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Sample records for large-scale transposon mutagenesis

  1. Large-Scale Mutagenesis of the Yeast Genome Using a Tn7-Derived Multipurpose Transposon

    PubMed Central

    Kumar, Anuj; Seringhaus, Michael; Biery, Matthew C.; Sarnovsky, Robert J.; Umansky, Lara; Piccirillo, Stacy; Heidtman, Matthew; Cheung, Kei-Hoi; Dobry, Craig J.; Gerstein, Mark B.; Craig, Nancy L.; Snyder, Michael

    2004-01-01

    We present here an unbiased and extremely versatile insertional library of yeast genomic DNA generated by in vitro mutagenesis with a multipurpose element derived from the bacterial transposon Tn7. This mini-Tn7 element has been engineered such that a single insertion can be used to generate a lacZ fusion, gene disruption, and epitope-tagged gene product. Using this transposon, we generated a plasmid-based library of ∼300,000 mutant alleles; by high-throughput screening in yeast, we identified and sequenced 9032 insertions affecting 2613 genes (45% of the genome). From analysis of 7176 insertions, we found little bias in Tn7 target-site selection in vitro. In contrast, we also sequenced 10,174 Tn3 insertions and found a markedly stronger preference for an AT-rich 5-base pair target sequence. We further screened 1327 insertion alleles in yeast for hypersensitivity to the chemotherapeutic cisplatin. Fifty-one genes were identified, including four functionally uncharacterized genes and 25 genes involved in DNA repair, replication, transcription, and chromatin structure. In total, the collection reported here constitutes the largest plasmid-based set of sequenced yeast mutant alleles to date and, as such, should be singularly useful for gene and genome-wide functional analysis. PMID:15466296

  2. Large-scale mutagenesis of the yeast genome using a Tn7-derived multipurpose transposon.

    PubMed

    Kumar, Anuj; Seringhaus, Michael; Biery, Matthew C; Sarnovsky, Robert J; Umansky, Lara; Piccirillo, Stacy; Heidtman, Matthew; Cheung, Kei-Hoi; Dobry, Craig J; Gerstein, Mark B; Craig, Nancy L; Snyder, Michael

    2004-10-01

    We present here an unbiased and extremely versatile insertional library of yeast genomic DNA generated by in vitro mutagenesis with a multipurpose element derived from the bacterial transposon Tn7. This mini-Tn7 element has been engineered such that a single insertion can be used to generate a lacZ fusion, gene disruption, and epitope-tagged gene product. Using this transposon, we generated a plasmid-based library of approximately 300,000 mutant alleles; by high-throughput screening in yeast, we identified and sequenced 9032 insertions affecting 2613 genes (45% of the genome). From analysis of 7176 insertions, we found little bias in Tn7 target-site selection in vitro. In contrast, we also sequenced 10,174 Tn3 insertions and found a markedly stronger preference for an AT-rich 5-base pair target sequence. We further screened 1327 insertion alleles in yeast for hypersensitivity to the chemotherapeutic cisplatin. Fifty-one genes were identified, including four functionally uncharacterized genes and 25 genes involved in DNA repair, replication, transcription, and chromatin structure. In total, the collection reported here constitutes the largest plasmid-based set of sequenced yeast mutant alleles to date and, as such, should be singularly useful for gene and genome-wide functional analysis.

  3. Transposon Mutagenesis in Mice

    PubMed Central

    Largaespada, David A.

    2010-01-01

    Understanding the functional landscape of the mammalian genome is the next big challenge of biomedical research. The completion of the first phases of the mouse and human genome projects, and expression analyses using microarray hybridization, generate critically important questions about the functional landscape and structure of the mammalian genome: how many genes, and of what type, are there; what kind of functional elements make up a properly functioning gene? One step in this process will be to create mutations in every identifiable mouse gene and analyze the resultant phenotypes. Transposons are being considered as tools to further initiatives to create a comprehensive resource of mutant mouse strains. Also, it may be possible to use transposons in true forward genetic screens in the mouse. The “Sleeping Beauty” (SB) transposon system is one such tool. Moreover, due to its tendency for local hopping, SB has been proposed as a method for regional saturation mutagenesis of the mouse genome. In this chapter, we review the tools and methods currently available to create mutant mice using in vivo, germline transposition in mice. PMID:19266336

  4. Large-scale transposon mutagenesis of Photobacterium profundum SS9 reveals new genetic loci important for growth at low temperature and high pressure.

    PubMed

    Lauro, Federico M; Tran, Khiem; Vezzi, Alessandro; Vitulo, Nicola; Valle, Giorgio; Bartlett, Douglas H

    2008-03-01

    Microorganisms adapted to piezopsychrophilic growth dominate the majority of the biosphere that is at relatively constant low temperatures and high pressures, but the genetic bases for the adaptations are largely unknown. Here we report the use of transposon mutagenesis with the deep-sea bacterium Photobacterium profundum strain SS9 to isolate dozens of mutant strains whose growth is impaired at low temperature and/or whose growth is altered as a function of hydrostatic pressure. In many cases the gene mutation-growth phenotype relationship was verified by complementation analysis. The largest fraction of loci associated with temperature sensitivity were involved in the biosynthesis of the cell envelope, in particular the biosynthesis of extracellular polysaccharide. The largest fraction of loci associated with pressure sensitivity were involved in chromosomal structure and function. Genes for ribosome assembly and function were found to be important for both low-temperature and high-pressure growth. Likewise, both adaptation to temperature and adaptation to pressure were affected by mutations in a number of sensory and regulatory loci, suggesting the importance of signal transduction mechanisms in adaptation to either physical parameter. These analyses were the first global analyses of genes conditionally required for low-temperature or high-pressure growth in a deep-sea microorganism.

  5. Construction of a large-scale Burkholderia cenocepacia J2315 transposon mutant library

    NASA Astrophysics Data System (ADS)

    Wong, Yee-Chin; Pain, Arnab; Nathan, Sheila

    2014-09-01

    Burkholderia cenocepacia, a pathogenic member of the Burkholderia cepacia complex (Bcc), has emerged as a significant threat towards cystic fibrosis patients, where infection often leads to the fatal clinical manifestation known as cepacia syndrome. Many studies have investigated the pathogenicity of B. cenocepacia as well as its ability to become highly resistant towards many of the antibiotics currently in use. In addition, studies have also been undertaken to understand the pathogen's capacity to adapt and survive in a broad range of environments. Transposon based mutagenesis has been widely used in creating insertional knock-out mutants and coupled with recent advances in sequencing technology, robust tools to study gene function in a genome-wide manner have been developed based on the assembly of saturated transposon mutant libraries. In this study, we describe the construction of a large-scale library of B. cenocepacia transposon mutants. To create transposon mutants of B. cenocepacia strain J2315, electrocompetent bacteria were electrotransformed with the EZ-Tn5 transposome. Tetracyline resistant colonies were harvested off selective agar and pooled. Mutants were generated in multiple batches with each batch consisting of ˜20,000 to 40,000 mutants. Transposon insertion was validated by PCR amplification of the transposon region. In conclusion, a saturated B. cenocepacia J2315 transposon mutant library with an estimated total number of 500,000 mutants was successfully constructed. This mutant library can now be further exploited as a genetic tool to assess the function of every gene in the genome, facilitating the discovery of genes important for bacterial survival and adaptation, as well as virulence.

  6. New transposon delivery plasmids for insertional mutagenesis in Bacillus anthracis

    PubMed Central

    Wilson, Adam C.; Perego, Marta; Hoch, James A.

    2007-01-01

    Two new transposon delivery vector systems utilizing Mariner and mini-Tn10 transposons have been developed for in vivo insertional mutagenesis in Bacillus anthracis and other compatible Gram-positive species. The utility of both systems was directly demonstrated through the mutagenesis of a widely used B. anthracis strain. PMID:17931726

  7. Large scale screen for transposon insertions into cloned genes.

    PubMed Central

    Hamilton, B A; Palazzolo, M J; Chang, J H; VijayRaghavan, K; Mayeda, C A; Whitney, M A; Meyerowitz, E M

    1991-01-01

    We describe a method of screening for transposon insertions in or near Drosophila loci that correspond to cloned DNA sequences. We mobilize a modified P element transposon that carries a bacterial plasmid origin of replication and a drug-resistance marker. The genomic sequences flanking each transposon insertion site can then be rescued as a plasmid in Escherichia coli. Libraries of such plasmids, representing pools of transposon-mutagenized individuals, are used as hybridization probes against cloned sequences to determine whether a transposon has inserted next to a particular site in the genome. The number of loci that can be screened simultaneously by this procedure is quite large. We have screened an array of cDNA clones representing almost 700 distinct loci against libraries representing 760 mutagenized flies, and we obtained hybridization signals to 7 different cDNAs. Three of these events have been analyzed in detail and represent genuine insertions near genomic sequences that correspond to the cDNAs. Images PMID:1849274

  8. Germline mutagenesis mediated by Sleeping Beauty transposon system in mice

    PubMed Central

    Takeda, Junji; Keng, Vincent W; Horie, Kyoji

    2007-01-01

    Following the descovery of its transposition activity in mammalian culture systems, the Sleeping Beauty (SB) transposon has since been applied to achieve germline mutagenesis in mice. Initially, the transposition efficiency was found to be low in cultured systems, but its activity in germ cells was unexpectedly high. This difference in transposition efficiency was found to be largely dependent on chromosomal status of the host genomic DNA and transposon vector design. The SB transposon system has been found to be suitable for comprehensive mutagenesis in mice. Therefore, it is an effective tool as a forward genetics screen for tagged insertional mutagenesis in mice. PMID:18047691

  9. Transposon Tn916 mutagenesis in Bacillus anthracis.

    PubMed Central

    Ivins, B E; Welkos, S L; Knudson, G B; Leblanc, D J

    1988-01-01

    Mutagenesis of Bacillus anthracis by the streptococcal tetracycline resistance transposon Tn916 is described. Tn916 was transferred from Streptococcus faecalis DS16C1 to B. anthracis VNR-1 by conjugation in a standard filter mating procedure. Tetracycline-resistant (Tcr) transconjugants were obtained at a frequency of 1.6 X 10(-8) per donor CFU. When donor and recipient cells were treated with nafcillin before conjugation, the frequency was increased nearly 10-fold. Nafcillin pretreatment of donor and recipient strains was used in all subsequent conjugation experiments. S.faecalis CG110, containing multiple chromosomal insertions of Tn916, transferred the transposon to B. anthracis VNR-1 at a frequency of 9.3 x 10(-5). A Tcr B. anthracis transconjugant, strain VNR-1-tet-1, transferred Tn916 to B. anthracis UM23-1 and Bacillus subtilis BST1 at frequencies of 2.1 x 10(-4) and 5.8 X 10(-6), respectively. The transfer of Tn916 occurred only on membrane filters, since no Tcr transconjugants were obtained when strains VNR-1-tet-1 and UM23-1 were mixed and incubated in broth culture. The presence of the Tn916-associated tetM gene in Tcr B. anthracis and B. subtilis transconjugants was confirmed in hybridization experiments by using a 5-kilobase-pair DNA fragment containing the tetM gene as a probe. Of 3,000 B. anthracis UM23-1 Tcr transconjugants tested, 21 were phenylalanine auxotrophs and 2 were auxotrophic for phenylalanine, tyrosine, and tryptophan. Images PMID:2826334

  10. A mariner transposon vector adapted for mutagenesis in oral streptococci

    PubMed Central

    Nilsson, Martin; Christiansen, Natalia; Høiby, Niels; Twetman, Svante; Givskov, Michael; Tolker-Nielsen, Tim

    2014-01-01

    This article describes the construction and characterization of a mariner-based transposon vector designed for use in oral streptococci, but with a potential use in other Gram-positive bacteria. The new transposon vector, termed pMN100, contains the temperature-sensitive origin of replication repATs-pWV01, a selectable kanamycin resistance gene, a Himar1 transposase gene regulated by a xylose-inducible promoter, and an erythromycin resistance gene flanked by himar inverted repeats. The pMN100 plasmid was transformed into Streptococcus mutans UA159 and transposon mutagenesis was performed via a protocol established to perform high numbers of separate transpositions despite a low frequency of transposition. The distribution of transposon inserts in 30 randomly picked mutants suggested that mariner transposon mutagenesis is unbiased in S. mutans. A generated transposon mutant library containing 5000 mutants was used in a screen to identify genes involved in the production of sucrose-dependent extracellular matrix components. Mutants with transposon inserts in genes encoding glycosyltransferases and the competence-related secretory locus were predominantly found in this screen. PMID:24753509

  11. Transposon mutagenesis in Actinobacillus pleuropneumoniae with a Tn10 derivative.

    PubMed Central

    Tascon, R I; Rodriguez-Ferri, E F; Gutierrez-Martin, C B; Rodriguez-Barbosa, I; Berche, P; Vazquez-Boland, J A

    1993-01-01

    A transposon mutagenesis procedure functional in the gram-negative swine pathogen Actinobacillus pleuropneumoniae was developed for the first time. The technique involved the use of a suicide conjugative plasmid, pLOF/Km, carrying a mini-Tn10 with an isopropyl-beta-D-thiogalactopyranoside (IPTG)-inducible transposase located outside the mobile element (M. Herrero, V. de Lorenzo, and K. N. Timmis, J. Bacteriol. 172:6557-6567, 1990). The plasmid was mobilized from Escherichia coli to A. pleuropneumoniae through the RP4-mediated broad-host-range conjugal transfer functions provided by the chromosome of the donor strain. When IPTG was present in the mating medium, A. pleuropneumoniae CM5 transposon mutants were obtained at a frequency of 10(-5), while no mutants were detected in the absence of IPTG. Since the frequency of conjugal transfer of the RP4 plasmid from E. coli to A. pleuropneumoniae CM5 was found to be as low as 10(-4), the above result indicated that the expression level of the transposase was a critical factor for obtaining a workable efficiency of transposon mutagenesis. The transposon insertions occurred at random, as determined by Southern blotting of chromosomal DNA of randomly selected mutants and by the ability to generate mutants defective for the selected phenotypes. Almost all the mutants analyzed resulted from a single insertion of the Tn10 element. About 1.2% of the mutants resulted from the cointegration of pLOF/Km into the A. pleuropneumoniae chromosome. The applicability of this transposon mutagenesis system was verified on other A. pleuropneumoniae strains of different serotypes. The usefulness of this transposon mutagenesis system in genetic studies of A. pleuropneumoniae is discussed. Images PMID:8396122

  12. Transposon-mediated mutagenesis of a baculovirus.

    PubMed

    Fraser, M J; Brusca, J S; Smith, G E; Summers, M D

    1985-09-01

    Spontaneous mutants of insect nuclear polyhedrosis viruses of Autographa californica (AcMNPV) and Galleria mellonella (GmMNPV) were analyzed. These mutants produce few polyhedra in infected cells and have insertions of host cell DNA. All the insertions mapped to two adjacent sites in the genome. The junctions between two host insertions and the viral DNA were sequenced. One of the insertions contained a perfect 7-bp inverted terminal repeat, and had caused a direct duplication of 4 bp of viral DNA at the point of insertion. Therefore, this insertion sequence has properties of a transposon of the host cell Trichoplusia ni.

  13. Sleeping Beauty transposon insertional mutagenesis based mouse models for cancer gene discovery

    PubMed Central

    Moriarity, Branden S; Largaespada, David A

    2016-01-01

    Large-scale genomic efforts to study human cancer, such as the cancer gene atlas (TCGA), have identified numerous cancer drivers in a wide variety of tumor types. However, there are limitations to this approach, the mutations and expression or copy number changes that are identified are not always clearly functionally relevant, and only annotated genes and genetic elements are thoroughly queried. The use of complimentary, nonbiased, functional approaches to identify drivers of cancer development and progression is ideal to maximize the rate at which cancer discoveries are achieved. One such approach that has been successful is the use of the Sleeping Beauty (SB) transposon-based mutagenesis system in mice. This system uses a conditionally expressed transposase and mutagenic transposon allele to target mutagenesis to somatic cells of a given tissue in mice to cause random mutations leading to tumor development. Analysis of tumors for transposon common insertion sites (CIS) identifies candidate cancer genes specific to that tumor type. While similar screens have been performed in mice with the PiggyBac (PB) transposon and viral approaches, we limit extensive discussion to SB. Here we discuss the basic structure of these screens, screens that have been performed, methods used to identify CIS. PMID:26051241

  14. Sleeping Beauty transposon insertional mutagenesis based mouse models for cancer gene discovery.

    PubMed

    Moriarity, Branden S; Largaespada, David A

    2015-02-01

    Large-scale genomic efforts to study human cancer, such as the cancer gene atlas (TCGA), have identified numerous cancer drivers in a wide variety of tumor types. However, there are limitations to this approach, the mutations and expression or copy number changes that are identified are not always clearly functionally relevant, and only annotated genes and genetic elements are thoroughly queried. The use of complimentary, nonbiased, functional approaches to identify drivers of cancer development and progression is ideal to maximize the rate at which cancer discoveries are achieved. One such approach that has been successful is the use of the Sleeping Beauty (SB) transposon-based mutagenesis system in mice. This system uses a conditionally expressed transposase and mutagenic transposon allele to target mutagenesis to somatic cells of a given tissue in mice to cause random mutations leading to tumor development. Analysis of tumors for transposon common insertion sites (CIS) identifies candidate cancer genes specific to that tumor type. While similar screens have been performed in mice with the PiggyBac (PB) transposon and viral approaches, we limit extensive discussion to SB. Here we discuss the basic structure of these screens, screens that have been performed, methods used to identify CIS.

  15. Mariner-based transposon mutagenesis for Bacteroides species.

    PubMed

    Ichimura, Minoru; Uchida, Keiko; Nakayama-Imaohji, Haruyuki; Hirakawa, Hideki; Tada, Tomoyo; Morita, Hidetoshi; Yasutomo, Koji; Okazaki, Katsuichiro; Kuwahara, Tomomi

    2014-06-01

    Bacteroides is one of the most predominant groups of human gut microbiota. Recent metagenomic analyses and studies on gnotobiotic mice demonstrated the tight association of Bacteroides with epithelial function, the gut immune system and systemic metabolism in the host. The mariner family transposon shows relatively low target site specificity and has hosts ranging from prokaryotes to eukaryotes. Thereby, random mutagenesis using the mariner family transposon is expected to identify key molecules for human-Bacteroides symbiosis. In this study, we constructed the plasmid pMI07 to deliver the gene cassette (ermF/ITR), which harbors the erythromycin resistant marker (ermF) and the inverted repeat sequences (ITRs) recognized by Himar1 transposase, to Bacteroides via electrotransformation. pMI07 successfully delivered ermF/ITR to the Bacteroides genomes and generated thousands of insertion mutants/μg of pMI07 in B. thetaiotaomicron, B. fragilis, B. ovatus, and also, although to a lesser extent, B. vulgatus. Analyses of the ermF/ITR insertion sites in B. thetaiotaomicron and B. vulgatus revealed that the cassette targeted the dinucleotide TA and integrated into the genomes in an unbiased manner. The data reported here will provide useful information for transposon mutagenesis in Bacteroides species, which will enable identification of the genes responsible for their unique phenotypes. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Large-scale analysis of the yeast genome by transposon tagging and gene disruption.

    PubMed

    Ross-Macdonald, P; Coelho, P S; Roemer, T; Agarwal, S; Kumar, A; Jansen, R; Cheung, K H; Sheehan, A; Symoniatis, D; Umansky, L; Heidtman, M; Nelson, F K; Iwasaki, H; Hager, K; Gerstein, M; Miller, P; Roeder, G S; Snyder, M

    1999-11-25

    Economical methods by which gene function may be analysed on a genomic scale are relatively scarce. To fill this need, we have developed a transposon-tagging strategy for the genome-wide analysis of disruption phenotypes, gene expression and protein localization, and have applied this method to the large-scale analysis of gene function in the budding yeast Saccharomyces cerevisiae. Here we present the largest collection of defined yeast mutants ever generated within a single genetic background--a collection of over 11,000 strains, each carrying a transposon inserted within a region of the genome expressed during vegetative growth and/or sporulation. These insertions affect nearly 2,000 annotated genes, representing about one-third of the 6,200 predicted genes in the yeast genome. We have used this collection to determine disruption phenotypes for nearly 8,000 strains using 20 different growth conditions; the resulting data sets were clustered to identify groups of functionally related genes. We have also identified over 300 previously non-annotated open reading frames and analysed by indirect immunofluorescence over 1,300 transposon-tagged proteins. In total, our study encompasses over 260,000 data points, constituting the largest functional analysis of the yeast genome ever undertaken.

  17. Sleeping Beauty transposon mutagenesis in rat spermatogonial stem cells.

    PubMed

    Ivics, Zoltán; Izsvák, Zsuzsanna; Medrano, Gerardo; Chapman, Karen M; Hamra, F Kent

    2011-09-08

    We describe an experimental approach for generating mutant alleles in rat spermatogonial stem cells (SSCs) using Sleeping Beauty (SB) transposon-mediated insertional mutagenesis. The protocol is based on mobilization of mutagenic gene-trap transposons from transfected plasmid vectors into the genomes of cultured stem cells. Cells with transposon insertions in expressed genes are selected on the basis of activation of an antibiotic-resistance gene encoded by the transposon. These gene-trap clones are transplanted into the testes of recipient males (either as monoclonal or polyclonal libraries); crossing of these founders with wild-type females allows the insertions to be passed to F(1) progeny. This simple, economic and user-friendly methodological pipeline enables screens for functional gene annotation in the rat, with applicability in other vertebrate models where germ line-competent stem cells have been established. The complete protocol from transfection of SSCs to the genotyping of heterozygous F(1) offspring that harbor genomic SB gene-trap insertions takes 5-6 months.

  18. Genes Necessary for Bacterial Magnetite Biomineralization Identified by Transposon Mutagenesis

    NASA Astrophysics Data System (ADS)

    Nash, C. Z.; Komeili, A.; Newman, D. K.; Kirschvink, J. L.

    2004-12-01

    Magnetic bacteria synthesize nanoscale crystals of magnetite in intracellular, membrane-bounded organelles (magnetosomes). These crystals are preserved in the fossil record at least as far back as the late Neoproterozoic and have been tentatively identified in much older rocks (1). This fossil record may provide deep time calibration points for molecular evolution studies once the genes involved in biologically controlled magnetic mineralization (BCMM) are known. Further, a genetic and biochemical understanding of BCMM will give insight into the depositional environment and biogeochemical cycles in which magnetic bacteria play a role. The BCMM process is not well understood, though proteins have been identified from the magnetosome membrane and genetic manipulation and biochemical characterization of these proteins are underway. Most of the proteins currently thought to be involved are encoded within the mam cluster, a large cluster of genes whose products localize to the magnetosome membrane and are conserved among magnetic bacteria (2). In an effort to identify all of the genes necessary for bacterial BCMM, we undertook a transposon mutagenesis of Magnetospirillum magneticum AMB-1. Non-magnetic mutants (MNMs) were identified by growth in liquid culture followed by a magnetic assay. The insertion site of the transposon was identified two ways. First MNMs were screened with a PCR assay to determine if the transposon had inserted into the mam cluster. Second, the transposon was rescued from the mutant DNA and cloned for sequencing. The majority insertion sites are located within the mam cluster. Insertion sites also occur in operons which have not previously been suspected to be involved in magnetite biomineralization. None of the insertion sites have occurred within genes reported from previous transposon mutagenesis studies of AMB-1 (3, 4). Two of the non-mam cluster insertion sites occur in operons containing genes conserved particularly between MS-1 and MC-1. We

  19. Statistical methods for building random transposon mutagenesis libraries.

    PubMed

    Will, Oliver

    2008-01-01

    During the construction of random transposon mutagenesis libraries, four essential statistical issues arise: (1) Computing basic probability results for number of open reading frame knockouts. (2) Estimating the number of new open reading frames that will be knockouts in the next set of clones. (3) Estimating the number of essential open reading frames. (4) Computing the probability that an open reading frame is essential given the distribution of insertions. This chapter examines these issues and evaluates potential solutions using three different approaches: Efron and Thisted's estimator, Will and Jacobs's parametric bootstrap, and Blades and Broman's Gibbs sampler. In doing so, this chapter provides guidance for using the R statistical project to solve these problems.

  20. Establishment of Tn5096-Based Transposon Mutagenesis in Gordonia polyisoprenivorans

    PubMed Central

    Banh, Quyen; Arenskötter, Matthias; Steinbüchel, Alexander

    2005-01-01

    The transposons Tn5, Tn10, Tn611, and Tn5096 were characterized regarding transposition in Gordonia polyisoprenivorans strain VH2. No insertional mutants were obtained employing Tn5 or Tn10. The thermosensitive plasmid pCG79 harboring Tn611 integrated into the chromosome of G. polyisoprenivorans; however, the insertional mutants were fairly unstable und reverted frequently to the wild-type phenotype. In contrast, various stable mutants were obtained employing Tn5096-mediated transposon mutagenesis. Auxotrophic mutants, mutants defective or deregulated in carotenoid biosynthesis, and mutants defective in utilization of rubber and/or highly branched isoprenoid hydrocarbons were obtained by integration of plasmid pMA5096 harboring Tn5096 as a whole into the genome. From about 25,000 isolated mutants, the insertion loci of pMA5096 were subsequently mapped in 20 independent mutants in genes which could be related to the above-mentioned metabolic pathways or to putative regulation proteins. Analyses of the genotypes of pMA5096-mediated mutants defective in biodegradation of poly(cis-1,4-isoprene) did not reveal homologues to recently identified genes coding for enzymes catalyzing the initial cleavage of poly(cis-1,4-isoprene). One rubber-negative mutant was disrupted in mcr, encoding an α-methylacyl-coenzyme A racemase. This mutant was defective in degradation of poly(cis-1,4-isoprene) and also of highly branched isoprenoid hydrocarbons. PMID:16151089

  1. An efficient strategy for large-scale high-throughput transposon-mediated sequencing of cDNA clones

    PubMed Central

    Butterfield, Yaron S. N.; Marra, Marco A.; Asano, Jennifer K.; Chan, Susanna Y.; Guin, Ranabir; Krzywinski, Martin I.; Lee, Soo Sen; MacDonald, Kim W. K.; Mathewson, Carrie A.; Olson, Teika E.; Pandoh, Pawan K.; Prabhu, Anna-Liisa; Schnerch, Angelique; Skalska, Ursula; Smailus, Duane E.; Stott, Jeff M.; Tsai, Miranda I.; Yang, George S.; Zuyderduyn, Scott D.; Schein, Jacqueline E.; Jones, Steven J. M.

    2002-01-01

    We describe an efficient high-throughput method for accurate DNA sequencing of entire cDNA clones. Developed as part of our involvement in the Mammalian Gene Collection full-length cDNA sequencing initiative, the method has been used and refined in our laboratory since September 2000. Amenable to large scale projects, we have used the method to generate >7 Mb of accurate sequence from 3695 candidate full-length cDNAs. Sequencing is accomplished through the insertion of Mu transposon into cDNAs, followed by sequencing reactions primed with Mu-specific sequencing primers. Transposon insertion reactions are not performed with individual cDNAs but rather on pools of up to 96 clones. This pooling strategy reduces the number of transposon insertion sequencing libraries that would otherwise be required, reducing the costs and enhancing the efficiency of the transposon library construction procedure. Sequences generated using transposon-specific sequencing primers are assembled to yield the full-length cDNA sequence, with sequence editing and other sequence finishing activities performed as required to resolve sequence ambiguities. Although analysis of the many thousands (22 785) of sequenced Mu transposon insertion events revealed a weak sequence preference for Mu insertion, we observed insertion of the Mu transposon into 1015 of the possible 1024 5mer candidate insertion sites. PMID:12034834

  2. The utility of transposon mutagenesis for cancer studies in the era of genome editing.

    PubMed

    DeNicola, Gina M; Karreth, Florian A; Adams, David J; Wong, Chi C

    2015-10-19

    The use of transposons as insertional mutagens to identify cancer genes in mice has generated a wealth of information over the past decade. Here, we discuss recent major advances in transposon-mediated insertional mutagenesis screens and compare this technology with other screening strategies.

  3. Generation of Enterobacter sp. YSU auxotrophs using transposon mutagenesis.

    PubMed

    Caguiat, Jonathan James

    2014-10-31

    Prototrophic bacteria grow on M-9 minimal salts medium supplemented with glucose (M-9 medium), which is used as a carbon and energy source. Auxotrophs can be generated using a transposome. The commercially available, Tn5-derived transposome used in this protocol consists of a linear segment of DNA containing an R6Kγ replication origin, a gene for kanamycin resistance and two mosaic sequence ends, which serve as transposase binding sites. The transposome, provided as a DNA/transposase protein complex, is introduced by electroporation into the prototrophic strain, Enterobacter sp. YSU, and randomly incorporates itself into this host's genome. Transformants are replica plated onto Luria-Bertani agar plates containing kanamycin, (LB-kan) and onto M-9 medium agar plates containing kanamycin (M-9-kan). The transformants that grow on LB-kan plates but not on M-9-kan plates are considered to be auxotrophs. Purified genomic DNA from an auxotroph is partially digested, ligated and transformed into a pir+ Escherichia coli (E. coli) strain. The R6Kγ replication origin allows the plasmid to replicate in pir+ E. coli strains, and the kanamycin resistance marker allows for plasmid selection. Each transformant possesses a new plasmid containing the transposon flanked by the interrupted chromosomal region. Sanger sequencing and the Basic Local Alignment Search Tool (BLAST) suggest a putative identity of the interrupted gene. There are three advantages to using this transposome mutagenesis strategy. First, it does not rely on the expression of a transposase gene by the host. Second, the transposome is introduced into the target host by electroporation, rather than by conjugation or by transduction and therefore is more efficient. Third, the R6Kγ replication origin makes it easy to identify the mutated gene which is partially recovered in a recombinant plasmid. This technique can be used to investigate the genes involved in other characteristics of Enterobacter sp. YSU or of a

  4. Random insertion and gene disruption via transposon mutagenesis of Ureaplasma parvum using a mini-transposon plasmid.

    PubMed

    Aboklaish, Ali F; Dordet-Frisoni, Emilie; Citti, Christine; Toleman, Mark A; Glass, John I; Spiller, O Brad

    2014-11-01

    While transposon mutagenesis has been successfully used for Mycoplasma spp. to disrupt and determine non-essential genes, previous attempts with Ureaplasma spp. have been unsuccessful. Using a polyethylene glycol-transformation enhancing protocol, we were able to transform three separate serovars of Ureaplasma parvum with a Tn4001-based mini-transposon plasmid containing a gentamicin resistance selection marker. Despite the large degree of homology between Ureaplasma parvum and Ureaplasma urealyticum, all attempts to transform the latter in parallel failed, with the exception of a single clinical U. urealyticum isolate. PCR probing and sequencing were used to confirm transposon insertion into the bacterial genome and identify disrupted genes. Transformation of prototype serovar 3 consistently resulted in transfer only of sequence between the mini-transposon inverted repeats, but some strains showed additional sequence transfer. Transposon insertion occurred randomly in the genome resulting in unique disruption of genes UU047, UU390, UU440, UU450, UU520, UU526, UU582 for single clones from a panel of screened clones. An intergenic insertion between genes UU187 and UU188 was also characterised. Two phenotypic alterations were observed in the mutated strains: Disruption of a DEAD-box RNA helicase (UU582) altered growth kinetics, while the U. urealyticum strain lost resistance to serum attack coincident with disruption of gene UUR10_137 and loss of expression of a 41 kDa protein. Transposon mutagenesis was used successfully to insert single copies of a mini-transposon into the genome and disrupt genes leading to phenotypic changes in Ureaplasma parvum strains. This method can now be used to deliver exogenous genes for expression and determine essential genes for Ureaplasma parvum replication in culture and experimental models.

  5. Transposon mutagenesis as an approach to improved understanding of Borrelia pathogenesis and biology

    PubMed Central

    Lin, Tao; Troy, Erin B.; Hu, Linden T.; Gao, Lihui; Norris, Steven J.

    2014-01-01

    Transposon insertion provides a method for near-random mutation of bacterial genomes, and has been utilized extensively for the study of bacterial pathogenesis and biology. This approach is particularly useful for organisms that are relatively refractory to genetic manipulation, including Lyme disease Borrelia. In this review, progress to date in the application of transposon mutagenesis to the study of Borrelia burgdorferi is reported. An effective Himar1-based transposon vector has been developed and used to acquire a sequence-defined library of nearly 4500 mutants in the infectious, moderately transformable B. burgdorferi B31 derivative 5A18NP1. Analysis of these transposon mutants using signature-tagged mutagenesis (STM) and Tn-seq approaches has begun to yield valuable information regarding the genes important in the pathogenesis and biology of this organism. PMID:24904839

  6. A highly efficient transposon mutagenesis system for the tomato pathogen Clavibacter michiganensis subsp. michiganensis.

    PubMed

    Kirchner, O; Gartemann, K H; Zellermann, E M; Eichenlaub, R; Burger, A

    2001-11-01

    A transposon mutagenesis system for Clavibacter michiganensis subsp. michiganensis was developed based on antibiotic resistance transposons that were derived from the insertion element IS1409 from Arthrobacter sp. strain TM1 NCIB12013. As a prerequisite, the electroporation efficiency was optimized by using unmethylated DNA and treatment of the cells with glycine such that about 5 x 10(6) transformants per microg of DNA were generally obtained. Electroporation of C. michiganensis subsp. michiganensis with a suicide vector carrying transposon Tn1409C resulted in approximately 1 x 10(3) transposon mutants per pg of DNA and thus is suitable for saturation mutagenesis. Analysis of Tn1409C insertion sites suggests a random mode of transposition. Transposition of Tn1409C was also demonstrated for other subspecies of C. michiganensis.

  7. Excavating the Genome: Large Scale Mutagenesis Screening for the Discovery of New Mouse Models

    PubMed Central

    Sundberg, John P.; Dadras, Soheil S.; Silva, Kathleen A.; Kennedy, Victoria E.; Murray, Stephen A.; Denegre, James; Schofield, Paul N.; King, Lloyd E.; Wiles, Michael; Pratt, C. Herbert

    2016-01-01

    Technology now exists for rapid screening of mutated laboratory mice to identify phenotypes associated with specific genetic mutations. Large repositories exist for spontaneous mutants and those induced by chemical mutagenesis, many of which have never been studied or comprehensively evaluated. To supplement these resources, a variety of techniques have been consolidated in an international effort to create mutations in all known protein coding genes in the mouse. With targeted embryonic stem cell lines now available for almost all protein coding genes and more recently CRISPR/Cas9 technology, large-scale efforts are underway to create novel mutant mouse strains and to characterize their phenotypes. However, accurate diagnosis of skin, hair, and nail diseases still relies on careful gross and histological analysis. While not automated to the level of the physiological phenotyping, histopathology provides the most direct and accurate diagnosis and correlation with human diseases. As a result of these efforts, many new mouse dermatological disease models are being developed. PMID:26551941

  8. Large-scale functional annotation and expanded implementations of the P{wHy} hybrid transposon in the Drosophila melanogaster genome.

    PubMed

    Myrick, Kyl V; Huet, François; Mohr, Stephanie E; Alvarez-García, Inés; Lu, Jeffrey T; Smith, Mark A; Crosby, Madeline A; Gelbart, William M

    2009-07-01

    Whole genome sequencing of the model organisms has created increased demand for efficient tools to facilitate the genome annotation efforts. Accordingly, we report the further implementations and analyses stemming from our publicly available P{wHy} library for Drosophila melanogaster. A two-step regime-large scale transposon mutagenesis followed by hobo-induced nested deletions-allows mutation saturation and provides significant enhancements to existing genomic coverage. We previously showed that, for a given starting insert, deletion saturation is readily obtained over a 60-kb interval; here, we perform a breakdown analysis of efficiency to identify rate-limiting steps in the process. Transrecombination, the hobo-induced recombination between two P{wHy} half molecules, was shown to further expand the P{wHy} mutational range, pointing to a potent, iterative process of transrecombination-reconstitution-transrecombination for alternating between very large and very fine-grained deletions in a self-contained manner. A number of strains also showed partial or complete repression of P{wHy} markers, depending on chromosome location, whereby asymmetric marker silencing allowed continuous phenotypic detection, indicating that P{wHy}-based saturational mutagenesis should be useful for the study of heterochromatin/positional effects.

  9. MouseNet database: digital management of a large-scale mutagenesis project.

    PubMed

    Pargent, W; Heffner, S; Schäble, K F; Soewarto, D; Fuchs, H; Hrabé de Angelis, M

    2000-07-01

    The Munich ENU Mouse Mutagenesis Screen is a large-scale mutant production, phenotyping, and mapping project. It encompasses two animal breeding facilities and a number of screening groups located in the general area of Munich. A central database is required to manage and process the immense amount of data generated by the mutagenesis project. This database, which we named MouseNet(c), runs on a Sybase platform and will finally store and process all data from the entire project. In addition, the system comprises a portfolio of functions needed to support the workflow management of the core facility and the screening groups. MouseNet(c) will make all of the data available to the participating screening groups, and later to the international scientific community. MouseNet(c) will consist of three major software components:* Animal Management System (AMS)* Sample Tracking System (STS)* Result Documentation System (RDS)MouseNet(c) provides the following major advantages:* being accessible from different client platforms via the Internet* being a full-featured multi-user system (including access restriction and data locking mechanisms)* relying on a professional RDBMS (relational database management system) which runs on a UNIX server platform* supplying workflow functions and a variety of plausibility checks.

