A Confirmation Study in Eastern Taiwan

    ERIC Educational Resources Information Center

    Hsu, Shih-Jang

    2009-01-01

    Two field studies form the basis of this article. The major purposes of Study 1 were to examine significant life experiences affecting the cultivation of environmental activists in eastern Taiwan, and to reconstruct the life paths followed by those active people who engaged in effective environmental action. 40 usable autobiographical memories…

  8. Changing Preschool Children's Attitudes into Behavior towards Selected Environmental Issues: An Action Research Study

    ERIC Educational Resources Information Center

    Ertürk Kara, Gözde; Aydos, E. Hande; Aydin, Özge

    2015-01-01

    The purpose of this study is to provide the transform of attitudes into behavior of 60-72 month of age children continued early childhood education toward environmental issues. Collaborative action research method of qualitative design was used. The whole participants of the study were 60-72 months of age children who were attending in an early…

  9. A Study of the Qualities Teachers Recommend in STS Issues Investigation and Action Instructional Materials.

    ERIC Educational Resources Information Center

    Rubba, Peter A.; Wiesenmayer, Randall L.

    This study was part of a project to develop science and technology (STS) issue investigation and action materials with the potential for use across science and social studies classes at the secondary level. Special emphasis was given to learning teachers' perspectives on the development of a unit on the impact of pesticides on society. Seven areas…

  10. Understanding the Impact of Affirmative Action Bans in Different Graduate Fields of Study

    ERIC Educational Resources Information Center

    Garces, Liliana M.

    2013-01-01

    This study examines the effects of affirmative action bans in four states (California, Florida, Texas, and Washington) on the enrollment of underrepresented students of color within six different graduate fields of study: the natural sciences, engineering, social sciences, business, education, and humanities. Findings show that affirmative action…

  11. Reconnecting with Your Passion: An Action Research Study Exploring Humanities and Professional Nursing

    ERIC Educational Resources Information Center

    Snyder, Melissa J.

    2012-01-01

    The purpose of this action research study was two-fold. The first purpose was to examine the process of how nurses engaged in a professional development program that drew upon reading and creative writing related to their lives and work as nurses. Secondly, this study examined the nurses' perspectives on how their involvement in the process…

  12. Reconstructing a Community, Reclaiming a Playground: A Participatory Action Research Study

    ERIC Educational Resources Information Center

    Hutzel, Karen

    2007-01-01

    This article describes a participatory action research study that examined participant's perceptions of community and of the West End neighborhood in Cincinnati, Ohio, where the study took place. It is argued that oppressive situations have developed strong collective identities and social capital among residents, which can lead to the development…

  13. Cancer, Employment, and American Indians: A Participatory Action Research Pilot Study

    ERIC Educational Resources Information Center

    Johnson, Sharon R.; Finifrock, DeAnna; Marshall, Catherine A.; Jaakola, Julia; Setterquist, Janette; Burross, Heidi L.; Hodge, Felicia Schanche

    2011-01-01

    American Indian cancer survivors are an underserved and understudied group. In this pilot study we attempted to address, through participatory action research, missing information about those factors that serve to either facilitate employment or hinder it for adult cancer survivors. One task of the study was to develop and/or modify…

  14. 48 CFR 970.3002 - CAS program requirements.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 48 Federal Acquisition Regulations System 5 2012-10-01 2012-10-01 false CAS program requirements. 970.3002 Section 970.3002 Federal Acquisition Regulations System DEPARTMENT OF ENERGY AGENCY....3002 CAS program requirements....

  15. 48 CFR 970.3002 - CAS program requirements.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 48 Federal Acquisition Regulations System 5 2013-10-01 2013-10-01 false CAS program requirements. 970.3002 Section 970.3002 Federal Acquisition Regulations System DEPARTMENT OF ENERGY AGENCY....3002 CAS program requirements....

  16. 48 CFR 970.3002 - CAS program requirements.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 48 Federal Acquisition Regulations System 5 2014-10-01 2014-10-01 false CAS program requirements. 970.3002 Section 970.3002 Federal Acquisition Regulations System DEPARTMENT OF ENERGY AGENCY....3002 CAS program requirements....

  17. Closure Report for Corrective Action Unit 357: Mud Pits and Waste Dump, Nevada Test Site, Nevada, Rev. No.: 0

    SciTech Connect

    Laura A. Pastor

    2005-04-01

    This Closure Report (CR) presents information supporting closure of Corrective Action Unit (CAU) 357: Mud Pits and Waste Dump, Nevada Test Site (NTS), Nevada. The CR complies with the requirements of the Federal Facility Agreement and Consent Order (FFACO) that was agreed to by the State of Nevada, U.S. Department of Energy (DOE), and the U.S. Department of Defense (FFACO, 1996). Corrective Action Unit 357 is comprised of 14 Corrective Action Sites (CASs) located in Areas 1, 4, 7, 8, 10, and 25 of the NTS (Figure 1-1). The NTS is located approximately 65 miles (mi) northwest of Las Vegas, Nevada. Corrective Action Unit 357 consists of 11 CASs that are mud pits located in Areas 7, 8, and 10. The mud pits were associated with drilling activities conducted on the NTS in support of the underground nuclear weapons testing. The remaining three CASs are boxes and pipes associated with Building 1-31.2el, lead bricks, and a waste dump. These CAS are located in Areas 1, 4, and 25, respectively. The following CASs are shown on Figure 1-1: CAS 07-09-02, Mud Pit; CAS 07-09-03, Mud Pit; CAS 07-09-04, Mud Pit; CAS 07-09-05, Mud Pit; CAS 08-09-01, Mud Pit; CAS 08-09-02, Mud Pit; CAS 08-09-03, Mud Pit; CAS 10-09-02, Mud Pit; CAS 10-09-04, Mud Pit; CAS 10-09-05, Mud Pit; CAS 10-09-06, Mud Pit, Stains, Material; CAS 01-99-01, Boxes, Pipes; CAS 04-26-03, Lead Bricks; and CAS 25-15-01, Waste Dump. The purpose of the corrective action activities was to obtain analytical data that supports the closure of CAU 357. Environmental samples were collected during the investigation to determine whether contaminants exist and if detected, their extent. The investigation and sampling strategy was designed to target locations and media most likely to be contaminated (biased sampling). A general site conceptual model was developed for each CAS to support and guide the investigation as outlined in the Streamlined Approach for Environmental Restoration (SAFER) Plan (NNSA/NSO, 2003b). This CR

  18. A Participatory Action Research Study of Nature Education in Nature: Towards Community-Based Eco-Pedagogy

    ERIC Educational Resources Information Center

    Eryaman, Mustafa Yunus; Yalcin-Ozdilek, Sukran; Okur, Emel; Cetinkaya, Zeynep; Uygun, Selcuk

    2010-01-01

    Contemporary nature education is exploring different ways to develop awareness for change and initiate action. Such educational activities go beyond creating understanding and awareness in order to develop a sense of commitment for individual and collective action. This participatory action research study aimed to improve teachers' sensitiveness…

  19. Engineered CRISPR-Cas9 nucleases with altered PAM specificities

    PubMed Central

    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

  20. Engineered CRISPR-Cas9 nucleases with altered PAM specificities.

    PubMed

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

    2015-07-23

    Although CRISPR-Cas9 nucleases are widely used for genome editing, the range of sequences that Cas9 can recognize is constrained by the need for a specific protospacer adjacent motif (PAM). 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 analysis. In addition, we identify and characterize 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 find 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.

  1. CRISPR-Cas immunity in prokaryotes.

    PubMed

    Marraffini, Luciano A

    2015-10-01

    Prokaryotic organisms are threatened by a large array of viruses and have developed numerous defence strategies. Among these, only clustered, regularly interspaced short palindromic repeat (CRISPR)-Cas systems provide adaptive immunity against foreign elements. Upon viral injection, a small sequence of the viral genome, known as a spacer, is integrated into the CRISPR locus to immunize the host cell. Spacers are transcribed into small RNA guides that direct the cleavage of the viral DNA by Cas nucleases. Immunization through spacer acquisition enables a unique form of evolution whereby a population not only rapidly acquires resistance to its predators but also passes this resistance mechanism vertically to its progeny.

  2. CRISPR-Cas: Revolutionising genome engineering.

    PubMed

    Nicholson, Samantha Anne; Pepper, Michael Sean

    2016-09-01

    The ability to permanently alter or repair the human genome has been the subject of a number of science fiction films, but with the recent advent of several customisable sequence-specific endonuclease technologies, genome engineering looks set to become a clinical reality in the near future. This article discusses recent advancements in the technology called 'clustered regularly interspaced palindromic repeat (CRISPR)-associated genes' (CRISPR-Cas), the potential of CRISPR-Cas to revolutionise molecular medicine, and the ethical and regulatory hurdles facing its application. PMID:27601107

  3. CRISPR-Cas: Revolutionising genome engineering.

    PubMed

    Nicholson, Samantha Anne; Pepper, Michael Sean

    2016-08-01

    The ability to permanently alter or repair the human genome has been the subject of a number of science fiction films, but with the recent advent of several customisable sequence-specific endonuclease technologies, genome engineering looks set to become a clinical reality in the near future. This article discusses recent advancements in the technology called 'clustered regularly interspaced palindromic repeat (CRISPR)-associated genes' (CRISPR-Cas), the potential of CRISPR-Cas to revolutionise molecular medicine, and the ethical and regulatory hurdles facing its application.

  4. Use of CAS in secondary school: a factor influencing the transition to university-level mathematics?

    NASA Astrophysics Data System (ADS)

    Varsavsky, Cristina

    2012-01-01

    Australian secondary school systems offer three levels of senior (year 12) mathematics studies, none of them compulsory: elementary, intermediate and advanced. The intermediate and advanced studies prepare students for further mathematics studies at university level. In the state of Victoria, there are two versions of intermediate mathematics: one where students learn and are examined with a computer algebra system (CAS) and another where students can only use scientific calculators. This study compares the performance of 1240 students as they transitioned to traditional university-level mathematics and according to whether they learned intermediate mathematics with or without the assistance of a CAS. This study concludes that students without CAS show a slight advantage, but the most important factor affecting student performance is the uptake of advanced-level mathematics studies in secondary school.

  5. Corrective Action Investigation Plan for Corrective Action Unit 555: Septic Systems Nevada Test Site, Nevada, Rev. No.: 0 with Errata

    SciTech Connect

    Pastor, Laura

    2005-12-01

    This Corrective Action Investigation Plan (CAIP) contains project-specific information including facility descriptions, environmental sample collection objectives, and criteria for conducting site investigation activities at Corrective Action Unit (CAU) 555: Septic Systems, Nevada Test Site (NTS), Nevada. This CAIP has been developed in accordance with the ''Federal Facility Agreement and Consent Order'' (FFACO) (1996) that was agreed to by the State of Nevada, the U.S. Department of Energy (DOE), and the U.S. Department of Defense. Corrective Action Unit 555 is located in Areas 1, 3 and 6 of the NTS, which is approximately 65 miles (mi) northwest of Las Vegas, Nevada, and is comprised of the five corrective action sites (CASs) shown on Figure 1-1 and listed below: (1) CAS 01-59-01, Area 1 Camp Septic System; (2) CAS 03-59-03, Core Handling Building Septic System; (3) CAS 06-20-05, Birdwell Dry Well; (4) CAS 06-59-01, Birdwell Septic System; and (5) CAS 06-59-02, National Cementers Septic System. An FFACO modification was approved on December 14, 2005, to include CAS 06-20-05, Birdwell Dry Well, as part of the scope of CAU 555. The work scope was expanded in this document to include the investigation of CAS 06-20-05. The Corrective Action Investigation (CAI) will include field inspections, radiological surveys, geophysical surveys, sampling of environmental media, analysis of samples, and assessment of investigation results, where appropriate. Data will be obtained to support corrective action alternative evaluations and waste management decisions. The CASs in CAU 555 are being investigated because hazardous and/or radioactive constituents may be present in concentrations that could potentially pose a threat to human health and the environment. Existing information on the nature and extent of potential contamination is insufficient to evaluate and recommend corrective action alternatives for the CASs. Additional information will be generated by conducting a CAI

  6. 48 CFR 30.201-1 - CAS applicability.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-1 CAS applicability. See 48 CFR 9903.201-1 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2012-10-01 2012-10-01 false CAS applicability....

  7. 48 CFR 30.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-2 Types of CAS coverage. See 48 CFR 9903.201-2 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2014-10-01 2014-10-01 false Types of CAS coverage....

  8. 48 CFR 30.201-1 - CAS applicability.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-1 CAS applicability. See 48 CFR 9903.201-1 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2013-10-01 2013-10-01 false CAS applicability....

  9. 48 CFR 30.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-2 Types of CAS coverage. See 48 CFR 9903.201-2 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2012-10-01 2012-10-01 false Types of CAS coverage....

  10. 48 CFR 9903.201-1 - CAS applicability.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 48 Federal Acquisition Regulations System 7 2012-10-01 2012-10-01 false CAS applicability. 9903... ACCOUNTING STANDARDS CONTRACT COVERAGE CAS Program Requirements 9903.201-1 CAS applicability. (a) This subsection describes the rules for determining whether a proposed contract or subcontract is exempt from...

  11. 48 CFR 30.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-2 Types of CAS coverage. See 48 CFR 9903.201-2 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2013-10-01 2013-10-01 false Types of CAS coverage....

  12. 48 CFR 9903.201-1 - CAS applicability.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 48 Federal Acquisition Regulations System 7 2010-10-01 2010-10-01 false CAS applicability. 9903... ACCOUNTING STANDARDS CONTRACT COVERAGE CAS Program Requirements 9903.201-1 CAS applicability. (a) This subsection describes the rules for determining whether a proposed contract or subcontract is exempt from...

  13. 48 CFR 30.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-2 Types of CAS coverage. See 48 CFR 9903.201-2 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2011-10-01 2011-10-01 false Types of CAS coverage....

  14. 48 CFR 30.201-1 - CAS applicability.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-1 CAS applicability. See 48 CFR 9903.201-1 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2011-10-01 2011-10-01 false CAS applicability....

  15. 48 CFR 9903.201-1 - CAS applicability.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 48 Federal Acquisition Regulations System 7 2014-10-01 2014-10-01 false CAS applicability. 9903... ACCOUNTING STANDARDS CONTRACT COVERAGE CAS Program Requirements 9903.201-1 CAS applicability. (a) This subsection describes the rules for determining whether a proposed contract or subcontract is exempt from...

  16. 48 CFR 30.201-1 - CAS applicability.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-1 CAS applicability. See 48 CFR 9903.201-1 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2014-10-01 2014-10-01 false CAS applicability....

  17. 48 CFR 9903.201-1 - CAS applicability.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 48 Federal Acquisition Regulations System 7 2013-10-01 2012-10-01 true CAS applicability. 9903.201... ACCOUNTING STANDARDS CONTRACT COVERAGE CAS Program Requirements 9903.201-1 CAS applicability. (a) This subsection describes the rules for determining whether a proposed contract or subcontract is exempt from...

  18. 48 CFR 30.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-2 Types of CAS coverage. See 48 CFR 9903.201-2 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Types of CAS coverage....

  19. 48 CFR 30.201-1 - CAS applicability.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... CONTRACTING REQUIREMENTS COST ACCOUNTING STANDARDS ADMINISTRATION CAS Program Requirements 30.201-1 CAS applicability. See 48 CFR 9903.201-1 (FAR appendix). ... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false CAS applicability....

  20. A Kinase-Independent Function of c-Src Mediates p130Cas Phosphorylation at the Serine-639 Site in Pressure Overloaded Myocardium.

    PubMed

    Palanisamy, Arun P; Suryakumar, Geetha; Panneerselvam, Kavin; Willey, Christopher D; Kuppuswamy, Dhandapani

    2015-12-01

    Early work in pressure overloaded (PO) myocardium shows that integrins mediate focal adhesion complex formation by recruiting the adaptor protein p130Cas (Cas) and nonreceptor tyrosine kinase c-Src. To explore c-Src role in Cas-associated changes during PO, we used a feline right ventricular in vivo PO model and a three-dimensional (3D) collagen-embedded adult cardiomyocyte in vitro model that utilizes a Gly-Arg-Gly-Asp-Ser (RGD) peptide for integrin stimulation. Cas showed slow electrophoretic mobility (band-shifting), recruitment to the cytoskeleton, and tyrosine phosphorylation at 165, 249, and 410 sites in both 48 h PO myocardium and 1 h RGD-stimulated cardiomyocytes. Adenoviral mediated expression of kinase inactive (negative) c-Src mutant with intact scaffold domains (KN-Src) in cardiomyocytes did not block the RGD stimulated changes in Cas. Furthermore, expression of KN-Src or kinase active c-Src mutant with intact scaffold function (A-Src) in two-dimensionally (2D) cultured cardiomyocytes was sufficient to cause Cas band-shifting, although tyrosine phosphorylation required A-Src. These data indicate that c-Src's adaptor function, but not its kinase function, is required for a serine/threonine specific phosphorylation(s) responsible for Cas band-shifting. To explore this possibility, Chinese hamster ovary cells that stably express Cas were infected with either β-gal or KN-Src adenoviruses and used for Cas immunoprecipitation combined with mass spectrometry analysis. In the KN-Src expressing cells, Cas showed phosphorylation at the serine-639 (human numbering) site. A polyclonal antibody raised against phospho-serine-639 detected Cas phosphorylation in 24-48 h PO myocardium. Our studies indicate that c-Src's adaptor function mediates serine-639 phosphorylation of Cas during integrin activation in PO myocardium.

  1. The Environmental Action and Philosophy Matrix: An Exploratory Study of the Environmental Attitudes of Recreation Management and Environmental Studies Students

    ERIC Educational Resources Information Center

    Schultz, Jeremy R.; Simpson, Steven; Elfessi, Abdulaziz M.

    2011-01-01

    This study is a comparative analysis of the environmental philosophies of college undergraduates enrolled in a Midwestern university. Two courses were used for the research, one from a recreation management curriculum and the other from environmental studies. The study utilized a survey instrument called the Environmental Action and Philosophy…

  2. Bridging the gap between knowledge and action for health: Case studies.

    PubMed Central

    Thamlikitkul, Visanu

    2006-01-01

    Biomedical discoveries could improve people's health only if they are suited to the diverse political and social contexts, health systems and population groups. Knowledge generated through evidence-informed health policy and practice when applied to the local situation enhances the quality and efficiency of health care. This article describes four case studies on bridging the gap between knowledge and action for health in a tertiary care hospital in Bangkok, Thailand. Gaps between knowledge and action for health are classified into "know-do" and "do-know" gaps with knowledge implementation and knowledge generation being the key measures for bridging the gap. PMID:16917646

  3. DFT/B3LYP study of tocopherols and chromans antioxidant action energetics

    NASA Astrophysics Data System (ADS)

    Klein, Erik; Lukeš, Vladimír; Ilčin, Michal

    2007-07-01

    Gas-phase reaction enthalpies related to the individual steps of three phenolic antioxidants action mechanisms - hydrogen atom transfer (HAT), single-electron transfer-proton transfer (SET-PT) and sequential proton loss electron transfer (SPLET) for four tocopherols and seven chromans - were calculated using DFT/B3LYP method. For α-tocopherol, one of the chromans and phenol, reaction enthalpies in water were computed. In comparison to gas phase, water causes severe changes in the energetics of studied compounds antioxidant action. From the thermodynamic point of view, entering SPLET mechanism represents the most probable process in water.

  4. Regulators of Androgen Action Resource: a one-stop shop for the comprehensive study of androgen receptor action.

    PubMed

    DePriest, Adam D; Fiandalo, Michael V; Schlanger, Simon; Heemers, Frederike; Mohler, James L; Liu, Song; Heemers, Hannelore V

    2016-01-01

    Androgen receptor (AR) is a ligand-activated transcription factor that is the main target for treatment of non-organ-confined prostate cancer (CaP). Failure of life-prolonging AR-targeting androgen deprivation therapy is due to flexibility in steroidogenic pathways that control intracrine androgen levels and variability in the AR transcriptional output. Androgen biosynthesis enzymes, androgen transporters and AR-associated coregulators are attractive novel CaP treatment targets. These proteins, however, are characterized by multiple transcript variants and isoforms, are subject to genomic alterations, and are differentially expressed among CaPs. Determining their therapeutic potential requires evaluation of extensive, diverse datasets that are dispersed over multiple databases, websites and literature reports. Mining and integrating these datasets are cumbersome, time-consuming tasks and provide only snapshots of relevant information. To overcome this impediment to effective, efficient study of AR and potential drug targets, we developed the Regulators of Androgen Action Resource (RAAR), a non-redundant, curated and user-friendly searchable web interface. RAAR centralizes information on gene function, clinical relevance, and resources for 55 genes that encode proteins involved in biosynthesis, metabolism and transport of androgens and for 274 AR-associated coregulator genes. Data in RAAR are organized in two levels: (i) Information pertaining to production of androgens is contained in a 'pre-receptor level' database, and coregulator gene information is provided in a 'post-receptor level' database, and (ii) an 'other resources' database contains links to additional databases that are complementary to and useful to pursue further the information provided in RAAR. For each of its 329 entries, RAAR provides access to more than 20 well-curated publicly available databases, and thus, access to thousands of data points. Hyperlinks provide direct access to gene

  5. Regulators of Androgen Action Resource: a one-stop shop for the comprehensive study of androgen receptor action

    PubMed Central

    Fiandalo, Michael V.; Schlanger, Simon; Heemers, Frederike; Mohler, James L.; Liu, Song; Heemers, Hannelore V.

    2016-01-01

    Androgen receptor (AR) is a ligand-activated transcription factor that is the main target for treatment of non-organ-confined prostate cancer (CaP). Failure of life-prolonging AR-targeting androgen deprivation therapy is due to flexibility in steroidogenic pathways that control intracrine androgen levels and variability in the AR transcriptional output. Androgen biosynthesis enzymes, androgen transporters and AR-associated coregulators are attractive novel CaP treatment targets. These proteins, however, are characterized by multiple transcript variants and isoforms, are subject to genomic alterations, and are differentially expressed among CaPs. Determining their therapeutic potential requires evaluation of extensive, diverse datasets that are dispersed over multiple databases, websites and literature reports. Mining and integrating these datasets are cumbersome, time-consuming tasks and provide only snapshots of relevant information. To overcome this impediment to effective, efficient study of AR and potential drug targets, we developed the Regulators of Androgen Action Resource (RAAR), a non-redundant, curated and user-friendly searchable web interface. RAAR centralizes information on gene function, clinical relevance, and resources for 55 genes that encode proteins involved in biosynthesis, metabolism and transport of androgens and for 274 AR-associated coregulator genes. Data in RAAR are organized in two levels: (i) Information pertaining to production of androgens is contained in a ‘pre-receptor level’ database, and coregulator gene information is provided in a ‘post-receptor level’ database, and (ii) an ‘other resources’ database contains links to additional databases that are complementary to and useful to pursue further the information provided in RAAR. For each of its 329 entries, RAAR provides access to more than 20 well-curated publicly available databases, and thus, access to thousands of data points. Hyperlinks provide direct access to

  6. We Did It Together: A Participatory Action Research Study on Poverty and Disability

    ERIC Educational Resources Information Center

    Buettgen, Alexis; Richardson, Jason; Beckham, Kristie; Richardson, Kathy; Ward, Michelle; Riemer, Manuel

    2012-01-01

    This article presents the perspective of both non-disabled and developmentally disabled people working together in a research project on poverty and disability. Our study used a participatory action research approach that challenges the norm of exclusion in the research process. Control of the research agenda has been inclusive and shared to…

  7. An Action Research Study: Using Classroom Guidance Lessons to Teach Middle School Students about Sexual Harassment

    ERIC Educational Resources Information Center

    Bates, Rebecca C.

    2006-01-01

    This article describes a three-part classroom guidance lesson that teaches middle school students the definition of sexual harassment, the difference between flirting and sexual harassment, and the harmful effects of sexual harassment. An action research study evaluated the effectiveness of the lessons in decreasing referrals for sexual harassment…

  8. Environmental Education in Action - III: Case Studies of Public Involvement in Environmental Policy.

    ERIC Educational Resources Information Center

    Schoenfeld, Clay, Comp.; Desinger, John F., Comp.

    Presented here are 27 case studies of public involvement in environmental policy. These are examples of environmental education or communications programs developed by local, state, regional or national environmental action groups. The reports tell how the groups have successfully, or unsuccessfully, mobilized public opinion in favor of beneficent…

  9. Prospects for the Future: The Use of Participatory Action Research to Study Educational Disadvantage

    ERIC Educational Resources Information Center

    James, Emily Alana

    2005-01-01

    Of all the varieties of educational disadvantage, issues involving students who are homeless, or who move frequently because of poverty, are perhaps some of the most difficult for public school educators in the US to address. Using a pragmatic mixed methods design, this study evaluates the efficacy of participatory action research (PAR): (1) as a…

  10. In-Service Teacher Training and Coaching on Marzano's Instructional Strategies: An Action Research Study

    ERIC Educational Resources Information Center

    Miller, Shenequa C.

    2014-01-01

    The focus of this action research study was the implementation of a professional development initiative comprised of two phases: a training program for teachers on Marzano's nine research-based instructional strategies, and the implementation, supported by follow-up coaching, during "Pear Mountain" High School's (a pseudonym) six-week…

  11. How Trainee Music Teachers Learn about Teaching by Talking to Each Other: An Action Research Study

    ERIC Educational Resources Information Center

    Cain, Tim

    2011-01-01

    This article presents an action research study into how trainee music teachers in England use a structured discussion process called "Collegial Consultation" to learn about teaching. The research shows that, in Collegial Consultation, trainees learn from each other by offering several solutions to a problem, offering reasons for their ideas,…

  12. An Action Research Study on the Effect of Interactive Technology and Active Learning on Student Performance

    ERIC Educational Resources Information Center

    Bear, Teresa J.