  10. Insertional mutagenesis by a hybrid piggyBac and sleeping beauty transposon in the rat.

    PubMed

    Furushima, Kenryo; Jang, Chuan-Wei; Chen, Diane W; Xiao, Ningna; Overbeek, Paul A; Behringer, Richard R

    2012-12-01

    A hybrid piggyBac/Sleeping Beauty transposon-based insertional mutagenesis system that can be mobilized by simple breeding was established in the rat. These transposons were engineered to include gene trap sequences and a tyrosinase (Tyr) pigmentation reporter to rescue the albinism of the genetic background used in the mutagenesis strategy. Single-copy transposon insertions were transposed into the rat genome by co-injection of plasmids carrying the transposon and RNA encoding piggyBac transposase into zygotes. The levels of transgenic Tyr expression were influenced by chromosomal context, leading to transgenic rats with different pigmentation that enabled visual genotyping. Transgenic rats designed to ubiquitously express either piggyBac or Sleeping Beauty transposase were generated by standard zygote injection also on an albino background. Bigenic rats carrying single-copy transposons at known loci and transposase transgenes exhibited coat color mosaicism, indicating somatic transposition. PiggyBac or Sleeping Beauty transposase bigenic rats bred with wild-type albino rats yielded offspring with pigmentation distinct from the initial transposon insertions as a consequence of germline transposition to new loci. The germline transposition frequency for Sleeping Beauty and piggyBac was ∼10% or about one new insertion per litter. Approximately 50% of the insertions occurred in introns. Chimeric transcripts containing endogenous and gene trap sequences were identified in Gabrb1 mutant rats. This mutagenesis system based on simple crosses and visual genotyping can be used to generate a collection of single-gene mutations in the rat.

  11. Insertional Mutagenesis by a Hybrid PiggyBac and Sleeping Beauty Transposon in the Rat

    PubMed Central

    Furushima, Kenryo; Jang, Chuan-Wei; Chen, Diane W.; Xiao, Ningna; Overbeek, Paul A.; Behringer, Richard R.

    2012-01-01

    A hybrid piggyBac/Sleeping Beauty transposon-based insertional mutagenesis system that can be mobilized by simple breeding was established in the rat. These transposons were engineered to include gene trap sequences and a tyrosinase (Tyr) pigmentation reporter to rescue the albinism of the genetic background used in the mutagenesis strategy. Single-copy transposon insertions were transposed into the rat genome by co-injection of plasmids carrying the transposon and RNA encoding piggyBac transposase into zygotes. The levels of transgenic Tyr expression were influenced by chromosomal context, leading to transgenic rats with different pigmentation that enabled visual genotyping. Transgenic rats designed to ubiquitously express either piggyBac or Sleeping Beauty transposase were generated by standard zygote injection also on an albino background. Bigenic rats carrying single-copy transposons at known loci and transposase transgenes exhibited coat color mosaicism, indicating somatic transposition. PiggyBac or Sleeping Beauty transposase bigenic rats bred with wild-type albino rats yielded offspring with pigmentation distinct from the initial transposon insertions as a consequence of germline transposition to new loci. The germline transposition frequency for Sleeping Beauty and piggyBac was ∼10% or about one new insertion per litter. Approximately 50% of the insertions occurred in introns. Chimeric transcripts containing endogenous and gene trap sequences were identified in Gabrb1 mutant rats. This mutagenesis system based on simple crosses and visual genotyping can be used to generate a collection of single-gene mutations in the rat. PMID:23023007

  12. Sleeping Beauty transposon mutagenesis identifies genes that cooperate with mutant Smad4 in gastric cancer development.

    PubMed

    Takeda, Haruna; Rust, Alistair G; Ward, Jerrold M; Yew, Christopher Chin Kuan; Jenkins, Nancy A; Copeland, Neal G

    2016-04-05

    Mutations in SMAD4 predispose to the development of gastrointestinal cancer, which is the third leading cause of cancer-related deaths. To identify genes driving gastric cancer (GC) development, we performed a Sleeping Beauty (SB) transposon mutagenesis screen in the stomach of Smad4(+/-) mutant mice. This screen identified 59 candidate GC trunk drivers and a much larger number of candidate GC progression genes. Strikingly, 22 SB-identified trunk drivers are known or candidate cancer genes, whereas four SB-identified trunk drivers, including PTEN, SMAD4, RNF43, and NF1, are known human GC trunk drivers. Similar to human GC, pathway analyses identified WNT, TGF-β, and PI3K-PTEN signaling, ubiquitin-mediated proteolysis, adherens junctions, and RNA degradation in addition to genes involved in chromatin modification and organization as highly deregulated pathways in GC. Comparative oncogenomic filtering of the complete list of SB-identified genes showed that they are highly enriched for genes mutated in human GC and identified many candidate human GC genes. Finally, by comparing our complete list of SB-identified genes against the list of mutated genes identified in five large-scale human GC sequencing studies, we identified LDL receptor-related protein 1B (LRP1B) as a previously unidentified human candidate GC tumor suppressor gene. In LRP1B, 129 mutations were found in 462 human GC samples sequenced, and LRP1B is one of the top 10 most deleted genes identified in a panel of 3,312 human cancers. SB mutagenesis has, thus, helped to catalog the cooperative molecular mechanisms driving SMAD4-induced GC growth and discover genes with potential clinical importance in human GC.

  13. Sleeping Beauty transposon mutagenesis identifies genes that cooperate with mutant Smad4 in gastric cancer development

    PubMed Central

    Takeda, Haruna; Rust, Alistair G.; Ward, Jerrold M.; Yew, Christopher Chin Kuan; Jenkins, Nancy A.; Copeland, Neal G.

    2016-01-01

    Mutations in SMAD4 predispose to the development of gastrointestinal cancer, which is the third leading cause of cancer-related deaths. To identify genes driving gastric cancer (GC) development, we performed a Sleeping Beauty (SB) transposon mutagenesis screen in the stomach of Smad4+/− mutant mice. This screen identified 59 candidate GC trunk drivers and a much larger number of candidate GC progression genes. Strikingly, 22 SB-identified trunk drivers are known or candidate cancer genes, whereas four SB-identified trunk drivers, including PTEN, SMAD4, RNF43, and NF1, are known human GC trunk drivers. Similar to human GC, pathway analyses identified WNT, TGF-β, and PI3K-PTEN signaling, ubiquitin-mediated proteolysis, adherens junctions, and RNA degradation in addition to genes involved in chromatin modification and organization as highly deregulated pathways in GC. Comparative oncogenomic filtering of the complete list of SB-identified genes showed that they are highly enriched for genes mutated in human GC and identified many candidate human GC genes. Finally, by comparing our complete list of SB-identified genes against the list of mutated genes identified in five large-scale human GC sequencing studies, we identified LDL receptor-related protein 1B (LRP1B) as a previously unidentified human candidate GC tumor suppressor gene. In LRP1B, 129 mutations were found in 462 human GC samples sequenced, and LRP1B is one of the top 10 most deleted genes identified in a panel of 3,312 human cancers. SB mutagenesis has, thus, helped to catalog the cooperative molecular mechanisms driving SMAD4-induced GC growth and discover genes with potential clinical importance in human GC. PMID:27006499

  14. Large-Scale Transposition Mutagenesis of Streptomyces coelicolor Identifies Hundreds of Genes Influencing Antibiotic Biosynthesis.

    PubMed

    Xu, Zhong; Wang, Yemin; Chater, Keith F; Ou, Hong-Yu; Xu, H Howard; Deng, Zixin; Tao, Meifeng

    2017-03-15

    Gram-positive Streptomyces bacteria produce thousands of bioactive secondary metabolites, including antibiotics. To systematically investigate genes affecting secondary metabolism, we developed a hyperactive transposase-based Tn5 transposition system and employed it to mutagenize the model species Streptomyces coelicolor, leading to the identification of 51,443 transposition insertions. These insertions were distributed randomly along the chromosome except for some preferred regions associated with relatively low GC content in the chromosomal core. The base composition of the insertion site and its flanking sequences compiled from the 51,443 insertions implied a 19-bp expanded target site surrounding the insertion site, with a slight nucleic acid base preference in some positions, suggesting a relative randomness of Tn5 transposition targeting in the high-GC Streptomyces genome. From the mutagenesis library, 724 mutants involving 365 genes had altered levels of production of the tripyrrole antibiotic undecylprodigiosin (RED), including 17 genes in the RED biosynthetic gene cluster. Genetic complementation revealed that most of the insertions (more than two-thirds) were responsible for the changed antibiotic production. Genes associated with branched-chain amino acid biosynthesis, DNA metabolism, and protein modification affected RED production, and genes involved in signaling, stress, and transcriptional regulation were overrepresented. Some insertions caused dramatic changes in RED production, identifying future targets for strain improvement.IMPORTANCE High-GC Gram-positive streptomycetes and related actinomycetes have provided more than 100 clinical drugs used as antibiotics, immunosuppressants, and antitumor drugs. Their genomes harbor biosynthetic genes for many more unknown compounds with potential as future drugs. Here we developed a useful genome-wide mutagenesis tool based on the transposon Tn5 for the study of secondary metabolism and its regulation

  15. A Plasmid-Transposon Hybrid Mutagenesis System Effective in a Broad Range of Enterobacteria

    PubMed Central

    Monson, Rita; Smith, Debra S.; Matilla, Miguel A.; Roberts, Kevin; Richardson, Elizabeth; Drew, Alison; Williamson, Neil; Ramsay, Josh; Welch, Martin; Salmond, George P. C.

    2015-01-01

    Random transposon mutagenesis is a powerful technique used to generate libraries of genetic insertions in many different bacterial strains. Here we develop a system facilitating random transposon mutagenesis in a range of different Gram-negative bacterial strains, including Pectobacterium atrosepticum, Citrobacter rodentium, Serratia sp. ATCC39006, Serratia plymuthica, Dickeya dadantii, and many more. Transposon mutagenesis was optimized in each of these strains and three studies are presented to show the efficacy of this system. Firstly, the important agricultural pathogen D. dadantii was mutagenized. Two mutants that showed reduced protease production and one mutant producing the previously cryptic pigment, indigoidine, were identified and characterized. Secondly, the enterobacterium, Serratia sp. ATCC39006 was mutagenized and mutants incapable of producing gas vesicles, proteinaceous intracellular organelles, were identified. One of these contained a β-galactosidase transcriptional fusion within the gene gvpA1, essential for gas vesicle production. Finally, the system was used to mutate the biosynthetic gene clusters of the antifungal, anti-oomycete and anticancer polyketide, oocydin A, in the plant-associated enterobacterium, Dickeya solani MK10. The mutagenesis system was developed to allow easy identification of transposon insertion sites by sequencing, after facile generation of a replicon encompassing the transposon and adjacent DNA, post-excision. Furthermore, the system can also create transcriptional fusions with either β-galactosidase or β-glucuronidase as reporters, and exploits a variety of drug resistance markers so that multiple selectable fusions can be generated in a single strain. This system of various transposons has wide utility and can be combined in many different ways. PMID:26733980

  16. A Plasmid-Transposon Hybrid Mutagenesis System Effective in a Broad Range of Enterobacteria.

    PubMed

    Monson, Rita; Smith, Debra S; Matilla, Miguel A; Roberts, Kevin; Richardson, Elizabeth; Drew, Alison; Williamson, Neil; Ramsay, Josh; Welch, Martin; Salmond, George P C

    2015-01-01

    Random transposon mutagenesis is a powerful technique used to generate libraries of genetic insertions in many different bacterial strains. Here we develop a system facilitating random transposon mutagenesis in a range of different Gram-negative bacterial strains, including Pectobacterium atrosepticum, Citrobacter rodentium, Serratia sp. ATCC39006, Serratia plymuthica, Dickeya dadantii, and many more. Transposon mutagenesis was optimized in each of these strains and three studies are presented to show the efficacy of this system. Firstly, the important agricultural pathogen D. dadantii was mutagenized. Two mutants that showed reduced protease production and one mutant producing the previously cryptic pigment, indigoidine, were identified and characterized. Secondly, the enterobacterium, Serratia sp. ATCC39006 was mutagenized and mutants incapable of producing gas vesicles, proteinaceous intracellular organelles, were identified. One of these contained a β-galactosidase transcriptional fusion within the gene gvpA1, essential for gas vesicle production. Finally, the system was used to mutate the biosynthetic gene clusters of the antifungal, anti-oomycete and anticancer polyketide, oocydin A, in the plant-associated enterobacterium, Dickeya solani MK10. The mutagenesis system was developed to allow easy identification of transposon insertion sites by sequencing, after facile generation of a replicon encompassing the transposon and adjacent DNA, post-excision. Furthermore, the system can also create transcriptional fusions with either β-galactosidase or β-glucuronidase as reporters, and exploits a variety of drug resistance markers so that multiple selectable fusions can be generated in a single strain. This system of various transposons has wide utility and can be combined in many different ways.

  17. Evaluating Risks of Insertional Mutagenesis by DNA Transposons in Gene Therapy

    PubMed Central

    Hackett, Perry B.; Largaespada, David A.; Switzer, Kirsten C.; Cooper, Laurence J.N.

    2013-01-01

    Investigational therapy can be successfully undertaken using viral- and non-viral-mediated ex vivo gene transfer. Indeed, recent clinical trials have established the potential for genetically modified T cells to improve and restore health. Recently the Sleeping Beauty (SB) transposon/transposase system has been applied in clinical trials to stably insert a chimeric antigen receptor (CAR) to redirect T-cell specificity. We discuss the context in which the SB system can be harnessed for gene therapy and describe the human application of SB-modified CAR+ T cells. We have focused on theoretical issues relating to insertional mutagenesis in the context of human genomes that are naturally subjected to remobilization of transposons and the experimental evidence over the last decade of employing SB transposons for defining genes that induce cancer. These findings are put into the context of the use of SB transposons in the treatment of human disease. PMID:23313630

  18. Development of an inducible transposon system for efficient random mutagenesis in Clostridium acetobutylicum

    PubMed Central

    Zhang, Ying; Xu, Shu; Chai, Changsheng; Yang, Sheng; Jiang, Weihong; Minton, Nigel P.; Gu, Yang

    2016-01-01

    Clostridium acetobutylicum is an industrially important Gram-positive organism, which is capable of producing economically important chemicals in the ABE (Acetone, Butanol and Ethanol) fermentation process. Renewed interests in the ABE process necessitate the availability of additional genetics tools to facilitate the derivation of a greater understanding of the underlying metabolic and regulatory control processes in operation through forward genetic strategies. In this study, a xylose inducible, mariner-based, transposon system was developed and shown to allow high-efficient random mutagenesis in the model strain ATCC 824. Of the thiamphenicol resistant colonies obtained, 91.9% were shown to be due to successful transposition of the catP-based mini-transposon element. Phenotypic screening of 200 transposon clones revealed a sporulation-defective clone with an insertion in spo0A, thereby demonstrating that this inducible transposon system can be used for forward genetic studies in C. acetobutylicum. PMID:27001972

  19. Evaluating risks of insertional mutagenesis by DNA transposons in gene therapy.

    PubMed

    Hackett, Perry B; Largaespada, David A; Switzer, Kirsten C; Cooper, Laurence J N

    2013-04-01

    Investigational therapy can be successfully undertaken using viral- and nonviral-mediated ex vivo gene transfer. Indeed, recent clinical trials have established the potential for genetically modified T cells to improve and restore health. Recently, the Sleeping Beauty (SB) transposon/transposase system has been applied in clinical trials to stably insert a chimeric antigen receptor (CAR) to redirect T-cell specificity. We discuss the context in which the SB system can be harnessed for gene therapy and describe the human application of SB-modified CAR(+) T cells. We have focused on theoretical issues relating to insertional mutagenesis in the context of human genomes that are naturally subjected to remobilization of transposons and the experimental evidence over the last decade of employing SB transposons for defining genes that induce cancer. These findings are put into the context of the use of SB transposons in the treatment of human disease.

  20. mariner-Based Transposon Mutagenesis of Rickettsia prowazekii▿ †

    PubMed Central

    Liu, Zhi-Mei; Tucker, Aimee M.; Driskell, Lonnie O.; Wood, David O.

    2007-01-01

    Rickettsia prowazekii, the causative agent of epidemic typhus, is an obligate intracellular bacterium that grows directly within the cytoplasm of its host cell, unbounded by a vacuolar membrane. The obligate intracytoplasmic nature of rickettsial growth places severe restrictions on the genetic analysis of this distinctive human pathogen. In order to expand the repertoire of genetic tools available for the study of this pathogen, we have employed the versatile mariner-based, Himar1 transposon system to generate insertional mutants of R. prowazekii. A transposon containing the R. prowazekii arr-2 rifampin resistance gene and a gene coding for a green fluorescent protein (GFPUV) was constructed and placed on a plasmid expressing the Himar1 transposase. Electroporation of this plasmid into R. prowazekii resulted in numerous transpositions into the rickettsial genome. Transposon insertion sites were identified by rescue cloning, followed by DNA sequencing. Random transpositions integrating at TA sites in both gene coding and intergenic regions were identified. Individual rickettsial clones were isolated by the limiting-dilution technique. Using both fixed and live-cell techniques, R. prowazekii transformants expressing GFPUV were easily visible by fluorescence microscopy. Thus, a mariner-based system provides an additional mechanism for generating rickettsial mutants that can be screened using GFPUV fluorescence. PMID:17720821

  1. Engineered Zymomonas mobilis for salt tolerance using EZ-Tn5-based transposon insertion mutagenesis system.

    PubMed

    Wang, Jing-Li; Wu, Bo; Qin, Han; You, Yang; Liu, Song; Shui, Zong-Xia; Tan, Fu-Rong; Wang, Yan-Wei; Zhu, Qi-Li; Li, Yan-Bin; Ruan, Zhi-Yong; Ma, Ke-Dong; Dai, Li-Chun; Hu, Guo-Quan; He, Ming-Xiong

    2016-06-10

    The cell growth and ethanol yield of Zymomonas mobilis may be detrimentally affected by salt stress frequently present in some biomass-based fermentation systems, leading to a decrease in the rate of sugar conversion to ethanol or other bioproducts. To address this problem, improving the salt tolerance of Z. mobilis is a desirable way. However, limited progress has been made in development of Z. mobilis with higher salt tolerance for some technical challenges in the past decades. Recently, transposon insertion mutant system has been widely used as a novel genetic tool in many organisms to develop mutant strains. In this study, Tn5-based transposon insertion mutagenesis system firstly used for construction of higher salt tolerance strain in Z. mobilis. Approximately 200 Z. mobilis ZM4 mutants were generated by using Tn5-based transposon mutagenesis system. The mutant strain ZMT2 with improved salt tolerance phenotype was obtained by screening on RM agar plates with additional 1 % NaCl. Strain ZMT2 was confirmed to exhibit better fermentation performance under NaCl stress than wild type of strain ZM4. The transposon insertion was located in ZMO1122 (himA) by genome walking. Discruption of himA gene showed that himA may play an important role in response to salt tolerance in Z. mobils. The mutant strain ZMT2 with a transposon insertion in himA gene of the genome showed obviously higher sugar conversion rate to ethonal under up to 2 % NaCl stress than did the wild ZM4 strain. Besides, ZMT2 exhibited shared fermentative capabilities with wild ZM4 strain under no or low NaCl stress. This report firstly showed that himA played a role in responding to NaCl stress. Furthermore, the result indicated that Tn5-based transposon mutagenesis system was a feasible tool not only for genetic engineering in Z. mobilis strain improvement, but also in tapping resistent genes.

  2. Isolation of Magnetospirillum magneticum AMB-1 mutants defective in bacterial magnetic particle synthesis by transposon mutagenesis.

    PubMed

    Wahyudi, A T; Takeyama, H; Matsunaga, T

    2001-01-01

    Nonmagnetic mutants of Magnetospirillum magneticum AMB-1 were recovered following mini-Tn5 transposon mutagenesis. Transconjugants with kanamycin resistance were obtained at a frequency of 2.7 x 10(-7) per recipient. Of 3327 transconjugants, 62 were defective for bacterial magnetic particle (BMP) synthesis. The frequency of independent transposition events for nonmagnetic mutants was about 1.4% in transconjugants. Further analysis of DNA sequences flanking transposon by inverted polymerase chain reaction allowed isolation of at least 10 genes or DNA sequences involved in BMP synthesis in M. magneticum AMB-1.

  3. Gene transfer and mutagenesis mediated by Sleeping Beauty transposon in Nile tilapia (Oreochromis niloticus).

    PubMed

    He, Xiaozhen; Li, Jie; Long, Yong; Song, Guili; Zhou, Peiyong; Liu, Qiuxiang; Zhu, Zuoyan; Cui, Zongbin

    2013-10-01

    The success of gene transfer has been demonstrated in many of vertebrate species, whereas the efficiency of producing transgenic animals remains pretty low due to the random integration of foreign genes into a recipient genome. The Sleeping Beauty (SB) transposon is able to improve the efficiency of gene transfer in zebrafish and mouse, but its activity in tilapia (Oreochromis niloticus) has yet to be characterized. Herein, we demonstrate the potential of using the SB transposon system as an effective tool for gene transfer and insertional mutagenesis in tilapia. A transgenic construct pT2/tiHsp70-SB11 was generated by subcloning the promoter of tilapia heat shock protein 70 (tiHsp70) gene, the SB11 transposase gene and the carp β-actin gene polyadenylation signal into the second generation of SB transposon. Transgenic tilapia was produced by microinjection of this construct with in vitro synthesized capped SB11 mRNA. SB11 transposon was detected in 28.89 % of founders, 12.9 % of F1 and 43.75 % of F2. Analysis of genomic sequences flanking integrated transposons indicates that this transgenic tilapia line carries two copies of SB transposon, which landed into two different endogenous genes. Induced expression of SB11 gene after heat shock was detected using reverse transcription PCR in F2 transgenic individuals. In addition, the Cre/loxP system was introduced to delete the SB11 cassette for stabilization of gene interruption and bio-safety. These findings suggest that the SB transposon system is active and can be used for efficient gene transfer and insertional mutagenesis in tilapia.

  4. Mutagenesis of barley malting quality QTLs with Ds transposons.

    PubMed

    Singh, Surinder; Tan, Han Qi; Singh, Jaswinder

    2012-03-01

    Various functional genomic tools are being used to identify and characterize genes in plants. The Activator/Dissociation (Ac/Ds) transposon-based approach offers great potential, especially in barley, due to its limited success of genetic transformation and its large genome size. The bias of the Ac/Ds system towards genic regions and its tendency toward localized transpositions can greatly enhance the discovery and tagging of genes linked to Ds. Barley is a key ingredient in malting and brewing industry; therefore, gene discovery in relation to malting has an industrial perspective. Malting quality in barley is a complex and quantitatively inherited trait. Two major quantitative trait loci (QTLs) affecting malting quality traits have been located on chromosome 4H. In this study, Ds was reactivated from parent transposants (TNP) lines, TNP-29 and TNP-79, where Ds was mapped in the vicinity of important malting QTLs. Reactivation of Ds was carried out both by conventional breeding and in vitro approaches. A threefold increase in reactivation frequency through the in vitro approach enabled the development of a new genomic resource for the dissection of malting QTL and gene discovery in barley. Identification of unique flanking sequences, using high-efficiency thermal asymmetric interlaced PCR and inverse PCR from these populations, has further emphasized the new location of Ds in the barley genome and provided new transposon mutants especially in β-GAL1, β-amylase-like gene and ABC transporter for functional genomic studies.

  5. Transposon mutagenesis identifies genes that cooperate with mutant Pten in breast cancer progression.

    PubMed

    Rangel, Roberto; Lee, Song-Choon; Hon-Kim Ban, Kenneth; Guzman-Rojas, Liliana; Mann, Michael B; Newberg, Justin Y; Kodama, Takahiro; McNoe, Leslie A; Selvanesan, Luxmanan; Ward, Jerrold M; Rust, Alistair G; Chin, Kuan-Yew; Black, Michael A; Jenkins, Nancy A; Copeland, Neal G

    2016-11-29

    Triple-negative breast cancer (TNBC) has the worst prognosis of any breast cancer subtype. To better understand the genetic forces driving TNBC, we performed a transposon mutagenesis screen in a phosphatase and tensin homolog (Pten) mutant mice and identified 12 candidate trunk drivers and a much larger number of progression genes. Validation studies identified eight TNBC tumor suppressor genes, including the GATA-like transcriptional repressor TRPS1 Down-regulation of TRPS1 in TNBC cells promoted epithelial-to-mesenchymal transition (EMT) by deregulating multiple EMT pathway genes, in addition to increasing the expression of SERPINE1 and SERPINB2 and the subsequent migration, invasion, and metastasis of tumor cells. Transposon mutagenesis has thus provided a better understanding of the genetic forces driving TNBC and discovered genes with potential clinical importance in TNBC.

  6. Transposon mutagenesis identifies genes that cooperate with mutant Pten in breast cancer progression

    PubMed Central

    Rangel, Roberto; Lee, Song-Choon; Hon-Kim Ban, Kenneth; Guzman-Rojas, Liliana; Mann, Michael B.; Newberg, Justin Y.; McNoe, Leslie A.; Selvanesan, Luxmanan; Ward, Jerrold M.; Rust, Alistair G.; Chin, Kuan-Yew; Black, Michael A.; Jenkins, Nancy A.; Copeland, Neal G.

    2016-01-01

    Triple-negative breast cancer (TNBC) has the worst prognosis of any breast cancer subtype. To better understand the genetic forces driving TNBC, we performed a transposon mutagenesis screen in a phosphatase and tensin homolog (Pten) mutant mice and identified 12 candidate trunk drivers and a much larger number of progression genes. Validation studies identified eight TNBC tumor suppressor genes, including the GATA-like transcriptional repressor TRPS1. Down-regulation of TRPS1 in TNBC cells promoted epithelial-to-mesenchymal transition (EMT) by deregulating multiple EMT pathway genes, in addition to increasing the expression of SERPINE1 and SERPINB2 and the subsequent migration, invasion, and metastasis of tumor cells. Transposon mutagenesis has thus provided a better understanding of the genetic forces driving TNBC and discovered genes with potential clinical importance in TNBC. PMID:27849608

  7. Random transposon mutagenesis of the Saccharopolyspora erythraea genome reveals additional genes influencing erythromycin biosynthesis

    PubMed Central

    Fedashchin, Andrij; Cernota, William H.; Gonzalez, Melissa C.; Leach, Benjamin I.; Kwan, Noelle; Wesley, Roy K.; Weber, J. Mark

    2015-01-01

    A single cycle of strain improvement was performed in Saccharopolyspora erythraea mutB and 15 genotypes influencing erythromycin production were found. Genotypes generated by transposon mutagenesis appeared in the screen at a frequency of ∼3%. Mutations affecting central metabolism and regulatory genes were found, as well as hydrolases, peptidases, glycosyl transferases and unknown genes. Only one mutant retained high erythromycin production when scaled-up from micro-agar plug fermentations to shake flasks. This mutant had a knockout of the cwh1 gene (SACE_1598), encoding a cell-wall-associated hydrolase. The cwh1 knockout produced visible growth and morphological defects on solid medium. This study demonstrated that random transposon mutagenesis uncovers strain improvement-related genes potentially useful for strain engineering. PMID:26468041

  8. An inducible tool for random mutagenesis in Aspergillus niger based on the transposon Vader.

    PubMed

    Paun, Linda; Nitsche, Benjamin; Homan, Tim; Ram, Arthur F; Kempken, Frank

    2016-07-01

    The ascomycete Aspergillus niger is widely used in the biotechnology, for instance in producing most of the world's citric acid. It is also known as a major food and feed contaminant. While generation of gene knockouts for functional genomics has become feasible in ku70 mutants, analyzing gene functions or metabolic pathways remains a laborious task. An unbiased transposon-based mutagenesis approach may aid this process of analyzing gene functions by providing mutant libraries in a short time. The Vader transposon is a non-autonomous DNA-transposon, which is activated by the homologous tan1-transposase. However, in the most commonly used lab strain of A. niger (N400 strain and derivatives), we found that the transposase, encoded by the tan1 gene, is mutated and inactive. To establish a Vader transposon-based mutagenesis system in the N400 background, we expressed the functional transposase of A. niger strain CBS 513.88 under the control of an inducible promoter based on the Tet-on system, which is activated in the presence of the antibiotic doxycycline (DOX). Increasing amounts of doxycycline lead to higher Vader excision frequencies, whereas little to none activity of Vader was observed without addition of doxycycline. Hence, this system appears to be suitable for producing stable mutants in the A. niger N400 background.

  9. Steady-state transposon mutagenesis in inbred maize.

    PubMed

    McCarty, Donald R; Settles, Andrew Mark; Suzuki, Masaharu; Tan, Bao Cai; Latshaw, Susan; Porch, Tim; Robin, Kevin; Baier, John; Avigne, Wayne; Lai, Jinsheng; Messing, Joachim; Koch, Karen E; Hannah, L Curtis

    2005-10-01

    We implement a novel strategy for harnessing the power of high-copy transposons for functional analysis of the maize genome, and report behavioral features of the Mutator system in a uniform inbred background. The unique UniformMu population and database facilitate high-throughput molecular analysis of Mu-tagged mutants and gene knockouts. Key features of the population include: (i) high mutation frequencies (7% independent seed mutations) and moderation of copy number (approximately 57 total Mu elements; 1-2 MuDR copies per plant) were maintained by continuous back-crossing into a phenotypically uniform inbred background; (ii) a bz1-mum9 marker enabled selection of stable lines (loss of MuDR), inhibiting further transpositions in lines selected for molecular analysis; (iii) build-up of mutation load was prevented by screening Mu-active parents to exclude plants carrying pre-existing seed mutations. To create a database of genomic sequences flanking Mu insertions, selected mutant lines were analyzed by sequencing of MuTAIL PCR clone libraries. These sequences were annotated and clustered to facilitate bioinformatic subtraction of ancestral elements and identification of insertions unique to mutant lines. New insertions targeted low-copy, gene-rich sequences, and in silico mapping revealed a random distribution of insertions over the genome. Our results indicate that Mu populations differ markedly in the occurrence of Mu insertion hotspots and the frequency of suppressible mutations. We suggest that controlled MuDR copy number in UniformMu lines is a key determinant of these differences. The public database (http://uniformmu.org; http://endosperm.info) includes pedigree and phenotypic data for over 2000 independent seed mutants selected from a population of 31 548 F2 lines and integrated with analyses of 34 255 MuTAIL sequences.

  10. Sleeping Beauty Transposon Mutagenesis of the Rat Genome in Spermatogonial Stem Cells

    PubMed Central

    Ivics, Zoltán; Izsvák, Zsuzsanna; Chapman, Karen M.; Hamra, F. Kent

    2011-01-01

    Since several aspects of physiology in rats has evolved to be more similar to humans than that of mice, it is highly desirable to link the rat into the process of annotating the human genome with function. However, the lack of technology for generating defined mutants in the rat genome has hindered the identification of causative relationships between genes and disease phenotypes. As an important step towards this goal, an approach of establishing transposon-mediated insertional mutagenesis in rat spermatogonial stem cells was recently developed. Transposons can be viewed as natural DNA transfer vehicles that, similar to integrating viruses, are capable of efficient genomic insertion. The mobility of transposons can be controlled by conditionally providing the transposase component of the transposition reaction. Thus, a DNA of interest such as a mutagenic gene trap cassette cloned between the inverted repeat sequences of a transposon-based vector can be utilized for stable genomic insertion in a regulated and highly efficient manner. Gene trap transposons integrate into the genome in a random fashion, and those mutagenic insertions that occurred in expressed genes can be selected in vitro based on activation of a reporter. Selected monoclonal as well as polyclonal libraries of gene trap clones are transplanted into the testes of recipient/founder male rats allowing passage of the mutation through the germline to F1 progeny after only a single cross with wild-type females. This paradigm enables a powerful methodological pipeline for forward genetic screens for functional gene annotation in the rat, as well as other vertebrate models. This article provides a detailed description on how to culturerat spermatogonial stem cell lines, their transfection with transposon plasmids, selection of gene trap insertions with antibiotics, transplantation of genetically modified stem cells and genotyping of knockout animals. PMID:21193047

  11. Sleeping Beauty transposon mutagenesis of the rat genome in spermatogonial stem cells.

    PubMed

    Ivics, Zoltán; Izsvák, Zsuzsanna; Chapman, Karen M; Hamra, F Kent

    2011-04-01

    Since several aspects of physiology in rats have evolved to be more similar to humans than that of mice, it is highly desirable to link the rat into the process of annotating the human genome with function. However, the lack of technology for generating defined mutants in the rat genome has hindered the identification of causative relationships between genes and disease phenotypes. As an important step towards this goal, an approach of establishing transposon-mediated insertional mutagenesis in rat spermatogonial stem cells was recently developed. Transposons can be viewed as natural DNA transfer vehicles that, similar to integrating viruses, are capable of efficient genomic insertion. The mobility of transposons can be controlled by conditionally providing the transposase component of the transposition reaction. Thus, a DNA of interest such as a mutagenic gene trap cassette cloned between the inverted repeat sequences of a transposon-based vector can be utilized for stable genomic insertion in a regulated and highly efficient manner. Gene-trap transposons integrate into the genome in a random fashion, and those mutagenic insertions that occurred in expressed genes can be selected in vitro based on activation of a reporter. Selected monoclonal as well as polyclonal libraries of gene trap clones are transplanted into the testes of recipient/founder male rats allowing passage of the mutation through the germline to F1 progeny after only a single cross with wild-type females. This paradigm enables a powerful methodological pipeline for forward genetic screens for functional gene annotation in the rat, as well as other vertebrate models. This article provides a detailed description on how to culture rat spermatogonial stem cell lines, their transfection with transposon plasmids, selection of gene-trap insertions with antibiotics, transplantation of genetically modified stem cells and genotyping of knockout animals. Copyright © 2011 Elsevier Inc. All rights reserved.

  12. Phenotypic mutants of the intracellular actinomycete Rhodococcus equi created by in vivo Himar1 transposon mutagenesis.

    PubMed

    Ashour, Joseph; Hondalus, Mary K

    2003-04-01

    Rhodococcus equi is a facultative intracellular opportunistic pathogen of immunocompromised people and a major cause of pneumonia in young horses. An effective live attenuated vaccine would be extremely useful in the prevention of R. equi disease in horses. Toward that end, we have developed an efficient transposon mutagenesis system that makes use of a Himar1 minitransposon delivered by a conditionally replicating plasmid for construction of R. equi mutants. We show that Himar1 transposition in R. equi is random and needs no apparent consensus sequence beyond the required TA dinucleotide. The diversity of the transposon library was demonstrated by the ease with which we were able to screen for auxotrophs and mutants with pigmentation and capsular phenotypes. One of the pigmentation mutants contained an insertion in a gene encoding phytoene desaturase, an enzyme of carotenoid biosynthesis, the pathway necessary for production of the characteristic salmon color of R. equi. We identified an auxotrophic mutant with a transposon insertion in the gene encoding a putative dual-functioning GTP cyclohydrolase II-3,4-dihydroxy-2-butanone-4-phosphate synthase, an enzyme essential for riboflavin biosynthesis. This mutant cannot grow in minimal medium in the absence of riboflavin supplementation. Experimental murine infection studies showed that, in contrast to wild-type R. equi, the riboflavin-requiring mutant is attenuated because it is unable to replicate in vivo. The mutagenesis methodology we have developed will allow the characterization of R. equi virulence mechanisms and the creation of other attenuated strains with vaccine potential.

  13. Phenotypic Mutants of the Intracellular Actinomycete Rhodococcus equi Created by In Vivo Himar1 Transposon Mutagenesis

    PubMed Central

    Ashour, Joseph; Hondalus, Mary K.

    2003-01-01

    Rhodococcus equi is a facultative intracellular opportunistic pathogen of immunocompromised people and a major cause of pneumonia in young horses. An effective live attenuated vaccine would be extremely useful in the prevention of R. equi disease in horses. Toward that end, we have developed an efficient transposon mutagenesis system that makes use of a Himar1 minitransposon delivered by a conditionally replicating plasmid for construction of R. equi mutants. We show that Himar1 transposition in R. equi is random and needs no apparent consensus sequence beyond the required TA dinucleotide. The diversity of the transposon library was demonstrated by the ease with which we were able to screen for auxotrophs and mutants with pigmentation and capsular phenotypes. One of the pigmentation mutants contained an insertion in a gene encoding phytoene desaturase, an enzyme of carotenoid biosynthesis, the pathway necessary for production of the characteristic salmon color of R. equi. We identified an auxotrophic mutant with a transposon insertion in the gene encoding a putative dual-functioning GTP cyclohydrolase II-3,4-dihydroxy-2-butanone-4-phosphate synthase, an enzyme essential for riboflavin biosynthesis. This mutant cannot grow in minimal medium in the absence of riboflavin supplementation. Experimental murine infection studies showed that, in contrast to wild-type R. equi, the riboflavin-requiring mutant is attenuated because it is unable to replicate in vivo. The mutagenesis methodology we have developed will allow the characterization of R. equi virulence mechanisms and the creation of other attenuated strains with vaccine potential. PMID:12670990

  14. Transposon mutagenesis in Mycoplasma hyopneumoniae using a novel mariner-based system for generating random mutations.