    2013-01-01

    This quantitative action science research study utilized a causal-comparative experimental research design in order to determine if the use of student response systems (clickers), as an active learning strategy in a community college course, improved student performance in the course. Students in the experimental group (n = 26) used clickers to…

  13. Learning How to Manage Bias: A Case Study of Youth Participatory Action Research

    ERIC Educational Resources Information Center

    Kirshner, Ben; Pozzoboni, Kristen; Jones, Hannah

    2011-01-01

    Youth programs that are organized around intellectually challenging, socially relevant projects create opportunities for deep cognitive engagement. One type of authentic project that deserves attention from applied developmental scientists is youth participatory action research (YPAR), in which participants study a problem relevant to young…

  14. Exploring Student Experiences of Belonging within an Urban High School Choral Ensemble: An Action Research Study

    ERIC Educational Resources Information Center

    Parker, Elizabeth Cassidy

    2010-01-01

    The purpose of this action research study was to describe adolescent singers' experiences of belonging within one urban high school choral ensemble. Understanding student perspectives on belonging within music ensembles can assist choral educators, parents and administrators in order to more fully support adolescent emotional and social…

  15. Authentic Tasks: A Participatory Action Research Study on a Teaching Method for the Inclusive Classroom

    ERIC Educational Resources Information Center

    Bruffy, Wendy Raso

    2012-01-01

    The way in which education is taught is forever changing. Therefore, a new method in which content is delivered is a central component in examining areas for improvement. In this paper, participatory action research (PAR) was used to investigate authentic tasks in the inclusive tenth and eleventh grade classroom. The purpose of this study was to…

  16. Business, Community, Schools, and University Perceptions of the Partnership Process: An Action Research Study.

    ERIC Educational Resources Information Center

    Borthwick, Arlene G.; And Others

    Findings from an action research study that sought to understand the partnership process from the perspectives of persons involved in a partnership project, the Cooperative Alliance for Gifted Education (CAGE), are presented in this paper. The CAGE project utilized three partners in Ohio--Kent State University, the Cleveland Public Schools, and…

  17. Characterizing Teaching Effectiveness in the Joint Action Theory in Didactics: An Exploratory Study in Primary School

    ERIC Educational Resources Information Center

    Sensevy, Gérard

    2014-01-01

    This paper presents an exploratory study of two consecutive reading sessions conducted in primary school by two different teachers. Our purpose is twofold. From a theoretical viewpoint, we propose a tentative set of conditions of teaching effectiveness by relying on the Joint Action Theory in Didactics. From a methodological viewpoint, drawing on…

  18. Joint Action in Didactics and Classroom Ecology: Comparing Theories Using a Case Study in Physical Education

    ERIC Educational Resources Information Center

    Amade-Escot, Chantal; Venturini, Patrice

    2015-01-01

    This paper examines the respective contribution of two theoretical approaches to teaching and learning: the classroom ecology (CE) framework from Anglo-American research and the joint action in didactics (JAD) framework, which is part of French "didactique" research. This theoretical comparison is grounded in data from a case study in a…

  19. Developing Pedagogical Content Knowledge of Science Teachers through Action Research: A Case Study from Pakistan

    ERIC Educational Resources Information Center

    Halai, Nelofer; Khan, Manzoor Ali

    2011-01-01

    The aim of this study is to analyze the action research undertaken by Khan as a teacher researcher in a private school as part of the degree requirement of his M.Ed. program in Teacher Education at the Aga Khan University, Institute for Educational Development. The purpose of this analysis undertaken by the first author and supervisor of the study…

  20. A Hermeneutic Phenomenological Study of Community Mural Making and Social Action Art Therapy

    ERIC Educational Resources Information Center

    Rossetto, Erica

    2012-01-01

    Through a hermeneutic phenomenological study of interview data from 8 community artists, the author sought to discover commonalities and differences in the worldviews and philosophies of self that underlie community mural making as they relate to art therapy as social action and art therapy practice within a traditional Western cultural framework.…

  1. A Community Organizes for Action: A Case Study of the Mon-Yough Region in Pennsylvania.

    ERIC Educational Resources Information Center

    Avery, Robert W.; Chesler, Herbert A.

    This case study examines the development and problems of the Mon-Yough Community Action Committee, Inc. (MYCAC), one of the local anti-poverty agencies in Allegheny County, Pennsylvania. The agency's major effort is to overcome problems created by the decline of the local steel industry by supporting existing welfare agencies, and through such…

  2. Implementing an Action Research Project: A Case Study in Making Decisions and Managing Challenges

    ERIC Educational Resources Information Center

    Greenbank, Paul

    2013-01-01

    This paper examines the challenges involved in implementing an action research project. It discusses a project which uses a series of interventions (unfreezing techniques, cases studies in conjunction with analogical encoding and lecturer input) to encourage students to critically reflect on their approach to career decision-making. This paper…

  3. Human Securitability: A Participatory Action Research Study Involving Novice Teachers and Youngsters

    ERIC Educational Resources Information Center

    Kravale-Paulina, Marite; Olehnovica, Eridiana

    2015-01-01

    Civic participation, initiative and interest in current events can bridge the alienation felt towards national and municipal institutions, thereby enabling individuals to improve their quality of life and contribute to all-round sustainable development of their resident state. This paper reports on a participatory action research study into civic…

  4. Scientist-Teacher Partnerships as Professional Development: An Action Research Study

    ERIC Educational Resources Information Center

    Willcuts, Meredith Harris

    2009-01-01

    The overall purpose of this action research study was to explore the experiences of ten middle school science teachers involved in a three-year partnership program between scientists and teachers at a Department of Energy national laboratory, including the impact of the program on their professional development, and to improve the partnership…

  5. An Intervention to Increase the Use of Asthma Action Plans in Schools: A MASNRN Study

    ERIC Educational Resources Information Center

    Pulcini, Joyce; DeSisto, Marie C.; McIntyre, C. Lynne

    2007-01-01

    School nurses, in collaboration with primary care providers (PCPs), can work to better manage asthma by using the Asthma Action Plan (AAP) with peak flow monitoring. The aim of this pilot study was to determine the effectiveness of an intervention to increase the number of AAPs in schools for students with asthma by having school nurses provide…

  6. A Case Study Examining Change in Teacher Beliefs through Collaborative Action Research

    ERIC Educational Resources Information Center

    Vaino, Katrin; Holbrook, Jack; Rannikmae, Miia

    2013-01-01

    The main goal of this study was to explore the role of collaborative action research in eliciting change in teacher beliefs. The beliefs were those of five chemistry teachers in implementing a new teaching approach, geared to enhancing students' scientific and technological literacy (STL). The teacher beliefs were analysed based on Ajzen's Theory…

  7. An Action Research Study in an Icelandic Preschool: Developing Consensus about Values and Values Education

    ERIC Educational Resources Information Center

    Sigurdardottir, Ingibjorg; Einarsdottir, Johanna

    2016-01-01

    Values education is embedded in the curricula of all the Nordic countries. However, values education remains a neglected area for research and practice in early childhood education and care. This article reports on the aspects of an action research project conducted in a preschool in Iceland, across a period of 18 months. The study focused on the…

  8. Self-Study in Teaching and Teacher Development: A Call to Action

    ERIC Educational Resources Information Center

    Pithouse, Kathleen; Mitchell, Claudia; Weber, Sandra

    2009-01-01

    This article maps out key features of self-study in teaching and teacher development, particularly in relation to social action. As teacher educator-researchers, we have become increasingly interested in how self-reflexivity in teaching and teacher development can illuminate social and educational challenges that have resonance beyond the self and…

  9. Applying the Participatory Action Research Model to the Study of Social Inclusion at Worksites.

    ERIC Educational Resources Information Center

    Park, Hyun-Sook; Gonsier-Gerdin, Jean; Hoffman, Stacey; Whaley, Susan; Yount, Michael

    1998-01-01

    A study used participatory action research (PAR) to explore social inclusion/relationships at worksites of 10 students (ages 17-21). The participatory intervention process assisted teachers and job coaches in making constructive changes in transition work experience programs to provide social opportunities for students and help them become part of…

  10. Interdisciplinary Literacy through Social Media in the Mathematics Classroom: An Action Research Study

    ERIC Educational Resources Information Center

    Casey, Gail

    2013-01-01

    This article looks at how social and participatory media can be used to strengthen interdisciplinary literacy and connects the multimodality of social environments with Middle-Years Mathematics curriculum and delivery. The article reports on part of an eighteen months action research study in an Australian public high school within the…

  11. Using Professional Development to Improve Elementary Teachers' Mathematics Teaching: An Action Research Study

    ERIC Educational Resources Information Center

    Good, Leslie

    2009-01-01

    Research indicates that it is common for elementary teachers to possess limited mathematics content knowledge, high levels of math anxiety, and low levels of mathematics teacher efficacy. Assuming action research as the chosen methodology, this research study was developed with the ultimate goal of improving how elementary teachers in my district…

  12. Resolving Leadership Dilemmas in New Zealand Kindergartens: An Action Research Study

    ERIC Educational Resources Information Center

    Cardno, Carol; Reynolds, Bronwyn

    2009-01-01

    Purpose: The purpose of this paper is to examine dilemmas encountered by kindergarten head teachers with the further aim of developing their capability to recognise and resolve "leadership dilemmas". Design/methodology/approach: Action research was used to conduct a three-phase study involving 16 kindergarten head teachers and six system managers…

  13. Exploring a Science Teacher's Uncertainty with Integrating Engineering Design: An Action Research Study

    ERIC Educational Resources Information Center

    Capobianco, Brenda M.

    2011-01-01

    This study examines a fifth grade science teacher's attempts at integrating engineering design using the construct of uncertainty. Collaborative action research served as a supportive mechanism to uncover and confront the teacher's uncertainties. Data were gathered through semi-structured interviews, reflections, classroom observations, lesson…

  14. Training and Work Organisation: An Action-Research Study in a Sales and Distribution Company

    ERIC Educational Resources Information Center

    Bernardes, Alda Cristina; Lopes, Albino Pedro

    2005-01-01

    This study seeks to define a method of designing work-linked training, based on day-to-day work practices and the collaboration between all those involved. From diagnosis to evaluation, no training is designed or given without considering the opinions and interests of the parties involved. The method used is based on action research (AR) and on…

  15. An Evaluation of Professional Development to Improve Teachers' Perspectives and Behaviors: An Action Research Study

    ERIC Educational Resources Information Center

    Beckford-Young, Paulette Vivienne

    2016-01-01

    The purpose of this action research study was to conduct a professional development activity to provide content-area teachers with academic vocabulary strategies to be implemented during instruction on a daily basis. Professional development is essential for teachers to gain new knowledge and skills in order to hone their craft to improve student…

  16. Education as Sustainability: An Action Research Study of the Burns Model of Sustainability Pedagogy

    ERIC Educational Resources Information Center

    Burns, Heather

    2009-01-01

    Postsecondary teaching and learning must be reoriented to equip learners with the knowledge, skills, and values they will need for creating a more sustainable world. This action research study examined the effects of implementing the "Burns model of sustainability pedagogy" in university courses taught by the researcher. This model is comprised of…

  17. Task-Based Language Learning and Teaching: An Action-Research Study

    ERIC Educational Resources Information Center

    Calvert, Megan; Sheen, Younghee

    2015-01-01

    The creation, implementation, and evaluation of language learning tasks remain a challenge for many teachers, especially those with limited experience with using tasks in their teaching. This action-research study reports on one teacher's experience of developing, implementing, critically reflecting on, and modifying a language learning task…

  18. Improving Middle School Parental Engagement in Transition to Common Core State Standards: An Action Research Study

    ERIC Educational Resources Information Center

    Harla, Donna K.

    2014-01-01

    Parental involvement in schools is an important potential contributor to improving American education and making the U.S. more globally competitive. This qualitative and quantitative mixed-methodology action research study probed the viability of engaging parents around issues of educational improvement by inviting them to participate in training…

  19. Emotional stimuli modulate readiness for action: a transcranial magnetic stimulation study.

    PubMed

    van Loon, Anouk M; van den Wildenberg, Wery P M; van Stegeren, Anda H; Hajcak, Greg; Ridderinkhof, K Richard

    2010-05-01

    Emotional stimuli may prime the motor system and facilitate action readiness. Direct evidence for this effect has been shown by recent studies using transcranial magnetic stimulation (TMS). When administered over the primary motor cortex involved in responding, TMS pulses elicit motor-evoked potentials (MEPs) in the represented muscles. The amplitudes of these MEPs reflect the state of corticospinal excitability. Here, we investigated the dynamic effects of induced emotions on action readiness, as reflected by corticospinal excitability. Subjects performed a choice task while viewing task-irrelevant emotional and neutral pictures. The pattern of MEP amplitudes showed a typical increase as the TMS pulse was presented closer in time to the imminent response. This dynamic pattern was amplified by both pleasant and unpleasant emotional stimuli, but more so when unpleasant pictures were viewed. These patterns present novel evidence in support of the notion that emotional stimuli modulate action readiness.

  20. Investigating the neural basis of cooperative joint action. An EEG hyperscanning study.

    PubMed

    Astolfi, L; Toppi, J; Vogel, P; Mattia, D; Babiloni, F; Ciaramidaro, A; Siniatchkin, M

    2014-01-01

    The aim of the present study is to investigate the neurophysiological basis of the cognitive functions underlying the execution of joint actions, by means of the recent technique called hyperscanning. Neuroelectrical hyperscanning is based on the simultaneous recording of brain activity from multiple subjects and includes the analysis of the functional relation between the brain activity of all the interacting individuals. We recorded simultaneous high density electroencephalography (hdEEG) from 16 pairs of subjects involved in a computerized joint action paradigm, with controlled levels of cooperation. Results of cortical connectivity analysis returned significant differences, in terms of inter-brain functional causal links, between the condition of cooperative joint action and a condition in which the subjects were told they were interacting with a PC, while actually interacting with another human subject. Such differences, described by selected brain connectivity indices, point toward an integration between the two subjects' brain activity in the cooperative condition, with respect to control conditions.

  1. A case study of the development of environmental action projects from the framework of participatory action research within two middle school classrooms

    NASA Astrophysics Data System (ADS)

    Charmatz, Kim

    The purpose of this study was to understand student and teacher empowerment through a socially critical environmental education perspective. The main research question guiding this study was: How do participants make sense of a learning experience in which students design and carry out an environmental action project in their community? This study used participatory action research and critical theory as practical and theoretical frameworks. These frameworks were relevant as this study sought to examine social change, power, and relationships through participants' experiences. The context of this study was within one seventh and one eighth grade classroom participating in environmental projects. The study was conducted in spring 2005 with an additional follow-up data collection period during spring 2006. The school was located in a densely populated metropolitan suburb. Fifty-three students, a teacher researcher, and three science teachers participated. Data sources were written surveys, scores on Middle School Environmental Literacy Survey Instrument (MSELI), observations, interviews, and student work. This study used a mixed methodological approach. Quantitative data analysis involved dependent samples t-test scores on the MSELI before and after the completion of the projects. Qualitative data were analyzed using an inductive analysis approach. This study has implications for educators interested in democratic education. Environmental action projects provide a context for students and teachers to learn interdisciplinary content knowledge, develop personal beliefs, and learn ways to take action in their communities. This pedagogy has the potential to increase cooperation, communication, and tensions within school communities. Students' participation in the development of environmental action projects may lead to feelings of empowerment or being able to make a difference in their community, as an individual or member of a group. Future research is needed to discern

  2. Molecular insights into DNA interference by CRISPR-associated nuclease-helicase Cas3.

    PubMed

    Gong, Bei; Shin, Minsang; Sun, Jiali; Jung, Che-Hun; Bolt, Edward L; van der Oost, John; Kim, Jeong-Sun

    2014-11-18

    Mobile genetic elements in bacteria are neutralized by a system based on clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins. Type I CRISPR-Cas systems use a "Cascade" ribonucleoprotein complex to guide RNA specifically to complementary sequence in invader double-stranded DNA (dsDNA), a process called "interference." After target recognition by Cascade, formation of an R-loop triggers recruitment of a Cas3 nuclease-helicase, completing the interference process by destroying the invader dsDNA. To elucidate the molecular mechanism of CRISPR interference, we analyzed crystal structures of Cas3 from the bacterium Thermobaculum terrenum, with and without a bound ATP analog. The structures reveal a histidine-aspartate (HD)-type nuclease domain fused to superfamily-2 (SF2) helicase domains and a distinct C-terminal domain. Binding of ATP analog at the interface of the SF2 helicase RecA-like domains rearranges a motif V with implications for the enzyme mechanism. The HD-nucleolytic site contains two metal ions that are positioned at the end of a proposed nucleic acid-binding tunnel running through the SF2 helicase structure. This structural alignment suggests a mechanism for 3' to 5' nucleolytic processing of the displaced strand of invader DNA that is coordinated with ATP-dependent 3' to 5' translocation of Cas3 along DNA. In agreement with biochemical studies, the presented Cas3 structures reveal important mechanistic details on the neutralization of genetic invaders by type I CRISPR-Cas systems.

  3. CRISPR/Cas9 produces anti-hepatitis B virus effect in hepatoma cells and transgenic mouse.

    PubMed

    Zhu, Wei; Xie, Kun; Xu, Yuanjian; Wang, Le; Chen, Kaiming; Zhang, Longzhen; Fang, Jianmin

    2016-06-01

    Chronic infection of hepatitis B virus (HBV) is at risk of liver cirrhosis and hepatocellular carcinoma and remains one of the major public health problems worldwide. It is a major barrier of persistence HBV cccDNA under current antiviral therapy as novel strategies of disrupting HBV cccDNA is pressing. The (CRISPR)/Cas9 system is presently emerging in gene editing and we also apply it for targeting and deleting the conserved regions of HBV genome. Two homologous sequences of HBV S and X genes were carried with CRISPR/Cas9 endonuclease to build pCas9 constructs, which may mediate anti-HBV effects of in vitro and in vivo systems in this study. The results showed the better anti-HBV productions by pCas9-2 and without significant differences in between Huh7 and HepG2 cells. CRISPR/Cas9 direct cleavage and mutagenesis were further analyzed of in vitro system. In the M-TgHBV mouse model of HBV, injection of pCas9 constructs by hydrodynamics decreased HBsAg of sera and liver HBcAg. In conclusion, this designed CRISPR/Cas9 system can induce anti-HBV effects and potentially consider as a novel therapeutic agent against chronic HBV infection.

  4. Generation of porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene by somatic cell nuclear transfer.

    PubMed

    Liu, Guoqian; Liu, Kai; Wei, Hengxi; Li, Li; Zhang, Shouquan

    2016-09-01

    Cas9 endonuclease, from so-called clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems of Streptococcus pyogenes, type II functions as an RNA-guided endonuclease and edits the genomes of prokaryotic and eukaryotic organisms, including deletion and insertion by DNA double‑stranded break repair mechanisms. In previous studies, it was observed that Cas9, with a genome‑scale lentiviral single‑guide RNA library, could be applied to a loss‑of‑function genetic screen, although the loss‑of‑function genes have yet to be verified in vitro and this approach has not been used in porcine cells. Based on these observations, lentiviral Cas9 was used to infect porcine primary fibroblasts to achieve cell colonies carrying Cas9 endonuclease. Subsequently, porcine fetal fibroblasts expressing the tetracycline‑inducible Cas9 gene were generated by somatic cell nuclear transfer, and three 30 day transgenic porcine fetal fibroblasts (PFFs) were obtained. Polymerase chain reaction (PCR), reverse transcription‑PCR and western blot analysis indicated that the PFFs were Cas9‑positive. In addition, one of the three integrations was located near to known functional genes in the PFF1 cell line, whereas neither of the integrations was located in the PFF1 or PFF2 cell lines. It was hypothesized that these transgenic PFFs may be useful for conditional genomic editing in pigs, and for generating ideal modified porcine models. PMID:27430306

  5. Generation of porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene by somatic cell nuclear transfer

    PubMed Central

    Liu, Guoqian; Liu, Kai; Wei, Hengxi; Li, Li; Zhang, Shouquan

    2016-01-01

    Cas9 endonuclease, from so-called clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems of Streptococcus pyogenes, type II functions as an RNA-guided endonuclease and edits the genomes of prokaryotic and eukaryotic organisms, including deletion and insertion by DNA double-stranded break repair mechanisms. In previous studies, it was observed that Cas9, with a genome-scale lentiviral single-guide RNA library, could be applied to a loss-of-function genetic screen, although the loss-of-function genes have yet to be verified in vitro and this approach has not been used in porcine cells. Based on these observations, lentiviral Cas9 was used to infect porcine primary fibroblasts to achieve cell colonies carrying Cas9 endonuclease. Subsequently, porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene were generated by somatic cell nuclear transfer, and three 30 day transgenic porcine fetal fibroblasts (PFFs) were obtained. Polymerase chain reaction (PCR), reverse transcription-PCR and western blot analysis indicated that the PFFs were Cas9-positive. In addition, one of the three integrations was located near to known functional genes in the PFF1 cell line, whereas neither of the integrations was located in the PFF1 or PFF2 cell lines. It was hypothesized that these transgenic PFFs may be useful for conditional genomic editing in pigs, and for generating ideal modified porcine models. PMID:27430306

  6. Closure Report for Corrective Action Unit 543: Liquid Disposal Units, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2008-01-01

    This Closure Report (CR) documents closure activities for Corrective Action Unit (CAU) 543, Liquid Disposal Units, according to the Federal Facility Agreement and Consent Order (FFACO, 1996) and the Corrective Action Plan (CAP) for CAU 543 (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2007). CAU 543 is located at the Nevada Test Site (NTS), Nevada (Figure 1), and consists of the following seven Corrective Action Sites (CASs): CAS 06-07-01, Decon Pad; CAS 15-01-03, Aboveground Storage Tank; CAS 15-04-01, Septic Tank; CAS 15-05-01, Leachfield; CAS 15-08-01, Liquid Manure Tank; CAS 15-23-01, Underground Radioactive Material Area; CAS 15-23-03, Contaminated Sump, Piping; and CAS 06-07-01 is located at the Decontamination Facility in Area 6, adjacent to Yucca Lake. The remaining CASs are located at the former U.S. Environmental Protection Agency (EPA) Farm in Area 15. The purpose of this CR is to provide a summary of the completed closure activities, to document waste disposal, and to present analytical data confirming that the remediation goals were met. The closure alternatives consisted of closure in place for two of the CASs, and no further action with implementation of best management practices (BMPs) for the remaining five CASs.

  7. 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

    PubMed Central

    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.

    2016-01-01

    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

  8. 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.

    PubMed

    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.

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

    PubMed

    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

  10. Closure Report for Corrective Action Unit 563: Septic Systems, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2010-02-28

    Corrective Action Unit (CAU) 563 is identified in the Federal Facility Agreement and Consent Order (FFACO) as “Septic Systems” and consists of the following four Corrective Action Sites (CASs), located in Areas 3 and 12 of the Nevada Test Site: · CAS 03-04-02, Area 3 Subdock Septic Tank · CAS 03-59-05, Area 3 Subdock Cesspool · CAS 12-59-01, Drilling/Welding Shop Septic Tanks · CAS 12-60-01, Drilling/Welding Shop Outfalls Closure activities were conducted from September to November 2009 in accordance with the FFACO (1996, as amended February 2008) and the Corrective Action Plan for CAU 563. The corrective action alternatives included No Further Action and Clean Closure.

  11. Studies on the mechanisms of action of the herbicide safener CGA-92194

    SciTech Connect

    Zama, P.

    1986-01-01

    CGA-92194 is a herbicide safener that is used as a seed dressing agent to protect grain sorghum against metolachlor injury. The potential adverse phytotoxic effects and the mechanisms of the protective action of this safener were studied in laboratory experiments. Adverse phytotoxicity was assessed by comparing CGA-92194 and the herbicide safeners cyometrinil and flurazole for their effects on CO/sub 2/ fixation, protein, DNA, RNA and lipid synthesis of enzymatically isolated leaf cells of soybean. The safening action mechanisms of CGA-92194 were studied by examining the potential interactions of this safener with metolachlor at the levels of uptake and macromolecular syntheses in enzymatically isolated leaf mesophyll protoplasts of grain sorghum. When CGA-92194 and metolachlor were given simultaneously, CGA-92194 enhanced /sup 14/C-metolachlor uptake into the sorghum protoplasts in a concentration-dependent pattern. Treatments with metolachlor and CGA-92194 in combination inhibited the incorporation of /sup 14/C-uracil, /sup 3/H-thymidine and /sup 14/C-acetate into sorghum protoplast macromolecules less than metolachlor given alone, suggesting the potential involvement of a competitive antagonism in CGA-92194 mechanism of action. The metabolic activity and growth of sorghum seedlings grown from CGA-92194-pretreated seeds were lower than that of seedlings grown from untreated seeds at 10 or 20 days after planting. These results indicate that a safener-induced stimulation of the spontaneous or enzymatic conjugation of metolachlor with GSH is most likely involved in CGA-92194 protective action.

  12. Micro grants as a stimulus for community action in residential health programmes: a case study.

    PubMed

    Schmidt, M; Plochg, T; Harting, J; Klazinga, N S; Stronks, K

    2009-09-01

    This paper aimed to explore the contribution of a micro grant financing scheme to community action in terms of residential health-promoting initiatives, interorganizational collaboration and public participation. The scheme was two-fold, consisting of (i) micro grants of 500-3500 Euros, which were easily obtainable by local organizations and (ii) neighbourhood health panels of community and health workers, functioning as a distributing mechanism. Data were collected using three methods: (i) observations of the neighbourhood-based health panels, (ii) in-depth interviews with policy-makers and professionals and (iii) analyses of documents and reports. This study demonstrated the three-fold role of micro grants as a vehicle to enable community action at an organizational level in terms of increased network activities between the local organizations, to set an agenda for the 'health topic' in non-traditional health agencies and to enable a number of health-promoting initiatives. Although these initiatives were attended by small groups of residents normally considered hard to reach, the actual public participation was limited. In their role as a distributing mechanism, the health panels were vital with regard to the achieved impact on the community action. However, certain limitations were also seen, which were related to the governance of the panels. This case study provides evidence to suggest that micro grants have the potential to stimulate community action at an organizational and a residential level, but with the prerequisite that grants be accompanied by increased investments in infrastructure.

  13. A Case Study Examining Change in Teacher Beliefs Through Collaborative Action Research

    NASA Astrophysics Data System (ADS)

    Vaino, Katrin; Holbrook, Jack; Rannikmäe, Miia

    2013-01-01

    The main goal of this study was to explore the role of collaborative action research in eliciting change in teacher beliefs. The beliefs were those of five chemistry teachers in implementing a new teaching approach, geared to enhancing students' scientific and technological literacy (STL). The teacher beliefs were analysed based on Ajzen's Theory of Planned Behaviour (2005) by looking at the teacher's (a) attitude towards implementing STL modules, (b) perceived subjective norms, and (c) behavioural control regarding the new teaching approach. After an introductory year, when teachers familiarised themselves with the new approach, a collaborative action research project was initiated in the second year of the study, helping teachers to minimise or overcome initially perceived constraints when implementing STL modules in their classroom. The processes of teacher change and the course of the project were investigated by teacher interviews, teacher informal commentaries, and meeting records. The formation of positive beliefs towards a STL approach increased continuously, although its extent and character varied depending on the teacher. The close cooperation, in the format of collaborative action research and especially through teacher group reflections and perceived collegial support, did support teacher professional development including change in their beliefs towards the new teaching approach. Additionally, positive feedback gained from other teachers through running a two-day in-service course in year three helped to strengthen all five teachers' existing beliefs towards the new approach. The current research demonstrated that perceived constraints, where identified, can be meaningfully addressed by teachers, through undertaking collaborative action research.

  14. Dual sgRNAs facilitate CRISPR/Cas9-mediated mouse genome targeting.

    PubMed

    Zhou, Jiankui; Wang, Jianying; Shen, Bin; Chen, Li; Su, Yang; Yang, Jing; Zhang, Wensheng; Tian, Xuemei; Huang, Xingxu

    2014-04-01

    The bacterial clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system is a versatile RNA-guided mammalian genome modification system. One-step generation of mouse genome targeting has been achieved by co-microinjection of one-cell stage embryos with Cas9 mRNA and small/single guide (sg)RNA. Many studies have focused on enhancing the efficiency of this system. In the present study, we report that simultaneous use of dual sgRNAs to target an individual gene significantly improved the Cas9-mediated genome targeting with a bi-allelic modification efficiency of up to 78%. We further observed that the target gene modifications were characterized by efficient germline transmission and site-dependent off-target effects, and also that the apolipoprotein E gene knockout-mediated defects in blood biochemical parameters were recapitulated by CRISPR/Cas9-mediated heritable gene modification. Our results provide a dual sgRNAs strategy to facilitate CRISPR/Cas9-mediated mouse genome targeting.

  15. Sensitivity of Alpha and Beta Oscillations to Sensorimotor Characteristics of Action: An EEG Study of Action Production and Gesture Observation

    PubMed Central

    Quandt, Lorna C.; Marshall, Peter J.; Shipley, Thomas F.; Beilock, Sian L.; Goldin-Meadow, Susan

    2012-01-01

    The sensorimotor experiences we gain when performing an action have been found to influence how our own motor systems are activated when we observe others performing that same action. Here we asked whether this phenomenon applies to the observation of gesture. Would the sensorimotor experiences we gain when performing an action on an object influence activation in our own motor systems when we observe others performing a gesture for that object? Participants were given sensorimotor experience with objects that varied in weight, and then observed video clips of an actor producing gestures for those objects. Electroencephalography (EEG) was recorded while participants first observed either an iconic gesture (pantomiming lifting an object) or a deictic gesture (pointing to an object) for an object, and then grasped and lifted the object indicated by the gesture. We analyzed EEG during gesture observation to determine whether oscillatory activity was affected by the observer’s sensorimotor experiences with the object represented in the gesture. Seeing a gesture for an object previously experienced as light was associated with a suppression of power in alpha and beta frequency bands, particularly at posterior electrodes. A similar pattern was found when participants lifted the light object, but over more diffuse electrodes. Moreover, alpha and beta bands at right parieto-occipital electrodes were sensitive to the type of gesture observed (iconic vs. deictic). These results demonstrate that sensorimotor experience with an object affects how a gesture for that object is processed, as measured by the gesture-observer’s EEG, and suggest that different types of gestures recruit the observer’s own motor system in different ways. PMID:22910276

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

    PubMed Central

    2011-01-01

    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 comparison continue to

  17. Can Discrete Joint Action Be Synergistic? Studying the Stabilization of Interpersonal Hand Coordination

    PubMed Central

    Romero, Veronica; Kallen, Rachel; Riley, Michael A.; Richardson, Michael J.

    2016-01-01

    The human perceptual-motor system is tightly coupled to the physical and informational dynamics of a task environment. These dynamics operate to constrain the high-dimensional order of the human movement system into low-dimensional, task-specific synergies—functional groupings of structural elements that are temporarily constrained to act as a single coordinated unit. The aim of the current study was to determine whether synergistic processes operate when coacting individuals coordinate to perform a discrete joint-action task. Pairs of participants sat next to each other and each used 1 arm to complete a pointer-to-target task. Using the uncontrolled manifold (UCM) analysis for the first time in a discrete joint action, the structure of joint-angle variance was examined to determine whether there was synergistic organization of the degrees of freedom employed at the interpersonal or intrapersonal levels. The results revealed that the motor actions performed by coactors were synergistically organized at both the interpersonal and intrapersonal levels. More importantly, however, the interpersonal synergy was found to be significantly stronger than the intrapersonal synergies. Accordingly, the results provide clear evidence that coacting individuals can become temporarily organized to form single synergistic 2-person systems during performance of a discrete joint action. PMID:26052696

  18. Overcoming doxorubicin resistance of cancer cells by Cas9-mediated gene disruption

    PubMed Central

    Ha, Jong Seong; Byun, Juyoung; Ahn, Dae-Ro

    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-toxic to the untreated cells. The potency of drug was enhanced in the cells treated with the protein-RNA complex as well as in those treated with plasmids, suggesting that mutation of the mdr1 gene by intracellular delivery of Cas9-sgRNA complex using proper protein delivery platforms could recover the drug susceptibility. Therefore, Cas9-mediated disruption of the drug resistance-related gene can be considered as a promising way to overcome multidrug resistance in cancer cells. PMID:26961701

  19. Pseudorabies virus can escape from CRISPR-Cas9-mediated inhibition.

    PubMed

    Peng, Zhiyuan; Ouyang, Ting; Pang, Daxin; Ma, Teng; Chen, Xinrong; Guo, Ning; Chen, Fuwang; Yuan, Lin; Ouyang, Hongsheng; Ren, Linzhu

    2016-09-01

    The CRISPR-Cas9 system is a newly developed genome-engineering tool used to inhibit virus infection by targeting the conserved regions of the viral genomic DNA. In the present study, we constructed a cell line stably expressing Cas9 endonuclease and sgRNA targeting the conserved UL30 gene of pseudorabies virus (PRV). During the PRV infection, the CRISPR-Cas9 system was efficient in cleaving the UL30 gene in each passage. However, deletions and insertions occurred at low passages, while substitutions were frequently observed at high passages. Furthermore, copy numbers and virus titers of PRV were significantly increased in a passage-dependent manner, indicating that viral genomic replication and assembly were more effective at the high passages than at low passages. These results demonstrated that PRV could escape from CRISPR-Cas9-mediated inhibition. Therefore, whether the CRISPR-Cas9 system is suitable for antiviral application should be considered and carefully verified. PMID:27507009

  20. Efficient Targeted Genome Modification in Maize Using CRISPR/Cas9 System.

    PubMed

    Feng, Chao; Yuan, Jing; Wang, Rui; Liu, Yang; Birchler, James A; Han, Fangpu

    2016-01-20

    CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 system, which is a newly developed technology for targeted genome modification, has been successfully used in a number of species. In this study, we applied this technology to carry out targeted genome modification in maize. A marker gene Zmzb7 was chosen for targeting. The sgRNA-Cas9 construct was transformed into maize protoplasts, and indel (insertion and deletion) mutations could be detected. A mutant seedling with an expected albino phenotype was obtained from screening 120 seedlings generated from 10 callus events. Mutation efficiency in maize heterochromatic regions was also investigated. Twelve sites with different expression levels in maize centromeres or pericentromere regions were selected. The sgRNA-Cas9 constructs were transformed into protoplasts followed by sequencing the transformed protoplast genomic DNA. The results show that the genes in heterochromatic regions could be targeted by the CRISPR/Cas9 system efficiently, no matter whether they are expressed or not. Meanwhile, off-target mutations were not found in the similar sites having no PAM (protospacer adjacent motif) or having more than two mismatches. Together, our results show that the CRISPR/Cas9 system is a robust and efficient tool for genome modification in both euchromatic and heterochromatic regions in maize. PMID:26842992

  1. Efficient Targeted Genome Modification in Maize Using CRISPR/Cas9 System.

    PubMed

    Feng, Chao; Yuan, Jing; Wang, Rui; Liu, Yang; Birchler, James A; Han, Fangpu

    2016-01-20

    CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 system, which is a newly developed technology for targeted genome modification, has been successfully used in a number of species. In this study, we applied this technology to carry out targeted genome modification in maize. A marker gene Zmzb7 was chosen for targeting. The sgRNA-Cas9 construct was transformed into maize protoplasts, and indel (insertion and deletion) mutations could be detected. A mutant seedling with an expected albino phenotype was obtained from screening 120 seedlings generated from 10 callus events. Mutation efficiency in maize heterochromatic regions was also investigated. Twelve sites with different expression levels in maize centromeres or pericentromere regions were selected. The sgRNA-Cas9 constructs were transformed into protoplasts followed by sequencing the transformed protoplast genomic DNA. The results show that the genes in heterochromatic regions could be targeted by the CRISPR/Cas9 system efficiently, no matter whether they are expressed or not. Meanwhile, off-target mutations were not found in the similar sites having no PAM (protospacer adjacent motif) or having more than two mismatches. Together, our results show that the CRISPR/Cas9 system is a robust and efficient tool for genome modification in both euchromatic and heterochromatic regions in maize.

  2. Mapping the Unshocked Interior Ejecta in the Cas A Supernova Remnant

    NASA Astrophysics Data System (ADS)

    Fesen, Robert; Milisavljevic, Dan

    2011-08-01

    Cassiopeia A (Cas A) provides perhaps our best look at the explosion dynamics of a core-collapse supernova (CCSN). Exploratory spectra of Cas A obtained by us in 2010 showed that the KPNO 4m + MARS' exceptional far-red throughput is capable of detecting the remnant's faint interior [S III] 9069,9531 and [Fe II] 8617 line emissions, thereby allowing one to study the structure of the whole remnant, from its core to the outer layers. Our November 2010 HST/WFC3 [S III] images show that this faint interior emission, which is presumably photoionized by UV and X-ray flux from Cas A's bright main shell, is extensive and seemingly highly structured. Here we request eight (8) nights using the KPNO 4m + MARS to obtain far-red spectroscopy of Cas A in order to survey the remnant's interior unshocked S- and Fe-rich ejecta...an observation that seemed impossible just a year ago. The proposed observations, when combined with our extensive set of spectra of Cas A's bright outer ejecta, will yield the first complete map of a CCSN - from its center to the outer, bright reverse-shock heated layers. These observations will give us an extraordinarily rare look at the geometry of a remnant's inner volume of expanding cloud of S- and Fe-rich debris unmodified by reverse shock instabilities, thus offering a unique and powerful test of high-mass progenitor supernova explosion models.

  3. Differential Distribution of Type II CRISPR-Cas Systems in Agricultural and Nonagricultural Campylobacter coli and Campylobacter jejuni Isolates Correlates with Lack of Shared Environments.

    PubMed

    Pearson, Bruce M; Louwen, Rogier; van Baarlen, Peter; van Vliet, Arnoud H M

    2015-09-02

    CRISPR (clustered regularly interspaced palindromic repeats)-Cas (CRISPR-associated) systems are sequence-specific adaptive defenses against phages and plasmids which are widespread in prokaryotes. Here we have studied whether phylogenetic relatedness or sharing of environmental niches affects the distribution and dissemination of Type II CRISPR-Cas systems, first in 132 bacterial genomes from 15 phylogenetic classes, ranging from Proteobacteria to Actinobacteria. There was clustering of distinct Type II CRISPR-Cas systems in phylogenetically distinct genera with varying G+C%, which share environmental niches. The distribution of CRISPR-Cas within a genus was studied using a large collection of genome sequences of the closely related Campylobacter species Campylobacter jejuni (N = 3,746) and Campylobacter coli (N = 486). The Cas gene cas9 and CRISPR-repeat are almost universally present in C. jejuni genomes (98.0% positive) but relatively rare in C. coli genomes (9.6% positive). Campylobacter jejuni and agricultural C. coli isolates share the C. jejuni CRISPR-Cas system, which is closely related to, but distinct from the C. coli CRISPR-Cas system found in C. coli isolates from nonagricultural sources. Analysis of the genomic position of CRISPR-Cas insertion suggests that the C. jejuni-type CRISPR-Cas has been transferred to agricultural C. coli. Conversely, the absence of the C. coli-type CRISPR-Cas in agricultural C. coli isolates may be due to these isolates not sharing the same environmental niche, and may be affected by farm hygiene and biosecurity practices in the agricultural sector. Finally, many CRISPR spacer alleles were linked with specific multilocus sequence types, suggesting that these can assist molecular epidemiology applications for C. jejuni and C. coli.

  4. The organization of the posterior parietal cortex devoted to upper limb actions: An fMRI study

    PubMed Central

    Ferri, Stefania; Rizzolatti, Giacomo

    2015-01-01

    Abstract The present fMRI study examined whether upper‐limb action classes differing in their motor goal are encoded by different PPC sectors. Action observation was used as a proxy for action execution. Subjects viewed actors performing object‐related (e.g., grasping), skin‐displacing (e.g., rubbing the skin), and interpersonal upper limb actions (e.g., pushing someone). Observation of the three action classes activated a three‐level network including occipito‐temporal, parietal, and premotor cortex. The parietal region common to observing all three action classes was located dorsally to the left intraparietal sulcus (DIPSM/DIPSA border). Regions specific for observing an action class were obtained by combining the interaction between observing action classes and stimulus types with exclusive masking for observing the other classes, while for regions considered preferentially active for a class the interaction was exclusively masked with the regions common to all observed actions. Left putative human anterior intraparietal was specific for observing manipulative actions, and left parietal operculum including putative human SII region, specific for observing skin‐displacing actions. Control experiments demonstrated that this latter activation depended on seeing the skin being moved and not simply on seeing touch. Psychophysiological interactions showed that the two specific parietal regions had similar connectivities. Finally, observing interpersonal actions preferentially activated a dorsal sector of left DIPSA, possibly the homologue of ventral intraparietal coding the impingement of the target person's body into the peripersonal space of the actor. These results support the importance of segregation according to the action class as principle of posterior parietal cortex organization for action observation and by implication for action execution. Hum Brain Mapp 36:3845–3866, 2015. © 2015 The Authors Human Brain Mapping Published by Wiley

  5. The organization of the posterior parietal cortex devoted to upper limb actions: An fMRI study.

    PubMed

    Ferri, Stefania; Rizzolatti, Giacomo; Orban, Guy A

    2015-10-01

    The present fMRI study examined whether upper-limb action classes differing in their motor goal are encoded by different PPC sectors. Action observation was used as a proxy for action execution. Subjects viewed actors performing object-related (e.g., grasping), skin-displacing (e.g., rubbing the skin), and interpersonal upper limb actions (e.g., pushing someone). Observation of the three action classes activated a three-level network including occipito-temporal, parietal, and premotor cortex. The parietal region common to observing all three action classes was located dorsally to the left intraparietal sulcus (DIPSM/DIPSA border). Regions specific for observing an action class were obtained by combining the interaction between observing action classes and stimulus types with exclusive masking for observing the other classes, while for regions considered preferentially active for a class the interaction was exclusively masked with the regions common to all observed actions. Left putative human anterior intraparietal was specific for observing manipulative actions, and left parietal operculum including putative human SII region, specific for observing skin-displacing actions. Control experiments demonstrated that this latter activation depended on seeing the skin being moved and not simply on seeing touch. Psychophysiological interactions showed that the two specific parietal regions had similar connectivities. Finally, observing interpersonal actions preferentially activated a dorsal sector of left DIPSA, possibly the homologue of ventral intraparietal coding the impingement of the target person's body into the peripersonal space of the actor. These results support the importance of segregation according to the action class as principle of posterior parietal cortex organization for action observation and by implication for action execution. PMID:26129732

  6. Closure Report for Corrective Action Unit 104: Area 7 Yucca Flat Atmospheric Test Sites, Nevada National Security Site, Nevada

    SciTech Connect

    2013-06-27

    This Closure Report (CR) presents information supporting closure of Corrective Action Unit (CAU) 104, Area 7 Yucca Flat Atmospheric Test Sites, and provides documentation supporting the completed corrective actions and confirmation that closure objectives for CAU 104 were met. This CR complies with the requirements of the Federal Facility Agreement and Consent Order (FFACO) that was agreed to by the State of Nevada; the U.S. Department of Energy (DOE), Environmental Management; the U.S. Department of Defense; and DOE, Legacy Management. CAU 104 consists of the following 15 Corrective Action Sites (CASs), located in Area 7 of the Nevada National Security Site: · CAS 07-23-03, Atmospheric Test Site T-7C · CAS 07-23-04, Atmospheric Test Site T7-1 · CAS 07-23-05, Atmospheric Test Site · CAS 07-23-06, Atmospheric Test Site T7-5a · CAS 07-23-07, Atmospheric Test Site - Dog (T-S) · CAS 07-23-08, Atmospheric Test Site - Baker (T-S) · CAS 07-23-09, Atmospheric Test Site - Charlie (T-S) · CAS 07-23-10, Atmospheric Test Site - Dixie · CAS 07-23-11, Atmospheric Test Site - Dixie · CAS 07-23-12, Atmospheric Test Site - Charlie (Bus) · CAS 07-23-13, Atmospheric Test Site - Baker (Buster) · CAS 07-23-14, Atmospheric Test Site - Ruth · CAS 07-23-15, Atmospheric Test Site T7-4 · CAS 07-23-16, Atmospheric Test Site B7-b · CAS 07-23-17, Atmospheric Test Site - Climax Closure activities began in October 2012 and were completed in April 2013. Activities were conducted according to the Corrective Action Decision Document/Corrective Action Plan for CAU 104. The corrective actions included No Further Action and Clean Closure. Closure activities generated sanitary waste, mixed waste, and recyclable material. Some wastes exceeded land disposal limits and required treatment prior to disposal. Other wastes met land disposal restrictions and were disposed in appropriate onsite landfills. The U.S. Department of Energy, National Nuclear Security Administration Nevada Field Office

  7. Efficient Generation of Myostatin Gene Mutated Rabbit by CRISPR/Cas9.

    PubMed

    Lv, Qingyan; Yuan, Lin; Deng, Jichao; Chen, Mao; Wang, Yong; Zeng, Jian; Li, Zhanjun; Lai, Liangxue

    2016-04-26

    CRISPR/Cas9 has been widely used in generating site-specific genetically modified animal models. Myostatin (MSTN) is a negative regulator of muscle mass, related to muscle growth and differentiation. The knockout of MSTN with the desired phenotype of double muscle has been successfully generated in mice, goats, pigs and cattle, but not in rabbits. In this study, the MSTN knockout (KO) rabbits were generated by co-injection of Cas9 mRNA and sgRNA into zygotes. The typical phenotype of double muscle with hyperplasia or hypertrophy of muscle fiber was observed in MSTN KO rabbits. Furthermore, a similar phenotype was found in the F1 generation, suggesting that the mutation of MSTN could be stably inherited in the MSTN KO rabbits. In summary, we have successfully generated MSTN KO rabbits using CRISPR/Cas9 system with high efficiency, which is a reliable and effective animal model for the study of muscle development and related diseases.

  8. Modeling disease in vivo with CRISPR/Cas9

    PubMed Central

    Dow, Lukas E.

    2015-01-01

    The recent advent of CRISPR/Cas9-mediated genome editing has created a wave of excitement across the scientific research community, carrying the promise of simple and effective genomic manipulation of nearly any cell type. CRISPR has quickly become the preferred tool for genetic manipulation, and shows incredible promise as a platform for studying gene function in vivo. Here, I discuss the current application of CRISPR technology to create new in vivo disease models, with a particular focus on how these tools, derived from an adaptive bacterial immune system, are helping us better model the complexity of human cancer. PMID:26432018

  9. CRISPR/Cas9 system as an innovative genetic engineering tool: Enhancements in sequence specificity and delivery methods.

    PubMed

    Jo, Young-Il; Suresh, Bharathi; Kim, Hyongbum; Ramakrishna, Suresh

    2015-12-01

    While human gene therapy has gained significant attention for its therapeutic promise, CRISPR/Cas9 technology has made a breakthrough as an efficient genome editing tool by emulating prokaryotic immune defense mechanisms. Although many studies have found that CRISPR/Cas9 technology is more efficient, specific and manipulable than previous generations of gene editing tools, it can be further improved by elevating its overall efficiency in a higher frequency of genome modifications and reducing its off-target effects. Here, we review the development of CRISPR/Cas9 technology, focusing on enhancement of its sequence specificity, reduction of off-target effects and delivery systems. Moreover, we describe recent successful applications of CRISPR/Cas9 technology in laboratory and clinical studies.

  10. CRISPR/Cas9 system as an innovative genetic engineering tool: Enhancements in sequence specificity and delivery methods.

    PubMed

    Jo, Young-Il; Suresh, Bharathi; Kim, Hyongbum; Ramakrishna, Suresh

    2015-12-01

    While human gene therapy has gained significant attention for its therapeutic promise, CRISPR/Cas9 technology has made a breakthrough as an efficient genome editing tool by emulating prokaryotic immune defense mechanisms. Although many studies have found that CRISPR/Cas9 technology is more efficient, specific and manipulable than previous generations of gene editing tools, it can be further improved by elevating its overall efficiency in a higher frequency of genome modifications and reducing its off-target effects. Here, we review the development of CRISPR/Cas9 technology, focusing on enhancement of its sequence specificity, reduction of off-target effects and delivery systems. Moreover, we describe recent successful applications of CRISPR/Cas9 technology in laboratory and clinical studies. PMID:26434948

  11. Gender difference in symptomatic radiographic knee osteoarthritis in the Knee Clinical Assessment – CAS(K): A prospective study in the general population

    PubMed Central

    Lacey, Rosie J; Thomas, Elaine; Duncan, Rachel C; Peat, George

    2008-01-01

    Background A recent study of adults aged ≥50 years reporting knee pain found an excess of radiographic knee osteoarthritis (knee ROA) in symptomatic males compared to females. This was independent of age, BMI and other clinical signs and symptoms. Since this finding contradicts many previous studies, our objective was to explore four possible explanations for this gender difference: X-ray views, selection, occupation and non-articular conditions. Methods A community-based prospective study. 819 adults aged ≥50 years reporting knee pain in the previous 12 months were recruited by postal questionnaires to a research clinic involving plain radiography (weight-bearing posteroanterior semiflexed, supine skyline and lateral views), clinical interview and physical examination. Any knee ROA, ROA severity, tibiofemoral joint osteoarthritis (TJOA) and patellofemoral joint osteoarthritis (PJOA) were defined using all three radiographic views. Occupational class was derived from current or last job title. Proportions of each gender with symptomatic knee ROA were expressed as percentages, stratified by age; differences between genders were expressed as percentage differences with 95% confidence intervals. Results 745 symptomatic participants were eligible and had complete X-ray data. Males had a higher occurrence (77%) of any knee ROA than females (61%). In 50–64 year olds, the excess in men was mild knee OA (particularly PJOA); in ≥65 year olds, the excess was both mild and moderate/severe knee OA (particularly combined TJOA/PJOA). This male excess persisted when using the posteroanterior view only (64% vs. 52%). The lowest level of participation in the clinic was symptomatic females aged 65+. Within each occupational class there were more males with symptomatic knee ROA than females. In those aged 50–64 years, non-articular conditions were equally common in both genders although, in those aged 65+, they occurred more frequently in symptomatic females (41%) than

  12. Closure Report for Corrective Action Unit 139: Waste Disposal Sites, Nevada Test Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2009-07-31

    Corrective Action Unit (CAU) 139 is identified in the Federal Facility Agreement and Consent Order (FFACO) as 'Waste Disposal Sites' and consists of the following seven Corrective Action Sites (CASs), located in Areas 3, 4, 6, and 9 of the Nevada Test Site: CAS 03-35-01, Burn Pit; CAS 04-08-02, Waste Disposal Site; CAS 04-99-01, Contaminated Surface Debris; CAS 06-19-02, Waste Disposal Site/Burn Pit; CAS 06-19-03, Waste Disposal Trenches; CAS 09-23-01, Area 9 Gravel Gertie; and CAS 09-34-01, Underground Detection Station. Closure activities were conducted from December 2008 to April 2009 according to the FFACO (1996, as amended February 2008) and the Corrective Action Plan for CAU 139 (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office, 2007b). The corrective action alternatives included No Further Action, Clean Closure, and Closure in Place with Administrative Controls. Closure activities are summarized. CAU 139, 'Waste Disposal Sites,' consists of seven CASs in Areas 3, 4, 6, and 9 of the NTS. The closure alternatives included No Further Action, Clean Closure, and Closure in Place with Administrative Controls. This CR provides a summary of completed closure activities, documentation of waste disposal, and confirmation that remediation goals were met. The following site closure activities were performed at CAU 139 as documented in this CR: (1) At CAS 03-35-01, Burn Pit, soil and debris were removed and disposed as LLW, and debris was removed and disposed as sanitary waste. (2) At CAS 04-08-02, Waste Disposal Site, an administrative UR was implemented. No postings or post-closure monitoring are required. (3) At CAS 04-99-01, Contaminated Surface Debris, soil and debris were removed and disposed as LLW, and debris was removed and disposed as sanitary waste. (4) At CAS 06-19-02, Waste Disposal Site/Burn Pit, no work was performed. (5) At CAS 06-19-03, Waste Disposal Trenches, a native soil cover was installed, and a UR was

  13. NTP toxicity report of reproductive dose range-finding study of Genistein (CAS No. 446-72-0) administered in feed to Sprague-Dawley rats.