    PubMed

    Maglennon, Gareth A; Cook, Beth S; Deeney, Alannah S; Bossé, Janine T; Peters, Sarah E; Langford, Paul R; Maskell, Duncan J; Tucker, Alexander W; Wren, Brendan W; Rycroft, Andrew N

    2013-12-21

    Mycoplasma hyopneumoniae is the cause of enzootic pneumonia in pigs, a chronic respiratory disease associated with significant economic losses to swine producers worldwide. The molecular pathogenesis of infection is poorly understood due to the lack of genetic tools to allow manipulation of the organism and more generally for the Mycoplasma genus. The objective of this study was to develop a system for generating random transposon insertion mutants in M. hyopneumoniae that could prove a powerful tool in enabling the pathogenesis of infection to be unraveled. A novel delivery vector was constructed containing a hyperactive C9 mutant of the Himar1 transposase along with a mini transposon containing the tetracycline resistance cassette, tetM. M. hyopneumoniae strain 232 was electroporated with the construct and tetM-expressing transformants selected on agar containing tetracycline. Individual transformants contained single transposon insertions that were stable upon serial passages in broth medium. The insertion sites of 44 individual transformants were determined and confirmed disruption of several M. hyopneumoniae genes. A large pool of over 10 000 mutants was generated that should allow saturation of the M. hyopneumoniae strain 232 genome. This is the first time that transposon mutagenesis has been demonstrated in this important pathogen and could be generally applied for other Mycoplasma species that are intractable to genetic manipulation. The ability to generate random mutant libraries is a powerful tool in the further study of the pathogenesis of this important swine pathogen.

  15. Transposon mutagenesis of Campylobacter jejuni identifies a bipartite energy taxis system required for motility.

    PubMed

    Hendrixson, D R; Akerley, B J; DiRita, V J

    2001-04-01

    Campylobacter jejuni constitutes the leading cause of bacterial gastroenteritis in the United States and a major cause of diarrhoea worldwide. Little is known about virulence mechanisms in this organism because of the scarcity of suitable genetic tools. We have developed an efficient system of in vitro transposon mutagenesis using a mariner-based transposon and purified mariner transposase. Through in vitro transposition of C. jejuni chromosomal DNA followed by natural transformation of the transposed DNA, large random transposon mutant libraries consisting of approximately 16 000 individual mutants were generated. The first genetic screen of C. jejuni using a transposon-generated mutant library identified 28 mutants defective for flagellar motility, one of the few known virulence determinants of this pathogen. We developed a second genetic system, which allows for the construction of defined chromosomal deletions in C. jejuni, and demonstrated the requirement of sigma28 and sigma54 for motility. In addition, we show that sigma28 is involved in the transcription of flaA and that sigma54 is required for transcription of three other flagellar genes, flaB and flgDE. We also identified two previously uncharacterized genes required for motility encoding proteins that we call CetA and CetB, which mediate energy taxis responses. Through our analysis of the Cet proteins, we propose a unique mechanism for sensing energy levels and mediating energy taxis in C. jejuni.

  16. Transposon Mutagenesis of Mycobacterium marinum Identifies a Locus Linking Pigmentation and Intracellular Survival

    PubMed Central

    Gao, Lian-Yong; Groger, Richard; Cox, Jeffery S.; Beverley, Stephen M.; Lawson, Elise H.; Brown, Eric J.

    2003-01-01

    Pathogenic mycobacteria survive and replicate within host macrophages, but the molecular mechanisms involved in this necessary step in the pathogenesis of infection are not completely understood. Mycobacterium marinum has recently been used as a model for aspects of the pathogenesis of tuberculosis because of its close genetic relationship to Mycobacterium tuberculosis and because of similarities in the pathology and course of infection caused by this organism in its natural hosts, fish and frogs, with tuberculosis in humans. In order to advance the utility of the M. marinum model, we have developed efficient transposon mutagenesis of the organism by using a Drosophila melanogaster mariner-based transposon. To determine the efficiency of transposition, we have analyzed pigmentation mutants from the transposon mutant library. In addition to insertions in four known genes in the pathway of pigment biosynthesis, two insertions in novel genes were identified in our mutant library. One of these is in a putative inhibitor of the carotenoid biosynthesis pathway. The second unexpected insertion is in an intergenic region between two genes homologous to Rv2603c and Rv2604c of M. tuberculosis. In addition to a pigmentation defect, this mutant showed increased susceptibility to singlet oxygen and grew poorly in murine macrophages. Complementation with M. tuberculosis genomic DNA encompassing Rv2603c to Rv2606c corrected the pigmentation and growth defects of the mutant. These data demonstrate the utility of mariner-based transposon mutagenesis of M. marinum and that M. marinum can be used to study the function of M. tuberculosis genes involved in intracellular survival and replication. PMID:12540574

  17. Transposon mutagenesis identifies genes driving hepatocellular carcinoma in a chronic hepatitis B mouse model

    PubMed Central

    Bard-Chapeau, Emilie A.; Nguyen, Anh-Tuan; Rust, Alistair G.; Sayadi, Ahmed; Lee, Philip; Chua, Belinda Q; New, Lee-Sun; de Jong, Johann; Ward, Jerrold M.; Chin, Christopher KY.; Chew, Valerie; Toh, Han Chong; Abastado, Jean-Pierre; Benoukraf, Touati; Soong, Richie; Bard, Frederic A.; Dupuy, Adam J.; Johnson, Randy L.; Radda, George K.; Chan, Eric CY.; Wessels, Lodewyk FA.; Adams, David J.

    2014-01-01

    The most common risk factor for developing hepatocellular carcinoma (HCC) is chronic infection with hepatitis B virus (HBV). To better understand the evolutionary forces driving HCC we performed a near saturating transposon mutagenesis screen in a mouse HBV model of HCC. This screen identified 21 candidate early stage drivers, and a bewildering number (2860) of candidate later stage drivers, that were enriched for genes mutated, deregulated, or that function in signaling pathways important for human HCC, with a striking 1199 genes linked to cellular metabolic processes. Our study provides a comprehensive overview of the genetic landscape of HCC. PMID:24316982

  18. Transposon mutagenesis in Desulfovibrio desulfuricans: Development of a random mutagenesis tool from Tn7

    SciTech Connect

    Wall, J.D.; Murnan, T.; Argyle, J.

    1996-10-01

    The transposons Tn5, Tn7, Tn9, and Tn10 or their derivatives have been examined for transposition in the sulfate-reducing bacterium Desulfovibrio desulfuricans G20. Tn7 inserted with a frequency of 10{sup {minus}4} to 10{sup {minus}3} into a unique attachment site that shows strong homology with those sites identified in other gram-negative bacteria. Inactivation of the tnsD gene in Tn7, encoding the function directing insertion into the unique site, yielded a derivative that transposed essentially randomly with a frequency of ca. 10{sup {minus}6} per donor. Derivatives of Tn5, but not wild-type Tn5, were also found to undergo random transposition at a similar frequency. No evidence was obtained for transposition of Tn9 or Tn10. 34 refs., 5 figs., 2 tabs.

  19. Sleeping Beauty Transposon Mutagenesis as a Tool for Gene Discovery in the NOD Mouse Model of Type 1 Diabetes

    PubMed Central

    Elso, Colleen M.; Chu, Edward P. F.; Alsayb, May A.; Mackin, Leanne; Ivory, Sean T.; Ashton, Michelle P.; Bröer, Stefan; Silveira, Pablo A.; Brodnicki, Thomas C.

    2015-01-01

    A number of different strategies have been used to identify genes for which genetic variation contributes to type 1 diabetes (T1D) pathogenesis. Genetic studies in humans have identified >40 loci that affect the risk for developing T1D, but the underlying causative alleles are often difficult to pinpoint or have subtle biological effects. A complementary strategy to identifying “natural” alleles in the human population is to engineer “artificial” alleles within inbred mouse strains and determine their effect on T1D incidence. We describe the use of the Sleeping Beauty (SB) transposon mutagenesis system in the nonobese diabetic (NOD) mouse strain, which harbors a genetic background predisposed to developing T1D. Mutagenesis in this system is random, but a green fluorescent protein (GFP)-polyA gene trap within the SB transposon enables early detection of mice harboring transposon-disrupted genes. The SB transposon also acts as a molecular tag to, without additional breeding, efficiently identify mutated genes and prioritize mutant mice for further characterization. We show here that the SB transposon is functional in NOD mice and can produce a null allele in a novel candidate gene that increases diabetes incidence. We propose that SB transposon mutagenesis could be used as a complementary strategy to traditional methods to help identify genes that, when disrupted, affect T1D pathogenesis. PMID:26438296

  20. Sleeping Beauty Transposon Mutagenesis as a Tool for Gene Discovery in the NOD Mouse Model of Type 1 Diabetes.

    PubMed

    Elso, Colleen M; Chu, Edward P F; Alsayb, May A; Mackin, Leanne; Ivory, Sean T; Ashton, Michelle P; Bröer, Stefan; Silveira, Pablo A; Brodnicki, Thomas C

    2015-10-04

    A number of different strategies have been used to identify genes for which genetic variation contributes to type 1 diabetes (T1D) pathogenesis. Genetic studies in humans have identified >40 loci that affect the risk for developing T1D, but the underlying causative alleles are often difficult to pinpoint or have subtle biological effects. A complementary strategy to identifying "natural" alleles in the human population is to engineer "artificial" alleles within inbred mouse strains and determine their effect on T1D incidence. We describe the use of the Sleeping Beauty (SB) transposon mutagenesis system in the nonobese diabetic (NOD) mouse strain, which harbors a genetic background predisposed to developing T1D. Mutagenesis in this system is random, but a green fluorescent protein (GFP)-polyA gene trap within the SB transposon enables early detection of mice harboring transposon-disrupted genes. The SB transposon also acts as a molecular tag to, without additional breeding, efficiently identify mutated genes and prioritize mutant mice for further characterization. We show here that the SB transposon is functional in NOD mice and can produce a null allele in a novel candidate gene that increases diabetes incidence. We propose that SB transposon mutagenesis could be used as a complementary strategy to traditional methods to help identify genes that, when disrupted, affect T1D pathogenesis. Copyright © 2015 Elso et al.

  1. Comprehensive Essentiality Analysis of the Mycobacterium tuberculosis Genome via Saturating Transposon Mutagenesis

    PubMed Central

    DeJesus, Michael A.; Gerrick, Elias R.; Xu, Weizhen; Park, Sae Woong; Long, Jarukit E.; Boutte, Cara C.; Rubin, Eric J.; Schnappinger, Dirk; Ehrt, Sabine; Fortune, Sarah M.; Sassetti, Christopher M.

    2017-01-01

    ABSTRACT   For decades, identifying the regions of a bacterial chromosome that are necessary for viability has relied on mapping integration sites in libraries of random transposon mutants to find loci that are unable to sustain insertion. To date, these studies have analyzed subsaturated libraries, necessitating the application of statistical methods to estimate the likelihood that a gap in transposon coverage is the result of biological selection and not the stochasticity of insertion. As a result, the essentiality of many genomic features, particularly small ones, could not be reliably assessed. We sought to overcome this limitation by creating a completely saturated transposon library in Mycobacterium tuberculosis. In assessing the composition of this highly saturated library by deep sequencing, we discovered that a previously unknown sequence bias of the Himar1 element rendered approximately 9% of potential TA dinucleotide insertion sites less permissible for insertion. We used a hidden Markov model of essentiality that accounted for this unanticipated bias, allowing us to confidently evaluate the essentiality of features that contained as few as 2 TA sites, including open reading frames (ORF), experimentally identified noncoding RNAs, methylation sites, and promoters. In addition, several essential regions that did not correspond to known features were identified, suggesting uncharacterized functions that are necessary for growth. This work provides an authoritative catalog of essential regions of the M. tuberculosis genome and a statistical framework for applying saturating mutagenesis to other bacteria. PMID:28096490

  2. Transposon and Deletion Mutagenesis of Genes Involved in Perchlorate Reduction in Azospira suillum PS

    PubMed Central

    Melnyk, Ryan A.; Clark, Iain C.; Liao, Annette; Coates, John D.

    2013-01-01

    ABSTRACT Although much work on the biochemistry of the key enzymes of bacterial perchlorate reduction, chlorite dismutase, and perchlorate reductase has been published, understanding of the molecular mechanisms of this metabolism has been somewhat hampered by the lack of a clear model system amenable to genetic manipulation. Using transposon mutagenesis and clean deletions, genes important for perchlorate reduction in Azospira suillum PS have been identified both inside and outside the previously described perchlorate reduction genomic island (PRI). Transposon mutagenesis identified 18 insertions in 11 genes that completely abrogate growth via reduction of perchlorate but have no phenotype during denitrification. Of the mutants deficient in perchlorate reduction, 14 had insertions that were mapped to eight different genes within the PRI, highlighting its importance in this metabolism. To further explore the role of these genes, we also developed systems for constructing unmarked deletions and for complementing these deletions. Using these tools, every core gene in the PRI was systematically deleted; 8 of the 17 genes conserved in the PRI are essential for perchlorate respiration, including 3 genes that comprise a unique histidine kinase system. Interestingly, the other 9 genes in the PRI are not essential for perchlorate reduction and may thus have unknown functions during this metabolism. We present a model detailing our current understanding of perchlorate reduction that incorporates new concepts about this metabolism. PMID:24381299

  3. Slingshot: a PiggyBac based transposon system for tamoxifen-inducible 'self-inactivating' insertional mutagenesis.

    PubMed

    Kong, Jun; Wang, Feng; Brenton, James D; Adams, David J

    2010-10-01

    We have developed a self-inactivating PiggyBac transposon system for tamoxifen inducible insertional mutagenesis from a stably integrated chromosomal donor. This system, which we have named 'Slingshot', utilizes a transposon carrying elements for both gain- and loss-of-function screens in vitro. We show that the Slingshot transposon can be efficiently mobilized from a range of chromosomal loci with high inducibility and low background generating insertions that are randomly dispersed throughout the genome. Furthermore, we show that once the Slingshot transposon has been mobilized it is not remobilized producing stable clonal integrants in all daughter cells. To illustrate the efficacy of Slingshot as a screening tool we set out to identify mediators of resistance to puromycin and the chemotherapeutic drug vincristine by performing genetrap screens in mouse embryonic stem cells. From these genome-wide screens we identified multiple independent insertions in the multidrug resistance transporter genes Abcb1a/b and Abcg2 conferring resistance to drug treatment. Importantly, we also show that the Slingshot transposon system is functional in other mammalian cell lines such as human HEK293, OVCAR-3 and PE01 cells suggesting that it may be used in a range of cell culture systems. Slingshot represents a flexible and potent system for genome-wide transposon-mediated mutagenesis with many potential applications.

  4. Transposon mutagenesis identifies genes and cellular processes driving epithelial-mesenchymal transition in hepatocellular carcinoma

    PubMed Central

    Kodama, Takahiro; Newberg, Justin Y.; Kodama, Michiko; Rangel, Roberto; Yoshihara, Kosuke; Tien, Jean C.; Parsons, Pamela H.; Wu, Hao; Finegold, Milton J.; Copeland, Neal G.; Jenkins, Nancy A.

    2016-01-01

    Epithelial-mesenchymal transition (EMT) is thought to contribute to metastasis and chemoresistance in patients with hepatocellular carcinoma (HCC), leading to their poor prognosis. The genes driving EMT in HCC are not yet fully understood, however. Here, we show that mobilization of Sleeping Beauty (SB) transposons in immortalized mouse hepatoblasts induces mesenchymal liver tumors on transplantation to nude mice. These tumors show significant down-regulation of epithelial markers, along with up-regulation of mesenchymal markers and EMT-related transcription factors (EMT-TFs). Sequencing of transposon insertion sites from tumors identified 233 candidate cancer genes (CCGs) that were enriched for genes and cellular processes driving EMT. Subsequent trunk driver analysis identified 23 CCGs that are predicted to function early in tumorigenesis and whose mutation or alteration in patients with HCC is correlated with poor patient survival. Validation of the top trunk drivers identified in the screen, including MET (MET proto-oncogene, receptor tyrosine kinase), GRB2-associated binding protein 1 (GAB1), HECT, UBA, and WWE domain containing 1 (HUWE1), lysine-specific demethylase 6A (KDM6A), and protein-tyrosine phosphatase, nonreceptor-type 12 (PTPN12), showed that deregulation of these genes activates an EMT program in human HCC cells that enhances tumor cell migration. Finally, deregulation of these genes in human HCC was found to confer sorafenib resistance through apoptotic tolerance and reduced proliferation, consistent with recent studies showing that EMT contributes to the chemoresistance of tumor cells. Our unique cell-based transposon mutagenesis screen appears to be an excellent resource for discovering genes involved in EMT in human HCC and potentially for identifying new drug targets. PMID:27247392

  5. Low-copy piggyBac transposon mutagenesis in mice identifies genes driving melanoma.

    PubMed

    Ni, Thomas K; Landrette, Sean F; Bjornson, Robert D; Bosenberg, Marcus W; Xu, Tian

    2013-09-17

    Despite considerable efforts to sequence hypermutated cancers such as melanoma, distinguishing cancer-driving genes from thousands of recurrently mutated genes remains a significant challenge. To circumvent the problematic background mutation rates and identify new melanoma driver genes, we carried out a low-copy piggyBac transposon mutagenesis screen in mice. We induced eleven melanomas with mutation burdens that were 100-fold lower relative to human melanomas. Thirty-eight implicated genes, including two known drivers of human melanoma, were classified into three groups based on high, low, or background-level mutation frequencies in human melanomas, and we further explored the functional significance of genes in each group. For two genes overlooked by prevailing discovery methods, we found that loss of membrane associated guanylate kinase, WW and PDZ domain containing 2 and protein tyrosine phosphatase, receptor type, O cooperated with the v-raf murine sarcoma viral oncogene homolog B (BRAF) recurrent V600E mutation to promote cellular transformation. Moreover, for infrequently mutated genes often disregarded by current methods, we discovered recurrent mitogen-activated protein kinase kinase kinase 1 (Map3k1)-activating insertions in our screen, mirroring recurrent MAP3K1 up-regulation in human melanomas. Aberrant expression of Map3k1 enabled growth factor-autonomous proliferation and drove BRAF-independent ERK signaling, thus shedding light on alternative means of activating this prominent signaling pathway in melanoma. In summary, our study contributes several previously undescribed genes involved in melanoma and establishes an important proof-of-principle for the utility of the low-copy transposon mutagenesis approach for identifying cancer-driving genes, especially those masked by hypermutation.

  6. Mammalian mutagenesis using a highly mobile somatic Sleeping Beauty transposon system.

    PubMed

    Dupuy, Adam J; Akagi, Keiko; Largaespada, David A; Copeland, Neal G; Jenkins, Nancy A

    2005-07-14

    Transposons have provided important genetic tools for functional genomic screens in lower eukaryotes but have proven less useful in higher eukaryotes because of their low transposition frequency. Here we show that Sleeping Beauty (SB), a member of the Tc1/mariner class of transposons, can be mobilized in mouse somatic cells at frequencies high enough to induce embryonic death and cancer in wild-type mice. Tumours are aggressive, with some animals developing two or even three different types of cancer within a few months of birth. The tumours result from SB insertional mutagenesis of cancer genes, thus facilitating the identification of genes and pathways that induce disease. SB transposition can easily be controlled to mutagenize any target tissue and can therefore, in principle, be used to induce many of the cancers affecting humans, including those for which little is known about the aetiology. The uses of SB are also not restricted to the mouse and could potentially be used for forward genetic screens in any higher eukaryote in which transgenesis is possible.

  7. Transposon-directed base-exchange mutagenesis (TDEM): a novel method for multiple-nucleotide substitutions within a target gene.

    PubMed

    Kim, Yun Cheol; Lee, Hui Sun; Yoon, Sukjoon; Morrison, Sherie L

    2009-06-01

    In this report we describe transposon-directed base-exchange mutagenesis (TDEM), an efficient and controllable method for introducing a mutation into a gene. Each round of TDEM can remove up to 11 base pairs from a randomly selected site within the target gene and replace them with any length of DNA of predetermined sequence. Therefore, the number of bases to be deleted and inserted can be independently regulated providing greater versatility than existing methods of transposon-based mutagenesis. Subsequently, multiple rounds of mutagenesis will provide a diverse mutant library that contains multiple mutations throughout the gene. Additionally, we developed a simple frame-checking procedure that eliminates nonfunctional mutants containing frameshifts or stop codons. As a proof of principle, we used TDEM to generate mutant lacZalpha lacking alpha-complementation activity and recovered active revertants using a second round of TDEM. Furthermore, a single round of TDEM yielded unique, inactive mutants of ccdB.

  8. Transposon mutagenesis identifies genetic drivers of BrafV600E melanoma

    PubMed Central

    Mann, Michael B; Black, Michael A; Jones, Devin J; Ward, Jerrold M; Yew, Christopher Chin Kuan; Newberg, Justin Y; Dupuy, Adam J; Rust, Alistair G; Bosenberg, Marcus W; McMahon, Martin; Print, Cristin G; Copeland, Neal G; Jenkins, Nancy A

    2016-01-01

    Although nearly half of human melanomas harbor oncogenic BRAFV600E mutations, the genetic events that cooperate with these mutations to drive melanogenesis are still largely unknown. Here we show that Sleeping Beauty (SB) transposon-mediated mutagenesis drives melanoma progression in BrafV600E mutant mice and identify 1,232 recurrently mutated candidate cancer genes (CCGs) from 70 SB-driven melanomas. CCGs are enriched in Wnt, PI3K, MAPK and netrin signaling pathway components and are more highly connected to one another than predicted by chance, indicating that SB targets cooperative genetic networks in melanoma. Human orthologs of >500 CCGs are enriched for mutations in human melanoma or showed statistically significant clinical associations between RNA abundance and survival of patients with metastatic melanoma. We also functionally validate CEP350 as a new tumor-suppressor gene in human melanoma. SB mutagenesis has thus helped to catalog the cooperative molecular mechanisms driving BRAFV600E melanoma and discover new genes with potential clinical importance in human melanoma. PMID:25848750

  9. Transposon mutagenesis identifies genetic drivers of Braf(V600E) melanoma.

    PubMed

    Mann, Michael B; Black, Michael A; Jones, Devin J; Ward, Jerrold M; Yew, Christopher Chin Kuan; Newberg, Justin Y; Dupuy, Adam J; Rust, Alistair G; Bosenberg, Marcus W; McMahon, Martin; Print, Cristin G; Copeland, Neal G; Jenkins, Nancy A

    2015-05-01

    Although nearly half of human melanomas harbor oncogenic BRAF(V600E) mutations, the genetic events that cooperate with these mutations to drive melanogenesis are still largely unknown. Here we show that Sleeping Beauty (SB) transposon-mediated mutagenesis drives melanoma progression in Braf(V600E) mutant mice and identify 1,232 recurrently mutated candidate cancer genes (CCGs) from 70 SB-driven melanomas. CCGs are enriched in Wnt, PI3K, MAPK and netrin signaling pathway components and are more highly connected to one another than predicted by chance, indicating that SB targets cooperative genetic networks in melanoma. Human orthologs of >500 CCGs are enriched for mutations in human melanoma or showed statistically significant clinical associations between RNA abundance and survival of patients with metastatic melanoma. We also functionally validate CEP350 as a new tumor-suppressor gene in human melanoma. SB mutagenesis has thus helped to catalog the cooperative molecular mechanisms driving BRAF(V600E) melanoma and discover new genes with potential clinical importance in human melanoma.

  10. In vivo growth characteristics of leucine and methionine auxotrophic mutants of Mycobacterium bovis BCG generated by transposon mutagenesis.

    PubMed Central

    McAdam, R A; Weisbrod, T R; Martin, J; Scuderi, J D; Brown, A M; Cirillo, J D; Bloom, B R; Jacobs, W R

    1995-01-01

    Insertional mutagenesis in Mycobacterium bovis BCG, a member of the slow-growing M. tuberculosis complex, was accomplished with transposons engineered from the Mycobacterium smegmatis insertion element IS1096. Transposons were created by placing a kanamycin resistance gene in several different positions in IS1096, and the resulting transposons were electroporated into BCG on nonreplicating plasmids. These analyses demonstrated that only one of the two open reading frames was necessary for transposition. A library of insertions was generated. Southern analysis of 23 kanamycin-resistant clones revealed that the transposons had inserted directly, with no evidence of cointegrate formation, into different restriction fragments in each clone. Sequence analysis of nine of the clones revealed junctional direct 8-bp repeats with only a slight similarity in target sites. These results suggest that IS1096-derived transposons transposed into the BCG genome in a relatively random fashion. Three auxotrophs, two for leucine and one for methionine, were isolated from the library of transposon insertions in BCG. They were characterized by sequencing and found to be homologous to the leuD gene of Escherichia coli and a sulfate-binding protein of cyanobacteria, respectively. When inoculated intravenously into C57BL/6 mice, the leucine auxotrophs, in contrast to the parent BCG strain or the methionine auxotroph, showed an inability to grow in vivo and were cleared within 7 weeks from the lungs and spleen. PMID:7868221

  11. Transposon and deletion mutagenesis of genes involved in perchlorate reduction in Azospira suillum PS.

    PubMed

    Melnyk, Ryan A; Clark, Iain C; Liao, Annette; Coates, John D

    2013-12-31

    Although much work on the biochemistry of the key enzymes of bacterial perchlorate reduction, chlorite dismutase, and perchlorate reductase has been published, understanding of the molecular mechanisms of this metabolism has been somewhat hampered by the lack of a clear model system amenable to genetic manipulation. Using transposon mutagenesis and clean deletions, genes important for perchlorate reduction in Azospira suillum PS have been identified both inside and outside the previously described perchlorate reduction genomic island (PRI). Transposon mutagenesis identified 18 insertions in 11 genes that completely abrogate growth via reduction of perchlorate but have no phenotype during denitrification. Of the mutants deficient in perchlorate reduction, 14 had insertions that were mapped to eight different genes within the PRI, highlighting its importance in this metabolism. To further explore the role of these genes, we also developed systems for constructing unmarked deletions and for complementing these deletions. Using these tools, every core gene in the PRI was systematically deleted; 8 of the 17 genes conserved in the PRI are essential for perchlorate respiration, including 3 genes that comprise a unique histidine kinase system. Interestingly, the other 9 genes in the PRI are not essential for perchlorate reduction and may thus have unknown functions during this metabolism. We present a model detailing our current understanding of perchlorate reduction that incorporates new concepts about this metabolism. Although perchlorate is generated naturally in the environment, groundwater contamination is largely a result of industrial activity. Bacteria capable of respiring perchlorate and remediating contaminated water have been isolated, but relatively little is known about the biochemistry and genetics of this process. Here we used two complementary approaches to identify genes involved in perchlorate reduction. Most of these genes are located on a genomic island

  12. Novel strategies for gene trapping and insertional mutagenesis mediated by Sleeping Beauty transposon.

    PubMed

    Song, Guili; Cui, Zongbin

    2013-09-01

    Gene and poly(A) trappings are high-throughput approaches to capture and interrupt the expression of endogenous genes within a target genome. Although a number of trapping vectors have been developed for investigation of gene functions in cells and vertebrate models, there is still room for the improvement of their efficiency and sensitivity. Recently, two novel trapping vectors mediated by Sleeping Beauty (SB) transposon have been generated by the combination of three functional cassettes that are required for finding endogenous genes, disrupting the expression of trapped genes, and inducing the excision of integrated traps from their original insertion sites and then inserting into another gene. In addition, several other strategies are utilized to improve the activities of two trapping vectors. First, activities of all components were examined in vitro before the generation of two vectors. Second, the inducible promoter from the tilapia Hsp70 gene was used to drive the expression of SB gene, which can mediate the excision of integrated transposons upon induction at 37 °C. Third, the Cre/LoxP system was introduced to delete the SB expression cassette for stabilization of gene interruption and bio-safety. Fourth, three stop codons in different reading frames were introduced downstream of a strong splice acceptor (SA) in the gene trapping vector to effectively terminate the translation of trapped endogenous genes. Fifth, the strong splicing donor (SD) and AU-rich RNA-destabilizing element exhibited no obvious insertion bias and markedly reduced SD read-through events, and the combination of an enhanced SA, a poly(A) signal and a transcript terminator in the poly(A) trapping vector efficiently disrupted the transcription of trapped genes. Thus, these two trapping vectors are alternative and effective tools for large-scale identification and disruption of endogenous genes in vertebrate cells and animals.

  13. Transposon insertional mutagenesis in mice identifies human breast cancer susceptibility genes and signatures for stratification.

    PubMed

    Chen, Liming; Jenjaroenpun, Piroon; Pillai, Andrea Mun Ching; Ivshina, Anna V; Ow, Ghim Siong; Efthimios, Motakis; Zhiqun, Tang; Tan, Tuan Zea; Lee, Song-Choon; Rogers, Keith; Ward, Jerrold M; Mori, Seiichi; Adams, David J; Jenkins, Nancy A; Copeland, Neal G; Ban, Kenneth Hon-Kim; Kuznetsov, Vladimir A; Thiery, Jean Paul

    2017-03-14

    Robust prognostic gene signatures and therapeutic targets are difficult to derive from expression profiling because of the significant heterogeneity within breast cancer (BC) subtypes. Here, we performed forward genetic screening in mice using Sleeping Beauty transposon mutagenesis to identify candidate BC driver genes in an unbiased manner, using a stabilized N-terminal truncated β-catenin gene as a sensitizer. We identified 134 mouse susceptibility genes from 129 common insertion sites within 34 mammary tumors. Of these, 126 genes were orthologous to protein-coding genes in the human genome (hereafter, human BC susceptibility genes, hBCSGs), 70% of which are previously reported cancer-associated genes, and ∼16% are known BC suppressor genes. Network analysis revealed a gene hub consisting of E1A binding protein P300 (EP300), CD44 molecule (CD44), neurofibromin (NF1) and phosphatase and tensin homolog (PTEN), which are linked to a significant number of mutated hBCSGs. From our survival prediction analysis of the expression of human BC genes in 2,333 BC cases, we isolated a six-gene-pair classifier that stratifies BC patients with high confidence into prognostically distinct low-, moderate-, and high-risk subgroups. Furthermore, we proposed prognostic classifiers identifying three basal and three claudin-low tumor subgroups. Intriguingly, our hBCSGs are mostly unrelated to cell cycle/mitosis genes and are distinct from the prognostic signatures currently used for stratifying BC patients. Our findings illustrate the strength and validity of integrating functional mutagenesis screens in mice with human cancer transcriptomic data to identify highly prognostic BC subtyping biomarkers.

  14. Transposon insertional mutagenesis in mice identifies human breast cancer susceptibility genes and signatures for stratification

    PubMed Central

    Chen, Liming; Jenjaroenpun, Piroon; Pillai, Andrea Mun Ching; Ivshina, Anna V.; Ow, Ghim Siong; Efthimios, Motakis; Zhiqun, Tang; Lee, Song-Choon; Rogers, Keith; Ward, Jerrold M.; Mori, Seiichi; Adams, David J.; Jenkins, Nancy A.; Copeland, Neal G.; Ban, Kenneth Hon-Kim; Kuznetsov, Vladimir A.; Thiery, Jean Paul

    2017-01-01

    Robust prognostic gene signatures and therapeutic targets are difficult to derive from expression profiling because of the significant heterogeneity within breast cancer (BC) subtypes. Here, we performed forward genetic screening in mice using Sleeping Beauty transposon mutagenesis to identify candidate BC driver genes in an unbiased manner, using a stabilized N-terminal truncated β-catenin gene as a sensitizer. We identified 134 mouse susceptibility genes from 129 common insertion sites within 34 mammary tumors. Of these, 126 genes were orthologous to protein-coding genes in the human genome (hereafter, human BC susceptibility genes, hBCSGs), 70% of which are previously reported cancer-associated genes, and ∼16% are known BC suppressor genes. Network analysis revealed a gene hub consisting of E1A binding protein P300 (EP300), CD44 molecule (CD44), neurofibromin (NF1) and phosphatase and tensin homolog (PTEN), which are linked to a significant number of mutated hBCSGs. From our survival prediction analysis of the expression of human BC genes in 2,333 BC cases, we isolated a six-gene-pair classifier that stratifies BC patients with high confidence into prognostically distinct low-, moderate-, and high-risk subgroups. Furthermore, we proposed prognostic classifiers identifying three basal and three claudin-low tumor subgroups. Intriguingly, our hBCSGs are mostly unrelated to cell cycle/mitosis genes and are distinct from the prognostic signatures currently used for stratifying BC patients. Our findings illustrate the strength and validity of integrating functional mutagenesis screens in mice with human cancer transcriptomic data to identify highly prognostic BC subtyping biomarkers. PMID:28251929

  15. Transposon mutagenesis reveals cooperation of ETS family transcription factors with signaling pathways in erythro-megakaryocytic leukemia

    PubMed Central

    Tang, Jian Zhong; Carmichael, Catherine L.; Shi, Wei; Metcalf, Donald; Ng, Ashley P.; Hyland, Craig D.; Jenkins, Nancy A.; Copeland, Neal G.; Howell, Viive M.; Zhao, Zhizhuang Joe; Smyth, Gordon K.; Kile, Benjamin T.; Alexander, Warren S.

    2013-01-01

    To define genetic lesions driving leukemia, we targeted cre-dependent Sleeping Beauty (SB) transposon mutagenesis to the blood-forming system using a hematopoietic-selective vav 1 oncogene (vav1) promoter. Leukemias of diverse lineages ensued, most commonly lymphoid leukemia and erythroleukemia. The inclusion of a transgenic allele of Janus kinase 2 (JAK2)V617F resulted in acceleration of transposon-driven disease and strong selection for erythroleukemic pathology with transformation of bipotential erythro-megakaryocytic cells. The genes encoding the E-twenty-six (ETS) transcription factors Ets related gene (Erg) and Ets1 were the most common sites for transposon insertion in SB-induced JAK2V617F-positive erythroleukemias, present in 87.5% and 65%, respectively, of independent leukemias examined. The role of activated Erg was validated by reproducing erythroleukemic pathology in mice transplanted with fetal liver cells expressing translocated in liposarcoma (TLS)-ERG, an activated form of ERG found in human leukemia. Via application of SB mutagenesis to TLS-ERG–induced erythroid transformation, we identified multiple loci as likely collaborators with activation of Erg. Jak2 was identified as a common transposon insertion site in TLS-ERG–induced disease, strongly validating the cooperation between JAK2V617F and transposon insertion at the Erg locus in the JAK2V617F-positive leukemias. Moreover, loci expressing other regulators of signal transduction pathways were conspicuous among the common transposon insertion sites in TLS-ERG–driven leukemia, suggesting that a key mechanism in erythroleukemia may be the collaboration of lesions disturbing erythroid maturation, most notably in genes of the ETS family, with mutations that reduce dependence on exogenous signals. PMID:23533276

  16. Transposon mutagenesis reveals cooperation of ETS family transcription factors with signaling pathways in erythro-megakaryocytic leukemia.

    PubMed

    Tang, Jian Zhong; Carmichael, Catherine L; Shi, Wei; Metcalf, Donald; Ng, Ashley P; Hyland, Craig D; Jenkins, Nancy A; Copeland, Neal G; Howell, Viive M; Zhao, Zhizhuang Joe; Smyth, Gordon K; Kile, Benjamin T; Alexander, Warren S

    2013-04-09

    To define genetic lesions driving leukemia, we targeted cre-dependent Sleeping Beauty (SB) transposon mutagenesis to the blood-forming system using a hematopoietic-selective vav 1 oncogene (vav1) promoter. Leukemias of diverse lineages ensued, most commonly lymphoid leukemia and erythroleukemia. The inclusion of a transgenic allele of Janus kinase 2 (JAK2)V617F resulted in acceleration of transposon-driven disease and strong selection for erythroleukemic pathology with transformation of bipotential erythro-megakaryocytic cells. The genes encoding the E-twenty-six (ETS) transcription factors Ets related gene (Erg) and Ets1 were the most common sites for transposon insertion in SB-induced JAK2V617F-positive erythroleukemias, present in 87.5% and 65%, respectively, of independent leukemias examined. The role of activated Erg was validated by reproducing erythroleukemic pathology in mice transplanted with fetal liver cells expressing translocated in liposarcoma (TLS)-ERG, an activated form of ERG found in human leukemia. Via application of SB mutagenesis to TLS-ERG-induced erythroid transformation, we identified multiple loci as likely collaborators with activation of Erg. Jak2 was identified as a common transposon insertion site in TLS-ERG-induced disease, strongly validating the cooperation between JAK2V617F and transposon insertion at the Erg locus in the JAK2V617F-positive leukemias. Moreover, loci expressing other regulators of signal transduction pathways were conspicuous among the common transposon insertion sites in TLS-ERG-driven leukemia, suggesting that a key mechanism in erythroleukemia may be the collaboration of lesions disturbing erythroid maturation, most notably in genes of the ETS family, with mutations that reduce dependence on exogenous signals.