    PubMed

    Delclos, K B; Newbold, Retha

    2007-11-01

    Genistein is a naturally occurring isoflavone that interacts with estrogen receptors and multiple other molecular targets. Human exposure to genistein is predominantly through consumption of soy products, including soy-based infant formula and dietary supplements. A series of short-term studies with genistein was conducted with two goals: 1) to obtain data necessary to establish dose levels for subsequent multigeneration reproductive and chronic toxicity studies and 2) to evaluate the effects of genistein on endpoints outside the reproductive tract. The data generated from these studies have been reported previously in the peer-reviewed literature or in technical reports (Appendix C). In addition, selected data from these studies were analyzed and discussed in the National Toxicology Program's Report of the Endocrine Disruptors Low-Dose Peer Review (NTP, 2001). The present report focuses on the reproductive and general toxicology endpoints evaluated. Data obtained in separate evaluations of behavioral, neuroanatomical, neurochemical, and immunological endpoints, as well as the assessment of serum genistein levels, are also discussed to put in better perspective the selection of doses for the multigenerational and chronic studies. Genistein was administered in an irradiated soy- and alfalfa-free diet (Purina 5K96) at exposure concentrations of 0, 5, 25, 100, 250, 625, or 1,250 ppm to 10 vaginal plug-positive, female Sprague-Dawley rats starting on gestation day 7 and continuing throughout pregnancy. These dietary exposure concentrations resulted in ingested doses of approximately 0.3, 1.7, 6.4, 16, 38, and 72 mg genistein/kg body weight to dams in the 5, 25, 100, 250, 625, and 1,250 ppm groups, respectively. Dietary exposure of the dams continued through lactation, during which time ingested doses were approximately 0.6, 3.5, 14, 37, 84, and 167 mg/kg per day. Pups from five litters, culled to eight per litter with an equal sex distribution on postnatal day (PND) 2

  14. NTP toxicity studies of carisoprodol (CAS No. 78-44-4) administered by Gavage to F344/N rats and B6C3F1 mice.

    PubMed

    Chan, P C

    2000-08-01

    [carisoprodol structure: see text] Carisoprodol is a widely used skeletal muscle relaxant and analgesic and is available as a prescription drug. Comparative studies were conducted to determine the toxicity of carisoprodol administered in corn oil and in 0.5% methylcellulose by gavage. Carisoprodol plasma concentrations of rats and mice were measured at the end of the 13-week studies; single-dose plasma carisoprodol analyses were also performed. Genetic toxicity studies were conducted in Salmonella typhimurium, L5178Y mouse lymphoma cells, cultured Chinese hamster ovary cells, and peripheral blood erythrocytes of mice. Groups of 10 male and 10 female F344/N rats received 0, 100, 200, 400, 800, or 1,600 mg carisoprodol per kilogram body weight in corn oil by gavage or 0, 100, 200, 400, or 800 mg/kg carisoprodol in 0.5% methylcellulose by gavage for 13 weeks. Groups of 10 male and 10 female B6C3F1 mice received 0, 75, 150, 300, 600, or 1,200 mg/kg carisoprodol in corn oil by gavage or 0, 600, 1,200, or 1,600 mg/kg carisoprodol in 0.5% methylcellulose by gavage for 13 weeks. Among rats that received carisoprodol in corn oil, survival was similar to that of the vehicle controls. Survival of rats administered carisoprodol in 0.5% methylcellulose was also similar to that of the vehicle controls after adjustment for deaths (two males and one female in the 800 mg/kg group and two females in the 400 mg/kg group). The final mean body weight gain of males administered 1,600 mg/kg carisoprodol in corn oil was significantly less than that of the vehicle controls; the final mean body weights and body weight gains of female rats in the 800 and 1,600 mg/kg groups were significantly greater. In the carisoprodol in 0.5% methylcellulose study, males in the 200 mg/kg group and females in the 100 and 800 mg/kg groups had significantly greater mean body weights and body weight gains than did the vehicle controls. Clinical findings in rats administered carisoprodol in corn oil or in 0

  15. Observation and imitation of actions performed by humans, androids, and robots: an EMG study.

    PubMed

    Hofree, Galit; Urgen, Burcu A; Winkielman, Piotr; Saygin, Ayse P

    2015-01-01

    Understanding others' actions is essential for functioning in the physical and social world. In the past two decades research has shown that action perception involves the motor system, supporting theories that we understand others' behavior via embodied motor simulation. Recently, empirical approach to action perception has been facilitated by using well-controlled artificial stimuli, such as robots. One broad question this approach can address is what aspects of similarity between the observer and the observed agent facilitate motor simulation. Since humans have evolved among other humans and animals, using artificial stimuli such as robots allows us to probe whether our social perceptual systems are specifically tuned to process other biological entities. In this study, we used humanoid robots with different degrees of human-likeness in appearance and motion along with electromyography (EMG) to measure muscle activity in participants' arms while they either observed or imitated videos of three agents produce actions with their right arm. The agents were a Human (biological appearance and motion), a Robot (mechanical appearance and motion), and an Android (biological appearance and mechanical motion). Right arm muscle activity increased when participants imitated all agents. Increased muscle activation was found also in the stationary arm both during imitation and observation. Furthermore, muscle activity was sensitive to motion dynamics: activity was significantly stronger for imitation of the human than both mechanical agents. There was also a relationship between the dynamics of the muscle activity and motion dynamics in stimuli. Overall our data indicate that motor simulation is not limited to observation and imitation of agents with a biological appearance, but is also found for robots. However we also found sensitivity to human motion in the EMG responses. Combining data from multiple methods allows us to obtain a more complete picture of action

  16. Observation and imitation of actions performed by humans, androids, and robots: an EMG study

    PubMed Central

    Hofree, Galit; Urgen, Burcu A.; Winkielman, Piotr; Saygin, Ayse P.

    2015-01-01

    Understanding others’ actions is essential for functioning in the physical and social world. In the past two decades research has shown that action perception involves the motor system, supporting theories that we understand others’ behavior via embodied motor simulation. Recently, empirical approach to action perception has been facilitated by using well-controlled artificial stimuli, such as robots. One broad question this approach can address is what aspects of similarity between the observer and the observed agent facilitate motor simulation. Since humans have evolved among other humans and animals, using artificial stimuli such as robots allows us to probe whether our social perceptual systems are specifically tuned to process other biological entities. In this study, we used humanoid robots with different degrees of human-likeness in appearance and motion along with electromyography (EMG) to measure muscle activity in participants’ arms while they either observed or imitated videos of three agents produce actions with their right arm. The agents were a Human (biological appearance and motion), a Robot (mechanical appearance and motion), and an Android (biological appearance and mechanical motion). Right arm muscle activity increased when participants imitated all agents. Increased muscle activation was found also in the stationary arm both during imitation and observation. Furthermore, muscle activity was sensitive to motion dynamics: activity was significantly stronger for imitation of the human than both mechanical agents. There was also a relationship between the dynamics of the muscle activity and motion dynamics in stimuli. Overall our data indicate that motor simulation is not limited to observation and imitation of agents with a biological appearance, but is also found for robots. However we also found sensitivity to human motion in the EMG responses. Combining data from multiple methods allows us to obtain a more complete picture of action

  17. [Study of the action of sultopride on psychomotor agitation. Apropos of 31 cases].

    PubMed

    Mouren, P; Poinso, Y; Guigou, G

    1976-06-23

    A study of sultopride, a new derivative of the substituted anisamides, has shown that this neuroleptic drug reduces markedly and rapidly psychomotor agitation, especially its aggressive component. It also has an action on hallucinations. Its indications, in order of efficacy, are states of acute agitation, psychotic states during the active phase, acute manifestations of alcoholism and behaviour disorders such as expansivity, impulsivity and agressivity. Its tolerance is good considering its strong activity.

  18. NTP Toxicity Studies of Cyclohexanone Oxime Administered by Drinking Water to B6C3F1 Mice (CAS No. 100-64-1).

    PubMed

    1996-04-01

    Cyclohexanone oxime is used primarily as a captive intermediate in the synthesis of caprolactam for the production of polycaprolactam (Nylon-6) fibers and plastics and also in a variety of industrial applications. Cyclohexanone oxime was selected for study because of the potential for human exposure and the interest in oximes as a chemical class. Toxicity studies of cyclohexanone oxime (approximately 99% pure) were carried out in male and female B6C3F1 mice; the compound was administered in drinking water for 2 weeks or 13 weeks. In addition, the genetic toxicity of cyclohexanone oxime was evaluated by determining mutagenicity in Salmonella typhimurium and induction of chromosomal aberrations in cultured Chinese hamster ovary cells in vitro, with and without S9 activation. The frequency of micronucleated normochromatic erythrocytes in the bone marrow and peripheral blood of mice from the 13-week study was also determined. In the 2-week study, groups of five male and five female mice were given drinking water containing 0, 106, 312, 625, 1,250, or 2,500 ppm cyclohexanone oxime. No deaths occurred, and there was no decrease in weight gain in any group. No gross lesions were observed; there were significant increases in relative spleen weights of males and females in the 2,500 ppm group and increases in the relative liver weight of male mice exposed to 312 ppm or greater. In the 13-week studies, groups of 10 male and 10 female mice were given drinking water containing 625, 1,250, 2,500, 5,000 or 10,000 ppm cyclohexanone oxime. Deaths occurred in the 10,000 ppm groups and weight gain was depressed in males and females given 10,000 ppm and in females given 5,000 ppm. There were significant increases in relative spleen weight at exposure levels of 5,000 and 10,000 ppm and significant increases in the relative liver weights of males and females that received 10,000 ppm. Microscopically, hematopoietic cell proliferation was observed in the spleen of males and females in

  19. Corrective Action Plan for Corrective Action Unit 335: Area 6 Injection Well and Drain Pit Nevada Test Site, Nevada

    SciTech Connect

    K. B. Campbell

    2002-10-01

    This Corrective Action Plan (CAP) has been prepared for Corrective Action Unit (CAU) 335, Area 6 Injection Well and Drain Pit, in accordance with the Federal Facility Agreement and Consent Order (FFACO) (1996). This CAP provides the methodology for implementing the approved corrective action alternative as listed in the Corrective Action Decision Document (CADD). However, there is one modification to the selected alternative. Due to the large area that would require fencing, it is proposed that instead of fencing, an appropriate number of warning signs attached to tee posts be used to delineate the use restriction area. CAU 335 is located in Area 6 of the Nevada Test Site (NTS) which is approximately 105 kilometers (km) (65 miles [mi]) northwest of Las Vegas, Nevada. CAU 335 is located in the Area 6 Well 3 Yard approximately 39 km (24 mi) north of Mercury, on the Mercury Highway and several hundred feet (ft) west along Road 6-06. CAU 335 consists of the following three Corrective Action Sites (CASs): CAS 06-20-01, Drums, Oil Waste, Spill; CAS 06-20-02, 20-inch Cased Hole; CAS 06-23-03, Drain Pit. The site history for CAU 335 is provided in the Corrective Action Investigation Plan (DOE/NV, 2000). Briefly, CAS 06-20-01, was used for storing material that was pumped out of CAS 06-20-02 and placed into four 208-liter (L) (55-gall [gal]) drums. The drums were taken to the NTS Area 5 Hazardous Waste Accumulation Site in 1991. CAS 06-20-01 will be closed with no further action required. Any spills associated with CAS 06-20-01 are addressed and considered part of CAS 06-20-02. CAS 06-20-02 was used for disposal of used motor oil, wastewater, and debris for an undetermined amount of time. In 1991, the casing was emptied of its contents, excavated, and backfilled. CAS 06-23-03 was used as a depository for effluent waste from truck-washing activities from 1960-1991.

  20. Corrective Action Plan for Corrective Action Unit 262: Area 25 Septic Systems and Underground Discharge Point, Nevada Test Site, Nevada

    SciTech Connect

    K. B. Campbell

    2002-06-01

    This Corrective Action Plan (CAP) provides selected corrective action alternatives and proposes the closure methodology for Corrective Action Unit (CAU) 262, Area 25 Septic Systems and Underground Discharge Point. CAU 262 is identified in the Federal Facility Agreement and Consent Order (FFACO) of 1996. Remediation of CAU 262 is required under the FFACO. CAU 262 is located in Area 25 of the Nevada Test Site (NTS), approximately 100 kilometers (km) (62 miles [mi]) northwest of Las Vegas, Nevada. The nine Corrective Action Sites (CASs) within CAU 262 are located in the Nuclear Rocket Development Station complex. Individual CASs are located in the vicinity of the Reactor Maintenance, Assembly, and Disassembly (R-MAD); Engine Maintenance, Assembly, and Disassembly (E-MAD); and Test Cell C compounds. CAU 262 includes the following CASs as provided in the FFACO (1996); CAS 25-02-06, Underground Storage Tank; CAS 25-04-06, Septic Systems A and B; CAS 25-04-07, Septic System; CAS 25-05-03, Leachfield; CAS 25-05-05, Leachfield; CAS 25-05-06, Leachfield; CAS 25-05-08, Radioactive Leachfield; CAS 25-05-12, Leachfield; and CAS 25-51-01, Dry Well. Figures 2, 3, and 4 show the locations of the R-MAD, the E-MAD, and the Test Cell C CASs, respectively. The facilities within CAU 262 supported nuclear rocket reactor engine testing. Activities associated with the program were performed between 1958 and 1973. However, several other projects used the facilities after 1973. A significant quantity of radioactive and sanitary waste was produced during routine operations. Most of the radioactive waste was managed by disposal in the posted leachfields. Sanitary wastes were disposed in sanitary leachfields. Septic tanks, present at sanitary leachfields (i.e., CAS 25-02-06,2504-06 [Septic Systems A and B], 25-04-07, 25-05-05,25-05-12) allowed solids to settle out of suspension prior to entering the leachfield. Posted leachfields do not contain septic tanks. All CASs located in CAU 262 are

  1. Toxicology and carcinogenesis studies of a binary mixture of 3,3',4,4',5-pentachlorobiphenyl (PCB 126) (Cas No. 57465-28-8) and 2,3',4,4',5-pentachlorobiphenyl (PCB 118) (Cas No. 31508-00-6) in female Harlan Sprague-Dawley rats (gavage studies).

    PubMed

    2006-11-01

    DIOXIN TOXIC EQUIVALENCY FACTOR EVALUATION OVERVIEW: Polyhalogenated aromatic hydrocarbons such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) have the ability to bind to and activate the ligand-activated transcription factor, the aryl hydrocarbon receptor (AhR). Structurally related compounds that bind to the AhR and exhibit biological actions similar to TCDD are commonly referred to as "dioxin-like compounds" (DLCs). Ambient human exposure to DLCs occurs through the ingestion of foods containing residues of DLCs that bioconcentrate through the food chain. Due to their lipophilicity and persistence, once internalized they accumulate in body tissue, mainly adipose, resulting in chronic lifetime human exposure. Since human exposure to DLCs always occurs as a complex mixture, the toxic equivalency factor (TEF) methodology has been developed as a mathematical tool to assess the health risk posed by complex mixtures of these compounds. The TEF methodology is a relative potency scheme that ranks the dioxin-like activity of a compound relative to TCDD, which is the most potent congener. This allows for the estimation of the potential dioxin-like activity of a mixture of chemicals, based on a common mechanism of action involving an initial binding of DLCs to the AhR. The toxic equivalency of DLCs was nominated for evaluation because of the widespread human exposure to DLCs and the lack of data on the adequacy of the TEF methodology for predicting relative potency for cancer risk. To address this, the National Toxicology Program conducted a series of 2-year bioassays in female Harlan Sprague-Dawley rats to evaluate the chronic toxicity and carcinogenicity of DLCs and structurally related polychlorinated biphenyls (PCBs) and mixtures of these compounds. Mixtures of polychlorinated biphenyls (PCBs) including 3,3',4,4',5-pentachlorobiphenyl (PCB 126) and 2,3',4,4',5-pentachlorobiphenyl (PCB 118) were produced commercially before 1977 for the electric industry as dielectric

  2. Increasing the efficiency of CRISPR/Cas9-mediated precise genome editing in rats by inhibiting NHEJ and using Cas9 protein.

    PubMed

    Ma, Yuanwu; Chen, Wei; Zhang, Xu; Yu, Lei; Dong, Wei; Pan, Shuo; Gao, Shan; Huang, Xingxu; Zhang, Lianfeng

    2016-07-01

    Precise modifications such as site mutation, codon replacement, insertion or precise targeted deletion are needed for studies of accurate gene function. The CRISPR/Cas9 system has been proved as a powerful tool to generate gene knockout and knockin animals. But the homologous recombination (HR)-directed precise genetic modification mediated by CRISPR/Cas9 is relatively lower compared with nonhomologous end-joining (NHEJ) pathway and extremely expected to be improved. Here, in this study 2 strategies were used to increase the precise genetic modification in rats. Scr7, a DNA ligase IV inhibitor, first identified as an anti-cancer compound, and considered as a potential NHEJ inhibitor, was used to increase the HR-mediated precise genetic modification. Meanwhile, the Cas9 protein instead of mRNA was used to save the mRNA to protein translation step to improve the precise modification efficiency. The Fabp2 and Dbndd1 loci were selected to knockin Cre and CreER(T2), respectively. Our result showed that both Scr7 and Cas9 protein can increase the precise modification. PMID:27163284

  3. Reforming primary science assessment practices: A case study of one teacher's professional development through action research

    NASA Astrophysics Data System (ADS)

    Briscoe, Carol; Wells, Elaine

    2002-05-01

    Calls for reform have suggested that classroom practice can best be changed by teachers who engage in their own research. This interpretive study examines the process of action research and how it contributes to the professional development of a first-grade teacher. The purpose of the study was to explore the research process experienced by the teacher as she examined whether portfolios could be used as an effective means for facilitating and assessing young children's development of science process skills. Data sources included a journal kept by the teacher, documents produced by the teacher and students as part of the portfolio implementation process, hand-written records of teacher's informal interviews with students, and anecdotal records from research team meetings during the study. Data analysis was designed to explore how the teacher's classroom practices and thinking evolved as she engaged in action research and attempted to solve the problems associated with deciding what to assess and how to implement portfolio assessment. We also examined the factors that supported the teacher's learning and change as she progressed through the research process. Data are presented in the form of four assertions that clarify how the action research process was influenced by various personal and contextual factors. Implications address factors that facilitated the teacher as researcher, and how this research project, initiated by the teacher, affected her professional development and professional life.

  4. Examination of mechanisms underlying enhanced memory performance in action video game players: a pilot study.

    PubMed

    Li, Xianchun; Cheng, Xiaojun; Li, Jiaying; Pan, Yafeng; Hu, Yi; Ku, Yixuan

    2015-01-01

    Previous studies have shown enhanced memory performance resulting from extensive action video game playing. The mechanisms underlying the cognitive benefit were investigated in the current study. We presented two types of retro-cues, with variable intervals to memory array (Task 1) or test array (Task 2), during the retention interval in a change detection task. In Task 1, action video game players demonstrated steady performance while non-action video game players showed decreased performance as cues occurred later, indicating their performance difference increased as the cue-to-memory-array intervals became longer. In Task 2, both participant groups increased their performance at similar rates as cues presented later, implying the performance difference in two groups were irrespective of the test-array-to-cue intervals. These findings suggested that memory benefit from game plays is not attributable to the higher ability of overcoming interference from the test array, but to the interactions between the two processes of protection from decay and resistance from interference, or from alternative hypotheses. Implications for future studies were discussed.

  5. Examination of mechanisms underlying enhanced memory performance in action video game players: a pilot study

    PubMed Central

    Li, Xianchun; Cheng, Xiaojun; Li, Jiaying; Pan, Yafeng; Hu, Yi; Ku, Yixuan

    2015-01-01

    Previous studies have shown enhanced memory performance resulting from extensive action video game playing. The mechanisms underlying the cognitive benefit were investigated in the current study. We presented two types of retro-cues, with variable intervals to memory array (Task 1) or test array (Task 2), during the retention interval in a change detection task. In Task 1, action video game players demonstrated steady performance while non-action video game players showed decreased performance as cues occurred later, indicating their performance difference increased as the cue-to-memory-array intervals became longer. In Task 2, both participant groups increased their performance at similar rates as cues presented later, implying the performance difference in two groups were irrespective of the test-array-to-cue intervals. These findings suggested that memory benefit from game plays is not attributable to the higher ability of overcoming interference from the test array, but to the interactions between the two processes of protection from decay and resistance from interference, or from alternative hypotheses. Implications for future studies were discussed. PMID:26136720

  6. Major bacterial lineages are essentially devoid of CRISPR-Cas viral defence systems

    PubMed Central

    Burstein, David; Sun, Christine L.; Brown, Christopher T.; Sharon, Itai; Anantharaman, Karthik; Probst, Alexander J.; Thomas, Brian C.; Banfield, Jillian F.

    2016-01-01

    Current understanding of microorganism–virus interactions, which shape the evolution and functioning of Earth's ecosystems, is based primarily on cultivated organisms. Here we investigate thousands of viral and microbial genomes recovered using a cultivation-independent approach to study the frequency, variety and taxonomic distribution of viral defence mechanisms. CRISPR-Cas systems that confer microorganisms with immunity to viruses are present in only 10% of 1,724 sampled microorganisms, compared with previous reports of 40% occurrence in bacteria and 81% in archaea. We attribute this large difference to the lack of CRISPR-Cas systems across major bacterial lineages that have no cultivated representatives. We correlate absence of CRISPR-Cas with lack of nucleotide biosynthesis capacity and a symbiotic lifestyle. Restriction systems are well represented in these lineages and might provide both non-specific viral defence and access to nucleotides. PMID:26837824

  7. Heritable multiplex genetic engineering in rats using CRISPR/Cas9.

    PubMed

    Ma, Yuanwu; Shen, Bin; Zhang, Xu; Lu, Yingdong; Chen, Wei; Ma, Jing; Huang, Xingxu; Zhang, Lianfeng

    2014-01-01

    The CRISPR/Cas9 system has been proven to be an efficient gene-editing tool for genome modification of cells and organisms. Multiplex genetic engineering in rat holds a bright future for the study of complex disease. Here, we show that this system enables the simultaneous disruption of four genes (ApoE, B2m, Prf1, and Prkdc) in rats in one-step, by co-injection of Cas9 mRNA and sgRNAs into fertilized eggs. We further observed the gene modifications are germline transmittable, and confirmed the off-target mutagenesis and mosaicism are rarely detected by comprehensive analysis. Thus, the CRISPR/Cas9 system makes it possible to efficiently and reliably generate gene knock-out rats.

  8. Transgene-free genome editing by germline injection of CRISPR/Cas RNA.

    PubMed

    Schwartz, Hillel T; Sternberg, Paul W

    2014-01-01

    Genome modification by CRISPR/Cas offers its users the ability to target endogenous sites in the genome for cleavage and for engineering precise genomic changes using template-directed repair, all with unprecedented ease and flexibility of targeting. As such, CRISPR/Cas is just part of a set of recently developed and rapidly improving tools that offer great potential for researchers to functionally access the genomes of organisms that have not previously been extensively used in a laboratory setting. We describe in detail protocols for using CRISPR/Cas to target genes of experimental organisms, in a manner that does not require transformation to obtain transgenic lines and that should be readily applicable to a wide range of previously little-studied species.

  9. Major bacterial lineages are essentially devoid of CRISPR-Cas viral defence systems

    DOE PAGES

    Burstein, David; Sun, Christine L.; Brown, Christopher T.; Sharon, Itai; Anantharaman, Karthik; Probst, Alexander J.; Thomas, Brian C.; Banfield, Jillian F.

    2016-02-03

    Here, current understanding of microorganism–virus interactions, which shape the evolution and functioning of Earth’s ecosystems, is based primarily on cultivated organisms. Here we investigate thousands of viral and microbial genomes recovered using a cultivation independent approach to study the frequency, variety and taxonomic distribution of viral defence mechanisms. CRISPR-Cas systems that confer microorganisms with immunity to viruses are present in only 10% of 1,724 sampled microorganisms, compared with previous reports of 40% occurrence in bacteria and 81% in archaea. We attribute this large difference to the lack of CRISPR-Cas systems across major bacterial lineages that have no cultivated representatives. Wemore » correlate absence of CRISPR-Cas with lack of nucleotide biosynthesis capacity and a symbiotic lifestyle. Restriction systems are well represented in these lineages and might provide both non-specific viral defence and access to nucleotides.« less

  10. Transgene-free genome editing by germline injection of CRISPR/Cas RNA.

    PubMed

    Schwartz, Hillel T; Sternberg, Paul W

    2014-01-01

    Genome modification by CRISPR/Cas offers its users the ability to target endogenous sites in the genome for cleavage and for engineering precise genomic changes using template-directed repair, all with unprecedented ease and flexibility of targeting. As such, CRISPR/Cas is just part of a set of recently developed and rapidly improving tools that offer great potential for researchers to functionally access the genomes of organisms that have not previously been extensively used in a laboratory setting. We describe in detail protocols for using CRISPR/Cas to target genes of experimental organisms, in a manner that does not require transformation to obtain transgenic lines and that should be readily applicable to a wide range of previously little-studied species. PMID:25398352

  11. Delivery and Specificity of CRISPR-Cas9 Genome Editing Technologies for Human Gene Therapy.

    PubMed

    Gori, Jennifer L; Hsu, Patrick D; Maeder, Morgan L; Shen, Shen; Welstead, G Grant; Bumcrot, David

    2015-07-01

    Genome editing using the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated 9 (Cas9) technology is revolutionizing the study of gene function and likely will give rise to an entire new class of therapeutics for a wide range of diseases. Achieving this goal requires not only characterization of the technology for efficacy and specificity but also optimization of its delivery to the target cells for each disease indication. In this review we survey the various methods by which the CRISPR-Cas9 components have been delivered to cells and highlight some of the more clinically relevant approaches. Additionally, we discuss the methods available for assessing the specificity of Cas9 editing; an important safety consideration for development of the technology.

  12. CRISPR/Cas9-Mediated Genomic Deletion of the Beta-1, 4 N-acetylgalactosaminyltransferase 1 Gene in Murine P19 Embryonal Carcinoma Cells Results in Low Sensitivity to Botulinum Neurotoxin Type C

    PubMed Central

    Tsukamoto, Kentaro; Ozeki, Chikako; Kohda, Tomoko; Tsuji, Takao

    2015-01-01

    Botulinum neurotoxins produced by Clostridium botulinum cause flaccid paralysis by inhibiting neurotransmitter release at peripheral nerve terminals. Previously, we found that neurons derived from the murine P19 embryonal carcinoma cell line exhibited high sensitivity to botulinum neurotoxin type C. In order to prove the utility of P19 cells for the study of the intracellular mechanism of botulinum neurotoxins, ganglioside-knockout neurons were generated by deletion of the gene encoding beta-1,4 N-acetylgalactosaminyltransferase 1 in P19 cells using the clustered regularly interspaced short palindromic repeats combined with Cas9 (CRISPR/Cas9) system. By using this system, knockout cells could be generated more easily than with previous methods. The sensitivity of the generated beta-1,4 N-acetylgalactosaminyltransferase 1-depleted P19 neurons to botulinum neurotoxin type C was decreased considerably, and the exogenous addition of the gangliosides GD1a, GD1b, and GT1b restored the susceptibility of P19 cells to botulinum neurotoxin type C. In particular, addition of a mixture of these three ganglioside more effectively recovered the sensitivity of knockout cells compared to independent addition of GD1a, GD1b, or GT1b. Consequently, the genome-edited P19 cells generated by the CRISPR/Cas9 system were useful for identifying and defining the intracellular molecules involved in the toxic action of botulinum neurotoxins. PMID:26177297

  13. Expanding the catalog of cas genes with metagenomes.

    PubMed

    Zhang, Quan; Doak, Thomas G; Ye, Yuzhen

    2014-02-01

    The CRISPR (clusters of regularly interspaced short palindromic repeats)-Cas adaptive immune system is an important defense system in bacteria, providing targeted defense against invasions of foreign nucleic acids. CRISPR-Cas systems consist of CRISPR loci and cas (CRISPR-associated) genes: sequence segments of invaders are incorporated into host genomes at CRISPR loci to generate specificity, while adjacent cas genes encode proteins that mediate the defense process. We pursued an integrated approach to identifying putative cas genes from genomes and metagenomes, combining similarity searches with genomic neighborhood analysis. Application of our approach to bacterial genomes and human microbiome datasets allowed us to significantly expand the collection of cas genes: the sequence space of the Cas9 family, the key player in the recently engineered RNA-guided platforms for genome editing in eukaryotes, is expanded by at least two-fold with metagenomic datasets. We found genes in cas loci encoding other functions, for example, toxins and antitoxins, confirming the recently discovered potential of coupling between adaptive immunity and the dormancy/suicide systems. We further identified 24 novel Cas families; one novel family contains 20 proteins, all identified from the human microbiome datasets, illustrating the importance of metagenomics projects in expanding the diversity of cas genes.

  14. Protein engineering of Cas9 for enhanced function

    PubMed Central

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

    2015-01-01

    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 challenges regarding Cas9 specificity, efficiency, fusion protein function, and spatiotemporal control within the cell remain. In this work, we develop a platform for constructing novel proteins to address these open questions. We demonstrate methods to either screen or select active Cas9 mutants and use the screening technique to isolate functional Cas9 variants with a heterologous PDZ domain inserted directly into the protein. As a proof of concept, these methods lay the groundwork for the future construction of diverse Cas9 proteins. Straightforward and accessible techniques for genetic editing are helping to elucidate biology in new and exciting ways; a platform to engineer new functionalities into Cas9 will help forge the next generation of genome modifying tools. PMID:25398355

  15. The CARN/ARNA Inaugural Study Day Inquiry: What Happens to Action Research after the Master's Degree?

    ERIC Educational Resources Information Center

    Shosh, Joseph M.; McAteer, Mary

    2016-01-01

    The Collaborative Action Research Network (CARN) held its first American study day on the east coast of the United States in conjunction with the Action Research Network of the Americas (ARNA) 2014 conference in Bethlehem, Pennsylvania, USA. Study day participants visited three American secondary schools, one each in Pennsylvania, New York, and…

  16. Participatory Action Research and the Reconstruction of Teachers' Practical Thinking: Lesson Studies and Core Reflection. An Experience in Spain

    ERIC Educational Resources Information Center

    Perez, Angel I.; Soto, Encarnacion; Servan, M. Jose

    2010-01-01

    Following the thoughts and topics we have discussed and worked on for a very long time with Bridget Somekh, we would like to present the theoretical relationship between lesson studies, action research and practical knowledge in teacher education. Inspired by the pedagogical philosophy of lesson studies, participatory action research, and core…

  17. Annotation and Classification of CRISPR-Cas Systems.

    PubMed

    Makarova, Kira S; Koonin, Eugene V

    2015-01-01

    The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas (CRISPR-associated proteins) is a prokaryotic adaptive immune system that is represented in most archaea and many bacteria. Among the currently known prokaryotic defense systems, the CRISPR-Cas genomic loci show unprecedented complexity and diversity. Classification of CRISPR-Cas variants that would capture their evolutionary relationships to the maximum possible extent is essential for comparative genomic and functional characterization of this theoretically and practically important system of adaptive immunity. To this end, a multipronged approach has been developed that combines phylogenetic analysis of the conserved Cas proteins with comparison of gene repertoires and arrangements in CRISPR-Cas loci. This approach led to the current classification of CRISPR-Cas systems into three distinct types and ten subtypes for each of which signature genes have been identified. Comparative genomic analysis of the CRISPR-Cas systems in new archaeal and bacterial genomes performed over the 3 years elapsed since the development of this classification makes it clear that new types and subtypes of CRISPR-Cas need to be introduced. Moreover, this classification system captures only part of the complexity of CRISPR-Cas organization and evolution, due to the intrinsic modularity and evolutionary mobility of these immunity systems, resulting in numerous recombinant variants. Moreover, most of the cas genes evolve rapidly, complicating the family assignment for many Cas proteins and the use of family profiles for the recognition of CRISPR-Cas subtype signatures. Further progress in the comparative analysis of CRISPR-Cas systems requires integration of the most sensitive sequence comparison tools, protein structure comparison, and refined approaches for comparison of gene neighborhoods.

  18. Precision Targeted Mutagenesis via Cas9 Paired Nickases in Rice

    PubMed Central

    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, off-target mutations can be suppressed by using Cas9 paired nickases together with paired guide RNAs (gRNAs). However, the performance of Cas9 paired nickases has not yet been fully assessed in plants. Here, we analyzed on- and off-target mutation frequency in rice calli and regenerated plants using Cas9 nuclease or Cas9 nickase with paired gRNAs. When Cas9 paired nickases were used, off-target mutations were fully suppressed in rice calli and regenerated plants. However, on-target mutation frequency also decreased compared with that induced by the Cas9 paired nucleases system. Since the gRNA sequence determines specific binding of Cas9 protein–gRNA ribonucleoproteins at the targeted sequence, the on-target mutation frequency of Cas9 paired nickases depends on the design of paired gRNAs. Our results suggest that a combination of gRNAs that can induce mutations at high efficiency with Cas9 nuclease should be used together with Cas9 nickase. Furthermore, we confirmed that a combination of gRNAs containing a one nucleotide (1 nt) mismatch toward the target sequence could not induce mutations when expressed with Cas9 nickase. Our results clearly show the effectiveness of Cas9 paired nickases in delivering on-target specific mutations. PMID:26936792

  19. ParticipACTION: Baseline assessment of the capacity available to the 'New ParticipACTION': A qualitative study of Canadian organizations

    PubMed Central

    2009-01-01

    Background Evaluation of the original ParticipACTION campaign effects focused on individual awareness, recall, and understanding. Less studied has been the impact such campaigns have had on the broader organizational capacity to mobilize and advocate for physical activity. With the relaunch of ParticipACTION, the purpose of this study was to qualitatively explore baseline organizational capacity to promote physical activity messages, programs, and services within the Canadian context. Methods Using a purposeful sampling strategy, we conducted semi-structured telephone interviews with 49 key informants representing a range of national, provincial, and local organizations with a mandate to promote physical activity. Interview data were analysed using a thematic analytic approach. Results Key informants painted a generally positive picture of current organizational capacity to promote physical activity messages, programs, and services in Canada. Will and leadership were clear strengths while infrastructure limitations remained the greatest concern. Some specific challenges included: 1) funding issues: the absence of core funding in a climate of shifting funding priorities; 2) the difficulty of working without a national physical activity policy (lack of leadership); 3) inconsistent provincial and educational sector level policies; and 4) a persistent focus on obesity rather than physical inactivity. Conclusion The data generated here can be utilized to monitor the future impact of ParticipACTION on enhancing and utilizing this organizational capacity. A range of indicators are suggested that could be used to illustrate ParticipACTION's impact on the broad field of physical activity promotion in the future. PMID:19995458

  20. Closure Report for Corrective Action Unit 548: Areas 9, 10, 18, 19, and 20 Housekeeping Sites, Nevada National Security Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2012-08-27

    This Closure Report (CR) documents closure activities for Corrective Action Unit (CAU) 548, Areas 9, 10, 18, 19, and 20 Housekeeping Sites, and complies with the Federal Facility Agreement and Consent Order (FFACO) that was agreed to by the State of Nevada; the U.S. Department of Energy (DOE), Environmental Management; the U.S. Department of Defense; and DOE, Legacy Management (FFACO, 1996 as amended). CAU 548 consists of the following Corrective Action Sites (CASs), located in Areas 9, 10, 12, 18, 19, and 20 of the Nevada National Security Site: · CAS 09-99-02, Material Piles (2) · CAS 09-99-04, Wax, Paraffin · CAS 09-99-05, Asbestos, Vermiculite · CAS 09-99-07, Tar Spill · CAS 10-22-02, Drums · CAS 10-22-05, Gas Block · CAS 10-22-07, Gas Block · CAS 10-22-34, Drum · CAS 10-22-38, Drum; Cable · CAS 12-99-04, Epoxy Tar Spill · CAS 12-99-08, Cement Spill · CAS 18-14-01, Transformers (3) · CAS 19-22-01, Drums · CAS 19-22-11, Gas Block (2) · CAS 19-44-01, Fuel Spill · CAS 20-22-07, Drums (2) · CAS 20-22-09, Drums (3) · CAS 20-22-14, Drums (2) · CAS 20-22-16, Drums (2) · CAS 20-24-09, Battery Closure activities began in July 2011 and were completed in December 2011 and included removal and disposal of material piles, spills, sanitary debris, a lead acid battery, lead and steel shot, and stained soil. Activities were conducted according to the Sectored Clean-up Work Plan for Housekeeping Category Waste Sites (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2003). Closure activities generated sanitary waste, hydrocarbon waste, low-level waste, hazardous waste, and mixed waste. Some wastes exceeded land disposal limits and required offsite treatment prior to disposal. Other wastes met land disposal restrictions and were disposed in appropriate onsite or offsite landfills. NNSA/NSO requests the following: · A Notice of Completion from the Nevada Division of Environmental Protection to NNSA/NSO for

  1. Carcinogenesis Bioassay of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (CAS No. 1746-01-6) in Swiss-Webster Mice (Dermal Study).

    PubMed

    1982-02-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin occurs as a highly toxic impurity found in herbicides containing 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and 2,4,5-T- derivatives, as well as in other chemicals synthesized using 2,4,5-trichlorophenol. The herbicide 2,4,5-T has been marketed in the United States since 1948. Production increased sharply between 1960 and 1970 when a 1:1 mixture of 2,4,5-T and 2,4-dichlorophenoxyacetic acid (2,4-D) was used as a defoliant in Vietnam under the names of "herbicide agent orange, herbicide orange, agent orange, and orange". During this 10-year period, about 106 million pounds of 2,4,5-T were sprayed. A carcinogenesis bioassay was conducted by applying an acetone suspension of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to the clipped backs of 30 male and female Swiss-Webster mice 3 days per week for 99 or 104 weeks. Similar groups were pretreated with 1 application of 50 &mgr;g dimethylbenzanthracene (DMBA) in 0.1 ml acetone 1 week before TCDD administration began. Female mice received 0.005 &mgr;g TCDD per application, and the male mice received 0.001 &mgr;g TCDD. As vehicle controls, 45 mice of each sex received 0.1 ml acetone three times per week. Thirty animals of each sex were used as untreated controls. Throughout the bioassay, mean body weights of the male and female mice administered TCDD, or TCDD following DMBA, were essentially the same as those of the corresponding vehicle control group. Mean body weights of dosed and vehicle control groups of males were less thanthose of the untreated control group throughout the study; for the females, mean body weights were less than the untreated controls during the first 80 weeks. In female mice, the incidences of fibrosarcoma in the integumentary system in dosed groups with TCDD were significantly (P=0.007) higher than that in the corresponding controls (2/41, 5%; 8/27, 30%). An increase in the same tumor type, although not statistically significant (P=0.084), was also observed in

  2. Carcinogenesis Bioassay of 2,3,7,8-Tetrachlorodibenzo-p-dioxin (CAS No. 1746-01-6) in Swiss-Webster Mice (Dermal Study).

    PubMed

    1982-02-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin occurs as a highly toxic impurity found in herbicides containing 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and 2,4,5-T- derivatives, as well as in other chemicals synthesized using 2,4,5-trichlorophenol. The herbicide 2,4,5-T has been marketed in the United States since 1948. Production increased sharply between 1960 and 1970 when a 1:1 mixture of 2,4,5-T and 2,4-dichlorophenoxyacetic acid (2,4-D) was used as a defoliant in Vietnam under the names of "herbicide agent orange, herbicide orange, agent orange, and orange". During this 10-year period, about 106 million pounds of 2,4,5-T were sprayed. A carcinogenesis bioassay was conducted by applying an acetone suspension of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to the clipped backs of 30 male and female Swiss-Webster mice 3 days per week for 99 or 104 weeks. Similar groups were pretreated with 1 application of 50 &mgr;g dimethylbenzanthracene (DMBA) in 0.1 ml acetone 1 week before TCDD administration began. Female mice received 0.005 &mgr;g TCDD per application, and the male mice received 0.001 &mgr;g TCDD. As vehicle controls, 45 mice of each sex received 0.1 ml acetone three times per week. Thirty animals of each sex were used as untreated controls. Throughout the bioassay, mean body weights of the male and female mice administered TCDD, or TCDD following DMBA, were essentially the same as those of the corresponding vehicle control group. Mean body weights of dosed and vehicle control groups of males were less thanthose of the untreated control group throughout the study; for the females, mean body weights were less than the untreated controls during the first 80 weeks. In female mice, the incidences of fibrosarcoma in the integumentary system in dosed groups with TCDD were significantly (P=0.007) higher than that in the corresponding controls (2/41, 5%; 8/27, 30%). An increase in the same tumor type, although not statistically significant (P=0.084), was also observed in

  3. Breaking-Cas-interactive design of guide RNAs for CRISPR-Cas experiments for ENSEMBL genomes.

    PubMed

    Oliveros, Juan C; Franch, Mònica; Tabas-Madrid, Daniel; San-León, David; Montoliu, Lluis; Cubas, Pilar; Pazos, Florencio

    2016-07-01

    The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unprecedented accuracy and specificity by using RNA-guided nucleases. A critical point when planning a CRISPR/Cas experiment is the design of the guide RNA (gRNA), which directs the nuclease and associated machinery to the desired genomic location. This gRNA has to fulfil the requirements of the nuclease and lack homology with other genome sites that could lead to off-target effects. Here we introduce the Breaking-Cas system for the design of gRNAs for CRISPR/Cas experiments, including those based in the Cas9 nuclease as well as others recently introduced. The server has unique features not available in other tools, including the possibility of using all eukaryotic genomes available in ENSEMBL (currently around 700), placing variable PAM sequences at 5' or 3' and setting the guide RNA length and the scores per nucleotides. It can be freely accessed at: http://bioinfogp.cnb.csic.es/tools/breakingcas, and the code is available upon request.

  4. Closure Report for Corrective Action Unit 562: Waste Systems, Nevada National Security Site, Nevada

    SciTech Connect

    NSTec Environmental Restoration

    2012-08-15

    This Closure Report (CR) presents information supporting closure of Corrective Action Unit (CAU) 562, Waste Systems, and provides documentation supporting the completed corrective actions and confirmation that closure objectives for CAU 562 were met. This CR complies with the requirements of the Federal Facility Agreement and Consent Order (FFACO) that was agreed to by the State of Nevada; the U.S. Department of Energy (DOE), Environmental Management; the U.S. Department of Defense; and DOE, Legacy Management (FFACO, 1996 as amended). CAU 562 consists of the following 13 Corrective Action Sites (CASs), located in Areas 2, 23, and 25 of the Nevada National Security Site: · CAS 02-26-11, Lead Shot · CAS 02-44-02, Paint Spills and French Drain · CAS 02-59-01, Septic System · CAS 02-60-01, Concrete Drain · CAS 02-60-02, French Drain · CAS 02-60-03, Steam Cleaning Drain · CAS 02-60-04, French Drain · CAS 02-60-05, French Drain · CAS 02-60-06, French Drain · CAS 02-60-07, French Drain · CAS 23-60-01, Mud Trap Drain and Outfall · CAS 23-99-06, Grease Trap · CAS 25-60-04, Building 3123 Outfalls Closure activities began in October 2011 and were completed in April 2012. Activities were conducted according to the Corrective Action Plan for CAU 562 (U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office [NNSA/NSO], 2011). The corrective actions included No Further Action and Clean Closure. Closure activities generated sanitary waste and hazardous waste. Some wastes exceeded land disposal limits and required offsite treatment prior to disposal. Other wastes met land disposal restrictions and were disposed in appropriate onsite or offsite landfills. NNSA/NSO requests the following: · A Notice of Completion from the Nevada Division of Environmental Protection to NNSA/NSO for closure of CAU 562 · The transfer of CAU 562 from Appendix III to Appendix IV, Closed Corrective Action Units, of the FFACO

  5. CERCLA interim action at the Par Pond unit: A case study

    SciTech Connect

    Hickey, H.M.; Matthews, S.S.; Neal, L.W.; Weiss, W.R.

    1993-11-01

    The Par Pond unit designated under CERCLA consists of sediments within a Savannah River Site (SRS) cooling water reservoir. The sediments are contaminated with radionuclides and nonradioactive constituents from nuclear production reactor operations. The mercury in Par Pond is believed to have originated from the Savannah River. Because of Par Pond Dam safety Issues, the water level of the reservoir was drawn down, exposing more than 1300 acres of contaminated sediments and triggering the need for CERCLA interim remedial action. This paper presents the interim action approach taken with Par Pond as a case study. The approach considered the complexity of the Par Pond ecosystem, the large size of Par Pond, the volume of contaminated sediments, and the institutional controls existing at SRS. The Environmental Protection Agency (EPA) considers units with large volumes of low-concentration wastes, as is the case with Par Pond, to be {open_quotes}special sites.{close_quotes} Accordingly, EPA guidance establishes that the range of alternatives developed focus primarily on containment options and other remedial approaches that mitigate potential risks associated with the {open_quotes}special site.{close_quotes} The remedial alternatives, according to EPA, are not to be prohibitively expensive or difficult to implement. This case study also is representative of the types of issues that will need to be addressed within the Department of Energy (DOE) complex as nuclear facilities are transitioned to inactive status and corrective/remedial actions are warranted.

  6. Visual exploration patterns of human figures in action: an eye tracker study with art paintings

    PubMed Central

    Villani, Daniela; Morganti, Francesca; Cipresso, Pietro; Ruggi, Simona; Riva, Giuseppe; Gilli, Gabriella

    2015-01-01

    Art exploration is a complex process conditioned by factors at different levels and includes both basic visual principles and complex cognitive factors. The human figure is considered a critical factor attracting the attention in art painting. Using an eye-tracking methodology, the goal of this study was to explore different elements of the human figure performing an action (face and body parts in action) in complex social scenes characterized by different levels of social interaction between agents depicted in scenes (individual vs. social). The sample included 44 laypersons, and the stimuli consisted of 10 fine art paintings representing the figurative style of classical art. The results revealed different scanning patterns of the human figure elements related to the level of social interaction of agents depicted in the scene. The agents’ face attracted eye movements in social interaction scenes while the agents’ body parts attracted eye movements only when the agents were involved in individual actions. These processes were confirmed specifically in participants with high empathic abilities who became immediately fixated on faces to develop a mimetic engagement with other agents. Future studies integrating other measures would help confirm the results obtained and strengthen their implication for embodiment processes. PMID:26579021

  7. Visual exploration patterns of human figures in action: an eye tracker study with art paintings.

    PubMed

    Villani, Daniela; Morganti, Francesca; Cipresso, Pietro; Ruggi, Simona; Riva, Giuseppe; Gilli, Gabriella

    2015-01-01

    Art exploration is a complex process conditioned by factors at different levels and includes both basic visual principles and complex cognitive factors. The human figure is considered a critical factor attracting the attention in art painting. Using an eye-tracking methodology, the goal of this study was to explore different elements of the human figure performing an action (face and body parts in action) in complex social scenes characterized by different levels of social interaction between agents depicted in scenes (individual vs. social). The sample included 44 laypersons, and the stimuli consisted of 10 fine art paintings representing the figurative style of classical art. The results revealed different scanning patterns of the human figure elements related to the level of social interaction of agents depicted in the scene. The agents' face attracted eye movements in social interaction scenes while the agents' body parts attracted eye movements only when the agents were involved in individual actions. These processes were confirmed specifically in participants with high empathic abilities who became immediately fixated on faces to develop a mimetic engagement with other agents. Future studies integrating other measures would help confirm the results obtained and strengthen their implication for embodiment processes.

  8. Atomic force microscopy study of the antimicrobial action of Sushi peptides on Gram negative bacteria.

    PubMed

    Li, A; Lee, P Y; Ho, B; Ding, J L; Lim, C T

    2007-03-01

    The antibacterial effect of the endotoxin-binding Sushi peptides against Gram-negative bacteria (GNB) is investigated in this study. Similar characteristics observed for Atomic force microscopy (AFM) images of peptide-treated Escherichia coli and Pseudomonas aeruginosa suggest that the Sushi peptides (S3) evoke comparable mechanism of action against different strains of GNB. The results also indicate that the Sushi peptides appear to act in three stages: damage of the bacterial outer membrane, permeabilization of the inner membrane and disintegration of both membranes. The AFM approach has provided vivid and detailed close-up images of the GNB undergoing various stages of antimicrobial peptide actions at the nanometer scale. The AFM results support our hypothesis that the S3 peptide perturbs the GNB membrane via the "carpet-model" and thus, provide important insights into their antimicrobial mechanisms.

  9. [The influence of selected antibiotics on the central action of aminophyllines--experimental studies].

    PubMed

    Szmygin, K

    1996-01-01

    Methylxanthines and some antibiotics can cause side effects, provoked by their central action, e.g. seizures. The epileptogenic effects of given drugs can be intensified during combined treatment, as a result of pharmacological interactions. In the present study the author investigated the influence of some commonly used antibiotics: benzylpenicillin, cefuroxime, doxycycline and amikacin upon central activity of methylxanthines in mice. The obtained results suggest, that all tested antibiotics, mainly benzylpenicillin, enhanced epileptogenicity of aminophylline in chemical seizures test. benzylpenicillin as only one among chosen antibiotics presented her own convulsant activity. During electrostimulation test, benzylpenicillin, doxycycline and amikacin intensified convulsions induced by methylxanthines. Only cefuroxime had no influence upon central action of methylxanthines in that experiments. Analysis of drugs' plasma levels, with using immunofluorescence methods, excluded pharmacokinetic interactions between them. Results of present investigation indicate, that there is a possibility of intensification of drugs' convulsant activity during combined treatment-aminophylline with some antibiotics in medical practice.

  10. Visualization of specific DNA sequences in living mouse embryonic stem cells with a programmable fluorescent CRISPR/Cas system.