  17. Functional characterization of the Sindbis virus E2 glycoprotein by transposon linker-insertion mutagenesis

    SciTech Connect

    Navaratnarajah, Chanakha K.; Kuhn, Richard J. . E-mail: kuhnr@purdue.edu

    2007-06-20

    The glycoprotein envelope of alphaviruses consists of two proteins, E1 and E2. E1 is responsible for fusion and E2 is responsible for receptor binding. An atomic structure is available for E1, but one for E2 has not been reported. In this study, transposon linker-insertion mutagenesis was used to probe the function of different domains of E2. A library of mutants, containing 19 amino acid insertions in the E2 glycoprotein sequence of the prototype alphavirus, Sindbis virus (SINV), was generated. Fifty-seven independent E2 insertions were characterized, of which more than half (67%) gave rise to viable virus. The wild-type-like mutants identify regions that accommodate insertions without perturbing virus production and can be used to insert targeting moieties to direct SINV to specific receptors. The defective and lethal mutants give insight into regions of E2 important for protein stability, transport to the cell membrane, E1-E2 contacts, and receptor binding.

  18. Characterization and Transposon Mutagenesis of the Maize (Zea mays) Pho1 Gene Family

    PubMed Central

    Salazar-Vidal, M. Nancy; Acosta-Segovia, Edith; Sánchez-León, Nidia; Ahern, Kevin R.; Brutnell, Thomas P.; Sawers, Ruairidh J. H.

    2016-01-01

    Phosphorus is an essential nutrient for all plants, but also one of the least mobile, and consequently least available, in the soil. Plants have evolved a series of molecular, metabolic and developmental adaptations to increase the acquisition of phosphorus and to maximize the efficiency of use within the plant. In Arabidopsis (Arabidopsis thaliana), the AtPHO1 protein regulates and facilitates the distribution of phosphorus. To investigate the role of PHO1 proteins in maize (Zea mays), the B73 reference genome was searched for homologous sequences, and four genes identified that were designated ZmPho1;1, ZmPho1;2a, ZmPho1;2b and ZmPho1;3. ZmPho1;2a and ZmPho1;2b are the most similar to AtPHO1, and represent candidate co-orthologs that we hypothesize to have been retained following whole genome duplication. Evidence was obtained for the production of natural anti-sense transcripts associated with both ZmPho1;2a and ZmPho1;2b, suggesting the possibility of regulatory crosstalk between paralogs. To characterize functional divergence between ZmPho1;2a and ZmPho1;2b, a program of transposon mutagenesis was initiated using the Ac/Ds system, and, here, we report the generation of novel alleles of ZmPho1;2a and ZmPho1;2b. PMID:27648940

  19. Characterization of the mgl operon of Escherichia coli by transposon mutagenesis and molecular cloning.

    PubMed

    Harayama, S; Bollinger, J; Iino, T; Hazelbauer, G L

    1983-01-01

    We used transposon insertion mutagenesis, molecular cloning, and a novel procedure for in vitro construction of polar and nonpolar insertion mutations to characterize the genetic organization and gene products of the beta-methylgalactoside (Mgl) transport system, which utilizes the galactose-binding protein. The data indicate that the mgl operon contained three genes, which were transcribed in the order mglB, mglA, and mglC. The first gene coded for the 31,000 Mr galactose-binding protein, which was synthesized as a 3,000-dalton-larger precursor form. The mglA product was a 50,000 Mr protein which was tightly associated with the membrane, and the mglC product was a 38,000 Mr protein which was apparently loosely associated with the membrane and was probably located on the internal face of the cytoplasmic membrane. Identification of gene products was facilitated by in vitro insertion of a fragment of Tn5 containing the gene conferring kanamycin resistance into a restriction site in the operon. The fragment proved to have a polar effect on the expression of promoter-distal genes only when inserted in one of the two possible orientations. The three identified gene products were necessary and apparently sufficient for transport activity, but only the binding protein was required for chemotaxis towards galactose. The transport system appeared to contain the minimum number of components for a binding protein-related system: a periplasmic recognition component, a transmembrane protein, and a peripheral membrane protein that may be involved in energy linkage.

  20. Characterization and Transposon Mutagenesis of the Maize (Zea mays) Pho1 Gene Family.

    PubMed

    Salazar-Vidal, M Nancy; Acosta-Segovia, Edith; Sánchez-León, Nidia; Ahern, Kevin R; Brutnell, Thomas P; Sawers, Ruairidh J H

    2016-01-01

    Phosphorus is an essential nutrient for all plants, but also one of the least mobile, and consequently least available, in the soil. Plants have evolved a series of molecular, metabolic and developmental adaptations to increase the acquisition of phosphorus and to maximize the efficiency of use within the plant. In Arabidopsis (Arabidopsis thaliana), the AtPHO1 protein regulates and facilitates the distribution of phosphorus. To investigate the role of PHO1 proteins in maize (Zea mays), the B73 reference genome was searched for homologous sequences, and four genes identified that were designated ZmPho1;1, ZmPho1;2a, ZmPho1;2b and ZmPho1;3. ZmPho1;2a and ZmPho1;2b are the most similar to AtPHO1, and represent candidate co-orthologs that we hypothesize to have been retained following whole genome duplication. Evidence was obtained for the production of natural anti-sense transcripts associated with both ZmPho1;2a and ZmPho1;2b, suggesting the possibility of regulatory crosstalk between paralogs. To characterize functional divergence between ZmPho1;2a and ZmPho1;2b, a program of transposon mutagenesis was initiated using the Ac/Ds system, and, here, we report the generation of novel alleles of ZmPho1;2a and ZmPho1;2b.

  1. A two-component enhancer-inhibitor transposon mutagenesis system for functional analysis of the Arabidopsis genome.

    PubMed Central

    Speulman, E; Metz, P L; van Arkel, G; te Lintel Hekkert, B; Stiekema, W J; Pereira, A

    1999-01-01

    A modified Enhancer-Inhibitor transposon system was used to generate a series of mutant lines by single-seed descent such that multiple I insertions occurred per plant. The distribution of original insertions in the population was assessed by isolating transposon-flanking DNA, and a database of insertion sites was created. Approximately three-quarters of the identified insertion sites show similarity to sequences stored in public databases, which demonstrates the power of this regimen of insertional mutagenesis. To isolate insertions in specific genes, we developed three-dimensional pooling and polymerase chain reaction strategies that we then validated by identifying mutants for the regulator genes APETALA1 and SHOOT MERISTEMLESS. The system then was used to identify inserts in a class of uncharacterized genes involved in lipid biosynthesis; one such insertion conferred a fiddlehead mutant phenotype. PMID:10521517

  2. Mutagenesis of dimeric plasmids by the transposon. gamma. delta. (Tn1000)

    SciTech Connect

    Liu, L.; Berg, C.M. )

    1990-05-01

    The Escherichia coli F factor mediates conjugal transfer of a plasmid such as pBR322 primarily by replicative transposition of transposon {gamma}{delta} (Tn1000) from F to that plasmid to form a cointegrate intermediate. Although resolution of this cointegrate always yields a plasmid containing a single {gamma}{delta} insertion, the occasional recovery of transposon-free plasmids after connuugal transfer has led to alternative hypotheses for F mobilization. The authors show here that {gamma}{delta}-free plasmids are found after F-mediated conjugal transfer only when the donor plasmid is a dimer and the recipient is Rec{sup +}.

  3. Use of a mariner-based transposon mutagenesis system to isolate Clostridium perfringens mutants deficient in gliding motility.

    PubMed

    Liu, Hualan; Bouillaut, Laurent; Sonenshein, Abraham L; Melville, Stephen B

    2013-02-01

    Clostridium perfringens is an anaerobic Gram-positive pathogen that causes many human and animal diseases, including food poisoning and gas gangrene. C. perfringens lacks flagella but possesses type IV pili (TFP). We have previously shown that C. perfringens can glide across an agar surface in long filaments composed of individual bacteria attached end to end and that two TFP-associated proteins, PilT and PilC, are needed for this. To discover additional gene products that play a role in gliding, we developed a plasmid-based mariner transposon mutagenesis system that works effectively in C. perfringens. More than 10,000 clones were screened for mutants that lacked the ability to move away from the edge of a colony. Twenty-four mutants (0.24%) were identified that fit the criteria. The genes containing insertions that affected gliding motility fell into nine different categories. One gene, CPE0278, which encodes a homolog of the SagA cell wall-dependent endopeptidase, acquired distinct transposon insertions in two independent mutants. sagA mutants were unable to form filaments due to a complete lack of end-to-end connections essential for gliding motility. Complementation of the sagA mutants with a wild-type copy of the gene restored gliding motility. We constructed an in-frame deletion mutation in the sagA gene and found that this mutant had a phenotype similar to those of the transposon mutants. We hypothesize that the sagA mutant strains are unable to form the molecular complexes which are needed to keep the cells in an end-to-end orientation, leading to separation of daughter cells and the inability to carry out gliding motility.

  4. A transposon toolkit for gene transfer and mutagenesis in protozoan parasites

    PubMed Central

    Damasceno, Jeziel D.; Beverley, Stephen M.; Tosi, Luiz R. O.

    2009-01-01

    Protozoan parasites affect millions of people around the world. Treatment and control of these diseases are complicated partly due to the intricate biology of these organisms. The interactions of species of Plasmodium, Leishmania and trypanosomes with their hosts are mediated by an unusual control of gene expression that is not fully understood. The availability of the genome sequence of these protozoa sets the stage for using more comprehensive, genome-wide strategies to study gene function. Transposons are effective tools for the systematic introduction of genetic alterations and different transposition systems have been adapted to study gene function in these human pathogens. A mariner transposon toolkit for use in vivo or in vitro in Leishmania parasites has been developed and can be used in a variety of applications. These modified mariner elements not only permit the inactivation of genes, but also mediate the rescue of translational gene fusions, bringing a major contribution to the investigation of Leishmania gene function. The piggyBac and Tn5 transposons have also been shown to mobilize across Plasmodium spp. genomes circumventing the current limitations in the genetic manipulation of these organisms. PMID:19763844

  5. A large-scale, gene-driven mutagenesis approach for the functional analysis of the mouse genome

    PubMed Central

    Hansen, Jens; Floss, Thomas; Van Sloun, Petra; Füchtbauer, Ernst-Martin; Vauti, Franz; Arnold, Hans-Hennig; Schnütgen, Frank; Wurst, Wolfgang; von Melchner, Harald; Ruiz, Patricia

    2003-01-01

    A major challenge of the postgenomic era is the functional characterization of every single gene within the mammalian genome. In an effort to address this challenge, we assembled a collection of mutations in mouse embryonic stem (ES) cells, which is the largest publicly accessible collection of such mutations to date. Using four different gene-trap vectors, we generated 5,142 sequences adjacent to the gene-trap integration sites (gene-trap sequence tags; http://genetrap.de) from >11,000 ES cell clones. Although most of the gene-trap vector insertions occurred randomly throughout the genome, we found both vector-independent and vector-specific integration “hot spots.” Because >50% of the hot spots were vector-specific, we conclude that the most effective way to saturate the mouse genome with gene-trap insertions is by using a combination of gene-trap vectors. When a random sample of gene-trap integrations was passaged to the germ line, 59% (17 of 29) produced an observable phenotype in transgenic mice, a frequency similar to that achieved by conventional gene targeting. Thus, gene trapping allows a large-scale and cost-effective production of ES cell clones with mutations distributed throughout the genome, a resource likely to accelerate genome annotation and the in vivo modeling of human disease. PMID:12904583

  6. Multipurpose Transposon-Insertion Libraries in Yeast.

    PubMed

    Kumar, Anuj

    2016-06-01

    Libraries of transposon-insertion alleles constitute powerful and versatile tools for large-scale analysis of yeast gene function. Transposon-insertion libraries are constructed most simply through mutagenesis of a plasmid-based genomic DNA library; modification of the mutagenizing transposon by incorporation of yeast selectable markers, recombination sites, and an epitope tag enables the application of insertion alleles for phenotypic screening and protein localization. In particular, yeast genomic DNA libraries have been mutagenized with modified bacterial transposons carrying the URA3 marker, lox recombination sites, and sequence encoding multiple copies of the hemagglutinin (HA) epitope. Mutagenesis with these transposons has yielded a large resource of insertion alleles affecting nearly 4000 yeast genes in total. Through well-established protocols, these insertion libraries can be introduced into the desired strain backgrounds and the resulting insertional mutants can be screened or systematically analyzed. Relative to alternative methods of UV irradiation or chemical mutagenesis, transposon-insertion alleles can be easily identified by PCR-based approaches or high-throughput sequencing. Transposon-insertion libraries also provide a cost-effective alternative to targeted deletion approaches, although, in contrast to start-codon to stop-codon deletions, insertion alleles might not represent true null-mutants. For protein-localization studies, transposon-insertion alleles can provide encoded epitope tags in-frame with internal codons; in many cases, these transposon-encoded epitope tags can provide a more accurate localization for proteins in which terminal sequences are crucial for intracellular targeting. Thus, overall, transposon-insertion libraries can be used quickly and economically and have a particular utility in screening for desired phenotypes and localization patterns in nonstandard genetic backgrounds. © 2016 Cold Spring Harbor Laboratory Press.

  7. Transposon activation mutagenesis as a screening tool for identifying resistance to cancer therapeutics

    PubMed Central

    2013-01-01

    Background The development of resistance to chemotherapies represents a significant barrier to successful cancer treatment. Resistance mechanisms are complex, can involve diverse and often unexpected cellular processes, and can vary with both the underlying genetic lesion and the origin or type of tumor. For these reasons developing experimental strategies that could be used to understand, identify and predict mechanisms of resistance in different malignant cells would be a major advance. Methods Here we describe a gain-of-function forward genetic approach for identifying mechanisms of resistance. This approach uses a modified piggyBac transposon to generate libraries of mutagenized cells, each containing transposon insertions that randomly activate nearby gene expression. Genes of interest are identified using next-gen high-throughput sequencing and barcode multiplexing is used to reduce experimental cost. Results Using this approach we successfully identify genes involved in paclitaxel resistance in a variety of cancer cell lines, including the multidrug transporter ABCB1, a previously identified major paclitaxel resistance gene. Analysis of co-occurring transposons integration sites in single cell clone allows for the identification of genes that might act cooperatively to produce drug resistance a level of information not accessible using RNAi or ORF expression screening approaches. Conclusion We have developed a powerful pipeline to systematically discover drug resistance in mammalian cells in vitro. This cost-effective approach can be readily applied to different cell lines, to identify canonical or context specific resistance mechanisms. Its ability to probe complex genetic context and non-coding genomic elements as well as cooperative resistance events makes it a good complement to RNAi or ORF expression based screens. PMID:23442791

  8. A large-scale mutant panel in wheat developed using heavy-ion beam mutagenesis and its application to genetic research

    NASA Astrophysics Data System (ADS)

    Murai, Koji; Nishiura, Aiko; Kazama, Yusuke; Abe, Tomoko

    2013-11-01

    Mutation analysis is a powerful tool for studying gene function. Heavy-ion beam mutagenesis is a comparatively new approach to inducing mutations in plants and is particularly efficient because of its high linear energy transfer (LET). High LET radiation induces a higher rate of DNA double-strand breaks than other mutagenic methods. Over the last 12 years, we have constructed a large-scale mutant panel in diploid einkorn wheat (Triticum monococcum) using heavy-ion beam mutagenesis. Einkorn wheat seeds were exposed to a heavy-ion beam and then sown in the field. Selfed seeds from each spike of M1 plants were used to generate M2 lines. Every year, we obtained approximately 1000 M2 lines and eventually developed a mutant panel with 10,000 M2 lines in total. This mutant panel is being systematically screened for mutations affecting reproductive growth, and especially for flowering-time mutants. To date, we have identified several flowering-time mutants of great interest: non-flowering mutants (mvp: maintained vegetative phase), late-flowering mutants, and early-flowering mutants. These novel mutations will be of value for investigations of the genetic mechanism of flowering in wheat.

  9. Scanning the Escherichia coli chromosome by random transposon mutagenesis and multiple phenotypic screening.

    PubMed

    Serina, Stefania; Nozza, Francesca; Nicastro, Giovanna; Faggioni, Federico; Mottl, Harald; Dehò, Gianni; Polissi, Alessandra

    2004-10-01

    Analysis of the complete DNA sequences of many microbial genomes available reveals a fair number of putative ORFs without an identified function. A systematic scan of the Escherichia coli chromosome was achieved by random transposition with a newly developed Tn5 minitransposon derivative carrying the arabinose-inducible araP(BAD) promoter oriented outward at one end (Tn5-araP(BAD)). The transposon insertion mutants obtained were assayed for conditional lethal phenotypes (arabinose dependence or sensitivity), for growth at two temperatures (37 and 15 degrees C) and in different media (rich and minimal medium). The Tn5-araP(BAD)-tagged genes were identified by sequencing the transposon insertion points. In this way we found a new essential gene cluster (yhbN-yhbG), produced conditional lethal (arabinose-dependent) mutations in already known essential genes (folD, frr, plsC, thiL, serS, thrS, and trpS) and provided a new phenotype (cold sensitivity) to other known genes (holD, ahpC, and tolA). Moreover, we identified eight putative ORFs (kch, ycaM, ycbQ, yddA, yddB, ydeK, ydeX, and yliF) that appear to be required in optimum growth conditions (rich medium at 37 degrees C) but not in the cold and in minimal medium.

  10. Identification of genes involved in biofilm formation and respiration via mini-Himar transposon mutagenesis of Geobacter sulfurreducens.

    PubMed

    Rollefson, Janet B; Levar, Caleb E; Bond, Daniel R

    2009-07-01

    Electron transfer from cells to metals and electrodes by the Fe(III)-reducing anaerobe Geobacter sulfurreducens requires proper expression of redox proteins and attachment mechanisms to interface bacteria with surfaces and neighboring cells. We hypothesized that transposon mutagenesis would complement targeted knockout studies in Geobacter spp. and identify novel genes involved in this process. Escherichia coli mating strains and plasmids were used to develop a conjugation protocol and deliver mini-Himar transposons, creating a library of over 8,000 mutants that was anaerobically arrayed and screened for a range of phenotypes, including auxotrophy for amino acids, inability to reduce Fe(III) citrate, and attachment to surfaces. Following protocol validation, mutants with strong phenotypes were further characterized in a three-electrode system to simultaneously quantify attachment, biofilm development, and respiratory parameters, revealing mutants defective in Fe(III) reduction but unaffected in electron transfer to electrodes (such as an insertion in GSU1330, a putative metal export protein) or defective in electrode reduction but demonstrating wild-type biofilm formation (due to an insertion upstream of the NHL domain protein GSU2505). An insertion in a putative ATP-dependent transporter (GSU1501) eliminated electrode colonization but not Fe(III) citrate reduction. A more complex phenotype was demonstrated by a mutant containing an insertion in a transglutaminase domain protein (GSU3361), which suddenly ceased to respire when biofilms reached approximately 50% of the wild-type levels. As most insertions were not in cytochromes but rather in transporters, two-component signaling proteins, and proteins of unknown function, this collection illustrates how biofilm formation and electron transfer are separate but complementary phenotypes, controlled by multiple loci not commonly studied in Geobacter spp.

  11. A single-copy Sleeping Beauty transposon mutagenesis screen identifies new PTEN-cooperating tumor suppressor genes

    PubMed Central

    de la Rosa, Jorge; Weber, Julia; Friedrich, Mathias Josef; Li, Yilong; Rad, Lena; Ponstingl, Hannes; Liang, Qi; de Quirós, Sandra Bernaldo; Noorani, Imran; Metzakopian, Emmanouil; Strong, Alexander; Li, Meng Amy; Astudillo, Aurora; Fernández-García, María Teresa; Fernández-García, María Soledad; Hoffman, Gary J.; Fuente, Rocío; Vassiliou, George S.; Rad, Roland; López-Otín, Carlos; Bradley, Allan; Cadiñanos, Juan

    2017-01-01

    The overwhelming amount of genetic alterations identified through cancer genome sequencing requires complementary approaches to interpret their significance and interactions. We developed a novel whole-body insertional mutagenesis screen in mice, designed for the discovery of Pten-cooperating tumor suppressors, in which mobilization of a single-copy inactivating Sleeping Beauty transposon is coupled to Pten disruption within the same genome. The analysis of 278 transposition-induced prostate, breast and skin tumors detected tissue-specific and shared datasets of known and candidate cancer genes. We validated ZBTB20, CELF2, PARD3, AKAP13 and WAC, identified by our screens in multiple cancer types, as new tumor suppressors in prostate cancer: we demonstrated their synergy with PTEN for preventing invasion in vitro and confirmed their clinical relevance. Further characterization of Wac in vivo revealed obligate haploinsufficiency for this autophagy-regulating gene in a Pten-deficient context. Our study identifies complex PTEN-cooperating tumor suppressor networks in different cancer types with potential clinical implications. PMID:28319090

  12. Transposon mutagenesis with coat color genotyping identifies an essential role for Skor2 in sonic hedgehog signaling and cerebellum development

    PubMed Central

    Wang, Baiping; Harrison, Wilbur; Overbeek, Paul A.; Zheng, Hui

    2011-01-01

    Correct development of the cerebellum requires coordinated sonic hedgehog (Shh) signaling from Purkinje to granule cells. How Shh expression is regulated in Purkinje cells is poorly understood. Using a novel tyrosinase minigene-tagged Sleeping Beauty transposon-mediated mutagenesis, which allows for coat color-based genotyping, we created mice in which the Ski/Sno family transcriptional co-repressor 2 (Skor2) gene is deleted. Loss of Skor2 leads to defective Purkinje cell development, a severe reduction of granule cell proliferation and a malformed cerebellum. Skor2 is specifically expressed in Purkinje cells in the brain, where it is required for proper expression of Shh. Skor2 overexpression suppresses BMP signaling in an HDAC-dependent manner and stimulates Shh promoter activity, suggesting that Skor2 represses BMP signaling to activate Shh expression. Our study identifies an essential function for Skor2 as a novel transcriptional regulator in Purkinje cells that acts upstream of Shh during cerebellum development. PMID:21937600

  13. A single-copy Sleeping Beauty transposon mutagenesis screen identifies new PTEN-cooperating tumor suppressor genes.

    PubMed

    de la Rosa, Jorge; Weber, Julia; Friedrich, Mathias Josef; Li, Yilong; Rad, Lena; Ponstingl, Hannes; Liang, Qi; de Quirós, Sandra Bernaldo; Noorani, Imran; Metzakopian, Emmanouil; Strong, Alexander; Li, Meng Amy; Astudillo, Aurora; Fernández-García, María Teresa; Fernández-García, María Soledad; Hoffman, Gary J; Fuente, Rocío; Vassiliou, George S; Rad, Roland; López-Otín, Carlos; Bradley, Allan; Cadiñanos, Juan

    2017-03-20

    The overwhelming number of genetic alterations identified through cancer genome sequencing requires complementary approaches to interpret their significance and interactions. Here we developed a novel whole-body insertional mutagenesis screen in mice, which was designed for the discovery of Pten-cooperating tumor suppressors. Toward this aim, we coupled mobilization of a single-copy inactivating Sleeping Beauty transposon to Pten disruption within the same genome. The analysis of 278 transposition-induced prostate, breast and skin tumors detected tissue-specific and shared data sets of known and candidate genes involved in cancer. We validated ZBTB20, CELF2, PARD3, AKAP13 and WAC, which were identified by our screens in multiple cancer types, as new tumor suppressor genes in prostate cancer. We demonstrated their synergy with PTEN in preventing invasion in vitro and confirmed their clinical relevance. Further characterization of Wac in vivo showed obligate haploinsufficiency for this gene (which encodes an autophagy-regulating factor) in a Pten-deficient context. Our study identified complex PTEN-cooperating tumor suppressor networks in different cancer types, with potential clinical implications.

  14. Mouse models of cancer: Sleeping Beauty transposons for insertional mutagenesis screens and reverse genetic studies.

    PubMed

    Tschida, Barbara R; Largaespada, David A; Keng, Vincent W

    2014-03-01

    The genetic complexity and heterogeneity of cancer has posed a problem in designing rationally targeted therapies effective in a large proportion of human cancer. Genomic characterization of many cancer types has provided a staggering amount of data that needs to be interpreted to further our understanding of this disease. Forward genetic screening in mice using Sleeping Beauty (SB) based insertional mutagenesis is an effective method for candidate cancer gene discovery that can aid in distinguishing driver from passenger mutations in human cancer. This system has been adapted for unbiased screens to identify drivers of multiple cancer types. These screens have already identified hundreds of candidate cancer-promoting mutations. These can be used to develop new mouse models for further study, which may prove useful for therapeutic testing. SB technology may also hold the key for rapid generation of reverse genetic mouse models of cancer, and has already been used to model glioblastoma and liver cancer. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Isolation of Rhodospirillum centenum Mutants Defective in Phototactic Colony Motility by Transposon Mutagenesis

    PubMed Central

    Jiang, Ze-Yu; Rushing, Brenda G.; Bai, Yong; Gest, Howard; Bauer, Carl E.

    1998-01-01

    The purple photosynthetic bacterium Rhodospirillum centenum is capable of forming swarm colonies that rapidly migrate toward or away from light, depending on the wavelength of excitation. To identify components specific for photoperception, we conducted mini-Tn5-mediated mutagenesis and screened approximately 23,000 transposition events for mutants that failed to respond to either continuous illumination or to a step down in light intensity. A majority of the ca. 250 mutants identified lost the ability to form motile swarm cells on an agar surface. These cells appeared to contain defects in the synthesis or assembly of surface-induced lateral flagella. Another large fraction of mutants that were unresponsive to light were shown to be defective in the formation of a functional photosynthetic apparatus. Several photosensory mutants also were obtained with defects in the perception and transmission of light signals. Twelve mutants in this class were shown to contain disruptions in a chemotaxis operon, and five mutants contained disruptions of components unique to photoperception. It was shown that screening for photosensory defective R. centenum swarm colonies is an effective method for genetic dissection of the mechanism of light sensing in eubacteria. PMID:9495765

  16. Identification and Characterization of Non-Cellulose-Producing Mutants of Gluconacetobacter hansenii Generated by Tn5 Transposon Mutagenesis

    PubMed Central

    Deng, Ying; Nagachar, Nivedita; Xiao, Chaowen; Tien, Ming

    2013-01-01

    The acs operon of Gluconacetobacter is thought to encode AcsA, AcsB, AcsC, and AcsD proteins that constitute the cellulose synthase complex, required for the synthesis and secretion of crystalline cellulose microfibrils. A few other genes have been shown to be involved in this process, but their precise role is unclear. We report here the use of Tn5 transposon insertion mutagenesis to identify and characterize six non-cellulose-producing (Cel−) mutants of Gluconacetobacter hansenii ATCC 23769. The genes disrupted were acsA, acsC, ccpAx (encoding cellulose-complementing protein [the subscript “Ax” indicates genes from organisms formerly classified as Acetobacter xylinum]), dgc1 (encoding guanylate dicyclase), and crp-fnr (encoding a cyclic AMP receptor protein/fumarate nitrate reductase transcriptional regulator). Protein blot analysis revealed that (i) AcsB and AcsC were absent in the acsA mutant, (ii) the levels of AcsB and AcsC were significantly reduced in the ccpAx mutant, and (iii) the level of AcsD was not affected in any of the Cel− mutants. Promoter analysis showed that the acs operon does not include acsD, unlike the organization of the acs operon of several strains of closely related Gluconacetobacter xylinus. Complementation experiments confirmed that the gene disrupted in each Cel− mutant was responsible for the phenotype. Quantitative real-time PCR and protein blotting results suggest that the transcription of bglAx (encoding β-glucosidase and located immediately downstream from acsD) was strongly dependent on Crp/Fnr. A bglAx knockout mutant, generated via homologous recombination, produced only ∼16% of the wild-type cellulose level. Since the crp-fnr mutant did not produce any cellulose, Crp/Fnr may regulate the expression of other gene(s) involved in cellulose biosynthesis. PMID:24013627

  17. Development of a high-frequency in vivo transposon mutagenesis system for Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942.

    PubMed

    Watabe, Kazuyuki; Mimuro, Mamoru; Tsuchiya, Tohru

    2014-11-01

    Synechocystis sp. PCC 6803 (Synechocystis) is the first sequenced photosynthetic organism and has two advantages: natural transformation and light-activated heterotrophic growth. Such characteristics have mainly promoted reverse genetic analysis in this organism, however, to date approximately 50% of genes are still annotated as 'unknown protein' or 'hypothetical protein'. Therefore, forward genetic analysis is required for the identification of significant genes responsible for photosynthesis and other physiological phenomena among the genes of unknown function. The in vivo transposon mutagenesis system is one of the major methods for random mutagenesis. However, present in vivo transposon mutagenesis systems for cyanobacteria face problems such as relatively low frequency of transposition and repeated transposition in the host cells. In this study, we constructed vectors based on a mini-Tn5-derived vector that was designed to prevent repeated transposition. Our vectors carry a hyperactive transposase and optimized recognition sequence of transposase, which were reported to enhance frequency of transposition. Using the vector, we succeeded in highly frequent transposition (9×10(-3) per recipient cell) in Synechocystis. Transposon insertion sites of 10 randomly selected mutants indicated that the insertion sites spread throughout the genome with low sequence dependency. Furthermore, one of the 10 mutants exhibited the slow-growing phenotype, and the mutant was functionally complemented by using our expression vector. Our system also worked with another model cyanobacterium, Synechococcus elongatus PCC 7942, with high frequency. These results indicate that the developed system can be applied to the forward genetic analysis of a broad range of cyanobacteria. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  18. Transposon Mutagenesis Paired with Deep Sequencing of Caulobacter crescentus under Uranium Stress Reveals Genes Essential for Detoxification and Stress Tolerance

    PubMed Central

    Yung, Mimi C.; Park, Dan M.; Overton, K. Wesley; Blow, Matthew J.; Hoover, Cindi A.; Smit, John; Murray, Sean R.; Ricci, Dante P.; Christen, Beat; Bowman, Grant R.

    2015-01-01

    ABSTRACT The ubiquitous aquatic bacterium Caulobacter crescentus is highly resistant to uranium (U) and facilitates U biomineralization and thus holds promise as an agent of U bioremediation. To gain an understanding of how C. crescentus tolerates U, we employed transposon (Tn) mutagenesis paired with deep sequencing (Tn-seq) in a global screen for genomic elements required for U resistance. Of the 3,879 annotated genes in the C. crescentus genome, 37 were found to be specifically associated with fitness under U stress, 15 of which were subsequently tested through mutational analysis. Systematic deletion analysis revealed that mutants lacking outer membrane transporters (rsaFa and rsaFb), a stress-responsive transcription factor (cztR), or a ppGpp synthetase/hydrolase (spoT) exhibited a significantly lower survival rate under U stress. RsaFa and RsaFb, which are homologues of TolC in Escherichia coli, have previously been shown to mediate S-layer export. Transcriptional analysis revealed upregulation of rsaFa and rsaFb by 4- and 10-fold, respectively, in the presence of U. We additionally show that rsaFa mutants accumulated higher levels of U than the wild type, with no significant increase in oxidative stress levels. Our results suggest a function for RsaFa and RsaFb in U efflux and/or maintenance of membrane integrity during U stress. In addition, we present data implicating CztR and SpoT in resistance to U stress. Together, our findings reveal novel gene targets that are key to understanding the molecular mechanisms of U resistance in C. crescentus. IMPORTANCE Caulobacter crescentus is an aerobic bacterium that is highly resistant to uranium (U) and has great potential to be used in U bioremediation, but its mechanisms of U resistance are poorly understood. We conducted a Tn-seq screen to identify genes specifically required for U resistance in C. crescentus. The genes that we identified have previously remained elusive using other omics approaches and thus

  19. Detection of a Putative TetR-Like Gene Related to Mycobacterium bovis BCG Growth in Cholesterol Using a gfp-Transposon Mutagenesis System.

    PubMed

    Otal, Isabel; Pérez-Herrán, Esther; Garcia-Morales, Lazaro; Menéndez, María C; Gonzalez-Y-Merchand, Jorge A; Martín, Carlos; García, María J

    2017-01-01

    In vitro transposition is a powerful genetic tool for identifying mycobacterial virulence genes and studying virulence factors in relation to the host. Transposon shuttle mutagenesis is a method for constructing stable insertions in the genome of different microorganisms including mycobacteria. Using an IS1096 derivative, we have constructed the Tngfp, a transposon containing a promoterless green fluorescent protein (gfp) gene. This transposon was able to transpose randomly in Mycobacterium bovis BCG. Bacteria with a single copy of the gfp gene per chromosome from an M. bovis BCG::Tngfp library were analyzed and cells exhibiting high levels of fluorescence were detected by flow cytometry. Application of this approach allowed for the selection of a mutant, BCG_2177c::Tngfp (BCG-Tn), on the basis of high level of long-standing fluorescence at stationary phase. This BCG-Tn mutant showed some particular phenotypic features compared to the wild type strain, mainly during stationary phase, when cholesterol was used as a sole carbon source, thus supporting the relationships of the targeted gene with the regulation of cholesterol metabolism in this bacteria. This approach showed that Tngfp is a potentially useful tool for studying the involvement of the targeted loci in metabolic pathways of mycobacteria.

  20. Detection of a Putative TetR-Like Gene Related to Mycobacterium bovis BCG Growth in Cholesterol Using a gfp-Transposon Mutagenesis System

    PubMed Central

    Otal, Isabel; Pérez-Herrán, Esther; Garcia-Morales, Lazaro; Menéndez, María C.; Gonzalez-y-Merchand, Jorge A.; Martín, Carlos; García, María J.

    2017-01-01

    In vitro transposition is a powerful genetic tool for identifying mycobacterial virulence genes and studying virulence factors in relation to the host. Transposon shuttle mutagenesis is a method for constructing stable insertions in the genome of different microorganisms including mycobacteria. Using an IS1096 derivative, we have constructed the Tngfp, a transposon containing a promoterless green fluorescent protein (gfp) gene. This transposon was able to transpose randomly in Mycobacterium bovis BCG. Bacteria with a single copy of the gfp gene per chromosome from an M. bovis BCG::Tngfp library were analyzed and cells exhibiting high levels of fluorescence were detected by flow cytometry. Application of this approach allowed for the selection of a mutant, BCG_2177c::Tngfp (BCG-Tn), on the basis of high level of long-standing fluorescence at stationary phase. This BCG-Tn mutant showed some particular phenotypic features compared to the wild type strain, mainly during stationary phase, when cholesterol was used as a sole carbon source, thus supporting the relationships of the targeted gene with the regulation of cholesterol metabolism in this bacteria. This approach showed that Tngfp is a potentially useful tool for studying the involvement of the targeted loci in metabolic pathways of mycobacteria. PMID:28321208

  1. Random mutagenesis in Corynebacterium glutamicum ATCC 13032 using an IS6100-based transposon vector identified the last unknown gene in the histidine biosynthesis pathway

    PubMed Central

    Mormann, Sascha; Lömker, Alexander; Rückert, Christian; Gaigalat, Lars; Tauch, Andreas; Pühler, Alfred; Kalinowski, Jörn

    2006-01-01

    Background Corynebacterium glutamicum, a Gram-positive bacterium of the class Actinobacteria, is an industrially relevant producer of amino acids. Several methods for the targeted genetic manipulation of this organism and rational strain improvement have been developed. An efficient transposon mutagenesis system for the completely sequenced type strain ATCC 13032 would significantly advance functional genome analysis in this bacterium. Results A comprehensive transposon mutant library comprising 10,080 independent clones was constructed by electrotransformation of the restriction-deficient derivative of strain ATCC 13032, C. glutamicum RES167, with an IS6100-containing non-replicative plasmid. Transposon mutants had stable cointegrates between the transposon vector and the chromosome. Altogether 172 transposon integration sites have been determined by sequencing of the chromosomal inserts, revealing that each integration occurred at a different locus. Statistical target site analyses revealed an apparent absence of a target site preference. From the library, auxotrophic mutants were obtained with a frequency of 2.9%. By auxanography analyses nearly two thirds of the auxotrophs were further characterized, including mutants with single, double and alternative nutritional requirements. In most cases the nutritional requirement observed could be correlated to the annotation of the mutated gene involved in the biosynthesis of an amino acid, a nucleotide or a vitamin. One notable exception was a clone mutagenized by transposition into the gene cg0910, which exhibited an auxotrophy for histidine. The protein sequence deduced from cg0910 showed high sequence similarities to inositol-1(or 4)-monophosphatases (EC 3.1.3.25). Subsequent genetic deletion of cg0910 delivered the same histidine-auxotrophic phenotype. Genetic complementation of the mutants as well as supplementation by histidinol suggests that cg0910 encodes the hitherto unknown essential L

  2. Resistance mechanisms to TP53-MDM2 inhibition identified by in vivo piggyBac transposon mutagenesis screen in an Arf(-/-) mouse model.