    PubMed

    Anton, Tobias; Bultmann, Sebastian; Leonhardt, Heinrich; Markaki, Yolanda

    2014-01-01

    Labeling and tracing of specific sequences in living cells has been a major challenge in studying the spatiotemporal dynamics of native chromatin. Here we repurposed the prokaryotic CRISPR/Cas adaptive immunity system to specifically detect endogenous genomic loci in mouse embryonic stem cells. We constructed a catalytically inactive version of the Cas9 endonuclease, fused it with eGFP (dCas9-eGFP) and co-expressed small guide RNAs (gRNAs) to target pericentric, centric, and telomeric repeats, which are enriched in distinct nuclear structures. With major satellite specific gRNAs we obtained a characteristic chromocenter (CC) pattern, while gRNAs targeting minor satellites and telomeres highlighted smaller foci coinciding with centromere protein B (CENP-B) and telomeric repeat-binding factor 2 (TRF2), respectively. DNA sequence specific labeling by gRNA/dCas9-eGFP complexes was directly shown with 3D-fluorescent in situ hybridization (3D-FISH). Structured illumination microscopy (3D-SIM) of gRNA/dCas9-eGFP expressing cells revealed chromatin ultrastructures and demonstrated the potential of this approach for chromatin conformation studies by super resolution microscopy. This programmable dCas9 labeling system opens new perspectives to study functional nuclear architecture.

  11. Visualization of specific DNA sequences in living mouse embryonic stem cells with a programmable fluorescent CRISPR/Cas system.

    PubMed

    Anton, Tobias; Bultmann, Sebastian; Leonhardt, Heinrich; Markaki, Yolanda

    2014-01-01

    Labeling and tracing of specific sequences in living cells has been a major challenge in studying the spatiotemporal dynamics of native chromatin. Here we repurposed the prokaryotic CRISPR/Cas adaptive immunity system to specifically detect endogenous genomic loci in mouse embryonic stem cells. We constructed a catalytically inactive version of the Cas9 endonuclease, fused it with eGFP (dCas9-eGFP) and co-expressed small guide RNAs (gRNAs) to target pericentric, centric, and telomeric repeats, which are enriched in distinct nuclear structures. With major satellite specific gRNAs we obtained a characteristic chromocenter (CC) pattern, while gRNAs targeting minor satellites and telomeres highlighted smaller foci coinciding with centromere protein B (CENP-B) and telomeric repeat-binding factor 2 (TRF2), respectively. DNA sequence specific labeling by gRNA/dCas9-eGFP complexes was directly shown with 3D-fluorescent in situ hybridization (3D-FISH). Structured illumination microscopy (3D-SIM) of gRNA/dCas9-eGFP expressing cells revealed chromatin ultrastructures and demonstrated the potential of this approach for chromatin conformation studies by super resolution microscopy. This programmable dCas9 labeling system opens new perspectives to study functional nuclear architecture. PMID:24637835

  12. Application of the genome editing tool CRISPR/Cas9 in non-human primates.

    PubMed

    Luo, Xin; Li, Min; Su, Bing

    2016-07-18

    In the past three years, RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeats (CRISPR) adaptive immune system has been used to facilitate efficient genome editing in many model and non-model animals. However, its application in nonhuman primates is still at the early stage, though in view of the similarities in anatomy, physiology, behavior and genetics, closely related nonhuman primates serve as optimal models for human biology and disease studies. In this review, we summarize the current proceedings of gene editing using CRISPR/Cas9 in nonhuman primates. PMID:27469252

  13. CRISPR/Cas9 for Human Genome Engineering and Disease Research.

    PubMed

    Xiong, Xin; Chen, Meng; Lim, Wendell A; Zhao, Dehua; Qi, Lei S

    2016-08-31

    The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system, a versatile RNA-guided DNA targeting platform, has been revolutionizing our ability to modify, manipulate, and visualize the human genome, which greatly advances both biological research and therapeutics development. Here, we review the current development of CRISPR/Cas9 technologies for gene editing, transcription regulation, genome imaging, and epigenetic modification. We discuss the broad application of this system to the study of functional genomics, especially genome-wide genetic screening, and to therapeutics development, including establishing disease models, correcting defective genetic mutations, and treating diseases.

  14. CRISPR/Cas9 for Human Genome Engineering and Disease Research.

    PubMed

    Xiong, Xin; Chen, Meng; Lim, Wendell A; Zhao, Dehua; Qi, Lei S

    2016-08-31

    The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system, a versatile RNA-guided DNA targeting platform, has been revolutionizing our ability to modify, manipulate, and visualize the human genome, which greatly advances both biological research and therapeutics development. Here, we review the current development of CRISPR/Cas9 technologies for gene editing, transcription regulation, genome imaging, and epigenetic modification. We discuss the broad application of this system to the study of functional genomics, especially genome-wide genetic screening, and to therapeutics development, including establishing disease models, correcting defective genetic mutations, and treating diseases. PMID:27216776

  15. Application of the genome editing tool CRISPR/Cas9 in non-human primates

    PubMed Central

    LUO, Xin; LI, Min; SU, Bing

    2016-01-01

    In the past three years, RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeats (CRISPR) adaptive immune system has been used to facilitate efficient genome editing in many model and non-model animals. However, its application in nonhuman primates is still at the early stage, though in view of the similarities in anatomy, physiology, behavior and genetics, closely related nonhuman primates serve as optimal models for human biology and disease studies. In this review, we summarize the current proceedings of gene editing using CRISPR/Cas9 in nonhuman primates. PMID:27469252

  16. Applications of the CRISPR-Cas9 system in cancer biology

    PubMed Central

    Sánchez-Rivera, Francisco J.; Jacks, Tyler

    2015-01-01

    Preface The prokaryotic type II clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system is rapidly revolutionizing the field of genetic engineering, allowing researchers to alter the genomes of a large variety of organisms with relative ease. Experimental approaches based on this versatile technology have the potential to transform the field of cancer genetics. Here we review current approaches based on CRISPR-Cas9 for functional studies of cancer genes, with emphasis on its applicability for the development of the next-generation models of human cancer. PMID:26040603

  17. Application of the genome editing tool CRISPR/Cas9 in non-human primates.

    PubMed

    Luo, Xin; Li, Min; Su, Bing

    2016-07-18

    In the past three years, RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeats (CRISPR) adaptive immune system has been used to facilitate efficient genome editing in many model and non-model animals. However, its application in nonhuman primates is still at the early stage, though in view of the similarities in anatomy, physiology, behavior and genetics, closely related nonhuman primates serve as optimal models for human biology and disease studies. In this review, we summarize the current proceedings of gene editing using CRISPR/Cas9 in nonhuman primates.

  18. The Influence of the CAS Standards on Academic Advisors and Advising Programs

    ERIC Educational Resources Information Center

    Keeling, Sarah

    2010-01-01

    Research on the influence of the Council for the Advancement of Standards in Higher Education (CAS) on academic advising is limited. Using a comparative case study method, I respond to this research gap by exploring how the standards influence practices of academic advising programs. Study results indicate that participating advisors knew little…

  19. Entrepreneurial Learning through Action: A Case Study of the Six-Squared Program

    ERIC Educational Resources Information Center

    Pittaway, Luke; Missing, Caroline; Hudson, Nigel; Maragh, Dean

    2009-01-01

    This paper explores the role of "action" in entrepreneurial learning and illustrates how programs designed to support action learning can enhance management development in entrepreneurial businesses. The paper begins by exploring action learning and the way "action" is conceived in different types of program. In the second part, the paper details…

  20. CSE1L/CAS, a microtubule-associated protein, inhibits taxol (paclitaxel)-induced apoptosis but enhances cancer cell apoptosis induced by various chemotherapeutic drugs.

    PubMed

    Liao, Ching-Fong; Luo, Shue-Fen; Shen, Tzu-Yun; Lin, Chin-Huang; Chien, Jung-Tsun; Du, Shin-Yi; Jiang, Ming-Chung

    2008-03-31

    CSE1L/CAS, a microtubule-associated, cellular apoptosis susceptibility protein, is highly expressed in various cancers. Microtubules are the target of paclitaxel-induced apoptosis. We studied the effects of increased or reduced CAS expression on cancer cell apoptosis induced by chemotherapeutic drugs including paclitaxel. Our results showed that CAS overexpression enhanced apoptosis induced by doxorubicin, 5-fluorouracil, cisplatin, and tamoxifen, but inhibited paclitaxel-induced apoptosis of cancer cells. Reductions in CAS produced opposite results. CAS overexpression enhanced p53 accumulation induced by doxorubicin, 5-fluorouracil, cisplatin, tamoxifen, and etoposide. CAS was associated with alpha-tubulin and beta-tubulin and enhanced the association between alpha-tubulin and beta-tubulin. Paclitaxel can induce G2/M phase cell cycle arrest and microtubule aster formation during apoptosis induction, but CAS overexpression reduced paclitaxel-induced G2/M phase cell cycle arrest and microtubule aster formation. Our results indicate that CAS may play an important role in regulating the cytotoxicities of chemotherapeutic drugs used in cancer chemotherapy against cancer cells.

  1. [Use of photonuclear reactions for studying the biological action of slow heavy ions].

    PubMed

    Kapchigashev, S P

    1984-01-01

    Since ranges of low-energy heavy ions are too little the study of their action on cells is only possible using indirect irradiation, for instance, with neurons which form low--energy heavy ions uniformly inside a cell. In our studies we used photonuclear reactions which occur upon irradiation of a bio-object with high energy bremsstrahlung. The advantage of this method over the method based on the usage of neutrons has been demonstrated. The data are submitted obtained after exposure of yeast cells to photonuclear products. PMID:6324271

  2. Effect of eplerenone on insulin action in essential hypertension: a randomised, controlled, crossover study.

    PubMed

    McMurray, E M; Wallace, I R; Ennis, C; Hunter, S J; Atkinson, A B; Bell, P M

    2014-10-01

    An association exists between hyperaldosteronism, hypertension and impaired insulin action. Eplerenone is a selective mineralocorticoid receptor antagonist; however, little is known about its effects on insulin action. The aim of this study was to determine the effect of eplerenone on insulin action in hypertensive adults, using the hyperinsulinaemic euglycaemic clamp. A randomised, controlled, double-blind, crossover design was employed. After a 6-week washout period, hypertensive, non-diabetic patients were treated with either eplerenone 25 mg twice daily or doxazosin 2 mg twice daily for 12 weeks. After each treatment period, insulin action was assessed by a hyperinsulinaemic euglycaemic clamp, with isotope dilution methodology. After washout, treatment groups were crossed over. Fifteen patients completed the study. There were no differences in fasting glucose, or fasting insulin between treatment with eplerenone or doxazosin. The measure of overall insulin sensitivity, exogenous glucose infusion rates during the last 30 min of the clamp, was similar with both treatments; 23.4 (3.9) μmol kg(-1) min(-1) after eplerenone and 23.3 (3.6) μmol kg(-1) min(-1) after doxazosin (P=0.83). Isotopically determined fasting endogenous glucose production rates were similar after both treatments (eplerenone 9.4 (0.6) μmol kg(-1) min(-1) vs doxazosin 10.6 (0.7) μmol kg(-1) min(-1)). There was a trend for lower endogenous glucose production rates during hyperinsulinaemia following eplerenone compared with doxazosin (2.0 (0.8) μmol kg(-1) min(-1) vs 4.1 (0.9) μmol kg(-1) min(-1)). There was no difference in insulin stimulated peripheral glucose utilisation rates after treatment with eplerenone or doxazosin (25.4 (3.6) μmol kg(-1) min(-1) vs 27.0 (3.9) μmol kg(-1) min(-1)). This study gives reassuring evidence of the neutral effect of eplerenone on insulin action in hypertensive, non-diabetic patients.

  3. Pedestrian and motorists' actions at pedestrian hybrid beacon sites: findings from a pilot study.

    PubMed

    Pulugurtha, Srinivas S; Self, Debbie R

    2015-01-01

    This paper focuses on an analysis of pedestrian and motorists' actions at sites with pedestrian hybrid beacons and assesses their effectiveness in improving the safety of pedestrians. Descriptive and statistical analyses (one-tail two-sample T-test and two-proportion Z-test) were conducted using field data collected during morning and evening peak hours at three study sites in the city of Charlotte, NC, before and after the installation of pedestrian hybrid beacons. Further, an analysis was conducted to assess the change in pedestrian and motorists' actions over time (before the installation; 1 month, 3 months, 6 months, and 12 months after the installation). Results showed an increase in average traffic speed at one of the pedestrian hybrid beacon sites while no specific trends were observed at the other two pedestrian hybrid beacon sites. A decrease in the number of motorists not yielding to pedestrians, pedestrians trapped in the middle of the street, and pedestrian-vehicle conflicts were observed at all the three pedestrian hybrid beacon sites. The installation of pedestrian hybrid beacons did not have a negative effect on pedestrian actions at two out of the three sites. Improvements seem to be relatively more consistent 3 months after the installation of the pedestrian hybrid beacon.

  4. Experimental study on bubble pulse features under the combined action of horizontal and vertical walls

    NASA Astrophysics Data System (ADS)

    Wang, Shi-ping; Chu, Wen-hua; Zhang, A.-man

    2014-06-01

    The pulse features of a bubble have a close connection with the boundary condition. When a bubble moves near a rigid wall, it will be attracted by the Bjerknes force of the wall, and a jet pointing at the wall will be generated. In real application, the bubble may move under the combined action of walls in different directions when it forms at the corner of a pipe or at the bottom of a dam. The motion of the bubble shows complex and nonlinear characteristics under these conditions. In order to investigate the bubble pulse features near complex walls, a horizontal wall and a vertical wall are put into the experimental water tank synchronously, and an electric circuit with 200 voltages is designed to generate discharge bubbles, and then experimental study on the bubble pulse features under the combined action of horizontal and vertical walls is carried out. The influences of the combined action of two walls on the bubble shape, pulse period, moving trace and inside jet are obtained by changing the distances from bubble center to the two walls. It aims at providing references for the relevant theoretical and numerical research.

  5. From recording discrete actions to studying continuous goal-directed behaviours in team sports.

    PubMed

    Correia, Vanda; Araújo, Duarte; Vilar, Luís; Davids, Keith

    2013-01-01

    This paper highlights the importance of examining interpersonal interactions in performance analysis of team sports, predicated on the relationship between perception and action, compared to the traditional cataloguing of actions by individual performers. We discuss how ecological dynamics may provide a potential unifying theoretical and empirical framework to achieve this re-emphasis in research. With reference to data from illustrative studies on performance analysis and sport expertise, we critically evaluate some of the main assumptions and methodological approaches with regard to understanding how information influences action and decision-making during team sports performance. Current data demonstrate how the understanding of performance behaviours in team sports by sport scientists and practitioners may be enhanced with a re-emphasis in research on the dynamics of emergent ongoing interactions. Ecological dynamics provides formal and theoretically grounded descriptions of player-environment interactions with respect to key performance goals and the unfolding information of competitive performance. Developing these formal descriptions and explanations of sport performance may provide a significant contribution to the field of performance analysis, supporting design and intervention in both research and practice.

  6. Enhancing local action planning through quantitative flood risk analysis: a case study in Spain

    NASA Astrophysics Data System (ADS)

    Castillo-Rodríguez, Jesica Tamara; Escuder-Bueno, Ignacio; Perales-Momparler, Sara; Ramón Porta-Sancho, Juan

    2016-07-01

    This article presents a method to incorporate and promote quantitative risk analysis to support local action planning against flooding. The proposed approach aims to provide a framework for local flood risk analysis, combining hazard mapping with vulnerability data to quantify risk in terms of expected annual affected population, potential injuries, number of fatalities, and economic damages. Flood risk is estimated combining GIS data of loads, system response, and consequences and using event tree modelling for risk calculation. The study area is the city of Oliva, located on the eastern coast of Spain. Results from risk modelling have been used to inform local action planning and to assess the benefits of structural and non-structural risk reduction measures. Results show the potential impact on risk reduction of flood defences and improved warning communication schemes through local action planning: societal flood risk (in terms of annual expected affected population) would be reduced up to 51 % by combining both structural and non-structural measures. In addition, the effect of seasonal population variability is analysed (annual expected affected population ranges from 82 to 107 %, compared with the current situation, depending on occupancy rates in hotels and campsites). Results highlight the need for robust and standardized methods for urban flood risk analysis replicability at regional and national scale.

  7. Nurses’ Empowerment in Self-Care Education to Stroke Patients: An Action Research Study

    PubMed Central

    Aslani, Zahra; Alimohammadi, Nasrollah; Taleghani, Fariba; Khorasani, Parvaneh

    2016-01-01

    ABSTRACT Background: Self-care needs are major problems among stroke patients. Nurses can support them through interventions such as education, a change in their attitude and emphasis on their remaining abilities. However, research has shown some weak points in the quality of care given to these patients. So the aim of this study was to improve the nurses’ practice in self-care education to stroke patients. Methods: The findings of evaluation phase showed that during action research, approaching the nurses’ empowerment in self-care education to stroke patients has been set in motion. The nursing practice improvement, knowledge based practice, nurses’ attitude change, ability to respond against routinization, and motivation promotion emphasize the success of change process. Facilitators and barriers of educating patients are acknowledged by the participants as a factor influencing the continuation of change. Results: The lack of nurses’ educating performance skills was overcome using action research and changes were made to improve the performance of nurses. Conclusions: The lack of nurses’ educating performance skills was overcome using action research and changes were made to improve the performance of nurses. PMID:27713896

  8. Data Integrity-A Study of Current Regulatory Thinking and Action.

    PubMed

    Shafiei, Nader; De Montardy, Regis; Rivera-Martinez, Edwin

    2015-01-01

    In reaction to breaches of data integrity in the pharmaceutical industry, regulatory authorities have introduced inspection approaches or initiatives with the aim of reducing occurrences of data integrity problems. This review article-based on study of 65 cases of regulatory action from 2002 to 2014-provides an overview of current regulatory thinking and action on breaches of data integrity affecting GxP (health-related regulations) processes supporting non-clinical studies, clinical studies, laboratory controls, and production controls. These case studies largely represent position of the U.S. Food and Drug Administration and the regulatory agencies affiliated with the European Medicines Agency. Also discussed is the role of human factors as a potential source of data integrity problems. The article concludes by recommending some remedial controls that could be established to avoid or reduce occurrences of data integrity problems.Lay Abstract: In fulfilling their mission to protect public health, regulatory agencies (e.g., U.S. Food and Drug Administration, European Medicines Agency) must establish confidence that medical products they approve are fit for their intended use. In so doing they rely on scientific and operational data generated during research, development, manufacturing, sales, marketing, distribution, and post-marketing surveillance activities. The level of confidence they build is directly proportional to the scientific validity and integrity of data presented to them by the sponsors of medical products. In this article we present analysis of 65 case studies that document regulatory action taken by various regulatory agencies on breach of data integrity between 2002 and 2014. The ensuing discussion on current trends largely represents position of the U.S. Food and Drug Administration and European Medicines Agency. The article concludes by proposing some remedial controls that could be established by pharmaceutical companies to avoid or reduce

  9. The CRISPR/Cas9 system for gene editing and its potential application in pain research

    PubMed Central

    Sun, Linlin; Lutz, Brianna Marie; Tao, Yuan-Xiang

    2016-01-01

    The CRISPR/Cas9 system is a research hotspot in genome editing and regulation. Currently, it is used in genomic silencing and knock-in experiments as well as transcriptional activation and repression. This versatile system consists of two components: a guide RNA (gRNA) and a Cas9 nuclease. Recognition of a genomic DNA target is mediated through base pairing with a 20-base gRNA. The latter further recruits the Cas9 endonuclease protein to the target site and creates double-stranded breaks in the target DNA. Compared with traditional genome editing directed by DNA-binding protein domains, this short RNA-directed Cas9 endonuclease system is simple and easily programmable. Although this system may have off-target effects and in vivo delivery and immune challenges, researchers have employed this system in vivo to establish disease models, study specific gene functions under certain disease conditions, and correct genomic information for disease treatment. In regards to pain research, the CRISPR/Cas9 system may act as a novel tool in gene correction therapy for pain-associated hereditary diseases and may be a new approach for RNA-guided transcriptional activation or repression of pain-related genes. In addition, this system is also applied to loss-of-function mutations in pain-related genes and knockin of reporter genes or loxP tags at pain-related genomic loci. The CRISPR/Cas9 system will likely be carried out widely in both bench work and clinical settings in the pain field. PMID:27500183

  10. CRISPR-Cas9-Mediated Single-Gene and Gene Family Disruption in Trypanosoma cruzi

    PubMed Central

    Peng, Duo; Kurup, Samarchith P.; Yao, Phil Y.; Minning, Todd A.

    2014-01-01

    ABSTRACT Trypanosoma cruzi is a protozoan parasite of humans and animals, affecting 10 to 20 million people and innumerable animals, primarily in the Americas. Despite being the largest cause of infection-induced heart disease worldwide, even among the neglected tropical diseases (NTDs) T. cruzi is considered one of the least well understood and understudied. The genetic complexity of T. cruzi as well as the limited set of efficient techniques for genome engineering contribute significantly to the relative lack of progress in and understanding of this pathogen. Here, we adapted the CRISPR-Cas9 system for the genetic engineering of T. cruzi, demonstrating rapid and efficient knockout of multiple endogenous genes, including essential genes. We observed that in the absence of a template, repair of the Cas9-induced double-stranded breaks (DSBs) in T. cruzi occurs exclusively by microhomology-mediated end joining (MMEJ) with various-sized deletions. When a template for DNA repair is provided, DSB repair by homologous recombination is achieved at an efficiency several orders of magnitude higher than that in the absence of CRISPR-Cas9-induced DSBs. We also demonstrate the high multiplexing capacity of CRISPR-Cas9 in T. cruzi by knocking down expression of an enzyme gene family consisting of 65 members, resulting in a significant reduction of enzymatic product with no apparent off-target mutations. Lastly, we show that Cas9 can mediate disruption of its own coding sequence, rescuing a growth defect in stable Cas9-expressing parasites. These results establish a powerful new tool for the analysis of gene functions in T. cruzi, enabling the study of essential genes and their functions and analysis of the many large families of related genes that occupy a substantial portion of the T. cruzi genome. PMID:25550322

  11. Less irritative action of wine and Japanese sake in rat stomachs: a comparative study with ethanol.

    PubMed

    Nakagiri, Akari; Fukushima, Kazuhiro; Kato, Shinichi; Takeuchi, Koji

    2006-02-01

    The ingestion of alcohol, especially in excess, causes acute gastric lesions and gastritis in humans, yet the mucosal irritative action of alcoholic beverages remains largely unknown. We examined the mucosal irritative action of whiskey, wine and Japanese sake in the rat stomach both ex vivo and in vitro, in comparison with ethanol. Under urethane anesthesia, a rat stomach was mounted in an ex vivo chamber, then superfused with saline, and the transmucosal potential difference (PD) was measured. After the basal PD had stabilized, the mucosa was exposed for 30 min to 2 ml of 15% ethanol, whiskey (containing 15% ethanol), white wine, or Japanese sake (the ethanol concentration of the latter two is 12-15%). In the in vitro study, rat epithelial cells (RGM1) were treated with the alcoholic beverages for 5 min, and the cell viability was determined with crystal violet. Ethanol or whiskey applied to the chamber caused a decrease in PD, while wine or Japanese sake did not. Histologically, surface epithelial damage was observed after exposure to both ethanol and whiskey, yet no damage was induced by white wine and Japanese sake. Likewise, both ethanol and whiskey markedly reduced the viability of RGM1 cells after 5 min of incubation, while neither white wine nor Japanese sake had any effect. In addition, supplementation of glucose significantly prevented the reduction in both PD and cell viability caused by ethanol. These results suggest that the mucosal irritative action of Japanese sake and white wine is much less pronounced than that of ethanol or whiskey and that the less damaging action of Japanese sake and white wine may be, at least partly, accounted for by the glucose contained in these alcoholic beverages.

  12. Action-Monitoring Dysfunction in Obstructive Sleep Apnea - A Pilot Study

    PubMed Central

    Chou, Ping-Song; Hsu, Chung-Yao; Wu, Meng-Ni; Liou, Li-Min; Lu, Shinag-Ru; Liu, Ching-Kuan; Lai, Chiou-Lian

    2016-01-01

    Obstructive sleep apnea (OSA) is associated with a broad range of frontal lobe dysfunctions. However, no study has investigated action monitoring, a crucial domain of frontal cognitive functions, in patients with OSA. By using the modified Flanker task, we tested the hypothesis that patients with OSA have an impaired action monitoring function. We recruited 25 untreated patients with moderate–severe OSA and 12 control participants who were matched for age, sex, apolipoprotein E4, and education level. Every enrolled participant underwent a standard overnight laboratory-based polysomnography and completed a modified Flanker task. Compared with the controls, the patients with OSA presented a significantly lower correct response rate in all trials (78.9% vs 95.9%, P = .008), congruent trials (84.7% vs 98.3%, P = .016), and incongruent trials (77.4% vs 94.7%, P = .009). The post-error correction rate was significantly lower in the patients with OSA than in the controls (74.9% vs 93.8%, P = .005). Furthermore, strong significant correlations were observed between the arousal index and correct rate in all trials (r = −0.390, P < .05) and in the incongruent trials (r = −0.429, P < .01), as well as between the arousal index and rate of post-error correction (r = −0.435, P < .01). We concluded that the action monitoring function was impaired in the patients with OSA. Sleep fragmentation was a major determinant of impaired action monitoring in these patients. PMID:27300504

  13. Less irritative action of wine and Japanese sake in rat stomachs: a comparative study with ethanol.

    PubMed

    Nakagiri, Akari; Fukushima, Kazuhiro; Kato, Shinichi; Takeuchi, Koji

    2006-02-01

    The ingestion of alcohol, especially in excess, causes acute gastric lesions and gastritis in humans, yet the mucosal irritative action of alcoholic beverages remains largely unknown. We examined the mucosal irritative action of whiskey, wine and Japanese sake in the rat stomach both ex vivo and in vitro, in comparison with ethanol. Under urethane anesthesia, a rat stomach was mounted in an ex vivo chamber, then superfused with saline, and the transmucosal potential difference (PD) was measured. After the basal PD had stabilized, the mucosa was exposed for 30 min to 2 ml of 15% ethanol, whiskey (containing 15% ethanol), white wine, or Japanese sake (the ethanol concentration of the latter two is 12-15%). In the in vitro study, rat epithelial cells (RGM1) were treated with the alcoholic beverages for 5 min, and the cell viability was determined with crystal violet. Ethanol or whiskey applied to the chamber caused a decrease in PD, while wine or Japanese sake did not. Histologically, surface epithelial damage was observed after exposure to both ethanol and whiskey, yet no damage was induced by white wine and Japanese sake. Likewise, both ethanol and whiskey markedly reduced the viability of RGM1 cells after 5 min of incubation, while neither white wine nor Japanese sake had any effect. In addition, supplementation of glucose significantly prevented the reduction in both PD and cell viability caused by ethanol. These results suggest that the mucosal irritative action of Japanese sake and white wine is much less pronounced than that of ethanol or whiskey and that the less damaging action of Japanese sake and white wine may be, at least partly, accounted for by the glucose contained in these alcoholic beverages. PMID:16534671

  14. The role of movement representation in episodic memory for actions: A study of patients with apraxia.

    PubMed

    Masumoto, Kouhei; Shirakawa, Masayuki; Higashiyama, Takeshi; Yokoyama, Kazumasa

    2015-01-01

    In attempting to memorize a sentence about an action, such as "Pick up the glass," performing the action (motor encoding) results in better memory performance than simply memorizing the words (verbal encoding). Such enhancement of memory is known as the enactment effect. Several theories have been proposed to explain this phenomenon using concepts such as physical motor information associated with speed, form, amplitude of movement and/or movement representations involved in movement imaging, knowledge on manipulating tools, and spatial relationships in the enactment effect. However, there have been no cognitive neuropsychological studies investigating whether the enactment effect is crucially influenced by physical motor information or movement representations. To clarify this issue, we compared healthy adult control participants with two different types of apraxia patients. One patient with left hemisphere lesions caused by cerebral infarction had a disability involving multiple movement representations. The other patient showed symptoms of corticobasal syndrome and was not able to benefit from feedback on the accuracy of her motor movements during enactment. Participants memorized action sentences via either verbal or motor encoding and responded to recall and recognition tests. Results indicated that the patient with the movement representation deficits exhibited worse memory performance than the other patient or control participants following both verbal and motor encoding. Although the enactment effect was present during recall in both patients, the effect was not observed for recognition in the patient with severe movement representation deficits. These results suggest that movement representations are involved in encoding episodic memories of action. Moreover, the role of movement representations appears to depend on the form of retrieval that is being used. PMID:25921790

  15. An Exploratory Study of Undergraduates' Attitudes toward Affirmative Action Policies for Asian Americans in College

    ERIC Educational Resources Information Center

    Hartlep, Nicholas Daniel; Lowinger, Robert Jay

    2014-01-01

    This exploratory study examined white undergraduate students' (a) racial attitudes towards Asian Americans, (b) principled policy attitudes toward affirmative action, and (c) self-interest in relation to their support for college-based affirmative action policies for Asian Americans at a Midwestern university. A sample (n = 264, 28% male, 72%…

  16. Using Reflective Practice to Incorporate Formative Assessment in a Middle School Science Classroom: A Participatory Action Research Study

    ERIC Educational Resources Information Center

    Trauth-Nare, Amy; Buck, Gayle

    2011-01-01

    Our purpose was to investigate the efficacy of using reflective practice to guide our action research study of incorporating formative assessment into middle school science teaching and learning. Using participatory action research, we worked collaboratively to incorporate formative assessment into two instructional units, and then engaged in…

  17. DNA targeting by the type I-G and type I-A CRISPR–Cas systems of Pyrococcus furiosus

    PubMed Central

    Elmore, Joshua; Deighan, Trace; Westpheling, Jan; Terns, Rebecca M.; Terns, Michael P.

    2015-01-01

    CRISPR–Cas systems silence plasmids and viruses in prokaryotes. CRISPR–Cas effector complexes contain CRISPR RNAs (crRNAs) that include sequences captured from invaders and direct CRISPR-associated (Cas) proteins to destroy corresponding invader nucleic acids. Pyrococcus furiosus (Pfu) harbors three CRISPR–Cas immune systems: a Cst (Type I-G) system with an associated Cmr (Type III-B) module at one locus, and a partial Csa (Type I-A) module (lacking known invader sequence acquisition and crRNA processing genes) at another locus. The Pfu Cmr complex cleaves complementary target RNAs, and Csa systems have been shown to target DNA, while the mechanism by which Cst complexes silence invaders is unknown. In this study, we investigated the function of the Cst as well as Csa system in Pfu strains harboring a single CRISPR–Cas system. Plasmid transformation assays revealed that the Cst and Csa systems both function by DNA silencing and utilize similar flanking sequence information (PAMs) to identify invader DNA. Silencing by each system specifically requires its associated Cas3 nuclease. crRNAs from the 7 shared CRISPR loci in Pfu are processed for use by all 3 effector complexes, and Northern analysis revealed that individual effector complexes dictate the profile of mature crRNA species that is generated. PMID:26519471

  18. Double-stranded endonuclease activity in Bacillus halodurans clustered regularly interspaced short palindromic repeats (CRISPR)-associated Cas2 protein.

    PubMed

    Nam, Ki Hyun; Ding, Fran; Haitjema, Charles; Huang, Qingqiu; DeLisa, Matthew P; Ke, Ailong

    2012-10-19

    The CRISPR (clustered regularly interspaced short palindromic repeats) system is a prokaryotic RNA-based adaptive immune system against extrachromosomal genetic elements. Cas2 is a universally conserved core CRISPR-associated protein required for the acquisition of new spacers for CRISPR adaptation. It was previously characterized as an endoribonuclease with preference for single-stranded (ss)RNA. Here, we show using crystallography, mutagenesis, and isothermal titration calorimetry that the Bacillus halodurans Cas2 (Bha_Cas2) from the subtype I-C/Dvulg CRISPR instead possesses metal-dependent endonuclease activity against double-stranded (ds)DNA. This activity is consistent with its putative function in producing new spacers for insertion into the 5'-end of the CRISPR locus. Mutagenesis and isothermal titration calorimetry studies revealed that a single divalent metal ion (Mg(2+) or Mn(2+)), coordinated by a symmetric Asp pair in the Bha_Cas2 dimer, is involved in the catalysis. We envision that a pH-dependent conformational change switches Cas2 into a metal-binding competent conformation for catalysis. We further propose that the distinct substrate preferences among Cas2 proteins may be determined by the sequence and structure in the β1-α1 loop.

  19. Tailor-made CRISPR/Cas system for highly efficient targeted gene replacement in the rice blast fungus.

    PubMed

    Arazoe, Takayuki; Miyoshi, Kennosuke; Yamato, Tohru; Ogawa, Tetsuo; Ohsato, Shuichi; Arie, Tsutomu; Kuwata, Shigeru

    2015-12-01

    CRISPR/Cas-derived RNA-guided nucleases (RGNs) that can generate DNA double-strand breaks (DSBs) at a specific sequence are widely used for targeted genome editing by induction of DSB repair in many organisms. The CRISPR/Cas system consists of two components: a single Cas9 nuclease and a single-guide RNA (sgRNA). Therefore, the system for constructing RGNs is simple and efficient, but the utilization of RGNs in filamentous fungi has not been validated. In this study, we established the CRISPR/Cas system in the model filamentous fungus, Pyricularia oryzae, using Cas9 that was codon-optimized for filamentous fungi, and the endogenous RNA polymerase (RNAP) III U6 promoter and a RNAP II fungal promoter for the expression of the sgRNA. We further demonstrated that RGNs could recognize the desired sequences and edit endogenous genes through homologous recombination-mediated targeted gene replacement with high efficiency. Our system will open the way for the development of various CRISPR/Cas-based applications in filamentous fungi. PMID:26039904

  20. Foreign DNA capture during CRISPR–Cas adaptive immunity

    PubMed Central

    Nuñez, James K.; Harrington, Lucas B.; Kranzusch, Philip J.; Engelman, Alan N.; Doudna, Jennifer A.

    2015-01-01

    Bacteria and archaea generate adaptive immunity against phages and plasmids by integrating foreign DNA of specific 30–40 base pair (bp) lengths into clustered regularly interspaced short palindromic repeats (CRISPR) loci as spacer segments1–6. The universally conserved Cas1–Cas2 integrase complex catalyzes spacer acquisition using a direct nucleophilic integration mechanism similar to retroviral integrases and transposases7–13. How the Cas1–Cas2 complex selects foreign DNA substrates for integration remains unknown. Here we present X-ray crystal structures of the Escherichia coli Cas1–Cas2 complex bound to cognate 33 nucleotide (nt) protospacer DNA substrates. The protein complex creates a curved binding surface spanning the length of the DNA and splays the ends of the protospacer to allow each terminal nucleophilic 3′–OH to enter a channel leading into the Cas1 active sites. Phosphodiester backbone interactions between the protospacer and the proteins explain the sequence-nonspecific substrate selection observed in vivo2–4. Our results uncover the structural basis for foreign DNA capture and the mechanism by which Cas1–Cas2 functions as a molecular ruler to dictate the sequence architecture of CRISPR loci. PMID:26503043

  1. Foreign DNA capture during CRISPR-Cas adaptive immunity.

    PubMed

    Nuñez, James K; Harrington, Lucas B; Kranzusch, Philip J; Engelman, Alan N; Doudna, Jennifer A

    2015-11-26

    Bacteria and archaea generate adaptive immunity against phages and plasmids by integrating foreign DNA of specific 30-40-base-pair lengths into clustered regularly interspaced short palindromic repeat (CRISPR) loci as spacer segments. The universally conserved Cas1-Cas2 integrase complex catalyses spacer acquisition using a direct nucleophilic integration mechanism similar to retroviral integrases and transposases. How the Cas1-Cas2 complex selects foreign DNA substrates for integration remains unknown. Here we present X-ray crystal structures of the Escherichia coli Cas1-Cas2 complex bound to cognate 33-nucleotide protospacer DNA substrates. The protein complex creates a curved binding surface spanning the length of the DNA and splays the ends of the protospacer to allow each terminal nucleophilic 3'-OH to enter a channel leading into the Cas1 active sites. Phosphodiester backbone interactions between the protospacer and the proteins explain the sequence-nonspecific substrate selection observed in vivo. Our results uncover the structural basis for foreign DNA capture and the mechanism by which Cas1-Cas2 functions as a molecular ruler to dictate the sequence architecture of CRISPR loci.

  2. Duration of activity and mode of action of modafinil: Studies on sleep and wakefulness in humans.

    PubMed

    Turner, C; Belyavin, A J; Nicholson, A N

    2014-07-01

    The duration of activity of modafinil was investigated in healthy male volunteers in two double-blind crossover studies. Mode of action was explored using a statistical model concerned with the relationship between total sleep duration and that of rapid eye movement (REM) sleep. Nocturnal sleep (23:00-07:00) followed by next-day performance (09:00-17:00) was studied in 12 subjects administered 100, 200, 300 mg modafinil and placebo, 0.5 h before bedtime. Performance overnight (19:00-08:45) followed by sleep (09:15-15:15) was studied in nine subjects administered 100, 200, 300, 400 mg modafinil, 300 mg caffeine and placebo at 22:15. Modafinil dose-dependently reduced sleep duration (nocturnal: 200 mg, p<0.05; 300 mg, p<0.001; morning: 300 and 400 mg, p<0.05) and REM sleep (nocturnal: 300 mg; morning: 400 mg; p<0.05). The statistical model revealed that reduced REM sleep was due to alerting activity, with no evidence of direct suppression of REM sleep, suggesting dopaminergic activity. Enhanced performance with modafinil during overnight work varied with dose (200 mg>100 mg; 300, 400 mg>200, 100 mg, caffeine). However, in the study of next-day performance, the enhancement was attenuated at the highest dose (300 mg) by the greater disturbance of prior sleep. These findings indicate that modafinil has a long duration of action, with alerting properties arising predominantly from dopaminergic activity. PMID:24306135

  3. Regulation of p130Cas/BCAR1 Expression in Tamoxifen-Sensitive and Tamoxifen-Resistant Breast Cancer Cells by EGR1 and NAB21

    PubMed Central

    Kumbrink, Joerg; Kirsch, Kathrin H

    2012-01-01

    Elevated levels of p130Cas/BCAR1 (Crk-associated substrate/breast cancer antiestrogen resistance 1) are found in aggressive breast tumors and are associated with tamoxifen resistance of mammary cancers. p130Cas promotes the integration of protein complexes involved in multiple signaling pathways frequently deregulated in breast cancer. To elucidate mechanisms leading to p130Cas up-regulation in mammary carcinomas and during acquired tamoxifen resistance, the regulation of p130Cas/BCAR1 was studied. Because multiple putative binding motifs for the inducible transcription factor EGR1 were identified in the 5′ region of BCAR1, the p130Cas/BCAR1 regulation by EGR1 and its coregulator NAB2 was investigated. Overexpression or short interfering RNA (siRNA)-mediated down-regulation of EGR1 or NAB2, and chromatin immunoprecipitations indicated that EGR1 and NAB2 act in concert to positively regulate p130Cas/BCAR1 expression in breast cancer cells. p130Cas depletion using siRNA showed that, in tamoxifen-sensitive MCF-7 cells, p130Cas regulates EGR1 and NAB2 expression, whereas in the derivative tamoxifen-resistant TAM-R cells, only NAB2 levels were influenced. BCAR1 messenger RNA and p130Cas protein were upregulated by phorbol esters following the kinetics of late response genes in MCF-7 but not in TAM-R cells. Thus, in MCF-7 cells, we identified a positive feedback loop where p130Cas positively regulates EGR1 and NAB2, which in turn induce p130Cas expression. Importantly, compared with MCF-7, enhanced NAB2 expression and increased EGR1 binding to the BCAR1 5′ region observed in TAM-R may lead to the constitutively increased p130Cas/BCAR1 levels in TAM-R cells. The uncovered differences in this EGR1/NAB2/p130Cas network in MCF-7 versus TAM-R cells may also contribute to p130Cas up-regulation during acquired tamoxifen resistance. PMID:22431919

  4. Quantum chemical study of the mechanism of action of vitamin K epoxide reductase (VKOR)

    NASA Astrophysics Data System (ADS)

    Deerfield, David, II; Davis, Charles H.; Wymore, Troy; Stafford, Darrel W.; Pedersen, Lee G.

    Possible model, but simplistic, mechanisms for the action of vitamin K epoxide reductase (VKOR) are investigated with quantum mechanical methods (B3LYP/6-311G**). The geometries of proposed model intermediates in the mechanisms are energy optimized. Finally, the energetics of the proposed (pseudo-enzymatic) pathways are compared. We find that the several pathways are all energetically feasible. These results will be useful for designing quantum mechanical/molecular mechanical method (QM/MM) studies of the enzymatic pathway once three-dimensional structural data are determined and available for VKOR.

  5. Corrective Action Plan for Corrective Action Unit 261: Area 25 Test Cell A Leachfield System, Nevada Test Site, Nevada

    SciTech Connect

    T. M. Fitzmaurice

    2000-08-01

    This Corrective Action Plan (CAP) has been prepared for the Corrective Action Unit (CAU)261 Area 25 Test Cell A Leachfield System in accordance with the Federal Facility and Consent Order (Nevada Division of Environmental Protection [NDEP] et al., 1996). This CAP provides the methodology for implementing the approved corrective action alternative as listed in the Corrective Action Decision Document (U.S. Department of Energy, Nevada Operations Office, 1999). Investigation of CAU 261 was conducted from February through May of 1999. There were no Constituents of Concern (COCs) identified at Corrective Action Site (CAS) 25-05-07 Acid Waste Leach Pit (AWLP). COCs identified at CAS 25-05-01 included diesel-range organics and radionuclides. The following closure actions will be implemented under this plan: Because COCs were not found at CAS 25-05-07 AWLP, no action is required; Removal of septage from the septic tank (CAS 25-05-01), the distribution box and the septic tank will be filled with grout; Removal of impacted soils identified near the initial outfall area; and Upon completion of this closure activity and approval of the Closure Report by NDEP, administrative controls, use restrictions, and site postings will be used to prevent intrusive activities at the site.

  6. An Analytical Framework for Categorizing the Use of CAS Symbolic Manipulation in Textbooks

    ERIC Educational Resources Information Center

    Davis, Jon D.; Fonger, Nicole L.

    2015-01-01

    The symbolic manipulation capabilities of computer algebra systems, which we refer to as CAS-S, are now becoming instantiated within secondary mathematics textbooks in the United States for the first time. While a number of research studies have examined how teachers use this technology in their classrooms, one of the most important factors in how…

  7. Effectiveness of an Integrated Phonological Awareness Approach for Children with Childhood Apraxia of Speech (CAS)

    ERIC Educational Resources Information Center

    McNeill, Brigid C.; Gillon, Gail T.; Dodd, Barbara

    2009-01-01

    This study investigated the effectiveness of an integrated phonological awareness approach for children with childhood apraxia of speech (CAS). Change in speech, phonological awareness, letter knowledge, word decoding, and spelling skills were examined. A controlled multiple single-subject design was employed. Twelve children aged 4-7 years with…

  8. Examinations in the Final Year of Transition to Mathematical Methods Computer Algebra System (CAS)

    ERIC Educational Resources Information Center

    Leigh-Lancaster, David; Les, Magdalena; Evans, Michael

    2010-01-01

    2009 was the final year of parallel implementation for Mathematical Methods Units 3 and 4 and Mathematical Methods (CAS) Units 3 and 4. From 2006-2009 there was a common technology-free short answer examination that covered the same function, algebra, calculus and probability content for both studies with corresponding expectations for key…

  9. 41 CFR 102-33.440 - Who must report CAS cost and utilization data?

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... (cas) Cost and Utilization Data § 102-33.440 Who must report CAS cost and utilization data? Executive agencies, except the Armed Forces and U.S. intelligence agencies, must report CAS cost and utilization...

  10. Dexfenfluramine and norfenfluramine: comparison of mechanism of action in feeding and brain Fos-ir studies.

    PubMed

    Rowland, N E; Roth, J D; McMullen, M R; Patel, A; Cespedes, A T

    2000-02-01

    Dexfenfluramine (dF) and dexnorfenfluramine (dNF), its metabolite, are anorectic agents that release serotonin (5-HT) and may have a direct postsynaptic action. The effects on the anorectic effects of dF and dNF of either acute (p-chlorophenylalanine, PCPA) or chronic (5,7-dihydroxytryptamine, 5,7-DHT) brain 5-HT depletions were studied in rats and compared with the actions of a 5-HT uptake inhibitor (fluoxetine) and 5-HT(1B/2C) receptor agonists [1-(3-trifluoromethyl-phenyl)-piperazine and 1-(3-chlorophenyl) piperazine]. The anorexia caused by these agonists was enhanced in rats with 5,7-DHT lesions, possibly a result of receptor supersensitivity. In contrast, fluoxetine anorexia was somewhat reduced in one study and was unchanged in a second. Both dF and dNF anorexias were enhanced in rats with 5,7-DHT lesions. In contrast, the anorectic effects of either dF or dNF were unchanged in PCPA-treated rats relative to controls. Compared with controls, 5, 7-DHT-lesion rats showed greatly increased dF- and dNF-induced Fos-like immunoreactivity (ir) in the paraventricular (PVN) and supraoptic (SON) hypothalamic nuclei, and in the median preoptic area (MnPO), but were similar to controls in most other areas. PCPA pretreatment increased dF- and dNF-induced Fos-ir in the PVN, SON, and MnPO. In controls, equianorectic doses of dF and dNF induced Fos-ir in similar brain regions, but dNF produced relatively larger effects than dF in SON, PVN, and MnPO. The data are discussed in terms of multiple pathways in the anorectic actions of dF and dNF.

  11. Targeted genome editing of sweet orange using Cas9/sgRNA.

    PubMed

    Jia, Hongge; Wang, Nian

    2014-01-01

    Genetic modification, including plant breeding, has been widely used to improve crop yield and quality, as well as to increase disease resistance. Targeted genome engineering is expected to contribute significantly to future varietal improvement, and genome editing technologies using zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9/single guide RNA (sgRNA) have already been successfully used to genetically modify plants. However, to date, there has been no reported use of any of the current genome editing approaches in sweet orange, an important fruit crop. In this study, we first developed a novel tool, Xcc-facilitated agroinfiltration, for enhancing transient protein expression in sweet orange leaves. We then successfully employed Xcc-facilitated agroinfiltration to deliver Cas9, along with a synthetic sgRNA targeting the CsPDS gene, into sweet orange. DNA sequencing confirmed that the CsPDS gene was mutated at the target site in treated sweet orange leaves. The mutation rate using the Cas9/sgRNA system was approximately 3.2 to 3.9%. Off-target mutagenesis was not detected for CsPDS-related DNA sequences in our study. This is the first report of targeted genome modification in citrus using the Cas9/sgRNA system-a system that holds significant promise for the study of citrus gene function and for targeted genetic modification.

  12. Targeted Genome Editing of Sweet Orange Using Cas9/sgRNA

    PubMed Central

    Jia, Hongge; Wang, Nian

    2014-01-01

    Genetic modification, including plant breeding, has been widely used to improve crop yield and quality, as well as to increase disease resistance. Targeted genome engineering is expected to contribute significantly to future varietal improvement, and genome editing technologies using zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9/single guide RNA (sgRNA) have already been successfully used to genetically modify plants. However, to date, there has been no reported use of any of the current genome editing approaches in sweet orange, an important fruit crop. In this study, we first developed a novel tool, Xcc-facilitated agroinfiltration, for enhancing transient protein expression in sweet orange leaves. We then successfully employed Xcc-facilitated agroinfiltration to deliver Cas9, along with a synthetic sgRNA targeting the CsPDS gene, into sweet orange. DNA sequencing confirmed that the CsPDS gene was mutated at the target site in treated sweet orange leaves. The mutation rate using the Cas9/sgRNA system was approximately 3.2 to 3.9%. Off-target mutagenesis was not detected for CsPDS-related DNA sequences in our study. This is the first report of targeted genome modification in citrus using the Cas9/sgRNA system—a system that holds significant promise for the study of citrus gene function and for targeted genetic modification. PMID:24710347

  13. Streamlined Approach for Environmental Restoration Plan for Corrective Action Unit 484: Surface Debris, Waste Sites, and Burn Area, Tonopah Test Range, Nevada

    SciTech Connect

    Bechel Nevada

    2004-05-01

    This Streamlined Approach for Environmental Restoration plan details the activities necessary to close Corrective Action Unit (CAU) 484: Surface Debris, Waste Sites, and Burn Area (Tonopah Test Range). CAU 484 consists of sites located at the Tonopah Test Range, Nevada, and is currently listed in Appendix III of the Federal Facility Agreement and Consent Order. CAU 484 consists of the following six Corrective Action Sites: (1) CAS RG-52-007-TAML, Davis Gun Penetrator Test; (2) CAS TA-52-001-TANL, NEDS Detonation Area; (3) CAS TA-52-004-TAAL, Metal Particle Dispersion Test; (4) CAS TA-52-005-TAAL, Joint Test Assembly DU Sites; (5) CAS TA-52-006-TAPL, Depleted Uranium Site; and (6) CAS TA-54-001-TANL, Containment Tank and Steel Structure

  14. Recent Advances in Genome Editing Using CRISPR/Cas9

    PubMed Central

    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

  15. Action Research as a Research Methodology for the Study of the Teaching and Learning of Science.

    ERIC Educational Resources Information Center

    Feldman, Allan; Minstrell, Jim

    This chapter addresses issues of action research from three perspectives. In the first section, what it means to engage in action research as a methodology for investigating teaching and learning in science education is overviewed and various conceptions of action research are explicitly made. The second perspective is that of an individual…

  16. The impact of inquiry-based instructional professional development upon instructional practice: An action research study

    NASA Astrophysics Data System (ADS)

    Broom, Frances A.

    This mixed method case study employs action research, conducted over a three month period with 11 elementary math and science practitioners. Inquiry as an instructional practice is a vital component of math and science instruction and STEM teaching. Teachers examined their beliefs and teaching practices with regard to those instructional factors that influence inquiry instruction. Video-taped lessons were compared to a rubric and pre and post questionnaires along with two interviews which informed the study. The results showed that while most beliefs were maintained, teachers implemented inquiry at a more advanced level after examining their teaching and reflecting on ways to increase inquiry practices. Because instructional practices provide only one component of inquiry-based instruction, other components need to be examined in a future study.

  17. Leiomyoblastome gastrique: à propos de trois cas

    PubMed Central

    Moujahid, Mountassir; Ennafaa, Issam; EL Rhari, Ahmed; Serghini, Issam; Chekoura, Khalid; Tahiri, Moulay Hassan

    2015-01-01

    Le leiomyoblastome gastrique est une tumeur musculaire rare qui touche essentiellement l'adulte. Son développement est souvent exogastrique. Le diagnostic positif repose sur l'histologie et le traitement est basé sur la chirurgie. Nous rapportons trois cas de leiomyoblastome gastrique colligés dans le service de chirurgie générale au 5ème Hôpital Militaire. L’âge moyen des patients est de 47 ans; le motif de consultation était représenté par une hémorragie digestive et l'imagerie médicale a posé le diagnostic de masse tumorale dans tous les cas. Le traitement chirurgical consistait en une gastrectomie partielle et le compte rendu anatomopathologique a confirmé le leiomyoblastome gastrique dans les trois cas. Le siège de la tumeur a été posé par la fibroscopie oeso gastroduodénale, le traitement était chirurgical et les suites post opératoires étaient simples avec un contrôle par des fibroscopies répétitives sans aucun signe de récidive. Le leiomyoblastome gastrique est une tumeur rare. L’écho endoscopie joue un rôle primordial dans le diagnostic positif ainsi que dans l’évaluation de l'extension pariétale de ces tumeurs. Le traitement est essentiellement chirurgical. PMID:26090000

  18. Distinctive laterality of neural networks supporting action understanding in left- and right-handed individuals: An EEG coherence study.