    PubMed

    Chapeau, Emilie A; Gembarska, Agnieszka; Durand, Eric Y; Mandon, Emeline; Estadieu, Claire; Romanet, Vincent; Wiesmann, Marion; Tiedt, Ralph; Lehar, Joseph; de Weck, Antoine; Rad, Roland; Barys, Louise; Jeay, Sebastien; Ferretti, Stephane; Kauffmann, Audrey; Sutter, Esther; Grevot, Armelle; Moulin, Pierre; Murakami, Masato; Sellers, William R; Hofmann, Francesco; Jensen, Michael Rugaard

    2017-03-21

    Inhibitors of double minute 2 protein (MDM2)-tumor protein 53 (TP53) interaction are predicted to be effective in tumors in which the TP53 gene is wild type, by preventing TP53 protein degradation. One such setting is represented by the frequent CDKN2A deletion in human cancer that, through inactivation of p14ARF, activates MDM2 protein, which in turn degrades TP53 tumor suppressor. Here we used piggyBac (PB) transposon insertional mutagenesis to anticipate resistance mechanisms occurring during treatment with the MDM2-TP53 inhibitor HDM201. Constitutive PB mutagenesis in Arf(-/-) mice provided a collection of spontaneous tumors with characterized insertional genetic landscapes. Tumors were allografted in large cohorts of mice to assess the pharmacologic effects of HDM201. Sixteen out of 21 allograft models were sensitive to HDM201 but ultimately relapsed under treatment. A comparison of tumors with acquired resistance to HDM201 and untreated tumors identified 87 genes that were differentially and significantly targeted by the PB transposon. Resistant tumors displayed a complex clonality pattern suggesting the emergence of several resistant subclones. Among the most frequent alterations conferring resistance, we observed somatic and insertional loss-of-function mutations in transformation-related protein 53 (Trp53) in 54% of tumors and transposon-mediated gain-of-function alterations in B-cell lymphoma-extra large (Bcl-xL), Mdm4, and two TP53 family members, resulting in expression of the TP53 dominant negative truncations ΔNTrp63 and ΔNTrp73. Enhanced BCL-xL and MDM4 protein expression was confirmed in resistant tumors, as well as in HDM201-resistant patient-derived tumor xenografts. Interestingly, concomitant inhibition of MDM2 and BCL-xL demonstrated significant synergy in p53 wild-type cell lines in vitro. Collectively, our findings identify several potential mechanisms by which TP53 wild-type tumors may escape MDM2-targeted therapy.

  3. Using Yeast Transposon-Insertion Libraries for Phenotypic Screening and Protein Localization.

    PubMed

    Kumar, Anuj

    2016-06-01

    This protocol details how to use a transposon-insertion library for phenotypic screening and protein localization. The insertion library was generated by mutagenesis of a plasmid-based yeast genomic DNA library by using a multipurpose transposon; the transposon produces gene disruptions, and, by Cre-mediated recombination at lox sites incorporated within the transposon, alleles with an in-frame insertion can be truncated to a residual transposon encoding multiple copies of the hemagglutinin epitope. Insertions are generated in yeast by shuttle mutagenesis. Yeast genomic DNA containing a transposon insertion is released from the library, and the mutagenized DNA sequences are introduced into a desired strain of yeast, where the insertion alleles replace native loci by homologous recombination. The insertion mutants can be screened for phenotypes, and the site of transposon insertion can subsequently be identified in selected mutants by inverse polymerase chain reaction (PCR). In-frame insertions within genes of interest can be truncated to an epitope-tagged allele by Cre-lox recombination, and the subcellular localization of the encoded protein product can be identified by standard methods of indirect immunofluorescence. In summary, the transposon-insertion libraries represent an informative resource for large-scale mutagenesis, presenting a straightforward alternative to labor-intensive targeted approaches for the construction of deletion alleles and fluorescent protein fusions.

  4. Identification of novel genes responsible for ethanol and/or thermotolerance by transposon mutagenesis in Saccharomyces cerevisiae.

    PubMed

    Kim, Hyun-Soo; Kim, Na-Rae; Yang, Jungwoo; Choi, Wonja

    2011-08-01

    Saccharomyces cerevisiae strains tolerant to ethanol and heat stresses are important for industrial ethanol production. In this study, five strains (Tn 1-5) tolerant to up to 15% ethanol were isolated by screening a transposon-mediated mutant library. Two of them displayed tolerance to heat (42 °C). The determination of transposon insertion sites and Northern blot analysis identified seven putative genes (CMP2, IMD4, SSK2, PPG1, DLD3, PAM1, and MSN2) and revealed simultaneous down-regulations of CMP2 and IMD4, and SSK2 and PPG1, down-regulation of DLD3, and disruptions of the open reading frame of PAM1 and MSN2, indicating that ethanol and/or heat tolerance can be conferred. Knockout mutants of these seven individual genes were ethanol tolerant and three of them (SSK2, PPG1, and PAM1) were tolerant to heat. Such tolerant phenotypes reverted to sensitive phenotypes by the autologous or overexpression of each gene. Five transposon mutants showed higher ethanol production and grew faster than the control strain when cultured in rich media containing 30% glucose and initial 6% ethanol at 30 °C. Of those, two thermotolerant transposon mutants (Tn 2 and Tn 3) exhibited significantly enhanced growth and ethanol production compared to the control at 42 °C. The genes identified in this study may provide a basis for the application in developing industrial yeast strains.

  5. Identification of genes involved in Mycoplasma gallisepticum biofilm formation using mini-Tn4001-SGM transposon mutagenesis.

    PubMed

    Wang, Yang; Yi, Li; Zhang, Fanqing; Qiu, Xusheng; Tan, Lei; Yu, Shengqing; Cheng, Xiangchao; Ding, Chan

    2017-01-01

    Mycoplasma gallisepticum (MG) is an important pathogen that can cause chronic respiratory disease in chickens and infectious sinusitis in turkeys. MG has the ability to form biofilms. The molecular mechanisms underlying MG biofilm formation are complex and poorly understood. To better understand the mechanisms involved in biofilm formation, mini-Tn4001-SGM, a novel transposon vector containing the gentamicin gene was constructed and electroporated into MG strain Rlow. Of the 738 mutants obtained, 12 had significantly reduced capacity to form biofilms in a polystyrene microtiter-plate biofilm assay. Ten different genes were identified as disrupted in these mutants using genomic walking from the transposon insertion sites and Southern bolt hybridization with a transposon-based probe. Four genes were associated with cellular processes, especially synthesis of extracellular polysaccharide and several lipoproteins encoded. Other genes were associated with translation, metabolism and gene regulation, and one had unknown function. Seven genes identified in this study have been previously associated with biofilm formation in MG or other bacterial species. The other three have not been previously reported to play a role in biofilm formation in MG. In conclusion, a new transposon vector was shown to be a powerful tool for future studies of MG pathogenesis. This study adds to our understanding of the molecular mechanisms involved in MG biofilm formation and may shed light on the persistence of MG infections.

  6. Transposon mutagenesis identified chromosomal and plasmid genes essential for adaptation of the marine bacterium Dinoroseobacter shibae to anaerobic conditions.

    PubMed

    Ebert, Matthias; Laaß, Sebastian; Burghartz, Melanie; Petersen, Jörn; Koßmehl, Sebastian; Wöhlbrand, Lars; Rabus, Ralf; Wittmann, Christoph; Tielen, Petra; Jahn, Dieter

    2013-10-01

    Anaerobic growth and survival are integral parts of the life cycle of many marine bacteria. To identify genes essential for the anoxic life of Dinoroseobacter shibae, a transposon library was screened for strains impaired in anaerobic denitrifying growth. Transposon insertions in 35 chromosomal and 18 plasmid genes were detected. The essential contribution of plasmid genes to anaerobic growth was confirmed with plasmid-cured D. shibae strains. A combined transcriptome and proteome approach identified oxygen tension-regulated genes. Transposon insertion sites of a total of 1,527 mutants without an anaerobic growth phenotype were determined to identify anaerobically induced but not essential genes. A surprisingly small overlap of only three genes (napA, phaA, and the Na(+)/Pi antiporter gene Dshi_0543) between anaerobically essential and induced genes was found. Interestingly, transposon mutations in genes involved in dissimilatory and assimilatory nitrate reduction (napA, nasA) and corresponding cofactor biosynthesis (genomic moaB, moeB, and dsbC and plasmid-carried dsbD and ccmH) were found to cause anaerobic growth defects. In contrast, mutation of anaerobically induced genes encoding proteins required for the later denitrification steps (nirS, nirJ, nosD), dimethyl sulfoxide reduction (dmsA1), and fermentation (pdhB1, arcA, aceE, pta, acs) did not result in decreased anaerobic growth under the conditions tested. Additional essential components (ferredoxin, cccA) of the anaerobic electron transfer chain and central metabolism (pdhB) were identified. Another surprise was the importance of sodium gradient-dependent membrane processes and genomic rearrangements via viruses, transposons, and insertion sequence elements for anaerobic growth. These processes and the observed contributions of cell envelope restructuring (lysM, mipA, fadK), C4-dicarboxylate transport (dctM1, dctM3), and protease functions to anaerobic growth require further investigation to unravel the

  7. Isolation of cyanobacterial mutants exhibiting growth defects under microoxic conditions by transposon tagging mutagenesis of Synechocystis sp. PCC 6803.

    PubMed

    Terauchi, Kazuki; Sobue, Riho; Furutani, Yuho; Aoki, Rina; Fujita, Yuichi

    2017-05-12

    Cyanobacteria are photosynthetic prokaryotes that perform oxygenic photosynthesis by extracting electrons from water, with the generation of oxygen as a byproduct. Cyanobacteria use oxygen not only for respiration to produce energy in the dark but also for biosynthesis of various metabolites, such as heme and chlorophyll. Oxygen levels dynamically fluctuate in the field environments, from hyperoxic at daytime to almost anaerobic at night. Thus, adaptation to anaerobiosis should be important for cyanobacteria to survive in low-oxygen and anaerobic environments. However, little is known about the molecular mechanisms of cyanobacterial anaerobiosis because cyanobacteria have been regarded as aerobic organisms. As a first step to elucidate cyanobacterial adaptation mechanisms to low-oxygen environments, we isolated five mutants, T-1-T-5, exhibiting growth defects under microoxic conditions. The mutants were obtained from a transposon-tagged mutant library of the cyanobacterium Synechocystis sp. PCC 6803, which was produced by in vitro transposon tagging of cyanobacterial genomic DNA. Southern blot analysis indicated that a kanamycin resistance gene was inserted in the genome as a single copy. We identified the chromosomal transposon-tagged locus in T-5. Two open reading frames (sll0577 and sll0578) were partially deleted by the insertion of the kanamycin resistance gene in T-5. A reverse transcription polymerase chain reaction suggested that these co-transcribed genes are constitutively expressed under both aerobic and microoxic conditions. Then, we isolated two mutants in which one of the two genes was individually disrupted. Only the mutants partially lacking an intact sll0578 gene showed growth defects under microoxic conditions, whereas it grew normally under aerobic conditions. sll0578 is annotated as purK encoding N(5)-carboxy-aminoimidazole ribonucleotide synthetase involved in purine metabolism. This result implies the unexpected physiological importance of Pur

  8. Case-oriented pathways analysis in pancreatic adenocarcinoma using data from a sleeping beauty transposon mutagenesis screen.

    PubMed

    Ho, Yen-Yi; Starr, Timothy K; LaRue, Rebecca S; Largaespada, David A

    2016-04-01

    Mutation studies of pancreatic ductal adenocarcinoma (PDA) have revealed complicated heterogeneous genomic landscapes of the disease. These studies cataloged a number of genes mutated at high frequencies, but also report a very large number of genes mutated in lower percentages of tumors. Taking advantage of a well-established forward genetic screening technique, with the Sleeping Beauty (SB) transposon, several studies produced PDA and discovered a number of common insertion sites (CIS) and associated genes that are recurrently mutated at high frequencies. As with human mutation studies, a very large number of genes were found to be altered by transposon insertion at low frequencies. These low frequency CIS associated genes may be very valuable to consider for their roles in cancer, since collectively they might emerge from a core group of genetic pathways. In this paper, we determined whether the genetic mutations in SB-accelerated PDA occur within a collated group of biological processes defined as gene sets. The approach considered both genes mutated in high and lower frequencies. We implemented a case-oriented, gene set enrichment analysis (CO-GSEA) on SB altered genes in PDA. Compared to traditional GSEA, CO-GSEA enables us to consider individual characteristics of mutation profiles of each PDA tumor. We identified genetic pathways with higher numbers of genetic mutations than expected by chance. We also present the correlations between these significant enriched genetic pathways, and their associations with CIS genes. These data suggest that certain pathway alterations cooperate in PDA development.

  9. Genetic Transformation of Hordeum vulgare ssp. spontaneum for the Development of a Transposon-Based Insertional Mutagenesis System.

    PubMed

    Cardinal, Marie-Josée; Kaur, Rajvinder; Singh, Jaswinder

    2016-10-01

    Domestication and intensive selective breeding of plants has triggered erosion of genetic diversity of important stress-related alleles. Researchers highlight the potential of using wild accessions as a gene source for improvement of cereals such as barley, which has major economic and social importance worldwide. Previously, we have successfully introduced the maize Ac/Ds transposon system for gene identification in cultivated barley. The objective of current research was to investigate the response of Hordeum vulgare ssp. spontaneum wild barley accessions in tissue culture to standardize parameters for introduction of Ac/Ds transposons through genetic transformation. We investigated the response of ten wild barley genotypes for callus induction, regenerative green callus induction and regeneration of fertile plants. The activity of exogenous Ac/Ds elements was observed through a transient assay on immature wild barley embryos/callus whereby transformed embryos/calli were identified by the expression of GUS. Transient Ds expression bombardment experiments were performed on 352 pieces of callus (3-5 mm each) or immature embryos in 4 genotypes of wild barley. The transformation frequency of putative transgenic callus lines based on transient GUS expression ranged between 72 and100 % in wild barley genotypes. This is the first report of a transformation system in H. vulgare ssp. spontaneum.

  10. Transposon mutagenesis of Salmonella enterica serovar Enteritidis identifies genes that contribute to invasiveness in human and chicken cells and survival in egg albumen.

    PubMed

    Shah, Devendra H; Zhou, Xiaohui; Kim, Hye-Young; Call, Douglas R; Guard, Jean

    2012-12-01

    Salmonella enterica serovar Enteritidis is an important food-borne pathogen, and chickens are a primary reservoir of human infection. While most knowledge about Salmonella pathogenesis is based on research conducted on Salmonella enterica serovar Typhimurium, S. Enteritidis is known to have pathobiology specific to chickens that impacts epidemiology in humans. Therefore, more information is needed about S. Enteritidis pathobiology in comparison to that of S. Typhimurium. We used transposon mutagenesis to identify S. Enteritidis virulence genes by assay of invasiveness in human intestinal epithelial (Caco-2) cells and chicken liver (LMH) cells and survival within chicken (HD-11) macrophages as a surrogate marker for virulence. A total of 4,330 transposon insertion mutants of an invasive G1 Nal(r) strain were screened using Caco-2 cells. This led to the identification of attenuating mutations in a total of 33 different loci, many of which include genes previously known to contribute to enteric infection (e.g., Salmonella pathogenicity island 1 [SPI-1], SPI-4, SPI-5, CS54, fliH, fljB, csgB, spvR, and rfbMN) in S. Enteritidis and other Salmonella serovars. Several genes or genomic islands that have not been reported previously (e.g., SPI-14, ksgA, SEN0034, SEN2278, and SEN3503) or that are absent in S. Typhimurium or in most other Salmonella serovars (e.g., pegD, SEN1152, SEN1393, and SEN1966) were also identified. Most mutants with reduced Caco-2 cell invasiveness also showed significantly reduced invasiveness in chicken liver cells and impaired survival in chicken macrophages and in egg albumen. Consequently, these genes may play an important role during infection of the chicken host and also contribute to successful egg contamination by S. Enteritidis.

  11. Transposon Mutagenesis of Salmonella enterica Serovar Enteritidis Identifies Genes That Contribute to Invasiveness in Human and Chicken Cells and Survival in Egg Albumen

    PubMed Central

    Zhou, Xiaohui; Kim, Hye-Young; Call, Douglas R.; Guard, Jean

    2012-01-01

    Salmonella enterica serovar Enteritidis is an important food-borne pathogen, and chickens are a primary reservoir of human infection. While most knowledge about Salmonella pathogenesis is based on research conducted on Salmonella enterica serovar Typhimurium, S. Enteritidis is known to have pathobiology specific to chickens that impacts epidemiology in humans. Therefore, more information is needed about S. Enteritidis pathobiology in comparison to that of S. Typhimurium. We used transposon mutagenesis to identify S. Enteritidis virulence genes by assay of invasiveness in human intestinal epithelial (Caco-2) cells and chicken liver (LMH) cells and survival within chicken (HD-11) macrophages as a surrogate marker for virulence. A total of 4,330 transposon insertion mutants of an invasive G1 Nalr strain were screened using Caco-2 cells. This led to the identification of attenuating mutations in a total of 33 different loci, many of which include genes previously known to contribute to enteric infection (e.g., Salmonella pathogenicity island 1 [SPI-1], SPI-4, SPI-5, CS54, fliH, fljB, csgB, spvR, and rfbMN) in S. Enteritidis and other Salmonella serovars. Several genes or genomic islands that have not been reported previously (e.g., SPI-14, ksgA, SEN0034, SEN2278, and SEN3503) or that are absent in S. Typhimurium or in most other Salmonella serovars (e.g., pegD, SEN1152, SEN1393, and SEN1966) were also identified. Most mutants with reduced Caco-2 cell invasiveness also showed significantly reduced invasiveness in chicken liver cells and impaired survival in chicken macrophages and in egg albumen. Consequently, these genes may play an important role during infection of the chicken host and also contribute to successful egg contamination by S. Enteritidis. PMID:22988017

  12. Large-scale insertional mutagenesis of Chlamydomonas supports phylogenomic functional prediction of photosynthetic genes and analysis of classical acetate-requiring mutants.

    PubMed

    Dent, Rachel M; Sharifi, Marina N; Malnoë, Alizée; Haglund, Cat; Calderon, Robert H; Wakao, Setsuko; Niyogi, Krishna K

    2015-04-01

    Chlamydomonas reinhardtii is a unicellular green alga that is a key model organism in the study of photosynthesis and oxidative stress. Here we describe the large-scale generation of a population of insertional mutants that have been screened for phenotypes related to photosynthesis and the isolation of 459 flanking sequence tags from 439 mutants. Recent phylogenomic analysis has identified a core set of genes, named GreenCut2, that are conserved in green algae and plants. Many of these genes are likely to be central to the process of photosynthesis, and they are over-represented by sixfold among the screened insertional mutants, with insertion events isolated in or adjacent to 68 of 597 GreenCut2 genes. This enrichment thus provides experimental support for functional assignments based on previous bioinformatic analysis. To illustrate one of the uses of the population, a candidate gene approach based on genome position of the flanking sequence of the insertional mutant CAL027_01_20 was used to identify the molecular basis of the classical C. reinhardtii mutation ac17. These mutations were shown to affect the gene PDH2, which encodes a subunit of the plastid pyruvate dehydrogenase complex. The mutants and associated flanking sequence data described here are publicly available to the research community, and they represent one of the largest phenotyped collections of algal insertional mutants to date.

  13. Silent Mischief: Bacteriophage Mu Insertions Contaminate Products of Escherichia coli Random Mutagenesis Performed Using Suicidal Transposon Delivery Plasmids Mobilized by Broad-Host-Range RP4 Conjugative Machinery ▿

    PubMed Central

    Ferrières, Lionel; Hémery, Gaëlle; Nham, Toan; Guérout, Anne-Marie; Mazel, Didier; Beloin, Christophe; Ghigo, Jean-Marc

    2010-01-01

    Random transposon mutagenesis is the strategy of choice for associating a phenotype with its unknown genetic determinants. It is generally performed by mobilization of a conditionally replicating vector delivering transposons to recipient cells using broad-host-range RP4 conjugative machinery carried by the donor strain. In the present study, we demonstrate that bacteriophage Mu, which was deliberately introduced during the original construction of the widely used donor strains SM10 λpir and S17-1 λpir, is silently transferred to Escherichia coli recipient cells at high frequency, both by hfr and by release of Mu particles by the donor strain. Our findings suggest that bacteriophage Mu could have contaminated many random-mutagenesis experiments performed on Mu-sensitive species with these popular donor strains, leading to potential misinterpretation of the transposon mutant phenotype and therefore perturbing analysis of mutant screens. To circumvent this problem, we precisely mapped Mu insertions in SM10 λpir and S17-1 λpir and constructed a new Mu-free donor strain, MFDpir, harboring stable hfr-deficient RP4 conjugative functions and sustaining replication of Π-dependent suicide vectors. This strain can therefore be used with most of the available transposon-delivering plasmids and should enable more efficient and easy-to-analyze mutant hunts in E. coli and other Mu-sensitive RP4 host bacteria. PMID:20935093

  14. Photo-biological hydrogen production by an acid tolerant mutant of Rhodovulum sulfidophilum P5 generated by transposon mutagenesis.

    PubMed

    Cai, Jinling; Wang, Guangce

    2014-02-01

    Most of the photosynthetic bacterial strains exhibit optimum hydrogen production at neutral initial pH, and lower initial pH resulted in a sharp decrease in hydrogen yield. Thus, screening of acid-tolerant hydrogen-producing photosynthetic bacteria is very important. To obtain acid tolerant mutants, a Tn7-based transposon was randomly inserted into the genomic DNA of Rhodovulum sulfidophilum P5. An acid tolerant mutant strain TH-102 exhibited increased hydrogen production in acidic environment (pH 4.5-6.5) and at higher temperatures (35 and 37°C) than the wild-type strain. At pH 5.5 and 35°C, the mutant strain TH-102 continuously produced hydrogen. The hydrogen yield and average rate were 2.16 ± 0.10 mol/mol acetate and 10.06 ± 0.47 mL/Lh, which was about 17.32 and 15.37-fold higher than that of the wild-type strain, respectively. This acid- and temperature-tolerant mutant strain TH-102 could be used in a cost-effective hydrogen production process employing both dark fermentative and photosynthetic bacteria. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Modification of nitrogen remobilization, grain fill and leaf senescence in maize (Zea mays) by transposon insertional mutagenesis in a protease gene.

    PubMed

    Donnison, Iain S; Gay, Alan P; Thomas, Howard; Edwards, Keith J; Edwards, David; James, Caron L; Thomas, Ann M; Ougham, Helen J

    2007-01-01

    A maize (Zea mays) senescence-associated legumain gene, See2beta, was characterized at the physiological and molecular levels to determine its role in senescence and resource allocation. A reverse-genetics screen of a maize Mutator (Mu) population identified a Mu insertion in See2beta. Maize plants homozygous for the insertion were produced. These See2 mutant and sibling wild-type plants were grown under high or low quantities of nitrogen (N). The early development of both genotypes was similar; however, tassel tip and collar emergence occurred earlier in the mutant. Senescence of the mutant leaves followed a similar pattern to that of wild-type leaves, but at later sampling points mutant plants contained more chlorophyll than wild-type plants and showed a small extension in photosynthetic activity. Total plant weight was higher in the wild-type than in the mutant, and there was a genotype x N interaction. Mutant plants under low N maintained cob weight, in contrast to wild-type plants under the same treatment. It is concluded, on the basis of transposon mutagenesis, that See2beta has an important role in N-use and resource allocation under N-limited conditions, and a minor but significant function in the later stages of senescence.

  16. Transposon Mutagenesis of the Plant-Associated Bacillus amyloliquefaciens ssp. plantarum FZB42 Revealed That the nfrA and RBAM17410 Genes Are Involved in Plant-Microbe-Interactions

    PubMed Central

    Dietel, Kristin; Beator, Barbara; Dolgova, Olga; Fan, Ben; Bleiss, Wilfrid; Ziegler, Jörg; Schmid, Michael; Hartmann, Anton; Borriss, Rainer

    2014-01-01

    Bacillus amyloliquefaciens ssp. plantarum FZB42 represents the prototype of Gram-positive plant growth promoting and biocontrol bacteria. In this study, we applied transposon mutagenesis to generate a transposon library, which was screened for genes involved in multicellular behavior and biofilm formation on roots as a prerequisite of plant growth promoting activity. Transposon insertion sites were determined by rescue-cloning followed by DNA sequencing. As in B. subtilis, the global transcriptional regulator DegU was identified as an activator of genes necessary for swarming and biofilm formation, and the DegU-mutant of FZB42 was found impaired in efficient root colonization. Direct screening of 3,000 transposon insertion mutants for plant-growth-promotion revealed the gene products of nfrA and RBAM_017140 to be essential for beneficial effects exerted by FZB42 on plants. We analyzed the performance of GFP-labeled wild-type and transposon mutants in the colonization of lettuce roots using confocal laser scanning microscopy. While the wild-type strain heavily colonized root surfaces, the nfrA mutant did not colonize lettuce roots, although it was not impaired in growth in laboratory cultures, biofilm formation and swarming motility on agar plates. The RBAM17410 gene, occurring in only a few members of the B. subtilis species complex, was directly involved in plant growth promotion. None of the mutant strains were affected in producing the plant growth hormone auxin. We hypothesize that the nfrA gene product is essential for overcoming the stress caused by plant response towards bacterial root colonization. PMID:24847778

  17. Precise excision and self-integration of a composite transposon as a model for spontaneous large-scale chromosome inversion/deletion of the Staphylococcus haemolyticus clinical strain JCSC1435.

    PubMed

    Watanabe, Shinya; Ito, Teruyo; Morimoto, Yuh; Takeuchi, Fumihiko; Hiramatsu, Keiichi

    2007-04-01

    Large-scale chromosomal inversions (455 to 535 kbp) or deletions (266 to 320 kbp) were found to accompany spontaneous loss of beta-lactam resistance during drug-free passage of the multiresistant Staphylococcus haemolyticus clinical strain JCSC1435. Identification and sequencing of the rearranged chromosomal loci revealed that ISSha1 of S. haemolyticus is responsible for the chromosome rearrangements.

  18. Mutator and MULE Transposons.

    PubMed

    Lisch, Damon

    2015-04-01

    The Mutator system of transposable elements (TEs) is a highly mutagenic family of transposons in maize. Because they transpose at high rates and target genic regions, these transposons can rapidly generate large numbers of new mutants, which has made the Mutator system a favored tool for both forward and reverse mutagenesis in maize. Low copy number versions of this system have also proved to be excellent models for understanding the regulation and behavior of Class II transposons in plants. Notably, the availability of a naturally occurring locus that can heritably silence autonomous Mutator elements has provided insights into the means by which otherwise active transposons are recognized and silenced. This chapter will provide a review of the biology, regulation, evolution and uses of this remarkable transposon system, with an emphasis on recent developments in our understanding of the ways in which this TE system is recognized and epigenetically silenced as well as recent evidence that Mu-like elements (MULEs) have had a significant impact on the evolution of plant genomes.

  19. Large scale dynamic systems

    NASA Technical Reports Server (NTRS)

    Doolin, B. F.

    1975-01-01

    Classes of large scale dynamic systems were discussed in the context of modern control theory. Specific examples discussed were in the technical fields of aeronautics, water resources and electric power.

  20. Genome-wide transposon mutagenesis of Proteus mirabilis: Essential genes, fitness factors for catheter-associated urinary tract infection, and the impact of polymicrobial infection on fitness requirements

    PubMed Central

    Smith, Sara N.; Zhao, Lili; Wu, Weisheng

    2017-01-01

    The Gram-negative bacterium Proteus mirabilis is a leading cause of catheter-associated urinary tract infections (CAUTIs), which are often polymicrobial. Numerous prior studies have uncovered virulence factors for P. mirabilis pathogenicity in a murine model of ascending UTI, but little is known concerning pathogenesis during CAUTI or polymicrobial infection. In this study, we utilized five pools of 10,000 transposon mutants each and transposon insertion-site sequencing (Tn-Seq) to identify the full arsenal of P. mirabilis HI4320 fitness factors for single-species versus polymicrobial CAUTI with Providencia stuartii BE2467. 436 genes in the input pools lacked transposon insertions and were therefore concluded to be essential for P. mirabilis growth in rich medium. 629 genes were identified as P. mirabilis fitness factors during single-species CAUTI. Tn-Seq from coinfection with P. stuartii revealed 217/629 (35%) of the same genes as identified by single-species Tn-Seq, and 1353 additional factors that specifically contribute to colonization during coinfection. Mutants were constructed in eight genes of interest to validate the initial screen: 7/8 (88%) mutants exhibited the expected phenotypes for single-species CAUTI, and 3/3 (100%) validated the expected phenotypes for polymicrobial CAUTI. This approach provided validation of numerous previously described P. mirabilis fitness determinants from an ascending model of UTI, the discovery of novel fitness determinants specifically for CAUTI, and a stringent assessment of how polymicrobial infection influences fitness requirements. For instance, we describe a requirement for branched-chain amino acid biosynthesis by P. mirabilis during coinfection due to high-affinity import of leucine by P. stuartii. Further investigation of genes and pathways that provide a competitive advantage during both single-species and polymicrobial CAUTI will likely provide robust targets for therapeutic intervention to reduce P. mirabilis

  1. Genome-wide transposon mutagenesis of Proteus mirabilis: Essential genes, fitness factors for catheter-associated urinary tract infection, and the impact of polymicrobial infection on fitness requirements.

    PubMed

    Armbruster, Chelsie E; Forsyth-DeOrnellas, Valerie; Johnson, Alexandra O; Smith, Sara N; Zhao, Lili; Wu, Weisheng; Mobley, Harry L T

    2017-06-01

    The Gram-negative bacterium Proteus mirabilis is a leading cause of catheter-associated urinary tract infections (CAUTIs), which are often polymicrobial. Numerous prior studies have uncovered virulence factors for P. mirabilis pathogenicity in a murine model of ascending UTI, but little is known concerning pathogenesis during CAUTI or polymicrobial infection. In this study, we utilized five pools of 10,000 transposon mutants each and transposon insertion-site sequencing (Tn-Seq) to identify the full arsenal of P. mirabilis HI4320 fitness factors for single-species versus polymicrobial CAUTI with Providencia stuartii BE2467. 436 genes in the input pools lacked transposon insertions and were therefore concluded to be essential for P. mirabilis growth in rich medium. 629 genes were identified as P. mirabilis fitness factors during single-species CAUTI. Tn-Seq from coinfection with P. stuartii revealed 217/629 (35%) of the same genes as identified by single-species Tn-Seq, and 1353 additional factors that specifically contribute to colonization during coinfection. Mutants were constructed in eight genes of interest to validate the initial screen: 7/8 (88%) mutants exhibited the expected phenotypes for single-species CAUTI, and 3/3 (100%) validated the expected phenotypes for polymicrobial CAUTI. This approach provided validation of numerous previously described P. mirabilis fitness determinants from an ascending model of UTI, the discovery of novel fitness determinants specifically for CAUTI, and a stringent assessment of how polymicrobial infection influences fitness requirements. For instance, we describe a requirement for branched-chain amino acid biosynthesis by P. mirabilis during coinfection due to high-affinity import of leucine by P. stuartii. Further investigation of genes and pathways that provide a competitive advantage during both single-species and polymicrobial CAUTI will likely provide robust targets for therapeutic intervention to reduce P. mirabilis

  2. Transposon mutagenesis of probiotic Lactobacillus casei identifies asnH, an asparagine synthetase gene involved in its immune-activating capacity.

    PubMed

    Ito, Masahiro; Kim, Yun-Gi; Tsuji, Hirokazu; Takahashi, Takuya; Kiwaki, Mayumi; Nomoto, Koji; Danbara, Hirofumi; Okada, Nobuhiko

    2014-01-01

    Lactobacillus casei ATCC 27139 enhances host innate immunity, and the J1 phage-resistant mutants of this strain lose the activity. A transposon insertion mutant library of L. casei ATCC 27139 was constructed, and nine J1 phage-resistant mutants out of them were obtained. Cloning and sequencing analyses identified three independent genes that were disrupted by insertion of the transposon element: asnH, encoding asparagine synthetase, and dnaJ and dnaK, encoding the molecular chaperones DnaJ and DnaK, respectively. Using an in vivo mouse model of Listeria infection, only asnH mutant showed deficiency in their ability to enhance host innate immunity, and complementation of the mutation by introduction of the wild-type asnH in the mutant strain recovered the immuno-augmenting activity. AsnH protein exhibited asparagine synthetase activity when the lysozyme-treated cell wall extracts of L. casei ATCC 27139 was added as substrate. The asnH mutants lost the thick and rigid peptidoglycan features that are characteristic to the wild-type cells, indicating that AsnH of L. casei is involved in peptidoglycan biosynthesis. These results indicate that asnH is required for the construction of the peptidoglycan composition involved in the immune-activating capacity of L. casei ATCC 27139.

  3. Genome Sequencing and Transposon Mutagenesis of Burkholderia seminalis TC3.4.2R3 Identify Genes Contributing to Suppression of Orchid Necrosis Caused by B. gladioli.

    PubMed

    Araújo, Welington L; Creason, Allison L; Mano, Emy T; Camargo-Neves, Aline A; Minami, Sonia N; Chang, Jeff H; Loper, Joyce E

    2016-06-01

    From a screen of 36 plant-associated strains of Burkholderia spp., we identified 24 strains that suppressed leaf and pseudobulb necrosis of orchid caused by B. gladioli. To gain insights into the mechanisms of disease suppression, we generated a draft genome sequence from one suppressive strain, TC3.4.2R3. The genome is an estimated 7.67 megabases in size, with three replicons, two chromosomes, and the plasmid pC3. Using a combination of multilocus sequence analysis and phylogenomics, we identified TC3.4.2R3 as B. seminalis, a species within the Burkholderia cepacia complex that includes opportunistic human pathogens and environmental strains. We generated and screened a library of 3,840 transposon mutants of strain TC3.4.2R3 on orchid leaves to identify genes contributing to plant disease suppression. Twelve mutants deficient in suppression of leaf necrosis were selected and the transposon insertions were mapped to eight loci. One gene is in a wcb cluster that is related to synthesis of extracellular polysaccharide, a key determinant in bacterial-host interactions in other systems, and the other seven are highly conserved among Burkholderia spp. The fundamental information developed in this study will serve as a resource for future research aiming to identify mechanisms contributing to biological control.

  4. Chemical and UV Mutagenesis.

    PubMed

    Bose, Jeffrey L

    2016-01-01

    The ability to create mutations is an important step towards understanding bacterial physiology and virulence. While targeted approaches are invaluable, the ability to produce genome-wide random mutations can lead to crucial discoveries. Transposon mutagenesis is a useful approach, but many interesting mutations can be missed by these insertions that interrupt coding and noncoding sequences due to the integration of an entire transposon. Chemical mutagenesis and UV-based random mutagenesis are alternate approaches to isolate mutations of interest with the potential of only single nucleotide changes. Once a standard method, difficulty in identifying mutation sites had decreased the popularity of this technique. However, thanks to the recent emergence of economical whole-genome sequencing, this approach to making mutations can once again become a viable option. Therefore, this chapter provides an overview protocol for random mutagenesis using UV light or DNA-damaging chemicals.

  5. Large scale scientific computing

    SciTech Connect

    Deuflhard, P. ); Engquist, B. )

    1987-01-01

    This book presents papers on large scale scientific computing. It includes: Initial value problems of ODE's and parabolic PDE's; Boundary value problems of ODE's and elliptic PDE's; Hyperbolic PDE's; Inverse problems; Optimization and optimal control problems; and Algorithm adaptation on supercomputers.

  6. Transposon facilitated DNA sequencing

    SciTech Connect

    Berg, D.E.; Berg, C.M.; Huang, H.V.

    1990-01-01

    The purpose of this research is to investigate and develop methods that exploit the power of bacterial transposable elements for large scale DNA sequencing: Our premise is that the use of transposons to put primer binding sites randomly in target DNAs should provide access to all portions of large DNA fragments, without the inefficiencies of methods involving random subcloning and attendant repetitive sequencing, or of sequential synthesis of many oligonucleotide primers that are used to match systematically along a DNA molecule. Two unrelated bacterial transposons, Tn5 and {gamma}{delta}, are being used because they have both proven useful for molecular analyses, and because they differ sufficiently in mechanism and specificity of transposition to merit parallel development.

  7. Large-Scale Disasters

    NASA Astrophysics Data System (ADS)

    Gad-El-Hak, Mohamed

    "Extreme" events - including climatic events, such as hurricanes, tornadoes, and drought - can cause massive disruption to society, including large death tolls and property damage in the billions of dollars. Events in recent years have shown the importance of being prepared and that countries need to work together to help alleviate the resulting pain and suffering. This volume presents a review of the broad research field of large-scale disasters. It establishes a common framework for predicting, controlling and managing both manmade and natural disasters. There is a particular focus on events caused by weather and climate change. Other topics include air pollution, tsunamis, disaster modeling, the use of remote sensing and the logistics of disaster management. It will appeal to scientists, engineers, first responders and health-care professionals, in addition to graduate students and researchers who have an interest in the prediction, prevention or mitigation of large-scale disasters.

  8. Rapid 96-well plates DNA extraction and sequencing procedures to identify genome-wide transposon insertion sites in a difficult to lyse bacterium: Lactobacillus casei.

    PubMed

    Scornec, Hélène; Tichit, Magali; Bouchier, Christiane; Pédron, Thierry; Cavin, Jean-François; Sansonetti, Philippe J; Licandro-Seraut, Hélène

    2014-11-01

    Random transposon mutagenesis followed by adequate screening methods is an unavoidable procedure to characterize genetics of bacterial adaptation to environmental changes. We have recently constructed a mutant library of Lactobacillus casei and we aimed to fully annotate it. However, we have observed that, for L. casei which is a difficult to lyse bacterium, methods used to identify the transposon insertion site in a few mutants (transposon rescue by restriction and recircularization or PCR-based methods) were not transposable for a larger number because they are too time-consuming and sometimes not reliable. Here, we describe a method for large-scale and reliable identification of transposon insertion sites in a L. casei mutant library of 9250 mutants. DNA extraction procedure based on silica membranes in 96-column format was optimized to obtain genomic DNA from a large number of mutants. Then reliable direct genomic sequencing was improved to fit the obtained genomic DNA extracts. Using this procedure, readable and identifiable sequences were obtained for 87% of the L. casei mutants. This method extends the applications of a library of this type, reduces the number of insertions needed to be screened, and allows selection of specific mutants from an arrayed and stored mutant library. This method is applicable to any already existing mutant library (obtained by transposon or insertional mutagenesis) and could be useful for other bacterial species, especially for highly lysis-resistant bacteria species such as lactic acid bacteria. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. Whole-body sleeping beauty mutagenesis can cause penetrant leukemia/lymphoma and rare high-grade glioma without associated embryonic lethality.

    PubMed

    Collier, Lara S; Adams, David J; Hackett, Christopher S; Bendzick, Laura E; Akagi, Keiko; Davies, Michael N; Diers, Miechaleen D; Rodriguez, Fausto J; Bender, Aaron M; Tieu, Christina; Matise, Ilze; Dupuy, Adam J; Copeland, Neal G; Jenkins, Nancy A; Hodgson, J Graeme; Weiss, William A; Jenkins, Robert B; Largaespada, David A

    2009-11-01

    The Sleeping Beauty (SB) transposon system has been used as a somatic mutagen to identify candidate cancer genes. In previous studies, efficient leukemia/lymphoma formation on an otherwise wild-type genetic background occurred in mice undergoing whole-body mobilization of transposons, but was accompanied by high levels of embryonic lethality. To explore the utility of SB for large-scale cancer gene discovery projects, we have generated mice that carry combinations of different transposon and transposase transgenes. We have identified a transposon/transposase combination that promotes highly penetrant leukemia/lymphoma formation on an otherwise wild-type genetic background, yet does not cause embryonic lethality. Infiltrating gliomas also occurred at lower penetrance in these mice. SB-induced or accelerated tumors do not harbor large numbers of chromosomal amplifications or deletions, indicating that transposon mobilization likely promotes tumor formation by insertional mutagenesis of cancer genes, and not by promoting wide-scale genomic instability. Cloning of transposon insertions from lymphomas/leukemias identified common insertion sites at known and candidate novel cancer genes. These data indicate that a high mutagenesis rate can be achieved using SB without high levels of embryonic lethality or genomic instability. Furthermore, the SB system could be used to identify new genes involved in lymphomagenesis/leukemogenesis.

  10. Large Scale Nonlinear Programming.

    DTIC Science & Technology

    1978-06-15

    KEY WORDS (Conhinu. as, t.n.t.. aid. if nic••iary aid ld.ntify by block n,a,b.r) L. In,~~~ IP!CIE LARGE SCALE OPTIMIZATION APPLICATIONS OF NONLINEAR ... NONLINEAR PROGRAMMING by Garth P. McCormick 1. Introduction The general mathematical programming ( optimization ) problem can be stated in the following form...because the difficulty in solving a general nonlinear optimization problem has a~ much to do with the nature of the functions involved as it does with the

  11. Characterization of the Pathogenicity of Streptococcus intermedius TYG1620 Isolated from a Human Brain Abscess Based on the Complete Genome Sequence with Transcriptome Analysis and Transposon Mutagenesis in a Murine Subcutaneous Abscess Model.

    PubMed

    Hasegawa, Noriko; Sekizuka, Tsuyoshi; Sugi, Yutaka; Kawakami, Nobuhiro; Ogasawara, Yumiko; Kato, Kengo; Yamashita, Akifumi; Takeuchi, Fumihiko; Kuroda, Makoto

    2017-02-01

    Streptococcus intermedius is known to cause periodontitis and pyogenic infections in the brain and liver. Here we report the complete genome sequence of strain TYG1620 (genome size, 2,006,877 bp; GC content, 37.6%; 2,020 predicted open reading frames [ORFs]) isolated from a brain abscess in an infant. Comparative analysis of S. intermedius genome sequences suggested that TYG1620 carries a notable type VII secretion system (T7SS), two long repeat regions, and 19 ORFs for cell wall-anchored proteins (CWAPs). To elucidate the genes responsible for the pathogenicity of TYG1620, transcriptome analysis was performed in a murine subcutaneous abscess model. The results suggest that the levels of expression of small hypothetical proteins similar to phenol-soluble modulin β1 (PSMβ1), a staphylococcal virulence factor, significantly increased in the abscess model. In addition, an experiment in a murine subcutaneous abscess model with random transposon (Tn) mutant attenuation suggested that Tn mutants with mutations in 212 ORFs in the Tn mutant library were attenuated in the murine abscess model (629 ORFs were disrupted in total); the 212 ORFs are putatively essential for abscess formation. Transcriptome analysis identified 37 ORFs, including paralogs of the T7SS and a putative glucan-binding CWAP in long repeat regions, to be upregulated and attenuated in vivo This study provides a comprehensive characterization of S. intermedius pathogenicity based on the complete genome sequence and a murine subcutaneous abscess model with transcriptome and Tn mutagenesis, leading to the identification of pivotal targets for vaccines or antimicrobial agents for the control of S. intermedius infections. Copyright © 2017 American Society for Microbiology.

  12. Characterization of the Pathogenicity of Streptococcus intermedius TYG1620 Isolated from a Human Brain Abscess Based on the Complete Genome Sequence with Transcriptome Analysis and Transposon Mutagenesis in a Murine Subcutaneous Abscess Model

    PubMed Central

    Hasegawa, Noriko; Sekizuka, Tsuyoshi; Sugi, Yutaka; Kawakami, Nobuhiro; Ogasawara, Yumiko; Kato, Kengo; Yamashita, Akifumi; Takeuchi, Fumihiko

    2016-01-01

    ABSTRACT Streptococcus intermedius is known to cause periodontitis and pyogenic infections in the brain and liver. Here we report the complete genome sequence of strain TYG1620 (genome size, 2,006,877 bp; GC content, 37.6%; 2,020 predicted open reading frames [ORFs]) isolated from a brain abscess in an infant. Comparative analysis of S. intermedius genome sequences suggested that TYG1620 carries a notable type VII secretion system (T7SS), two long repeat regions, and 19 ORFs for cell wall-anchored proteins (CWAPs). To elucidate the genes responsible for the pathogenicity of TYG1620, transcriptome analysis was performed in a murine subcutaneous abscess model. The results suggest that the levels of expression of small hypothetical proteins similar to phenol-soluble modulin β1 (PSMβ1), a staphylococcal virulence factor, significantly increased in the abscess model. In addition, an experiment in a murine subcutaneous abscess model with random transposon (Tn) mutant attenuation suggested that Tn mutants with mutations in 212 ORFs in the Tn mutant library were attenuated in the murine abscess model (629 ORFs were disrupted in total); the 212 ORFs are putatively essential for abscess formation. Transcriptome analysis identified 37 ORFs, including paralogs of the T7SS and a putative glucan-binding CWAP in long repeat regions, to be upregulated and attenuated in vivo. This study provides a comprehensive characterization of S. intermedius pathogenicity based on the complete genome sequence and a murine subcutaneous abscess model with transcriptome and Tn mutagenesis, leading to the identification of pivotal targets for vaccines or antimicrobial agents for the control of S. intermedius infections. PMID:27895128

  13. Large scale tracking algorithms

    SciTech Connect

    Hansen, Ross L.; Love, Joshua Alan; Melgaard, David Kennett; Karelitz, David B.; Pitts, Todd Alan; Zollweg, Joshua David; Anderson, Dylan Z.; Nandy, Prabal; Whitlow, Gary L.; Bender, Daniel A.; Byrne, Raymond Harry

    2015-01-01

    Low signal-to-noise data processing algorithms for improved detection, tracking, discrimination and situational threat assessment are a key research challenge. As sensor technologies progress, the number of pixels will increase signi cantly. This will result in increased resolution, which could improve object discrimination, but unfortunately, will also result in a significant increase in the number of potential targets to track. Many tracking techniques, like multi-hypothesis trackers, suffer from a combinatorial explosion as the number of potential targets increase. As the resolution increases, the phenomenology applied towards detection algorithms also changes. For low resolution sensors, "blob" tracking is the norm. For higher resolution data, additional information may be employed in the detection and classfication steps. The most challenging scenarios are those where the targets cannot be fully resolved, yet must be tracked and distinguished for neighboring closely spaced objects. Tracking vehicles in an urban environment is an example of such a challenging scenario. This report evaluates several potential tracking algorithms for large-scale tracking in an urban environment.

  14. Large scale traffic simulations

    SciTech Connect

    Nagel, K.; Barrett, C.L. |; Rickert, M. |

    1997-04-01

    Large scale microscopic (i.e. vehicle-based) traffic simulations pose high demands on computational speed in at least two application areas: (i) real-time traffic forecasting, and (ii) long-term planning applications (where repeated {open_quotes}looping{close_quotes} between the microsimulation and the simulated planning of individual person`s behavior is necessary). As a rough number, a real-time simulation of an area such as Los Angeles (ca. 1 million travellers) will need a computational speed of much higher than 1 million {open_quotes}particle{close_quotes} (= vehicle) updates per second. This paper reviews how this problem is approached in different projects and how these approaches are dependent both on the specific questions and on the prospective user community. The approaches reach from highly parallel and vectorizable, single-bit implementations on parallel supercomputers for Statistical Physics questions, via more realistic implementations on coupled workstations, to more complicated driving dynamics implemented again on parallel supercomputers. 45 refs., 9 figs., 1 tab.

  15. Tol2 transposon-mediated transgenesis in Xenopus tropicalis.

    PubMed

    Hamlet, Michelle R Johnson; Yergeau, Donald A; Kuliyev, Emin; Takeda, Masatoshi; Taira, Masanori; Kawakami, Koichi; Mead, Paul E

    2006-09-01

    The diploid frog Xenopus tropicalis is becoming a powerful developmental genetic model system. Sequencing of the X. tropicalis genome is nearing completion and several labs are embarking on mutagenesis screens. We are interested in developing insertional mutagenesis strategies in X. tropicalis. Transposon-mediated insertional mutagenesis, once used exclusively in plants and invertebrate systems, is now more widely applicable to vertebrates. The first step in developing transposons as tools for mutagenesis is to demonstrate that these mobile elements function efficiently in the target organism. Here, we show that the Medaka fish transposon, Tol2, is able to stably integrate into the X. tropicalis genome and will serve as a powerful tool for insertional mutagenesis strategies in the frog.

  16. Transposon tools: worldwide landscape of intellectual property and technological developments.

    PubMed

    Palazzoli, Fabien; Testu, François-Xavier; Merly, Franck; Bigot, Yves

    2010-03-01

    DNA transposons are considered to be good candidates for developing tools for genome engineering, insertional mutagenesis and gene delivery for therapeutic purposes, as illustrated by the recent first clinical trial of a transposon. In this article we set out to highlight the interest of patent information, and to develop a strategy for the technological development of transposon tools, similar to what has been done in many other fields. We propose a patent landscape for transposon tools, including the changes in international patent applications, and review the leading inventors and applicants. We also provide an overview of the potential patent portfolio for the prokaryotic and eukaryotic transposons that are exploited by spin-off companies. Finally, we discuss the difficulties involved in tracing relevant state-of-the-art of articles and patent documents, based on the example of one of the most promising transposon systems, including all the impacts on the technological development of transposon tools.

  17. Haploinsufficiency-based large-scale forward genetic analysis of filamentous growth in the diploid human fungal pathogen C.albicans

    PubMed Central

    Uhl, M.Andrew; Biery, Matt; Craig, Nancy; Johnson, Alexander D.

    2003-01-01

    Candida albicans is the most prevalent human fungal pathogen. Here, we take advantage of haploinsufficiency and transposon mutagenesis to perform large-scale loss-of-function genetic screen in this organism. We identified mutations in 146 genes that affect the switch between its single-cell (yeast) form and filamentous forms of growth; this switch appears central to the virulence of C.albicans. The encoded proteins include those involved in nutrient sensing, signal transduction, transcriptional control, cytoskeletal organization and cell wall construction. Approxim ately one-third of the genes identified in the screen lack homologs in Saccharomyces cerevisiae and other model organisms and thus constitute candidate antifungal drug targets. These results illustrate the value of performing forward genetic studies in bona fide pathogens. PMID:12773383

  18. Haploinsufficiency-based large-scale forward genetic analysis of filamentous growth in the diploid human fungal pathogen C.albicans.

    PubMed

    Uhl, M Andrew; Biery, Matt; Craig, Nancy; Johnson, Alexander D

    2003-06-02

    Candida albicans is the most prevalent human fungal pathogen. Here, we take advantage of haploinsufficiency and transposon mutagenesis to perform large-scale loss-of-function genetic screen in this organism. We identified mutations in 146 genes that affect the switch between its single-cell (yeast) form and filamentous forms of growth; this switch appears central to the virulence of C.albicans. The encoded proteins include those involved in nutrient sensing, signal transduction, transcriptional control, cytoskeletal organization and cell wall construction. Approximately one-third of the genes identified in the screen lack homologs in Saccharomyces cerevisiae and other model organisms and thus constitute candidate antifungal drug targets. These results illustrate the value of performing forward genetic studies in bona fide pathogens.

  19. Large-scale structural optimization

    NASA Technical Reports Server (NTRS)

    Sobieszczanski-Sobieski, J.

    1983-01-01

    Problems encountered by aerospace designers in attempting to optimize whole aircraft are discussed, along with possible solutions. Large scale optimization, as opposed to component-by-component optimization, is hindered by computational costs, software inflexibility, concentration on a single, rather than trade-off, design methodology and the incompatibility of large-scale optimization with single program, single computer methods. The software problem can be approached by placing the full analysis outside of the optimization loop. Full analysis is then performed only periodically. Problem-dependent software can be removed from the generic code using a systems programming technique, and then embody the definitions of design variables, objective function and design constraints. Trade-off algorithms can be used at the design points to obtain quantitative answers. Finally, decomposing the large-scale problem into independent subproblems allows systematic optimization of the problems by an organization of people and machines.

  20. Large-scale circuit simulation

    NASA Astrophysics Data System (ADS)

    Wei, Y. P.

    1982-12-01

    The simulation of VLSI (Very Large Scale Integration) circuits falls beyond the capabilities of conventional circuit simulators like SPICE. On the other hand, conventional logic simulators can only give the results of logic levels 1 and 0 with the attendent loss of detail in the waveforms. The aim of developing large-scale circuit simulation is to bridge the gap between conventional circuit simulation and logic simulation. This research is to investigate new approaches for fast and relatively accurate time-domain simulation of MOS (Metal Oxide Semiconductors), LSI (Large Scale Integration) and VLSI circuits. New techniques and new algorithms are studied in the following areas: (1) analysis sequencing (2) nonlinear iteration (3) modified Gauss-Seidel method (4) latency criteria and timestep control scheme. The developed methods have been implemented into a simulation program PREMOS which could be used as a design verification tool for MOS circuits.

  1. Large Scale Dynamos in Stars

    NASA Astrophysics Data System (ADS)

    Vishniac, Ethan T.

    2015-01-01

    We show that a differentially rotating conducting fluid automatically creates a magnetic helicity flux with components along the rotation axis and in the direction of the local vorticity. This drives a rapid growth in the local density of current helicity, which in turn drives a large scale dynamo. The dynamo growth rate derived from this process is not constant, but depends inversely on the large scale magnetic field strength. This dynamo saturates when buoyant losses of magnetic flux compete with the large scale dynamo, providing a simple prediction for magnetic field strength as a function of Rossby number in stars. Increasing anisotropy in the turbulence produces a decreasing magnetic helicity flux, which explains the flattening of the B/Rossby number relation at low Rossby numbers. We also show that the kinetic helicity is always a subdominant effect. There is no kinematic dynamo in real stars.

  2. Very Large Scale Integration (VLSI).

    ERIC Educational Resources Information Center

    Yeaman, Andrew R. J.

    Very Large Scale Integration (VLSI), the state-of-the-art production techniques for computer chips, promises such powerful, inexpensive computing that, in the future, people will be able to communicate with computer devices in natural language or even speech. However, before full-scale VLSI implementation can occur, certain salient factors must be…

  3. Galaxy clustering on large scales.

    PubMed Central

    Efstathiou, G

    1993-01-01

    I describe some recent observations of large-scale structure in the galaxy distribution. The best constraints come from two-dimensional galaxy surveys and studies of angular correlation functions. Results from galaxy redshift surveys are much less precise but are consistent with the angular correlations, provided the distortions in mapping between real-space and redshift-space are relatively weak. The galaxy two-point correlation function, rich-cluster two-point correlation function, and galaxy-cluster cross-correlation function are all well described on large scales ( greater, similar 20h-1 Mpc, where the Hubble constant, H0 = 100h km.s-1.Mpc; 1 pc = 3.09 x 10(16) m) by the power spectrum of an initially scale-invariant, adiabatic, cold-dark-matter Universe with Gamma = Omegah approximately 0.2. I discuss how this fits in with the Cosmic Background Explorer (COBE) satellite detection of large-scale anisotropies in the microwave background radiation and other measures of large-scale structure in the Universe. PMID:11607400

  4. Fighting an old war with a new weapon--silencing transposons by Piwi-interacting RNA.

    PubMed

    Guo, Manhong; Wu, Yuliang

    2013-09-01

    Discovered six decades ago, transposons are known to selfishly multiply within and between chromosomes. Although they may play a creative role in building new functional parts of the genome, transposons usually cause insertional mutagenesis and/or turn nearby genes on or off. To maintain genome integrity, cells use a variety of strategies to defend against the proliferation of transposons. A class of small noncoding RNA, discovered seven years ago and called piRNA, is a new player in the war to silence transposons. piRNA is made via two biogenesis pathways: the primary processing pathway and the ping-pong amplification loop. These pathways are critically involved in transposon RNA degradation, DNA methylation, and histone modification machinery that represses transposons. In this review, we briefly introduce transposon-caused genomic instability and summarize our current understanding of the piRNA pathway, focusing on its key function in transposon silencing.

  5. DNA transposons: nature and applications in genomics.

    PubMed

    Muñoz-López, Martín; García-Pérez, José L

    2010-04-01

    Repeated DNA makes up a large fraction of a typical mammalian genome, and some repetitive elements are able to move within the genome (transposons and retrotransposons). DNA transposons move from one genomic location to another by a cut-and-paste mechanism. They are powerful forces of genetic change and have played a significant role in the evolution of many genomes. As genetic tools, DNA transposons can be used to introduce a piece of foreign DNA into a genome. Indeed, they have been used for transgenesis and insertional mutagenesis in different organisms, since these elements are not generally dependent on host factors to mediate their mobility. Thus, DNA transposons are useful tools to analyze the regulatory genome, study embryonic development, identify genes and pathways implicated in disease or pathogenesis of pathogens, and even contribute to gene therapy. In this review, we will describe the nature of these elements and discuss recent advances in this field of research, as well as our evolving knowledge of the DNA transposons most widely used in these studies.

  6. Transformation frequency of a mariner-based transposon in Rickettsia rickettsii.

    PubMed

    Clark, Tina R; Lackey, Amanda M; Kleba, Betsy; Driskell, Lonnie O; Lutter, Erika I; Martens, Craig; Wood, David O; Hackstadt, Ted

    2011-09-01

    Transformation frequencies of a mariner-based transposon system in Rickettsia rickettsii were determined using a plaque assay system for enumeration and isolation of mutants. Sequence analysis of insertion sites in both R. rickettsii and R. prowazekii indicated that insertions were random. Transposon mutagenesis provides a useful tool for rickettsial research.

  7. Cosmology with Large Scale Structure

    NASA Astrophysics Data System (ADS)

    Ho, Shirley; Cuesta, A.; Ross, A.; Seo, H.; DePutter, R.; Padmanabhan, N.; White, M.; Myers, A.; Bovy, J.; Blanton, M.; Hernandez, C.; Mena, O.; Percival, W.; Prada, F.; Ross, N. P.; Saito, S.; Schneider, D.; Skibba, R.; Smith, K.; Slosar, A.; Strauss, M.; Verde, L.; Weinberg, D.; Bachall, N.; Brinkmann, J.; da Costa, L. A.

    2012-01-01

    The Sloan Digital Sky Survey I-III surveyed 14,000 square degrees, and delivered over a trillion pixels of imaging data. I present cosmological results from this unprecedented data set which contains over a million galaxies distributed between redshift of 0.45 to 0.70. With such a large volume of data set, high precision cosmological constraints can be obtained given a careful control and understanding of observational systematics. I present a novel treatment of observational systematics and its application to the clustering signals from the data set. I will present cosmological constraints on dark components of the Universe and tightest constraints of the non-gaussianity of early Universe to date utilizing Large Scale Structure.

  8. Large scale biomimetic membrane arrays.

    PubMed

    Hansen, Jesper S; Perry, Mark; Vogel, Jörg; Groth, Jesper S; Vissing, Thomas; Larsen, Marianne S; Geschke, Oliver; Emneús, Jenny; Bohr, Henrik; Nielsen, Claus H

    2009-10-01

    To establish planar biomimetic membranes across large scale partition aperture arrays, we created a disposable single-use horizontal chamber design that supports combined optical-electrical measurements. Functional lipid bilayers could easily and efficiently be established across CO(2) laser micro-structured 8 x 8 aperture partition arrays with average aperture diameters of 301 +/- 5 microm. We addressed the electro-physical properties of the lipid bilayers established across the micro-structured scaffold arrays by controllable reconstitution of biotechnological and physiological relevant membrane peptides and proteins. Next, we tested the scalability of the biomimetic membrane design by establishing lipid bilayers in rectangular 24 x 24 and hexagonal 24 x 27 aperture arrays, respectively. The results presented show that the design is suitable for further developments of sensitive biosensor assays, and furthermore demonstrate that the design can conveniently be scaled up to support planar lipid bilayers in large square-centimeter partition arrays.

  9. Challenges for Large Scale Simulations

    NASA Astrophysics Data System (ADS)

    Troyer, Matthias

    2010-03-01

    With computational approaches becoming ubiquitous the growing impact of large scale computing on research influences both theoretical and experimental work. I will review a few examples in condensed matter physics and quantum optics, including the impact of computer simulations in the search for supersolidity, thermometry in ultracold quantum gases, and the challenging search for novel phases in strongly correlated electron systems. While only a decade ago such simulations needed the fastest supercomputers, many simulations can now be performed on small workstation clusters or even a laptop: what was previously restricted to a few experts can now potentially be used by many. Only part of the gain in computational capabilities is due to Moore's law and improvement in hardware. Equally impressive is the performance gain due to new algorithms - as I will illustrate using some recently developed algorithms. At the same time modern peta-scale supercomputers offer unprecedented computational power and allow us to tackle new problems and address questions that were impossible to solve numerically only a few years ago. While there is a roadmap for future hardware developments to exascale and beyond, the main challenges are on the algorithmic and software infrastructure side. Among the problems that face the computational physicist are: the development of new algorithms that scale to thousands of cores and beyond, a software infrastructure that lifts code development to a higher level and speeds up the development of new simulation programs for large scale computing machines, tools to analyze the large volume of data obtained from such simulations, and as an emerging field provenance-aware software that aims for reproducibility of the complete computational workflow from model parameters to the final figures. Interdisciplinary collaborations and collective efforts will be required, in contrast to the cottage-industry culture currently present in many areas of computational

  10. Transposons for cancer gene discovery: Sleeping Beauty and beyond

    PubMed Central

    Collier, Lara S; Largaespada, David A

    2007-01-01

    The use of Sleeping Beauty transposons as somatic mutagens to discover cancer genes in hematopoietic tumors and sarcomas has been documented. Here, we discuss the future of Sleeping Beauty for cancer genetic studies and the potential use of additional transposable elements for somatic mutagenesis. PMID:18047692

  11. Large-scale PACS implementation.

    PubMed

    Carrino, J A; Unkel, P J; Miller, I D; Bowser, C L; Freckleton, M W; Johnson, T G

    1998-08-01

    The transition to filmless radiology is a much more formidable task than making the request for proposal to purchase a (Picture Archiving and Communications System) PACS. The Department of Defense and the Veterans Administration have been pioneers in the transformation of medical diagnostic imaging to the electronic environment. Many civilian sites are expected to implement large-scale PACS in the next five to ten years. This presentation will related the empirical insights gleaned at our institution from a large-scale PACS implementation. Our PACS integration was introduced into a fully operational department (not a new hospital) in which work flow had to continue with minimal impact. Impediments to user acceptance will be addressed. The critical components of this enormous task will be discussed. The topics covered during this session will include issues such as phased implementation, DICOM (digital imaging and communications in medicine) standard-based interaction of devices, hospital information system (HIS)/radiology information system (RIS) interface, user approval, networking, workstation deployment and backup procedures. The presentation will make specific suggestions regarding the implementation team, operating instructions, quality control (QC), training and education. The concept of identifying key functional areas is relevant to transitioning the facility to be entirely on line. Special attention must be paid to specific functional areas such as the operating rooms and trauma rooms where the clinical requirements may not match the PACS capabilities. The printing of films may be necessary for certain circumstances. The integration of teleradiology and remote clinics into a PACS is a salient topic with respect to the overall role of the radiologists providing rapid consultation. A Web-based server allows a clinician to review images and reports on a desk-top (personal) computer and thus reduce the number of dedicated PACS review workstations. This session

  12. Large scale cluster computing workshop

    SciTech Connect

    Dane Skow; Alan Silverman

    2002-12-23

    Recent revolutions in computer hardware and software technologies have paved the way for the large-scale deployment of clusters of commodity computers to address problems heretofore the domain of tightly coupled SMP processors. Near term projects within High Energy Physics and other computing communities will deploy clusters of scale 1000s of processors and be used by 100s to 1000s of independent users. This will expand the reach in both dimensions by an order of magnitude from the current successful production facilities. The goals of this workshop were: (1) to determine what tools exist which can scale up to the cluster sizes foreseen for the next generation of HENP experiments (several thousand nodes) and by implication to identify areas where some investment of money or effort is likely to be needed. (2) To compare and record experimences gained with such tools. (3) To produce a practical guide to all stages of planning, installing, building and operating a large computing cluster in HENP. (4) To identify and connect groups with similar interest within HENP and the larger clustering community.

  13. Large-Scale Sequence Comparison.

    PubMed

    Lal, Devi; Verma, Mansi

    2017-01-01

    There are millions of sequences deposited in genomic databases, and it is an important task to categorize them according to their structural and functional roles. Sequence comparison is a prerequisite for proper categorization of both DNA and protein sequences, and helps in assigning a putative or hypothetical structure and function to a given sequence. There are various methods available for comparing sequences, alignment being first and foremost for sequences with a small number of base pairs as well as for large-scale genome comparison. Various tools are available for performing pairwise large sequence comparison. The best known tools either perform global alignment or generate local alignments between the two sequences. In this chapter we first provide basic information regarding sequence comparison. This is followed by the description of the PAM and BLOSUM matrices that form the basis of sequence comparison. We also give a practical overview of currently available methods such as BLAST and FASTA, followed by a description and overview of tools available for genome comparison including LAGAN, MumMER, BLASTZ, and AVID.

  14. Large Scale Homing in Honeybees

    PubMed Central

    Pahl, Mario; Zhu, Hong; Tautz, Jürgen; Zhang, Shaowu

    2011-01-01

    Honeybee foragers frequently fly several kilometres to and from vital resources, and communicate those locations to their nest mates by a symbolic dance language. Research has shown that they achieve this feat by memorizing landmarks and the skyline panorama, using the sun and polarized skylight as compasses and by integrating their outbound flight paths. In order to investigate the capacity of the honeybees' homing abilities, we artificially displaced foragers to novel release spots at various distances up to 13 km in the four cardinal directions. Returning bees were individually registered by a radio frequency identification (RFID) system at the hive entrance. We found that homing rate, homing speed and the maximum homing distance depend on the release direction. Bees released in the east were more likely to find their way back home, and returned faster than bees released in any other direction, due to the familiarity of global landmarks seen from the hive. Our findings suggest that such large scale homing is facilitated by global landmarks acting as beacons, and possibly the entire skyline panorama. PMID:21602920

  15. Large Scale Magnetostrictive Valve Actuator

    NASA Technical Reports Server (NTRS)

    Richard, James A.; Holleman, Elizabeth; Eddleman, David

    2008-01-01

    Marshall Space Flight Center's Valves, Actuators and Ducts Design and Development Branch developed a large scale magnetostrictive valve actuator. The potential advantages of this technology are faster, more efficient valve actuators that consume less power and provide precise position control and deliver higher flow rates than conventional solenoid valves. Magnetostrictive materials change dimensions when a magnetic field is applied; this property is referred to as magnetostriction. Magnetostriction is caused by the alignment of the magnetic domains in the material s crystalline structure and the applied magnetic field lines. Typically, the material changes shape by elongating in the axial direction and constricting in the radial direction, resulting in no net change in volume. All hardware and testing is complete. This paper will discuss: the potential applications of the technology; overview of the as built actuator design; discuss problems that were uncovered during the development testing; review test data and evaluate weaknesses of the design; and discuss areas for improvement for future work. This actuator holds promises of a low power, high load, proportionally controlled actuator for valves requiring 440 to 1500 newtons load.

  16. Transposon transgenesis in Xenopus.

    PubMed

    Yergeau, Donald A; Kelley, Clair M; Zhu, Haiqing; Kuliyev, Emin; Mead, Paul E

    2010-05-01

    Transposon-mediated integration strategies in Xenopus offer simple and robust methods for the generation of germline transgenic animals. Co-injection of fertilized one-cell embryos with plasmid DNA harboring a transposon transgene and synthetic mRNA encoding the cognate transposase enzyme results in mosaic integration of the transposon at early cleavage stages that are frequently passed through the germline in the adult animal. Micro-injection of fertilized embryos is a routine procedure used by many laboratories that use Xenopus as a developmental model and, as such, the transposon transgenesis method can be performed without additional equipment or specialized methodologies. The methods for injecting Xenopus embryos are well documented in the literature so here we provide a step-by-step guide to other aspects of transposon transgenesis, including screening mosaic founders for germline transmission of the transgene and general husbandry considerations related to management of populations of transgenic frogs.

  17. Transposon transgenesis in Xenopus

    PubMed Central

    Yergeau, Donald A.; Kelley, Clair M.; Zhu, Haiqing; Kuliyev, Emin; Mead, Paul E.

    2010-01-01

    Transposon-mediated integration strategies in Xenopus offer simple and robust methods for the generation of germline transgenic animals. Co-injection of fertilized one-cell embryos with plasmid DNA harboring a transposon transgene and synthetic mRNA encoding the cognate transposase enzyme results in mosaic integration of the transposon at early cleavage stages that are frequently passed through the germline in the adult animal. Micro-injection of fertilized embryos is a routine procedure used by many laboratories that use Xenopus as a developmental model and, as such, the transposon transgenesis method can be performed without additional equipment or specialized methodologies. The methods for injecting Xenopus embryos are well documented in the literature so here we provide a step-by-step guide to other aspects of transposon transgenesis, including screening mosaic founders for germline transmission of the transgene and general husbandry considerations related to management of populations of transgenic frogs. PMID:20211730

  18. Transposons in C. elegans.

    PubMed

    Bessereau, Jean-Louis

    2006-01-18

    Transposons are discrete segments of DNA capable of moving through the genome of their host via an RNA intermediate in the case of class I retrotransposon or via a "cut-and-paste" mechanism for class II DNA transposons. Since transposons take advantage of their host's cellular machinery to proliferate in the genome and enter new hosts, transposable elements can be viewed as parasitic or "selfish DNA". However, transposons may have been beneficial for their hosts as genome evolution drivers, thus providing an example of molecular mutualism. Interactions between transposon and C. elegans research were undoubtedly mutualistic, leading to the advent of needed genomic tools to drive C. elegans research while providing insights into the transposition field. Tc1, the first C. elegans transposon to be identified, turned out to be the founding member of a widespread family of mobile elements: the Tc1/mariner superfamily. The investigation into transposition regulation in C. elegans has uncovered an unforeseen link between transposition, genome surveillance and RNA interference. Conversely, transposons were utilized soon after their identification to inactivate and clone genes, providing some of the first molecular identities of C. elegans genes. Recent results suggest that transposons might provide a means to engineer site-directed mutations into the C. elegans genome. This article describes the different transposons present in the C. elegans genome with a specific emphasis on the ones that proved to be mobile under laboratory conditions. Mechanisms and control of transposition are discussed briefly. Some tools based on the use of transposons for C. elegans research are presented at the end of this review.

  19. Methane emissions on large scales

    NASA Astrophysics Data System (ADS)

    Beswick, K. M.; Simpson, T. W.; Fowler, D.; Choularton, T. W.; Gallagher, M. W.; Hargreaves, K. J.; Sutton, M. A.; Kaye, A.

    with previous results from the area, indicating that this method of data analysis provided good estimates of large scale methane emissions.

  20. Large Scale Nanolaminate Deformable Mirror

    SciTech Connect

    Papavasiliou, A; Olivier, S; Barbee, T; Miles, R; Chang, K

    2005-11-30

    This work concerns the development of a technology that uses Nanolaminate foils to form light-weight, deformable mirrors that are scalable over a wide range of mirror sizes. While MEMS-based deformable mirrors and spatial light modulators have considerably reduced the cost and increased the capabilities of adaptive optic systems, there has not been a way to utilize the advantages of lithography and batch-fabrication to produce large-scale deformable mirrors. This technology is made scalable by using fabrication techniques and lithography that are not limited to the sizes of conventional MEMS devices. Like many MEMS devices, these mirrors use parallel plate electrostatic actuators. This technology replicates that functionality by suspending a horizontal piece of nanolaminate foil over an electrode by electroplated nickel posts. This actuator is attached, with another post, to another nanolaminate foil that acts as the mirror surface. Most MEMS devices are produced with integrated circuit lithography techniques that are capable of very small line widths, but are not scalable to large sizes. This technology is very tolerant of lithography errors and can use coarser, printed circuit board lithography techniques that can be scaled to very large sizes. These mirrors use small, lithographically defined actuators and thin nanolaminate foils allowing them to produce deformations over a large area while minimizing weight. This paper will describe a staged program to develop this technology. First-principles models were developed to determine design parameters. Three stages of fabrication will be described starting with a 3 x 3 device using conventional metal foils and epoxy to a 10-across all-metal device with nanolaminate mirror surfaces.

  1. Large-Scale Information Systems

    SciTech Connect

    D. M. Nicol; H. R. Ammerlahn; M. E. Goldsby; M. M. Johnson; D. E. Rhodes; A. S. Yoshimura

    2000-12-01

    Large enterprises are ever more dependent on their Large-Scale Information Systems (LSLS), computer systems that are distinguished architecturally by distributed components--data sources, networks, computing engines, simulations, human-in-the-loop control and remote access stations. These systems provide such capabilities as workflow, data fusion and distributed database access. The Nuclear Weapons Complex (NWC) contains many examples of LSIS components, a fact that motivates this research. However, most LSIS in use grew up from collections of separate subsystems that were not designed to be components of an integrated system. For this reason, they are often difficult to analyze and control. The problem is made more difficult by the size of a typical system, its diversity of information sources, and the institutional complexities associated with its geographic distribution across the enterprise. Moreover, there is no integrated approach for analyzing or managing such systems. Indeed, integrated development of LSIS is an active area of academic research. This work developed such an approach by simulating the various components of the LSIS and allowing the simulated components to interact with real LSIS subsystems. This research demonstrated two benefits. First, applying it to a particular LSIS provided a thorough understanding of the interfaces between the system's components. Second, it demonstrated how more rapid and detailed answers could be obtained to questions significant to the enterprise by interacting with the relevant LSIS subsystems through simulated components designed with those questions in mind. In a final, added phase of the project, investigations were made on extending this research to wireless communication networks in support of telemetry applications.

  2. Insertional mutagenesis and illegitimate recombination in mycobacteria.

    PubMed Central

    Kalpana, G V; Bloom, B R; Jacobs, W R

    1991-01-01

    Mycobacteria, particularly Mycobacterium tuberculosis, Mycobacterium leprae, and Mycobacterium avium, are major pathogens of man. Although insertional mutagenesis has been an invaluable genetic tool for analyzing the mechanisms of microbial pathogenesis, it has not yet been possible to apply it to the mycobacteria. To overcome intrinsic difficulties in directly manipulating the genetics of slow-growing mycobacteria, including M. tuberculosis and bacille Calmette-Guérin (BCG) vaccine strains, we developed a system for random shuttle mutagenesis. A genomic library of Mycobacterium smegmatis was subjected to transposon mutagenesis with Tn5 seq1, a derivative of Tn5, in Escherichia coli and these transposon-containing recombinant plasmids were reintroduced into mycobacterial chromosomes by homologous recombination. This system has allowed us to isolate several random auxotrophic mutants of M. smegmatis. To extend this strategy to M. tuberculosis and BCG, targeted mutagenesis was performed using a cloned BCG methionine gene that was subjected to Tn5 seq1 mutagenesis in E. coli and reintroduced into the mycobacteria. Surprisingly for prokaryotes, both BCG and M. tuberculosis were found to incorporate linear DNA fragments into illegitimate sites throughout the mycobacterial genomes at a frequency of 10(-5) to 10(-4) relative to the number of transformants obtained with autonomously replicating vectors. Thus the efficient illegitimate recombination of linear DNA fragments provides the basis for an insertional mutagenesis system for M. tuberculosis and BCG. Images PMID:2052623

  3. Phylogenetic and Functional Characterization of the hAT Transposon Superfamily

    PubMed Central

    Arensburger, Peter; Hice, Robert H.; Zhou, Liqin; Smith, Ryan C.; Tom, Ariane C.; Wright, Jennifer A.; Knapp, Joshua; O'Brochta, David A.; Craig, Nancy L.; Atkinson, Peter W.