    PubMed

    Kelly, Rachel; Mizelle, J C; Wheaton, Lewis A

    2015-08-01

    Prior work has demonstrated that perspective and handedness of observed actions can affect action understanding differently in right and left-handed persons, suggesting potential differences in the neural networks underlying action understanding between right and left-handed individuals. We sought to evaluate potential differences in these neural networks using electroencephalography (EEG). Right- and left-handed participants observed images of tool-use actions from egocentric and allocentric perspectives, with right- and left-handed actors performing the actions. Participants judged the outcome of the observed actions, and response accuracy and latency were recorded. Behaviorally, the highest accuracy and shortest latency was found in the egocentric perspective for right- and left-handed observers. Handedness of subject showed an effect on accuracy and latency also, where right-handed observers were faster to respond than left-handed observers, but on average were less accurate. Mu band (8-10 Hz) cortico-cortical coherence analysis indicated that right-handed observers have coherence in the motor dominant left parietal-premotor networks when looking at an egocentric right or allocentric left hands. When looking in an egocentric perspective at a left hand or allocentric right hand, coherence was lateralized to right parietal-premotor areas. In left-handed observers, bilateral parietal-premotor coherence patterns were observed regardless of actor handedness. These findings suggest that the cortical networks involved in understanding action outcomes are dependent on hand dominance, and notably right handed participants seem to utilize motor systems based on the limb seen performing the action. The decreased accuracy for right-handed participants on allocentric images could be due to asymmetrical lateralization of encoding action and motoric dominance, which may interfere with translating allocentric limb action outcomes. Further neurophysiological studies will

  19. Studies on the mechanisms of action of picrotoxin, quercetin and pregnanolone at the GABAρ1 receptor

    PubMed Central

    Goutman, Juan D; Calvo, Daniel J

    2004-01-01

    The mechanisms of action of antagonists of the γ-aminobutyric acid C (GABAC) receptor picrotoxin, quercetin and pregnanolone were studied. Ionic currents (chloride), mediated through human homomeric GABAρ1 receptors expressed in Xenopus oocytes, were recorded by two-electrode voltage clamp. Dose–response (D–R) curves and kinetic measurements of GABAρ1 currents were carried out in the presence or absence of antagonists. Use-dependent actions were also evaluated. Picrotoxin, quercetin and pregnanolone exerted noncompetitive actions. IC50 values measured at the EC50 for GABA (1 μM) were as follows: picrotoxin 0.6±0.1 μM (Hill coefficient n=1.0±0.2); quercetin 4.4±0.4 μM (n=1.5±0.2); pregnanolone 2.1±0.5 μM (n=0.8±0.1). These antagonists produced changes only in the slope of the linear current–voltage relationships, which was indicative of voltage-independent effects. The effect of picrotoxin on GABAρ1 currents was use-dependent, strongly relied on agonist concentration and showed a slow onset and offset. The mechanism was compatible with an allosteric inhibition and receptor activation was a prerequisite for antagonism. The effect of quercetin was use-independent, showed relatively fast onset and offset, and resulted in a slowed time course of the GABA-evoked currents. The effect of pregnanolone was use-independent, presented fast onset and a very slow washout, and did not affect current activation. All the antagonists accelerated the time course of deactivation of the GABAρ1 currents. PMID:14732759

  20. CORRECTIVE ACTION PLAN FOR CORRECTIVE ACTION UNIT 536: AREA 3 RELEASE SITE, NEVADA TEST SITE, NEVADA

    SciTech Connect

    2005-09-01

    CAU 536 consists of CAS 03-44-02, Steam Jenny Discharge, located in Area 3 of the NTS. The site was characterized in 2004 according to the approved CAIP and the site characterization results are reported in the CAU 536 CADD. The purpose of this Corrective Action Plan (CAP) is to provide the detailed scope of work required to implement the recommended corrective actions as specified in the approved CAU 536 CADD.

  1. La tuberculose cutanée: observation de six cas confirmés au CHU Souro SANOU (CHUSS) de Bobo-Dioulasso (Burkina Faso)

    PubMed Central

    Andonaba, Jean Baptiste; Barro-Traoré, Fatou; Yaméogo, Téné; Diallo, Boukary; Korsaga-Somé, Nina; Traoré, Adama

    2013-01-01

    La localisation cutanée de la maladie tuberculeuse demeure une forme rare et représente seulement 2,1% des localisations. L'objet de cette étude est de rapporter le profil épidémiologique, anatomoclinique et évolutif des cas de tuberculose ganglio-cutanée diagnostiqués dans un CHU au Burkina Faso. La fréquence de la tuberculose cutanée est très faible au CHUSS. Six cas ont été diagnostiqués entre 2004 et 2010, soit une fréquence de un cas par an. La durée d’évolution des cas allait de deux jusqu’à dix ans avant leur diagnostic. Les lésions observées étaient: trois scrofulodermes, trois gommes, une tuberculose testiculaire associée à un mal de Pott, un cas de polyadénopathies et des cicatrices atropho-rétractiles dans la plupart des cas. Sur le plan anatomopathologique, des granulomes tuberculoïdes ont été mis en évidence dans tous les cas avec une forte réaction tuberculinique à l'IDR. Sous antituberculeux pendant six mois, l’évolution a été bonne dans tous les cas mais au prix de séquelles cutanées cicatricielles inesthétiques. Son ampleur reste peut-être encore méconnue. Le renforcement du plateau technique du CHU et une bonne collaboration interdisciplinaire contribuerait à un meilleur diagnostic et prise en charge de cette affection. PMID:24648863

  2. Functional analysis of Bombyx Wnt1 during embryogenesis using the CRISPR/Cas9 system.

    PubMed

    Zhang, Zhongjie; Aslam, Abu F M; Liu, Xiaojing; Li, Muwang; Huang, Yongping; Tan, Anjiang

    2015-08-01

    Recently established, custom-designed nuclease technologies such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated system provide attractive genome editing tools. Targeted gene mutagenesis using the CRISPR/Cas9 system has been achieved in several orders of insects. However, outside of studies on Drosophila melanogaster and the lepidopteron model insect Bombyx mori, little success has been reported, which is largely due to a lack of effective genetic manipulation tools that can be used in other insect orders. To create a simple and effective method of gene knockout analysis, especially for dissecting gene functioning during insect embryogenesis, we performed a functional analysis of the Bombyx Wnt1 (BmWnt1) gene using Cas9/sgRNA-mediated gene mutagenesis. The Wnt1 gene is required for embryonic patterning in various organisms, and its crucial roles during embryogenesis have been demonstrated in several insect orders. Direct injection of Cas9 mRNA and BmWnt1-specific sgRNA into Bombyx embryos induced a typical Wnt-deficient phenotype: injected embryos could not hatch and exhibited severe defects in body segmentation and pigmentation in a dose-dependent manner. Quantitative real-time PCR (qRT-PCR) analysis revealed that Hox genes were down-regulated after BmWnt1 depletion. Furthermore, large deletion, up to 18Kb, ware generated. The current study demonstrates that using the CRISPR/Cas9 system is a promising approach to achieve targeted gene mutagenesis during insect embryogenesis. PMID:26070541

  3. Heritable CRISPR/Cas9-mediated genome editing in the yellow fever mosquito, Aedes aegypti.

    PubMed

    Dong, Shengzhang; Lin, Jingyi; Held, Nicole L; Clem, Rollie J; Passarelli, A Lorena; Franz, Alexander W E

    2015-01-01

    In vivo targeted gene disruption is a powerful tool to study gene function. Thus far, two tools for genome editing in Aedes aegypti have been applied, zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN). As a promising alternative to ZFN and TALEN, which are difficult to produce and validate using standard molecular biological techniques, the clustered regularly interspaced short palindromic repeats/CRISPR-associated sequence 9 (CRISPR/Cas9) system has recently been discovered as a "do-it-yourself" genome editing tool. Here, we describe the use of CRISPR/Cas9 in the mosquito vector, Aedes aegypti. In a transgenic mosquito line expressing both Dsred and enhanced cyan fluorescent protein (ECFP) from the eye tissue-specific 3xP3 promoter in separated but tightly linked expression cassettes, we targeted the ECFP nucleotide sequence for disruption. When supplying the Cas9 enzyme and two sgRNAs targeting different regions of the ECFP gene as in vitro transcribed mRNAs for germline transformation, we recovered four different G1 pools (5.5% knockout efficiency) where individuals still expressed DsRed but no longer ECFP. PCR amplification, cloning, and sequencing of PCR amplicons revealed indels in the ECFP target gene ranging from 2-27 nucleotides. These results show for the first time that CRISPR/Cas9 mediated gene editing is achievable in Ae. aegypti, paving the way for further functional genomics related studies in this mosquito species. PMID:25815482

  4. Genome Editing by CRISPR/Cas9: A Game Change in the Genetic Manipulation of Protists.

    PubMed

    Lander, Noelia; Chiurillo, Miguel A; Docampo, Roberto

    2016-09-01

    Genome editing by CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated gene 9) system has been transformative in biology. Originally discovered as an adaptive prokaryotic immune system, CRISPR/Cas9 has been repurposed for genome editing in a broad range of model organisms, from yeast to mammalian cells. Protist parasites are unicellular organisms producing important human diseases that affect millions of people around the world. For many of these diseases, such as malaria, Chagas disease, leishmaniasis and cryptosporidiosis, there are no effective treatments or vaccines available. The recent adaptation of the CRISPR/Cas9 technology to several protist models will be playing a key role in the functional study of their proteins, in the characterization of their metabolic pathways, and in the understanding of their biology, and will facilitate the search for new chemotherapeutic targets. In this work we review recent studies where the CRISPR/Cas9 system was adapted to protist parasites, particularly to Apicomplexans and trypanosomatids, emphasizing the different molecular strategies used for genome editing of each organism, as well as their advantages. We also discuss the potential usefulness of this technology in the green alga Chlamydomonas reinhardtii.

  5. Genome Editing by CRISPR/Cas9: A Game Change in the Genetic Manipulation of Protists.

    PubMed

    Lander, Noelia; Chiurillo, Miguel A; Docampo, Roberto

    2016-09-01

    Genome editing by CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated gene 9) system has been transformative in biology. Originally discovered as an adaptive prokaryotic immune system, CRISPR/Cas9 has been repurposed for genome editing in a broad range of model organisms, from yeast to mammalian cells. Protist parasites are unicellular organisms producing important human diseases that affect millions of people around the world. For many of these diseases, such as malaria, Chagas disease, leishmaniasis and cryptosporidiosis, there are no effective treatments or vaccines available. The recent adaptation of the CRISPR/Cas9 technology to several protist models will be playing a key role in the functional study of their proteins, in the characterization of their metabolic pathways, and in the understanding of their biology, and will facilitate the search for new chemotherapeutic targets. In this work we review recent studies where the CRISPR/Cas9 system was adapted to protist parasites, particularly to Apicomplexans and trypanosomatids, emphasizing the different molecular strategies used for genome editing of each organism, as well as their advantages. We also discuss the potential usefulness of this technology in the green alga Chlamydomonas reinhardtii. PMID:27315329

  6. Functional analysis of Bombyx Wnt1 during embryogenesis using the CRISPR/Cas9 system.

    PubMed

    Zhang, Zhongjie; Aslam, Abu F M; Liu, Xiaojing; Li, Muwang; Huang, Yongping; Tan, Anjiang

    2015-08-01

    Recently established, custom-designed nuclease technologies such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated system provide attractive genome editing tools. Targeted gene mutagenesis using the CRISPR/Cas9 system has been achieved in several orders of insects. However, outside of studies on Drosophila melanogaster and the lepidopteron model insect Bombyx mori, little success has been reported, which is largely due to a lack of effective genetic manipulation tools that can be used in other insect orders. To create a simple and effective method of gene knockout analysis, especially for dissecting gene functioning during insect embryogenesis, we performed a functional analysis of the Bombyx Wnt1 (BmWnt1) gene using Cas9/sgRNA-mediated gene mutagenesis. The Wnt1 gene is required for embryonic patterning in various organisms, and its crucial roles during embryogenesis have been demonstrated in several insect orders. Direct injection of Cas9 mRNA and BmWnt1-specific sgRNA into Bombyx embryos induced a typical Wnt-deficient phenotype: injected embryos could not hatch and exhibited severe defects in body segmentation and pigmentation in a dose-dependent manner. Quantitative real-time PCR (qRT-PCR) analysis revealed that Hox genes were down-regulated after BmWnt1 depletion. Furthermore, large deletion, up to 18Kb, ware generated. The current study demonstrates that using the CRISPR/Cas9 system is a promising approach to achieve targeted gene mutagenesis during insect embryogenesis.

  7. Bystander Action in Situations of Dating and Sexual Aggression: A Mixed Methodological Study of High School Youth.

    PubMed

    Edwards, Katie M; Rodenhizer-Stämpfli, Kara Anne; Eckstein, Robert P

    2015-12-01

    Bystander action is a critical component of dating and sexual aggression prevention; however, little is known about barriers and facilitators of bystander action among high school youth and in what situations youth are willing to engage in bystander action. The current study examined bystander action in situations of dating and sexual aggression using a mixed methodological design. Participants included primarily Caucasian (83.0%, n = 181) male (54.6%, n = 119) and female (44.5%, n = 97) high school youth (N = 218). Most (93.6%) students had the opportunity to take action during the past year in situations of dating or sexual aggression; being female and histories of dating and sexual aggression related to bystander action. Thematic analysis of the focus group data identified barriers (e.g., the aggression not meeting a certain threshold, anticipated negative consequences) to bystander action, as well as insight on promising forms of action (e.g., verbally telling the perpetrator to stop, getting a teacher); problematic intervention methods (e.g., threatening or using physical violence to stop the perpetrator) were also noted. Implications for programming are discussed.

  8. Corrective Action Decision Document/Closure Report for Corrective Action Unit 367: Area 10 Sedan, Ess and Uncle Unit Craters Nevada National Security Site, Nevada, Revision 0

    SciTech Connect

    Patrick Matthews

    2011-06-01

    Corrective Action Unit 367 comprises four corrective action sites (CASs): • 10-09-03, Mud Pit • 10-45-01, U-10h Crater (Sedan) • 10-45-02, Ess Crater Site • 10-45-03, Uncle Crater Site The purpose of this Corrective Action Decision Document/Closure Report is to provide justification and documentation of the corrective actions and site closure activities implemented at CAU 367. A corrective action of closure in place with use restrictions was completed at each of the three crater CASs (10-45-01, 10-45-02, and 10-45-03); corrective actions were not required at CAS 10-09-03. In addition, a limited soil removal corrective action was conducted at the location of a potential source material release. Based on completion of these correction actions, no additional corrective action is required at CAU 367, and site closure is considered complete. Corrective action investigation (CAI) activities were performed from February 2010 through March 2011, as set forth in the Corrective Action Investigation Plan for Corrective Action Unit 367: Area 10 Sedan, Ess and Uncle Unit Craters, Nevada Test Site, Nevada. The approach for the CAI was divided into two facets: investigation of the primary release of radionuclides, and investigation of non-test or other releases (e.g., migration in washes and potential source material). Based on the proximity of the Uncle, Ess, and Sedan craters, the impact of the Sedan test on the fallout deposited from the two earlier tests, and aerial radiological surveys, the CAU 367 investigation was designed to study the releases from the three crater CASs as one combined release (primary release). Corrective Action Site 10-09-03, Mud Pit, consists of two mud pits identified at CAU 367. The mud pits are considered non-test releases or other releases and were investigated independent of the three crater CASs. The purpose of the CAI was to fulfill data needs as defined during the data quality objective (DQO) process. The CAU 367 dataset of

  9. Involvement of the Motor System in Comprehension of Non-Literal Action Language: A Meta-Analysis Study.

    PubMed

    Yang, Jie; Shu, Hua

    2016-01-01

    Although numerous studies have shown that the sensory-motor system is involved in semantic processing of language stimuli, it is still unclear whether comprehension of abstract concepts is embodied, and whether the involvement of the sensory-motor system is context-dependent. Investigation of how the motor system is activated during comprehension of non-literal action languages can help address these issues. So far several studies have reported brain activations during non-literal action language comprehension, but the findings are highly inconsistent because of different types of non-literal action language stimuli. To clarify how the motor system is involved in comprehension of different types of non-literal languages, the current study conducted quantitative meta-analyses on fMRI findings about comprehension of sentences describing fictive motions, metaphoric actions, and idiomatic actions. Results showed that fictive motion sentences elicited activation in the right parahippocampal gyrus, an area important for spatial processing. For metaphoric actions, the left precentral gyrus (BA 6) was strongly activated, suggesting a link between metaphoric and literal meanings. For idiomatic actions, activity was found in the left inferior frontal gyrus (BA 44/45), highlighting semantic selection and inhibition. No premotor or motor activity was found in idiom condition. These results together suggest that the involvement of the sensory-motor system in abstract concepts processing is flexible, depending on semantic features of the language stimuli and links between abstract and literal meanings.

  10. Factors affecting collective action for forest fire management: a comparative study of community forest user groups in central Siwalik, Nepal.

    PubMed

    Sapkota, Lok Mani; Shrestha, Rajendra Prasad; Jourdain, Damien; Shivakoti, Ganesh P

    2015-01-01

    The attributes of social ecological systems affect the management of commons. Strengthening and enhancing social capital and the enforcement of rules and sanctions aid in the collective action of communities in forest fire management. Using a set of variables drawn from previous studies on the management of commons, we conducted a study across 20 community forest user groups in Central Siwalik, Nepal, by dividing the groups into two categories based on the type and level of their forest fire management response. Our study shows that the collective action in forest fire management is consistent with the collective actions in other community development activities. However, the effectiveness of collective action is primarily dependent on the complex interaction of various variables. We found that strong social capital, strong enforcement of rules and sanctions, and users' participation in crafting the rules were the major variables that strengthen collective action in forest fire management. Conversely, users' dependency on a daily wage and a lack of transparency were the variables that weaken collective action. In fire-prone forests such as the Siwalik, our results indicate that strengthening social capital and forming and enforcing forest fire management rules are important variables that encourage people to engage in collective action in fire management.

  11. Factors Affecting Collective Action for Forest Fire Management: A Comparative Study of Community Forest User Groups in Central Siwalik, Nepal

    NASA Astrophysics Data System (ADS)

    Sapkota, Lok Mani; Shrestha, Rajendra Prasad; Jourdain, Damien; Shivakoti, Ganesh P.

    2015-01-01

    The attributes of social ecological systems affect the management of commons. Strengthening and enhancing social capital and the enforcement of rules and sanctions aid in the collective action of communities in forest fire management. Using a set of variables drawn from previous studies on the management of commons, we conducted a study across 20 community forest user groups in Central Siwalik, Nepal, by dividing the groups into two categories based on the type and level of their forest fire management response. Our study shows that the collective action in forest fire management is consistent with the collective actions in other community development activities. However, the effectiveness of collective action is primarily dependent on the complex interaction of various variables. We found that strong social capital, strong enforcement of rules and sanctions, and users' participation in crafting the rules were the major variables that strengthen collective action in forest fire management. Conversely, users' dependency on a daily wage and a lack of transparency were the variables that weaken collective action. In fire-prone forests such as the Siwalik, our results indicate that strengthening social capital and forming and enforcing forest fire management rules are important variables that encourage people to engage in collective action in fire management.

  12. Using CAS to Solve a Mathematics Task: A Deconstruction

    ERIC Educational Resources Information Center

    Berger, Margot

    2010-01-01

    I investigate how and whether a heterogeneous group of first-year university mathematics students in South Africa harness the potential power of a computer algebra system (CAS) when doing a specific mathematics task. In order to do this, I develop a framework for deconstructing a mathematics task requiring the use of CAS, into its primary…

  13. 48 CFR 9903.201-2 - Types of CAS coverage.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ..., OFFICE OF FEDERAL PROCUREMENT POLICY, OFFICE OF MANAGEMENT AND BUDGET PROCUREMENT PRACTICES AND COST... 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—...

  14. 48 CFR 970.3002-1 - CAS applicability.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ....3002-1 CAS applicability. The provisions of 48 CFR part 30 and 48 CFR chapter 99 (FAR Appendix) shall... 48 Federal Acquisition Regulations System 5 2014-10-01 2014-10-01 false CAS applicability. 970.3002-1 Section 970.3002-1 Federal Acquisition Regulations System DEPARTMENT OF ENERGY...

  15. 48 CFR 970.3002-1 - CAS applicability.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ....3002-1 CAS applicability. The provisions of 48 CFR part 30 and 48 CFR chapter 99 (FAR Appendix) shall... 48 Federal Acquisition Regulations System 5 2013-10-01 2013-10-01 false CAS applicability. 970.3002-1 Section 970.3002-1 Federal Acquisition Regulations System DEPARTMENT OF ENERGY...

  16. 48 CFR 970.3002-1 - CAS applicability.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ....3002-1 CAS applicability. The provisions of 48 CFR part 30 and 48 CFR chapter 99 (FAR Appendix) shall... 48 Federal Acquisition Regulations System 5 2012-10-01 2012-10-01 false CAS applicability. 970.3002-1 Section 970.3002-1 Federal Acquisition Regulations System DEPARTMENT OF ENERGY...

  17. Transformation of OODT CAS to Perform Larger Tasks

    NASA Technical Reports Server (NTRS)

    Mattmann, Chris; Freeborn, Dana; Crichton, Daniel; Hughes, John; Ramirez, Paul; Hardman, Sean; Woollard, David; Kelly, Sean

    2008-01-01

    A computer program denoted OODT CAS has been transformed to enable performance of larger tasks that involve greatly increased data volumes and increasingly intensive processing of data on heterogeneous, geographically dispersed computers. Prior to the transformation, OODT CAS (also alternatively denoted, simply, 'CAS') [wherein 'OODT' signifies 'Object-Oriented Data Technology' and 'CAS' signifies 'Catalog and Archive Service'] was a proven software component used to manage scientific data from spaceflight missions. In the transformation, CAS was split into two separate components representing its canonical capabilities: file management and workflow management. In addition, CAS was augmented by addition of a resource-management component. This third component enables CAS to manage heterogeneous computing by use of diverse resources, including high-performance clusters of computers, commodity computing hardware, and grid computing infrastructures. CAS is now more easily maintainable, evolvable, and reusable. These components can be used separately or, taking advantage of synergies, can be used together. Other elements of the transformation included addition of a separate Web presentation layer that supports distribution of data products via Really Simple Syndication (RSS) feeds, and provision for full Resource Description Framework (RDF) exports of metadata.

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

    PubMed

    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.

  19. Cas Proteins in Normal and Pathological Cell Growth Control

    PubMed Central

    Tikhmyanova, Nadezhda; Little, Joy L.; Golemis, Erica A.

    2009-01-01

    Proteins of the CAS (Crk-Associated Substrate) family (BCAR1/p130Cas, NEDD9/HEF1/Cas-L, EFS/SIN and CASS4/HEPL) are integral players in normal and pathological cell biology. CAS proteins act as scaffolds to regulate protein complexes controlling migration and chemotaxis, apoptosis, cell cycle, and differentiation, and have more recently been linked to a role in progenitor cell function. Reflecting these complex functions, over-expression of CAS proteins has now been strongly linked to poor prognosis and increased metastasis in cancer, as well as resistance to first-line chemotherapeutics in multiple tumor types including breast and lung cancers, glioblastoma, and melanoma. Further, CAS proteins have also been linked to additional pathological conditions including inflammatory disorders, Alzheimer’s and Parkinson’s disease, as well as developmental defects. This review will explore the roles of the CAS proteins in normal and pathological states in the context of the many mechanistic insights into CAS protein function that have emerged in the past decade. PMID:19937461

  20. Optimization of genome editing through CRISPR-Cas9 engineering.

    PubMed

    Zhang, Jian-Hua; Adikaram, Poorni; Pandey, Mritunjay; Genis, Allison; Simonds, William F

    2016-04-01

    CRISPR (Clustered Regularly-Interspaced Short Palindromic Repeats)-Cas9 (CRISPR associated protein 9) has rapidly become the most promising genome editing tool with great potential to revolutionize medicine. Through guidance of a 20 nucleotide RNA (gRNA), CRISPR-Cas9 finds and cuts target protospacer DNA precisely 3 base pairs upstream of a PAM (Protospacer Adjacent Motif). The broken DNA ends are repaired by either NHEJ (Non-Homologous End Joining) resulting in small indels, or by HDR (Homology Directed Repair) for precise gene or nucleotide replacement. Theoretically, CRISPR-Cas9 could be used to modify any genomic sequences, thereby providing a simple, easy, and cost effective means of genome wide gene editing. However, the off-target activity of CRISPR-Cas9 that cuts DNA sites with imperfect matches with gRNA have been of significant concern because clinical applications require 100% accuracy. Additionally, CRISPR-Cas9 has unpredictable efficiency among different DNA target sites and the PAM requirements greatly restrict its genome editing frequency. A large number of efforts have been made to address these impeding issues, but much more is needed to fully realize the medical potential of CRISPR-Cas9. In this article, we summarize the existing problems and current advances of the CRISPR-Cas9 technology and provide perspectives for the ultimate perfection of Cas9-mediated genome editing.