    2011-01-01

    Transposons are found in virtually all organisms and play fundamental roles in genome evolution. They can also acquire new functions in the host organism and some have been developed as incisive genetic tools for transformation and mutagenesis. The hAT transposon superfamily contains members from the plant and animal kingdoms, some of which are active when introduced into new host organisms. We have identified two new active hAT transposons, AeBuster1, from the mosquito Aedes aegypti and TcBuster from the red flour beetle Tribolium castaneum. Activity of both transposons is illustrated by excision and transposition assays performed in Drosophila melanogaster and Ae. aegypti and by in vitro strand transfer assays. These two active insect transposons are more closely related to the Buster sequences identified in humans than they are to the previously identified active hAT transposons, Ac, Tam3, Tol2, hobo, and Hermes. We therefore reexamined the structural and functional relationships of hAT and hAT-like transposase sequences extracted from genome databases and found that the hAT superfamily is divided into at least two families. This division is supported by a difference in target-site selections generated by active transposons of each family. We name these families the Ac and Buster families after the first identified transposon or transposon-like sequence in each. We find that the recently discovered SPIN transposons of mammals are located within the family of Buster elements. PMID:21368277

  4. Large-Scale Reform Comes of Age

    ERIC Educational Resources Information Center

    Fullan, Michael

    2009-01-01

    This article reviews the history of large-scale education reform and makes the case that large-scale or whole system reform policies and strategies are becoming increasingly evident. The review briefly addresses the pre 1997 period concluding that while the pressure for reform was mounting that there were very few examples of deliberate or…

  5. Automating large-scale reactor systems

    SciTech Connect

    Kisner, R.A.

    1985-01-01

    This paper conveys a philosophy for developing automated large-scale control systems that behave in an integrated, intelligent, flexible manner. Methods for operating large-scale systems under varying degrees of equipment degradation are discussed, and a design approach that separates the effort into phases is suggested. 5 refs., 1 fig.

  6. Remobilization of Sleeping Beauty transposons in the germline of Xenopus tropicalis

    PubMed Central

    2011-01-01

    Background The Sleeping Beauty (SB) transposon system has been used for germline transgenesis of the diploid frog, Xenopus tropicalis. Injecting one-cell embryos with plasmid DNA harboring an SB transposon substrate together with mRNA encoding the SB transposase enzyme resulted in non-canonical integration of small-order concatemers of the transposon. Here, we demonstrate that SB transposons stably integrated into the frog genome are effective substrates for remobilization. Results Transgenic frogs that express the SB10 transposase were bred with SB transposon-harboring animals to yield double-transgenic 'hopper' frogs. Remobilization events were observed in the progeny of the hopper frogs and were verified by Southern blot analysis and cloning of the novel integrations sites. Unlike the co-injection method used to generate founder lines, transgenic remobilization resulted in canonical transposition of the SB transposons. The remobilized SB transposons frequently integrated near the site of the donor locus; approximately 80% re-integrated with 3 Mb of the donor locus, a phenomenon known as 'local hopping'. Conclusions In this study, we demonstrate that SB transposons integrated into the X. tropicalis genome are effective substrates for excision and re-integration, and that the remobilized transposons are transmitted through the germline. This is an important step in the development of large-scale transposon-mediated gene- and enhancer-trap strategies in this highly tractable developmental model system. PMID:22115366

  7. Remobilization of Sleeping Beauty transposons in the germline of Xenopus tropicalis.

    PubMed

    Yergeau, Donald A; Kelley, Clair M; Kuliyev, Emin; Zhu, Haiqing; Johnson Hamlet, Michelle R; Sater, Amy K; Wells, Dan E; Mead, Paul E

    2011-11-24

    The Sleeping Beauty (SB) transposon system has been used for germline transgenesis of the diploid frog, Xenopus tropicalis. Injecting one-cell embryos with plasmid DNA harboring an SB transposon substrate together with mRNA encoding the SB transposase enzyme resulted in non-canonical integration of small-order concatemers of the transposon. Here, we demonstrate that SB transposons stably integrated into the frog genome are effective substrates for remobilization. Transgenic frogs that express the SB10 transposase were bred with SB transposon-harboring animals to yield double-transgenic 'hopper' frogs. Remobilization events were observed in the progeny of the hopper frogs and were verified by Southern blot analysis and cloning of the novel integrations sites. Unlike the co-injection method used to generate founder lines, transgenic remobilization resulted in canonical transposition of the SB transposons. The remobilized SB transposons frequently integrated near the site of the donor locus; approximately 80% re-integrated with 3 Mb of the donor locus, a phenomenon known as 'local hopping'. In this study, we demonstrate that SB transposons integrated into the X. tropicalis genome are effective substrates for excision and re-integration, and that the remobilized transposons are transmitted through the germline. This is an important step in the development of large-scale transposon-mediated gene- and enhancer-trap strategies in this highly tractable developmental model system.

  8. A large-scale complex haploinsufficiency-based genetic interaction screen in Candida albicans: analysis of the RAM network during morphogenesis.

    PubMed

    Bharucha, Nike; Chabrier-Rosello, Yeissa; Xu, Tao; Johnson, Cole; Sobczynski, Sarah; Song, Qingxuan; Dobry, Craig J; Eckwahl, Matthew J; Anderson, Christopher P; Benjamin, Andrew J; Kumar, Anju; Krysan, Damian J

    2011-04-01

    The morphogenetic transition between yeast and filamentous forms of the human fungal pathogen Candida albicans is regulated by a variety of signaling pathways. How these pathways interact to orchestrate morphogenesis, however, has not been as well characterized. To address this question and to identify genes that interact with the Regulation of Ace2 and Morphogenesis (RAM) pathway during filamentation, we report the first large-scale genetic interaction screen in C. albicans.Our strategy for this screen was based on the concept of complex haploinsufficiency (CHI). A heterozygous mutant of CBK1(cbk1Δ/CBK1), a key RAM pathway protein kinase, was subjected to transposon-mediated, insertional mutagenesis. The resulting double heterozygous mutants (6,528 independent strains) were screened for decreased filamentation on SpiderMedium (SM). From the 441 mutants showing altered filamentation, 139 transposon insertion sites were sequenced,yielding 41 unique CBK1-interacting genes. This gene set was enriched in transcriptional targets of Ace2 and, strikingly, the cAMP-dependent protein kinase A (PKA) pathway, suggesting an interaction between these two pathways. Further analysis indicates that the RAM and PKA pathways co-regulate a common set of genes during morphogenesis and that hyperactivation of the PKA pathway may compensate for loss of RAM pathway function. Our data also indicate that the PKA–regulated transcription factor Efg1 primarily localizes to yeast phase cells while the RAM–pathway regulated transcription factor Ace2 localizes to daughter nuclei of filamentous cells, suggesting that Efg1 and Ace2 regulate a common set of genes at separate stages of morphogenesis. Taken together, our observations indicate that CHI–based screening is a useful approach to genetic interaction analysis in C. albicans and support a model in which these two pathways regulate a common set of genes at different stages of filamentation.

  9. Chromosomal mobilization and reintegration of Sleeping Beauty and PiggyBac transposons.

    PubMed

    Liang, Qi; Kong, Jun; Stalker, James; Bradley, Allan

    2009-06-01

    The Sleeping Beauty and PiggyBac DNA transposon systems have recently been developed as tools for insertional mutagenesis. We have compared the chromosomal mobilization efficiency and insertion site preference of the two transposons mobilized from the same donor site in mouse embryonic stem (ES) cells under conditions in which there were no selective constraints on the transposons' insertion sites. Compared with Sleeping Beauty, PiggyBac exhibits higher transposition efficiencies, no evidence for local hopping and a significant bias toward reintegration in intragenic regions, which demonstrate its utility for insertional mutagenesis. Although Sleeping Beauty had no detectable genomic bias with respect to insertions in genes or intergenic regions, both Sleeping Beauty and PiggyBac transposons displayed preferential integration into actively transcribed loci. Copyright 2009 Wiley-Liss, Inc.

  10. Transgenesis in Xenopus using the Sleeping Beauty transposon system

    PubMed Central

    Yergeau, Donald A.; Johnson Hamlet, Michelle R.; Kuliyev, Emin; Zhu, Haiqing; Doherty, Joanne R.; Archer, Taylor D.; Subhawong, Andrea P.; Valentine, Marc B.; Kelley, Clair M.; Mead, Paul E.

    2010-01-01

    Transposon-based integration systems have been widely used for genetic manipulation of invertebrate and plant model systems. In the past decade, these powerful tools have begun to be used in vertebrates for transgenesis, insertional mutagenesis and gene therapy applications. Sleeping Beauty (SB) is a member of Tc1/mariner class of transposases and is derived from an inactive form of the gene isolated from Atlantic salmon. SB has been used extensively in human cell lines and in whole animal vertebrate model systems such as the mouse, rat and zebrafish. In this study, we describe the use of SB in the diploid frog Xenopus tropicalis to generate stable transgenic lines. SB transposon transgenes integrate into the X. tropicalis genome by a non-canonical process and are passed through the germline. We compare the activity of SB in this model organism with that of Tol2, a hAT (hobo, Ac1, TAM)-like transposon system. PMID:19517568

  11. Large Scale Metal Additive Techniques Review

    SciTech Connect

    Nycz, Andrzej; Adediran, Adeola I; Noakes, Mark W; Love, Lonnie J

    2016-01-01

    In recent years additive manufacturing made long strides toward becoming a main stream production technology. Particularly strong progress has been made in large-scale polymer deposition. However, large scale metal additive has not yet reached parity with large scale polymer. This paper is a review study of the metal additive techniques in the context of building large structures. Current commercial devices are capable of printing metal parts on the order of several cubic feet compared to hundreds of cubic feet for the polymer side. In order to follow the polymer progress path several factors are considered: potential to scale, economy, environment friendliness, material properties, feedstock availability, robustness of the process, quality and accuracy, potential for defects, and post processing as well as potential applications. This paper focuses on current state of art of large scale metal additive technology with a focus on expanding the geometric limits.

  12. Large-scale regions of antimatter

    SciTech Connect

    Grobov, A. V. Rubin, S. G.

    2015-07-15

    Amodified mechanism of the formation of large-scale antimatter regions is proposed. Antimatter appears owing to fluctuations of a complex scalar field that carries a baryon charge in the inflation era.

  13. The Large -scale Distribution of Galaxies

    NASA Astrophysics Data System (ADS)

    Flin, Piotr

    A review of the Large-scale structure of the Universe is given. A connection is made with the titanic work by Johannes Kepler in many areas of astronomy and cosmology. A special concern is made to spatial distribution of Galaxies, voids and walls (cellular structure of the Universe). Finaly, the author is concluding that the large scale structure of the Universe can be observed in much greater scale that it was thought twenty years ago.

  14. Multiple independent defective suppressor-mutator transposon insertions in Arabidopsis: a tool for functional genomics.

    PubMed Central

    Tissier, A F; Marillonnet, S; Klimyuk, V; Patel, K; Torres, M A; Murphy, G; Jones, J D

    1999-01-01

    A new system for insertional mutagenesis based on the maize Enhancer/Suppressor-mutator (En/Spm) element was introduced into Arabidopsis. A single T-DNA construct carried a nonautonomous defective Spm (dSpm) element with a phosphinothricin herbicide resistance (BAR) gene, a transposase expression cassette, and a counterselectable gene. This construct was used to select for stable dSpm transpositions. Treatments for both positive (BAR) and negative selection markers were applicable to soil-grown plants, allowing the recovery of new transpositions on a large scale. To date, a total of 48,000 lines in pools of 50 have been recovered, of which approximately 80% result from independent insertion events. DNA extracted from these pools was used in reverse genetic screens, either by polymerase chain reaction (PCR) using primers from the transposon and the targeted gene or by the display of insertions whereby inverse PCR products of insertions from the DNA pools are spotted on a membrane that is then hybridized with the probe of interest. By sequencing PCR-amplified fragments adjacent to insertion sites, we established a sequenced insertion-site database of 1200 sequences. This database permitted a comparison of the chromosomal distribution of transpositions from various T-DNA locations. PMID:10521516

  15. Development of hyperactive sleeping beauty transposon vectors by mutational analysis.

    PubMed

    Zayed, Hatem; Izsvák, Zsuzsanna; Walisko, Oliver; Ivics, Zoltán

    2004-02-01

    The Sleeping Beauty (SB) transposable element is a promising vector for transgenesis in vertebrates and is being developed as a novel, nonviral system for gene therapeutic purposes. A mutagenesis approach was undertaken to improve various aspects of the transposon, including safety and overall efficiency of gene transfer in human cells. Deletional analysis of transposon sequences within first-generation SB vectors showed that the inverted repeats of the element are necessary and sufficient to mediate high-efficiency transposition. We constructed a "sandwich" transposon, in which the DNA to be mobilized is flanked by two complete SB elements arranged in an inverted orientation. The sandwich element has superior ability to transpose >10-kb transgenes, thereby extending the cloning capacity of SB-based vectors. We derived hyperactive versions of the SB transposase by single-amino-acid substitutions. These mutations act synergistically and result in an almost fourfold enhancement of activity compared to the wild-type transposase. When combined with hyperactive transposons and transiently overexpressed HMGB1, a cellular cofactor of SB transposition, hyperactive transposases elevate transposition by almost an order of magnitude compared to the first-generation transposon system. The improved vector system should prove useful for efficient gene transfer in vertebrates.

  16. Large-scale cortical networks and cognition.

    PubMed

    Bressler, S L

    1995-03-01

    The well-known parcellation of the mammalian cerebral cortex into a large number of functionally distinct cytoarchitectonic areas presents a problem for understanding the complex cortical integrative functions that underlie cognition. How do cortical areas having unique individual functional properties cooperate to accomplish these complex operations? Do neurons distributed throughout the cerebral cortex act together in large-scale functional assemblages? This review examines the substantial body of evidence supporting the view that complex integrative functions are carried out by large-scale networks of cortical areas. Pathway tracing studies in non-human primates have revealed widely distributed networks of interconnected cortical areas, providing an anatomical substrate for large-scale parallel processing of information in the cerebral cortex. Functional coactivation of multiple cortical areas has been demonstrated by neurophysiological studies in non-human primates and several different cognitive functions have been shown to depend on multiple distributed areas by human neuropsychological studies. Electrophysiological studies on interareal synchronization have provided evidence that active neurons in different cortical areas may become not only coactive, but also functionally interdependent. The computational advantages of synchronization between cortical areas in large-scale networks have been elucidated by studies using artificial neural network models. Recent observations of time-varying multi-areal cortical synchronization suggest that the functional topology of a large-scale cortical network is dynamically reorganized during visuomotor behavior.

  17. Evolution of complex resistance transposons from an ancestral mercury transposon.

    PubMed

    Tanaka, M; Yamamoto, T; Sawai, T

    1983-03-01

    The molecular interrelationship of a transposon family which confers multiple antibiotic resistance and is assumed to have been generated from an ancestral mercury transposon was analyzed. Initially, the transposons Tn2613 (7.2 kilobases), encoding mercury resistance, and Tn2608 (13.5 kilobases), encoding mercury, streptomycin, and sulfonamide resistances, were isolated and their structures were analyzed. Next, the following transposons were compared with respect to their genetic and physical maps: Tn2613 and Tn501, encoding mercury resistance; Tn2608 and Tn21, encoding mercury, streptomycin, and sulfonamide resistance; Tn2607 and Tn4, encoding streptomycin, sulfonamide, and ampicillin resistance; and Tn2603, encoding mercury, streptomycin, sulfonamide, and ampicillin resistance. The results suggest that the transposons encoding multiple resistance were evolved from an ancestral mercury transposon.

  18. Survey on large scale system control methods

    NASA Technical Reports Server (NTRS)

    Mercadal, Mathieu

    1987-01-01

    The problem inherent to large scale systems such as power network, communication network and economic or ecological systems were studied. The increase in size and flexibility of future spacecraft has put those dynamical systems into the category of large scale systems, and tools specific to the class of large systems are being sought to design control systems that can guarantee more stability and better performance. Among several survey papers, reference was found to a thorough investigation on decentralized control methods. Especially helpful was the classification made of the different existing approaches to deal with large scale systems. A very similar classification is used, even though the papers surveyed are somehow different from the ones reviewed in other papers. Special attention is brought to the applicability of the existing methods to controlling large mechanical systems like large space structures. Some recent developments are added to this survey.

  19. Large-scale nanophotonic phased array.

    PubMed

    Sun, Jie; Timurdogan, Erman; Yaacobi, Ami; Hosseini, Ehsan Shah; Watts, Michael R

    2013-01-10

    Electromagnetic phased arrays at radio frequencies are well known and have enabled applications ranging from communications to radar, broadcasting and astronomy. The ability to generate arbitrary radiation patterns with large-scale phased arrays has long been pursued. Although it is extremely expensive and cumbersome to deploy large-scale radiofrequency phased arrays, optical phased arrays have a unique advantage in that the much shorter optical wavelength holds promise for large-scale integration. However, the short optical wavelength also imposes stringent requirements on fabrication. As a consequence, although optical phased arrays have been studied with various platforms and recently with chip-scale nanophotonics, all of the demonstrations so far are restricted to one-dimensional or small-scale two-dimensional arrays. Here we report the demonstration of a large-scale two-dimensional nanophotonic phased array (NPA), in which 64 × 64 (4,096) optical nanoantennas are densely integrated on a silicon chip within a footprint of 576 μm × 576 μm with all of the nanoantennas precisely balanced in power and aligned in phase to generate a designed, sophisticated radiation pattern in the far field. We also show that active phase tunability can be realized in the proposed NPA by demonstrating dynamic beam steering and shaping with an 8 × 8 array. This work demonstrates that a robust design, together with state-of-the-art complementary metal-oxide-semiconductor technology, allows large-scale NPAs to be implemented on compact and inexpensive nanophotonic chips. In turn, this enables arbitrary radiation pattern generation using NPAs and therefore extends the functionalities of phased arrays beyond conventional beam focusing and steering, opening up possibilities for large-scale deployment in applications such as communication, laser detection and ranging, three-dimensional holography and biomedical sciences, to name just a few.

  20. Probe mapping to facilitate transposon-based DNA sequencing

    SciTech Connect

    Strausbaugh, L.D.; Bourke, M.T.; Sommer, M.T.; Coon, M.E.; Berg, C.M. )

    1990-08-01

    A promising strategy for DNA sequencing exploits transposons to provide mobile sites for the binding of sequencing primers. For such a strategy to be maximally efficient, the location and orientation of the transposon must be readily determined and the insertion sites should be randomly distributed. The authors demonstrate an efficient probe-based method for the localization and orientation of transposon-borne primer sites, which is adaptable to large-scale sequencing strategies. This approach requires no prior restriction enzyme mapping or knowledge of the cloned sequence and eliminates the inefficiency inherent in totally random sequencing methods. To test the efficiency of probe mapping, 49 insertions of the transposon {gamma}{delta} (Tn1000) in a cloned fragment of Drosophila melanogaster DNA were mapped and oriented. In addition, oligonucleotide primers specific for unique subterminal {gamma}{delta} segments were used to prime dideoxynucleotide double-stranded sequencing. These data provided an opportunity to rigorously examine {gamma}{delta} insertion sites. The insertions were quire randomly distributed, even though the target DNA fragment had both A+T-rich and G+C-rich regions; in G+C-rich DNA, the insertions were found in A+T-rich valleys. These data demonstrate that {gamma}{delta} is an excellent choice for supplying mobile primer binding sites to cloned DNA and that transposon-based probe mapping permits the sequences of large cloned segments to be determined without any subcloning.

  1. The large-scale distribution of galaxies

    NASA Technical Reports Server (NTRS)

    Geller, Margaret J.

    1989-01-01

    The spatial distribution of galaxies in the universe is characterized on the basis of the six completed strips of the Harvard-Smithsonian Center for Astrophysics redshift-survey extension. The design of the survey is briefly reviewed, and the results are presented graphically. Vast low-density voids similar to the void in Bootes are found, almost completely surrounded by thin sheets of galaxies. Also discussed are the implications of the results for the survey sampling problem, the two-point correlation function of the galaxy distribution, the possibility of detecting large-scale coherent flows, theoretical models of large-scale structure, and the identification of groups and clusters of galaxies.

  2. Transposons for gene therapy!

    PubMed

    Ivics, Zoltán; Izsvák, Zsuzsanna

    2006-10-01

    Gene therapy is a promising strategy for the treatment of several inherited and acquired human diseases. Several vector platforms exist for the delivery of therapeutic nucleic acids into cells. Vectors based on viruses are very efficient at introducing gene constructs into cells, but their use has been associated with genotoxic effects of vector integration or immunological complications due to repeated administration in vivo. Non-viral vectors are easier to engineer and manufacture, but their efficient delivery into cells is a major challenge, and the lack of their chromosomal integration precludes long-term therapeutic effects. Transposable elements are non-viral gene delivery vehicles found ubiquitously in nature. Transposon-based vectors have the capacity of stable genomic integration and long-lasting expression of transgene constructs in cells. Molecular reconstruction of Sleeping Beauty, an ancient transposon in fish, represents a cornerstone in applying transposition-mediated gene delivery in vertebrate species, including humans. This review summarizes the state-of-the-art in the application of transposable elements for therapeutic gene transfer, and identifies key targets for the development of transposon-based gene vectors with enhanced efficacy and safety for human applications.

  3. Management of large-scale technology

    NASA Technical Reports Server (NTRS)

    Levine, A.

    1985-01-01

    Two major themes are addressed in this assessment of the management of large-scale NASA programs: (1) how a high technology agency was a decade marked by a rapid expansion of funds and manpower in the first half and almost as rapid contraction in the second; and (2) how NASA combined central planning and control with decentralized project execution.

  4. Large-scale multimedia modeling applications

    SciTech Connect

    Droppo, J.G. Jr.; Buck, J.W.; Whelan, G.; Strenge, D.L.; Castleton, K.J.; Gelston, G.M.

    1995-08-01

    Over the past decade, the US Department of Energy (DOE) and other agencies have faced increasing scrutiny for a wide range of environmental issues related to past and current practices. A number of large-scale applications have been undertaken that required analysis of large numbers of potential environmental issues over a wide range of environmental conditions and contaminants. Several of these applications, referred to here as large-scale applications, have addressed long-term public health risks using a holistic approach for assessing impacts from potential waterborne and airborne transport pathways. Multimedia models such as the Multimedia Environmental Pollutant Assessment System (MEPAS) were designed for use in such applications. MEPAS integrates radioactive and hazardous contaminants impact computations for major exposure routes via air, surface water, ground water, and overland flow transport. A number of large-scale applications of MEPAS have been conducted to assess various endpoints for environmental and human health impacts. These applications are described in terms of lessons learned in the development of an effective approach for large-scale applications.

  5. Evaluating Large-Scale Interactive Radio Programmes

    ERIC Educational Resources Information Center

    Potter, Charles; Naidoo, Gordon

    2009-01-01

    This article focuses on the challenges involved in conducting evaluations of interactive radio programmes in South Africa with large numbers of schools, teachers, and learners. It focuses on the role such large-scale evaluation has played during the South African radio learning programme's development stage, as well as during its subsequent…

  6. Evaluating Large-Scale Interactive Radio Programmes

    ERIC Educational Resources Information Center

    Potter, Charles; Naidoo, Gordon

    2009-01-01

    This article focuses on the challenges involved in conducting evaluations of interactive radio programmes in South Africa with large numbers of schools, teachers, and learners. It focuses on the role such large-scale evaluation has played during the South African radio learning programme's development stage, as well as during its subsequent…

  7. Dissection of Filamentous Growth by Transposon Mutagenesis in Saccharomyces Cerevisiae

    PubMed Central

    Mosch, H. U.; Fink, G. R.

    1997-01-01

    Diploid Saccharomyces cerevisiae strains starved for nitrogen undergo a developmental transition from growth as single yeast form (YF) cells to a multicellular form consisting of filaments of pseudohyphal (PH) cells. Filamentous growth is regulated by an evolutionarily conserved signaling pathway that includes the small GTP-binding proteins Ras2p and Cdc42p, the protein kinases Ste20p, Ste11p and Ste7p, and the transcription factor Ste12p. Here, we designed a genetic screen for mutant strains defective for filamentous growth (dfg) to identify novel targets of the filamentation signaling pathway, and we thereby identified 16 different genes, CDC39, STE12, TEC1, WHI3, NAB1, DBR1, CDC55, SRV2, TPM1, SPA2, BNI1, DFG5, DFG9, DFG10, BUD8 and DFG16, mutations that block filamentous growth. Phenotypic analysis of dfg mutant strains genetically dissects filamentous growth into the cellular processes of signal transduction, bud site selection, cell morphogenesis and invasive growth. Epistasis tests between dfg mutant alleles and dominant activated alleles of the RAS2 and STE11 genes, RAS2(Val19) and STE11-4, respectively, identify putative targets for the filamentation signaling pathway. Several of the genes described here have homologues in filamentous fungi, where they also regulate fungal development. PMID:9055077

  8. Condition Monitoring of Large-Scale Facilities

    NASA Technical Reports Server (NTRS)

    Hall, David L.

    1999-01-01

    This document provides a summary of the research conducted for the NASA Ames Research Center under grant NAG2-1182 (Condition-Based Monitoring of Large-Scale Facilities). The information includes copies of view graphs presented at NASA Ames in the final Workshop (held during December of 1998), as well as a copy of a technical report provided to the COTR (Dr. Anne Patterson-Hine) subsequent to the workshop. The material describes the experimental design, collection of data, and analysis results associated with monitoring the health of large-scale facilities. In addition to this material, a copy of the Pennsylvania State University Applied Research Laboratory data fusion visual programming tool kit was also provided to NASA Ames researchers.

  9. Large-scale Advanced Propfan (LAP) program

    NASA Technical Reports Server (NTRS)

    Sagerser, D. A.; Ludemann, S. G.

    1985-01-01

    The propfan is an advanced propeller concept which maintains the high efficiencies traditionally associated with conventional propellers at the higher aircraft cruise speeds associated with jet transports. The large-scale advanced propfan (LAP) program extends the research done on 2 ft diameter propfan models to a 9 ft diameter article. The program includes design, fabrication, and testing of both an eight bladed, 9 ft diameter propfan, designated SR-7L, and a 2 ft diameter aeroelastically scaled model, SR-7A. The LAP program is complemented by the propfan test assessment (PTA) program, which takes the large-scale propfan and mates it with a gas generator and gearbox to form a propfan propulsion system and then flight tests this system on the wing of a Gulfstream 2 testbed aircraft.

  10. Large-scale fibre-array multiplexing

    SciTech Connect

    Cheremiskin, I V; Chekhlova, T K

    2001-05-31

    The possibility of creating a fibre multiplexer/demultiplexer with large-scale multiplexing without any basic restrictions on the number of channels and the spectral spacing between them is shown. The operating capacity of a fibre multiplexer based on a four-fibre array ensuring a spectral spacing of 0.7 pm ({approx} 10 GHz) between channels is demonstrated. (laser applications and other topics in quantum electronics)

  11. Modeling Human Behavior at a Large Scale

    DTIC Science & Technology

    2012-01-01

    Discerning intentions in dynamic human action. Trends in Cognitive Sciences , 5(4):171 – 178, 2001. Shirli Bar-David, Israel Bar-David, Paul C. Cross, Sadie...Limits of predictability in human mobility. Science , 327(5968):1018, 2010. S.A. Stouffer. Intervening opportunities: a theory relating mobility and...Modeling Human Behavior at a Large Scale by Adam Sadilek Submitted in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy

  12. Large-Scale Aerosol Modeling and Analysis

    DTIC Science & Technology

    2008-09-30

    aerosol species up to six days in advance anywhere on the globe. NAAPS and COAMPS are particularly useful for forecasts of dust storms in areas...impact cloud processes globally. With increasing dust storms due to climate change and land use changes in desert regions, the impact of the...bacteria in large-scale dust storms is expected to significantly impact warm ice cloud formation, human health, and ecosystems globally. In Niemi et al

  13. Large-scale instabilities of helical flows

    NASA Astrophysics Data System (ADS)

    Cameron, Alexandre; Alexakis, Alexandros; Brachet, Marc-Étienne

    2016-10-01

    Large-scale hydrodynamic instabilities of periodic helical flows of a given wave number K are investigated using three-dimensional Floquet numerical computations. In the Floquet formalism the unstable field is expanded in modes of different spacial periodicity. This allows us (i) to clearly distinguish large from small scale instabilities and (ii) to study modes of wave number q of arbitrarily large-scale separation q ≪K . Different flows are examined including flows that exhibit small-scale turbulence. The growth rate σ of the most unstable mode is measured as a function of the scale separation q /K ≪1 and the Reynolds number Re. It is shown that the growth rate follows the scaling σ ∝q if an AKA effect [Frisch et al., Physica D: Nonlinear Phenomena 28, 382 (1987), 10.1016/0167-2789(87)90026-1] is present or a negative eddy viscosity scaling σ ∝q2 in its absence. This holds both for the Re≪1 regime where previously derived asymptotic results are verified but also for Re=O (1 ) that is beyond their range of validity. Furthermore, for values of Re above a critical value ReSc beyond which small-scale instabilities are present, the growth rate becomes independent of q and the energy of the perturbation at large scales decreases with scale separation. The nonlinear behavior of these large-scale instabilities is also examined in the nonlinear regime where the largest scales of the system are found to be the most dominant energetically. These results are interpreted by low-order models.

  14. Economically viable large-scale hydrogen liquefaction

    NASA Astrophysics Data System (ADS)

    Cardella, U.; Decker, L.; Klein, H.

    2017-02-01

    The liquid hydrogen demand, particularly driven by clean energy applications, will rise in the near future. As industrial large scale liquefiers will play a major role within the hydrogen supply chain, production capacity will have to increase by a multiple of today’s typical sizes. The main goal is to reduce the total cost of ownership for these plants by increasing energy efficiency with innovative and simple process designs, optimized in capital expenditure. New concepts must ensure a manageable plant complexity and flexible operability. In the phase of process development and selection, a dimensioning of key equipment for large scale liquefiers, such as turbines and compressors as well as heat exchangers, must be performed iteratively to ensure technological feasibility and maturity. Further critical aspects related to hydrogen liquefaction, e.g. fluid properties, ortho-para hydrogen conversion, and coldbox configuration, must be analysed in detail. This paper provides an overview on the approach, challenges and preliminary results in the development of efficient as well as economically viable concepts for large-scale hydrogen liquefaction.

  15. Large-Scale Visual Data Analysis

    NASA Astrophysics Data System (ADS)

    Johnson, Chris

    2014-04-01

    Modern high performance computers have speeds measured in petaflops and handle data set sizes measured in terabytes and petabytes. Although these machines offer enormous potential for solving very large-scale realistic computational problems, their effectiveness will hinge upon the ability of human experts to interact with their simulation results and extract useful information. One of the greatest scientific challenges of the 21st century is to effectively understand and make use of the vast amount of information being produced. Visual data analysis will be among our most most important tools in helping to understand such large-scale information. Our research at the Scientific Computing and Imaging (SCI) Institute at the University of Utah has focused on innovative, scalable techniques for large-scale 3D visual data analysis. In this talk, I will present state- of-the-art visualization techniques, including scalable visualization algorithms and software, cluster-based visualization methods and innovate visualization techniques applied to problems in computational science, engineering, and medicine. I will conclude with an outline for a future high performance visualization research challenges and opportunities.

  16. Large-scale neuromorphic computing systems

    NASA Astrophysics Data System (ADS)

    Furber, Steve

    2016-10-01

    Neuromorphic computing covers a diverse range of approaches to information processing all of which demonstrate some degree of neurobiological inspiration that differentiates them from mainstream conventional computing systems. The philosophy behind neuromorphic computing has its origins in the seminal work carried out by Carver Mead at Caltech in the late 1980s. This early work influenced others to carry developments forward, and advances in VLSI technology supported steady growth in the scale and capability of neuromorphic devices. Recently, a number of large-scale neuromorphic projects have emerged, taking the approach to unprecedented scales and capabilities. These large-scale projects are associated with major new funding initiatives for brain-related research, creating a sense that the time and circumstances are right for progress in our understanding of information processing in the brain. In this review we present a brief history of neuromorphic engineering then focus on some of the principal current large-scale projects, their main features, how their approaches are complementary and distinct, their advantages and drawbacks, and highlight the sorts of capabilities that each can deliver to neural modellers.

  17. Effective Gene Trapping Mediated by Sleeping Beauty Transposon

    PubMed Central

    Song, Guili; Li, Qing; Long, Yong; Gu, Qilin; Hackett, Perry B.; Cui, Zongbin

    2012-01-01

    Gene trapping is a high-throughput approach to elucidate gene functions by disrupting and recapitulating expression of genes in a target genome. A number of transposon-based gene-trapping systems are developed for mutagenesis in cells and model organisms, but there is still much room for the improvement of their efficiency in gene disruption and mutation. Herein, a gene-trapping system mediated by Sleeping Beauty (SB) transposon was developed by inclusion of three functional cassettes. The mutation cassette can abrogate the splice of trapped genes and terminate their translation. Once an endogenous gene is captured, the finding cassette independently drives the translation of reporter gene in HeLa cells and zebrafish embryos. The efficiency cassette controls the remobilization of integrated traps through inducible expression of SB gene. Analysis of transposon-genome junctions indicate that most of trap cassettes are integrated into an intron without an obvious 3′ bias. The transcription of trapped genes was abrogated by alternative splicing of the mutation cassette. In addition, integrated transposons can be induced to excise from their original insertion sites. Furthermore, the Cre/LoxP system was introduced to delete the efficiency cassette for stabilization of gene interruption and bio-safety. Thus, this gene-trap vector is an alternative and effective tool for the capture and disruption of endogenous genes in vitro and in vivo. PMID:22952894

  18. Experimental Simulations of Large-Scale Collisions

    NASA Technical Reports Server (NTRS)

    Housen, Kevin R.

    2002-01-01

    This report summarizes research on the effects of target porosity on the mechanics of impact cratering. Impact experiments conducted on a centrifuge provide direct simulations of large-scale cratering on porous asteroids. The experiments show that large craters in porous materials form mostly by compaction, with essentially no deposition of material into the ejecta blanket that is a signature of cratering in less-porous materials. The ratio of ejecta mass to crater mass is shown to decrease with increasing crater size or target porosity. These results are consistent with the observation that large closely-packed craters on asteroid Mathilde appear to have formed without degradation to earlier craters.

  19. What is a large-scale dynamo?

    NASA Astrophysics Data System (ADS)

    Nigro, G.; Pongkitiwanichakul, P.; Cattaneo, F.; Tobias, S. M.

    2017-01-01

    We consider kinematic dynamo action in a sheared helical flow at moderate to high values of the magnetic Reynolds number (Rm). We find exponentially growing solutions which, for large enough shear, take the form of a coherent part embedded in incoherent fluctuations. We argue that at large Rm large-scale dynamo action should be identified by the presence of structures coherent in time, rather than those at large spatial scales. We further argue that although the growth rate is determined by small-scale processes, the period of the coherent structures is set by mean-field considerations.

  20. Large-scale brightenings associated with flares

    NASA Technical Reports Server (NTRS)

    Mandrini, Cristina H.; Machado, Marcos E.

    1992-01-01

    It is shown that large-scale brightenings (LSBs) associated with solar flares, similar to the 'giant arches' discovered by Svestka et al. (1982) in images obtained by the SSM HXIS hours after the onset of two-ribbon flares, can also occur in association with confined flares in complex active regions. For these events, a clear link between the LSB and the underlying flare is clearly evident from the active-region magnetic field topology. The implications of these findings are discussed within the framework of the interacting loops of flares and the giant arch phenomenology.

  1. Large scale phononic metamaterials for seismic isolation

    SciTech Connect

    Aravantinos-Zafiris, N.; Sigalas, M. M.

    2015-08-14

    In this work, we numerically examine structures that could be characterized as large scale phononic metamaterials. These novel structures could have band gaps in the frequency spectrum of seismic waves when their dimensions are chosen appropriately, thus raising the belief that they could be serious candidates for seismic isolation structures. Different and easy to fabricate structures were examined made from construction materials such as concrete and steel. The well-known finite difference time domain method is used in our calculations in order to calculate the band structures of the proposed metamaterials.

  2. Large-scale planar lightwave circuits

    NASA Astrophysics Data System (ADS)

    Bidnyk, Serge; Zhang, Hua; Pearson, Matt; Balakrishnan, Ashok

    2011-01-01

    By leveraging advanced wafer processing and flip-chip bonding techniques, we have succeeded in hybrid integrating a myriad of active optical components, including photodetectors and laser diodes, with our planar lightwave circuit (PLC) platform. We have combined hybrid integration of active components with monolithic integration of other critical functions, such as diffraction gratings, on-chip mirrors, mode-converters, and thermo-optic elements. Further process development has led to the integration of polarization controlling functionality. Most recently, all these technological advancements have been combined to create large-scale planar lightwave circuits that comprise hundreds of optical elements integrated on chips less than a square inch in size.

  3. Large-Scale PV Integration Study

    SciTech Connect

    Lu, Shuai; Etingov, Pavel V.; Diao, Ruisheng; Ma, Jian; Samaan, Nader A.; Makarov, Yuri V.; Guo, Xinxin; Hafen, Ryan P.; Jin, Chunlian; Kirkham, Harold; Shlatz, Eugene; Frantzis, Lisa; McClive, Timothy; Karlson, Gregory; Acharya, Dhruv; Ellis, Abraham; Stein, Joshua; Hansen, Clifford; Chadliev, Vladimir; Smart, Michael; Salgo, Richard; Sorensen, Rahn; Allen, Barbara; Idelchik, Boris

    2011-07-29

    This research effort evaluates the impact of large-scale photovoltaic (PV) and distributed generation (DG) output on NV Energy’s electric grid system in southern Nevada. It analyzes the ability of NV Energy’s generation to accommodate increasing amounts of utility-scale PV and DG, and the resulting cost of integrating variable renewable resources. The study was jointly funded by the United States Department of Energy and NV Energy, and conducted by a project team comprised of industry experts and research scientists from Navigant Consulting Inc., Sandia National Laboratories, Pacific Northwest National Laboratory and NV Energy.

  4. Colloquium: Large scale simulations on GPU clusters

    NASA Astrophysics Data System (ADS)

    Bernaschi, Massimo; Bisson, Mauro; Fatica, Massimiliano

    2015-06-01

    Graphics processing units (GPU) are currently used as a cost-effective platform for computer simulations and big-data processing. Large scale applications require that multiple GPUs work together but the efficiency obtained with cluster of GPUs is, at times, sub-optimal because the GPU features are not exploited at their best. We describe how it is possible to achieve an excellent efficiency for applications in statistical mechanics, particle dynamics and networks analysis by using suitable memory access patterns and mechanisms like CUDA streams, profiling tools, etc. Similar concepts and techniques may be applied also to other problems like the solution of Partial Differential Equations.

  5. Neutrinos and large-scale structure

    SciTech Connect

    Eisenstein, Daniel J.

    2015-07-15

    I review the use of cosmological large-scale structure to measure properties of neutrinos and other relic populations of light relativistic particles. With experiments to measure the anisotropies of the cosmic microwave anisotropies and the clustering of matter at low redshift, we now have securely measured a relativistic background with density appropriate to the cosmic neutrino background. Our limits on the mass of the neutrino continue to shrink. Experiments coming in the next decade will greatly improve the available precision on searches for the energy density of novel relativistic backgrounds and the mass of neutrinos.

  6. Large-scale Heterogeneous Network Data Analysis

    DTIC Science & Technology

    2012-07-31

    Data for Multi-Player Influence Maximization on Social Networks.” KDD 2012 (Demo).  Po-Tzu Chang , Yen-Chieh Huang, Cheng-Lun Yang, Shou-De Lin, Pu...Jen Cheng. “Learning-Based Time-Sensitive Re-Ranking for Web Search.” SIGIR 2012 (poster)  Hung -Che Lai, Cheng-Te Li, Yi-Chen Lo, and Shou-De Lin...Exploiting and Evaluating MapReduce for Large-Scale Graph Mining.” ASONAM 2012 (Full, 16% acceptance ratio).  Hsun-Ping Hsieh , Cheng-Te Li, and Shou

  7. Internationalization Measures in Large Scale Research Projects

    NASA Astrophysics Data System (ADS)

    Soeding, Emanuel; Smith, Nancy

    2017-04-01

    Internationalization measures in Large Scale Research Projects Large scale research projects (LSRP) often serve as flagships used by universities or research institutions to demonstrate their performance and capability to stakeholders and other interested parties. As the global competition among universities for the recruitment of the brightest brains has increased, effective internationalization measures have become hot topics for universities and LSRP alike. Nevertheless, most projects and universities are challenged with little experience on how to conduct these measures and make internationalization an cost efficient and useful activity. Furthermore, those undertakings permanently have to be justified with the Project PIs as important, valuable tools to improve the capacity of the project and the research location. There are a variety of measures, suited to support universities in international recruitment. These include e.g. institutional partnerships, research marketing, a welcome culture, support for science mobility and an effective alumni strategy. These activities, although often conducted by different university entities, are interlocked and can be very powerful measures if interfaced in an effective way. On this poster we display a number of internationalization measures for various target groups, identify interfaces between project management, university administration, researchers and international partners to work together, exchange information and improve processes in order to be able to recruit, support and keep the brightest heads to your project.

  8. Local gravity and large-scale structure

    NASA Technical Reports Server (NTRS)

    Juszkiewicz, Roman; Vittorio, Nicola; Wyse, Rosemary F. G.

    1990-01-01

    The magnitude and direction of the observed dipole anisotropy of the galaxy distribution can in principle constrain the amount of large-scale power present in the spectrum of primordial density fluctuations. This paper confronts the data, provided by a recent redshift survey of galaxies detected by the IRAS satellite, with the predictions of two cosmological models with very different levels of large-scale power: the biased Cold Dark Matter dominated model (CDM) and a baryon-dominated model (BDM) with isocurvature initial conditions. Model predictions are investigated for the Local Group peculiar velocity, v(R), induced by mass inhomogeneities distributed out to a given radius, R, for R less than about 10,000 km/s. Several convergence measures for v(R) are developed, which can become powerful cosmological tests when deep enough samples become available. For the present data sets, the CDM and BDM predictions are indistinguishable at the 2 sigma level and both are consistent with observations. A promising discriminant between cosmological models is the misalignment angle between v(R) and the apex of the dipole anisotropy of the microwave background.

  9. Large-scale Intelligent Transporation Systems simulation

    SciTech Connect

    Ewing, T.; Canfield, T.; Hannebutte, U.; Levine, D.; Tentner, A.

    1995-06-01

    A prototype computer system has been developed which defines a high-level architecture for a large-scale, comprehensive, scalable simulation of an Intelligent Transportation System (ITS) capable of running on massively parallel computers and distributed (networked) computer systems. The prototype includes the modelling of instrumented ``smart`` vehicles with in-vehicle navigation units capable of optimal route planning and Traffic Management Centers (TMC). The TMC has probe vehicle tracking capabilities (display position and attributes of instrumented vehicles), and can provide 2-way interaction with traffic to provide advisories and link times. Both the in-vehicle navigation module and the TMC feature detailed graphical user interfaces to support human-factors studies. The prototype has been developed on a distributed system of networked UNIX computers but is designed to run on ANL`s IBM SP-X parallel computer system for large scale problems. A novel feature of our design is that vehicles will be represented by autonomus computer processes, each with a behavior model which performs independent route selection and reacts to external traffic events much like real vehicles. With this approach, one will be able to take advantage of emerging massively parallel processor (MPP) systems.

  10. Local gravity and large-scale structure

    NASA Technical Reports Server (NTRS)

    Juszkiewicz, Roman; Vittorio, Nicola; Wyse, Rosemary F. G.

    1990-01-01

    The magnitude and direction of the observed dipole anisotropy of the galaxy distribution can in principle constrain the amount of large-scale power present in the spectrum of primordial density fluctuations. This paper confronts the data, provided by a recent redshift survey of galaxies detected by the IRAS satellite, with the predictions of two cosmological models with very different levels of large-scale power: the biased Cold Dark Matter dominated model (CDM) and a baryon-dominated model (BDM) with isocurvature initial conditions. Model predictions are investigated for the Local Group peculiar velocity, v(R), induced by mass inhomogeneities distributed out to a given radius, R, for R less than about 10,000 km/s. Several convergence measures for v(R) are developed, which can become powerful cosmological tests when deep enough samples become available. For the present data sets, the CDM and BDM predictions are indistinguishable at the 2 sigma level and both are consistent with observations. A promising discriminant between cosmological models is the misalignment angle between v(R) and the apex of the dipole anisotropy of the microwave background.

  11. Large-scale Globally Propagating Coronal Waves.

    PubMed

    Warmuth, Alexander

    Large-scale, globally propagating wave-like disturbances have been observed in the solar chromosphere and by inference in the corona since the 1960s. However, detailed analysis of these phenomena has only been conducted since the late 1990s. This was prompted by the availability of high-cadence coronal imaging data from numerous spaced-based instruments, which routinely show spectacular globally propagating bright fronts. Coronal waves, as these perturbations are usually referred to, have now been observed in a wide range of spectral channels, yielding a wealth of information. Many findings have supported the "classical" interpretation of the disturbances: fast-mode MHD waves or shocks that are propagating in the solar corona. However, observations that seemed inconsistent with this picture have stimulated the development of alternative models in which "pseudo waves" are generated by magnetic reconfiguration in the framework of an expanding coronal mass ejection. This has resulted in a vigorous debate on the physical nature of these disturbances. This review focuses on demonstrating how the numerous observational findings of the last one and a half decades can be used to constrain our models of large-scale coronal waves, and how a coherent physical understanding of these disturbances is finally emerging.

  12. Xenopus transgenics: methods using transposons.

    PubMed

    Kelley, Clair M; Yergeau, Donald A; Zhu, Haiqing; Kuliyev, Emin; Mead, Paul E

    2012-01-01

    The generation of transgenic animals is an essential tool for many genetic strategies. DNA "cut-and-paste" transposon systems can be used to efficiently modify the Xenopus genome. The DNA transposon substrate, harbored on a circularized plasmid, is co-injected into fertilized Xenopus embryos at the one-cell stage together with mRNA encoding the cognate transposase enzyme. The cellular machinery rapidly translates the exogenous mRNA to produce active transposase enzyme that catalyzes excision of the transposon substrate from the plasmid and stable integration into the genomic DNA.

  13. Efficient, large scale separation of coal macerals

    SciTech Connect

    Dyrkacz, G.R.; Bloomquist, C.A.A.

    1988-01-01

    The authors believe that the separation of macerals by continuous flow centrifugation offers a simple technique for the large scale separation of macerals. With relatively little cost (/approximately/ $10K), it provides an opportunity for obtaining quite pure maceral fractions. Although they have not completely worked out all the nuances of this separation system, they believe that the problems they have indicated can be minimized to pose only minor inconvenience. It cannot be said that this system completely bypasses the disagreeable tedium or time involved in separating macerals, nor will it by itself overcome the mental inertia required to make maceral separation an accepted necessary fact in fundamental coal science. However, they find their particular brand of continuous flow centrifugation is considerably faster than sink/float separation, can provide a good quality product with even one separation cycle, and permits the handling of more material than a conventional sink/float centrifuge separation.

  14. Primer design for large scale sequencing.

    PubMed Central

    Haas, S; Vingron, M; Poustka, A; Wiemann, S

    1998-01-01

    We have developed PRIDE, a primer design program that automatically designs primers in single contigs or whole sequencing projects to extend the already known sequence and to double strand single-stranded regions. The program is fully integrated into the Staden package (GAP4) and accessible with a graphical user interface. PRIDE uses a fuzzy logic-based system to calculate primer qualities. The computational performance of PRIDE is enhanced by using suffix trees to store the huge amount of data being produced. A test set of 110 sequencing primers and 11 PCR primer pairs has been designed on genomic templates, cDNAs and sequences containing repetitive elements to analyze PRIDE's success rate. The high performance of PRIDE, combined with its minimal requirement of user interaction and its fast algorithm, make this program useful for the large scale design of primers, especially in large sequencing projects. PMID:9611248

  15. Grid sensitivity capability for large scale structures

    NASA Technical Reports Server (NTRS)

    Nagendra, Gopal K.; Wallerstein, David V.

    1989-01-01

    The considerations and the resultant approach used to implement design sensitivity capability for grids into a large scale, general purpose finite element system (MSC/NASTRAN) are presented. The design variables are grid perturbations with a rather general linking capability. Moreover, shape and sizing variables may be linked together. The design is general enough to facilitate geometric modeling techniques for generating design variable linking schemes in an easy and straightforward manner. Test cases have been run and validated by comparison with the overall finite difference method. The linking of a design sensitivity capability for shape variables in MSC/NASTRAN with an optimizer would give designers a powerful, automated tool to carry out practical optimization design of real life, complicated structures.

  16. Large-Scale Organization of Glycosylation Networks

    NASA Astrophysics Data System (ADS)

    Kim, Pan-Jun; Lee, Dong-Yup; Jeong, Hawoong

    2009-03-01

    Glycosylation is a highly complex process to produce a diverse repertoire of cellular glycans that are frequently attached to proteins and lipids. Glycans participate in fundamental biological processes including molecular trafficking and clearance, cell proliferation and apoptosis, developmental biology, immune response, and pathogenesis. N-linked glycans found on proteins are formed by sequential attachments of monosaccharides with the help of a relatively small number of enzymes. Many of these enzymes can accept multiple N-linked glycans as substrates, thus generating a large number of glycan intermediates and their intermingled pathways. Motivated by the quantitative methods developed in complex network research, we investigate the large-scale organization of such N-glycosylation pathways in a mammalian cell. The uncovered results give the experimentally-testable predictions for glycosylation process, and can be applied to the engineering of therapeutic glycoproteins.

  17. Large-scale optimization of neuron arbors

    NASA Astrophysics Data System (ADS)

    Cherniak, Christopher; Changizi, Mark; Won Kang, Du

    1999-05-01

    At the global as well as local scales, some of the geometry of types of neuron arbors-both dendrites and axons-appears to be self-organizing: Their morphogenesis behaves like flowing water, that is, fluid dynamically; waterflow in branching networks in turn acts like a tree composed of cords under tension, that is, vector mechanically. Branch diameters and angles and junction sites conform significantly to this model. The result is that such neuron tree samples globally minimize their total volume-rather than, for example, surface area or branch length. In addition, the arbors perform well at generating the cheapest topology interconnecting their terminals: their large-scale layouts are among the best of all such possible connecting patterns, approaching 5% of optimum. This model also applies comparably to arterial and river networks.

  18. Engineering management of large scale systems

    NASA Technical Reports Server (NTRS)

    Sanders, Serita; Gill, Tepper L.; Paul, Arthur S.

    1989-01-01

    The organization of high technology and engineering problem solving, has given rise to an emerging concept. Reasoning principles for integrating traditional engineering problem solving with system theory, management sciences, behavioral decision theory, and planning and design approaches can be incorporated into a methodological approach to solving problems with a long range perspective. Long range planning has a great potential to improve productivity by using a systematic and organized approach. Thus, efficiency and cost effectiveness are the driving forces in promoting the organization of engineering problems. Aspects of systems engineering that provide an understanding of management of large scale systems are broadly covered here. Due to the focus and application of research, other significant factors (e.g., human behavior, decision making, etc.) are not emphasized but are considered.

  19. Large scale cryogenic fluid systems testing

    NASA Technical Reports Server (NTRS)

    1992-01-01

    NASA Lewis Research Center's Cryogenic Fluid Systems Branch (CFSB) within the Space Propulsion Technology Division (SPTD) has the ultimate goal of enabling the long term storage and in-space fueling/resupply operations for spacecraft and reusable vehicles in support of space exploration. Using analytical modeling, ground based testing, and on-orbit experimentation, the CFSB is studying three primary categories of fluid technology: storage, supply, and transfer. The CFSB is also investigating fluid handling, advanced instrumentation, and tank structures and materials. Ground based testing of large-scale systems is done using liquid hydrogen as a test fluid at the Cryogenic Propellant Tank Facility (K-site) at Lewis' Plum Brook Station in Sandusky, Ohio. A general overview of tests involving liquid transfer, thermal control, pressure control, and pressurization is given.

  20. Large scale preparation of pure phycobiliproteins.

    PubMed

    Padgett, M P; Krogmann, D W

    1987-01-01

    This paper describes simple procedures for the purification of large amounts of phycocyanin and allophycocyanin from the cyanobacterium Microcystis aeruginosa. A homogeneous natural bloom of this organism provided hundreds of kilograms of cells. Large samples of cells were broken by freezing and thawing. Repeated extraction of the broken cells with distilled water released phycocyanin first, then allophycocyanin, and provides supporting evidence for the current models of phycobilisome structure. The very low ionic strength of the aqueous extracts allowed allophycocyanin release in a particulate form so that this protein could be easily concentrated by centrifugation. Other proteins in the extract were enriched and concentrated by large scale membrane filtration. The biliproteins were purified to homogeneity by chromatography on DEAE cellulose. Purity was established by HPLC and by N-terminal amino acid sequence analysis. The proteins were examined for stability at various pHs and exposures to visible light.

  1. Primer design for large scale sequencing.

    PubMed

    Haas, S; Vingron, M; Poustka, A; Wiemann, S

    1998-06-15

    We have developed PRIDE, a primer design program that automatically designs primers in single contigs or whole sequencing projects to extend the already known sequence and to double strand single-stranded regions. The program is fully integrated into the Staden package (GAP4) and accessible with a graphical user interface. PRIDE uses a fuzzy logic-based system to calculate primer qualities. The computational performance of PRIDE is enhanced by using suffix trees to store the huge amount of data being produced. A test set of 110 sequencing primers and 11 PCR primer pairs has been designed on genomic templates, cDNAs and sequences containing repetitive elements to analyze PRIDE's success rate. The high performance of PRIDE, combined with its minimal requirement of user interaction and its fast algorithm, make this program useful for the large scale design of primers, especially in large sequencing projects.

  2. Large-scale synthesis of peptides.

    PubMed

    Andersson, L; Blomberg, L; Flegel, M; Lepsa, L; Nilsson, B; Verlander, M

    2000-01-01

    Recent advances in the areas of formulation and delivery have rekindled the interest of the pharmaceutical community in peptides as drug candidates, which, in turn, has provided a challenge to the peptide industry to develop efficient methods for the manufacture of relatively complex peptides on scales of up to metric tons per year. This article focuses on chemical synthesis approaches for peptides, and presents an overview of the methods available and in use currently, together with a discussion of scale-up strategies. Examples of the different methods are discussed, together with solutions to some specific problems encountered during scale-up development. Finally, an overview is presented of issues common to all manufacturing methods, i.e., methods used for the large-scale purification and isolation of final bulk products and regulatory considerations to be addressed during scale-up of processes to commercial levels. Copyright 2000 John Wiley & Sons, Inc. Biopolymers (Pept Sci) 55: 227-250, 2000

  3. Large Scale Quantum Simulations of Nuclear Pasta

    NASA Astrophysics Data System (ADS)

    Fattoyev, Farrukh J.; Horowitz, Charles J.; Schuetrumpf, Bastian

    2016-03-01

    Complex and exotic nuclear geometries collectively referred to as ``nuclear pasta'' are expected to naturally exist in the crust of neutron stars and in supernovae matter. Using a set of self-consistent microscopic nuclear energy density functionals we present the first results of large scale quantum simulations of pasta phases at baryon densities 0 . 03 < ρ < 0 . 10 fm-3, proton fractions 0 . 05

  4. Jovian large-scale stratospheric circulation

    NASA Technical Reports Server (NTRS)

    West, R. A.; Friedson, A. J.; Appleby, J. F.

    1992-01-01

    An attempt is made to diagnose the annual-average mean meridional residual Jovian large-scale stratospheric circulation from observations of the temperature and reflected sunlight that reveal the morphology of the aerosol heating. The annual mean solar heating, total radiative flux divergence, mass stream function, and Eliassen-Palm flux divergence are shown. The stratospheric radiative flux divergence is dominated the high latitudes by aerosol absorption. Between the 270 and 100 mbar pressure levels, where there is no aerosol heating in the model, the structure of the circulation at low- to midlatitudes is governed by the meridional variation of infrared cooling in association with the variation of zonal mean temperatures observed by IRIS. The principal features of the vertical velocity profile found by Gierasch et al. (1986) are recovered in the present calculation.

  5. Large-scale parametric survival analysis.

    PubMed

    Mittal, Sushil; Madigan, David; Cheng, Jerry Q; Burd, Randall S

    2013-10-15

    Survival analysis has been a topic of active statistical research in the past few decades with applications spread across several areas. Traditional applications usually consider data with only a small numbers of predictors with a few hundreds or thousands of observations. Recent advances in data acquisition techniques and computation power have led to considerable interest in analyzing very-high-dimensional data where the number of predictor variables and the number of observations range between 10(4) and 10(6). In this paper, we present a tool for performing large-scale regularized parametric survival analysis using a variant of the cyclic coordinate descent method. Through our experiments on two real data sets, we show that application of regularized models to high-dimensional data avoids overfitting and can provide improved predictive performance and calibration over corresponding low-dimensional models.

  6. Large-Scale Parametric Survival Analysis†

    PubMed Central

    Mittal, Sushil; Madigan, David; Cheng, Jerry; Burd, Randall S.

    2013-01-01

    Survival analysis has been a topic of active statistical research in the past few decades with applications spread across several areas. Traditional applications usually consider data with only small numbers of predictors with a few hundreds or thousands of observations. Recent advances in data acquisition techniques and computation power has led to considerable interest in analyzing very high-dimensional data where the number of predictor variables and the number of observations range between 104 – 106. In this paper, we present a tool for performing large-scale regularized parametric survival analysis using a variant of cyclic coordinate descent method. Through our experiments on two real data sets, we show that application of regularized models to high-dimensional data avoids overfitting and can provide improved predictive performance and calibration over corresponding low-dimensional models. PMID:23625862

  7. Large scale study of tooth enamel

    SciTech Connect

    Bodart, F.; Deconninck, G.; Martin, M.Th.

    1981-04-01

    Human tooth enamel contains traces of foreign elements. The presence of these elements is related to the history and the environment of the human body and can be considered as the signature of perturbations which occur during the growth of a tooth. A map of the distribution of these traces on a large scale sample of the population will constitute a reference for further investigations of environmental effects. One hundred eighty samples of teeth were first analysed using PIXE, backscattering and nuclear reaction techniques. The results were analysed using statistical methods. Correlations between O, F, Na, P, Ca, Mn, Fe, Cu, Zn, Pb and Sr were observed and cluster analysis was in progress. The techniques described in the present work have been developed in order to establish a method for the exploration of very large samples of the Belgian population.

  8. The challenge of large-scale structure

    NASA Astrophysics Data System (ADS)

    Gregory, S. A.

    1996-03-01

    The tasks that I have assumed for myself in this presentation include three separate parts. The first, appropriate to the particular setting of this meeting, is to review the basic work of the founding of this field; the appropriateness comes from the fact that W. G. Tifft made immense contributions that are not often realized by the astronomical community. The second task is to outline the general tone of the observational evidence for large scale structures. (Here, in particular, I cannot claim to be complete. I beg forgiveness from any workers who are left out by my oversight for lack of space and time.) The third task is to point out some of the major aspects of the field that may represent the clues by which some brilliant sleuth will ultimately figure out how galaxies formed.

  9. Modeling the Internet's large-scale topology

    PubMed Central

    Yook, Soon-Hyung; Jeong, Hawoong; Barabási, Albert-László

    2002-01-01

    Network generators that capture the Internet's large-scale topology are crucial for the development of efficient routing protocols and modeling Internet traffic. Our ability to design realistic generators is limited by the incomplete understanding of the fundamental driving forces that affect the Internet's evolution. By combining several independent databases capturing the time evolution, topology, and physical layout of the Internet, we identify the universal mechanisms that shape the Internet's router and autonomous system level topology. We find that the physical layout of nodes form a fractal set, determined by population density patterns around the globe. The placement of links is driven by competition between preferential attachment and linear distance dependence, a marked departure from the currently used exponential laws. The universal parameters that we extract significantly restrict the class of potentially correct Internet models and indicate that the networks created by all available topology generators are fundamentally different from the current Internet. PMID:12368484

  10. Large-scale sequential quadratic programming algorithms

    SciTech Connect

    Eldersveld, S.K.

    1992-09-01

    The problem addressed is the general nonlinear programming problem: finding a local minimizer for a nonlinear function subject to a mixture of nonlinear equality and inequality constraints. The methods studied are in the class of sequential quadratic programming (SQP) algorithms, which have previously proved successful for problems of moderate size. Our goal is to devise an SQP algorithm that is applicable to large-scale optimization problems, using sparse data structures and storing less curvature information but maintaining the property of superlinear convergence. The main features are: 1. The use of a quasi-Newton approximation to the reduced Hessian of the Lagrangian function. Only an estimate of the reduced Hessian matrix is required by our algorithm. The impact of not having available the full Hessian approximation is studied and alternative estimates are constructed. 2. The use of a transformation matrix Q. This allows the QP gradient to be computed easily when only the reduced Hessian approximation is maintained. 3. The use of a reduced-gradient form of the basis for the null space of the working set. This choice of basis is more practical than an orthogonal null-space basis for large-scale problems. The continuity condition for this choice is proven. 4. The use of incomplete solutions of quadratic programming subproblems. Certain iterates generated by an active-set method for the QP subproblem are used in place of the QP minimizer to define the search direction for the nonlinear problem. An implementation of the new algorithm has been obtained by modifying the code MINOS. Results and comparisons with MINOS and NPSOL are given for the new algorithm on a set of 92 test problems.

  11. Supporting large-scale computational science

    SciTech Connect

    Musick, R., LLNL

    1998-02-19

    Business needs have driven the development of commercial database systems since their inception. As a result, there has been a strong focus on supporting many users, minimizing the potential corruption or loss of data, and maximizing performance metrics like transactions per second, or TPC-C and TPC-D results. It turns out that these optimizations have little to do with the needs of the scientific community, and in particular have little impact on improving the management and use of large-scale high-dimensional data. At the same time, there is an unanswered need in the scientific community for many of the benefits offered by a robust DBMS. For example, tying an ad-hoc query language such as SQL together with a visualization toolkit would be a powerful enhancement to current capabilities. Unfortunately, there has been little emphasis or discussion in the VLDB community on this mismatch over the last decade. The goal of the paper is to identify the specific issues that need to be resolved before large-scale scientific applications can make use of DBMS products. This topic is addressed in the context of an evaluation of commercial DBMS technology applied to the exploration of data generated by the Department of Energy`s Accelerated Strategic Computing Initiative (ASCI). The paper describes the data being generated for ASCI as well as current capabilities for interacting with and exploring this data. The attraction of applying standard DBMS technology to this domain is discussed, as well as the technical and business issues that currently make this an infeasible solution.

  12. Improving Recent Large-Scale Pulsar Surveys

    NASA Astrophysics Data System (ADS)

    Cardoso, Rogerio Fernando; Ransom, S.

    2011-01-01

    Pulsars are unique in that they act as celestial laboratories for precise tests of gravity and other extreme physics (Kramer 2004). There are approximately 2000 known pulsars today, which is less than ten percent of pulsars in the Milky Way according to theoretical models (Lorimer 2004). Out of these 2000 known pulsars, approximately ten percent are known millisecond pulsars, objects used for their period stability for detailed physics tests and searches for gravitational radiation (Lorimer 2008). As the field and instrumentation progress, pulsar astronomers attempt to overcome observational biases and detect new pulsars, consequently discovering new millisecond pulsars. We attempt to improve large scale pulsar surveys by examining three recent pulsar surveys. The first, the Green Bank Telescope 350MHz Drift Scan, a low frequency isotropic survey of the northern sky, has yielded a large number of candidates that were visually inspected and identified, resulting in over 34.000 thousands candidates viewed, dozens of detections of known pulsars, and the discovery of a new low-flux pulsar, PSRJ1911+22. The second, the PALFA survey, is a high frequency survey of the galactic plane with the Arecibo telescope. We created a processing pipeline for the PALFA survey at the National Radio Astronomy Observatory in Charlottesville- VA, in addition to making needed modifications upon advice from the PALFA consortium. The third survey examined is a new GBT 820MHz survey devoted to find new millisecond pulsars by observing the target-rich environment of unidentified sources in the FERMI LAT catalogue. By approaching these three pulsar surveys at different stages, we seek to improve the success rates of large scale surveys, and hence the possibility for ground-breaking work in both basic physics and astrophysics.

  13. Introducing Large-Scale Innovation in Schools

    NASA Astrophysics Data System (ADS)

    Sotiriou, Sofoklis; Riviou, Katherina; Cherouvis, Stephanos; Chelioti, Eleni; Bogner, Franz X.

    2016-08-01

    Education reform initiatives tend to promise higher effectiveness in classrooms especially when emphasis is given to e-learning and digital resources. Practical changes in classroom realities or school organization, however, are lacking. A major European initiative entitled Open Discovery Space (ODS) examined the challenge of modernizing school education via a large-scale implementation of an open-scale methodology in using technology-supported innovation. The present paper describes this innovation scheme which involved schools and teachers all over Europe, embedded technology-enhanced learning into wider school environments and provided training to teachers. Our implementation scheme consisted of three phases: (1) stimulating interest, (2) incorporating the innovation into school settings and (3) accelerating the implementation of the innovation. The scheme's impact was monitored for a school year using five indicators: leadership and vision building, ICT in the curriculum, development of ICT culture, professional development support, and school resources and infrastructure. Based on about 400 schools, our study produced four results: (1) The growth in digital maturity was substantial, even for previously high scoring schools. This was even more important for indicators such as vision and leadership" and "professional development." (2) The evolution of networking is presented graphically, showing the gradual growth of connections achieved. (3) These communities became core nodes, involving numerous teachers in sharing educational content and experiences: One out of three registered users (36 %) has shared his/her educational resources in at least one community. (4) Satisfaction scores ranged from 76 % (offer of useful support through teacher academies) to 87 % (good environment to exchange best practices). Initiatives such as ODS add substantial value to schools on a large scale.

  14. Supporting large-scale computational science

    SciTech Connect

    Musick, R

    1998-10-01

    A study has been carried out to determine the feasibility of using commercial database management systems (DBMSs) to support large-scale computational science. Conventional wisdom in the past has been that DBMSs are too slow for such data. Several events over the past few years have muddied the clarity of this mindset: 1. 2. 3. 4. Several commercial DBMS systems have demonstrated storage and ad-hoc quer access to Terabyte data sets. Several large-scale science teams, such as EOSDIS [NAS91], high energy physics [MM97] and human genome [Kin93] have adopted (or make frequent use of) commercial DBMS systems as the central part of their data management scheme. Several major DBMS vendors have introduced their first object-relational products (ORDBMSs), which have the potential to support large, array-oriented data. In some cases, performance is a moot issue. This is true in particular if the performance of legacy applications is not reduced while new, albeit slow, capabilities are added to the system. The basic assessment is still that DBMSs do not scale to large computational data. However, many of the reasons have changed, and there is an expiration date attached to that prognosis. This document expands on this conclusion, identifies the advantages and disadvantages of various commercial approaches, and describes the studies carried out in exploring this area. The document is meant to be brief, technical and informative, rather than a motivational pitch. The conclusions within are very likely to become outdated within the next 5-7 years, as market forces will have a significant impact on the state of the art in scientific data management over the next decade.

  15. Voids in the Large-Scale Structure

    NASA Astrophysics Data System (ADS)

    El-Ad, Hagai; Piran, Tsvi

    1997-12-01

    Voids are the most prominent feature of the large-scale structure of the universe. Still, their incorporation into quantitative analysis of it has been relatively recent, owing essentially to the lack of an objective tool to identify the voids and to quantify them. To overcome this, we present here the VOID FINDER algorithm, a novel tool for objectively quantifying voids in the galaxy distribution. The algorithm first classifies galaxies as either wall galaxies or field galaxies. Then, it identifies voids in the wall-galaxy distribution. Voids are defined as continuous volumes that do not contain any wall galaxies. The voids must be thicker than an adjustable limit, which is refined in successive iterations. In this way, we identify the same regions that would be recognized as voids by the eye. Small breaches in the walls are ignored, avoiding artificial connections between neighboring voids. We test the algorithm using Voronoi tesselations. By appropriate scaling of the parameters with the selection function, we apply it to two redshift surveys, the dense SSRS2 and the full-sky IRAS 1.2 Jy. Both surveys show similar properties: ~50% of the volume is filled by voids. The voids have a scale of at least 40 h-1 Mpc and an average -0.9 underdensity. Faint galaxies do not fill the voids, but they do populate them more than bright ones. These results suggest that both optically and IRAS-selected galaxies delineate the same large-scale structure. Comparison with the recovered mass distribution further suggests that the observed voids in the galaxy distribution correspond well to underdense regions in the mass distribution. This confirms the gravitational origin of the voids.

  16. Defining essential genes and identifying virulence factors of Porphyromonas gingivalis by massively-parallel sequencing of transposon libraries (Tn-seq)

    PubMed Central

    Klein, Brian A.; Duncan, Margaret J.; Hu, Linden T.

    2016-01-01

    Summary Porphyromonas gingivalis is a keystone pathogen in the development and progression of periodontal disease. Obstacles to the development of saturated transposon libraries have previously limited transposon mutant-based screens as well as essential gene studies. We have developed a system for efficient transposon mutagenesis of P. gingivalis using a modified mariner transposon. Tn-seq is a technique that allows for quantitative assessment of individual mutants within a transposon mutant library by sequencing the transposon-genome junctions and then compiling mutant presence by mapping to a base genome. Using Tn-seq, it is possible to quickly define all the insertional mutants in a library and thus identify non-essential genes under the conditions in which the library was produced. Identification of fitness of individual mutants under specific conditions can be performed by exposing the library to selective pressures. PMID:25636611

  17. Transposon tagging of disease resistance genes

    SciTech Connect

    Michelmore, R.W. . Dept. of Physics)

    1989-01-01

    We are developing a transposon mutagenesis system for lettuce to clone genes for resistance to the fungal pathogen, Bremia lactucae. Activity of heterologous transposons is being studied in transgenic plants. Southern analysis of T{sub 1} and T{sub 2} plants containing Tam3 from Antirrhinum provided ambiguous results. Multiple endonuclease digests indicated that transposition had occurred; however, in no plant were all endonuclease digests consistent with a simple excision event. Southern or PCR analysis of over 50 plans containing Ac from maize have also failed to reveal clear evidence of transposition; this is contrast to experiments by others with the same constructs who have observed high rates of Ac excision in other plant species. Nearly all of 65 T{sub 2} families containing Ac interrupting a chimeric streptomycin resistance gene (Courtesy J. Jones, Sainsbury Lab., UK) clearly segregated for streptomycin resistance. Southern analyses, however, showed no evidence of transposition, indicating restoration of a functional message by other mechanisms, possibly mRNA processing. Transgenic plants have also been generated containing CaMV 35S or hsp70 promoters fused to transposase coding sequences or a Ds element interrupting a chimeric GUS gene (Courtesy M. Lassner, UC Davis). F{sub 1} plants containing both constructs were analyzed for transposition. Only two plants containing both constructs were obtained from 48 progeny, far fewer than expected, and neither showed evidence of transposition in Southerns and GUS assays. We are currently constructing further chimeric transposase fusions. To test for the stability of the targeted disease resistance genes, 50,000 F{sub 1} plants heterozygous for three resistance genes were generated; no mutants have been identified in the 5000 so far screened.

  18. Identification of virulence determinants for endocarditis in Streptococcus sanguinis by signature-tagged mutagenesis.

    PubMed

    Paik, Sehmi; Senty, Lauren; Das, Sankar; Noe, Jody C; Munro, Cindy L; Kitten, Todd

    2005-09-01

    Streptococcus sanguinis is a gram-positive, facultative anaerobe and a normal inhabitant of the human oral cavity. It is also one of the most common agents of infective endocarditis, a serious endovascular infection. To identify virulence factors for infective endocarditis, signature-tagged mutagenesis (STM) was applied to the SK36 strain of S. sanguinis, whose genome is being sequenced. STM allows the large-scale creation, in vivo screening, and recovery of a series of mutants with altered virulence. Screening of 800 mutants by STM identified 38 putative avirulent and 5 putative hypervirulent mutants. Subsequent molecular analysis of a subset of these mutants identified genes encoding undecaprenol kinase, homoserine kinase, anaerobic ribonucleotide reductase, adenylosuccinate lyase, and a hypothetical protein. Virulence reductions ranging from 2-to 150-fold were confirmed by competitive index assays. One putatively hypervirulent strain with a transposon insertion in an intergenic region was identified, though increased virulence was not confirmed in competitive index assays. All mutants grew comparably to SK36 in aerobic broth culture except for the homoserine kinase mutant. Growth of this mutant was restored by the addition of threonine to the medium. Mutants containing an insertion or in-frame deletion in the anaerobic ribonucleotide reductase gene failed to grow under strictly anaerobic conditions. The results suggest that housekeeping functions such as cell wall synthesis, amino acid and nucleic acid synthesis, and the ability to survive under anaerobic conditions are important virulence factors in S. sanguinis endocarditis.

  19. Statistical Measures of Large-Scale Structure

    NASA Astrophysics Data System (ADS)

    Vogeley, Michael; Geller, Margaret; Huchra, John; Park, Changbom; Gott, J. Richard

    1993-12-01

    \\inv Mpc} To quantify clustering in the large-scale distribution of galaxies and to test theories for the formation of structure in the universe, we apply statistical measures to the CfA Redshift Survey. This survey is complete to m_{B(0)}=15.5 over two contiguous regions which cover one-quarter of the sky and include ~ 11,000 galaxies. The salient features of these data are voids with diameter 30-50\\hmpc and coherent dense structures with a scale ~ 100\\hmpc. Comparison with N-body simulations rules out the ``standard" CDM model (Omega =1, b=1.5, sigma_8 =1) at the 99% confidence level because this model has insufficient power on scales lambda >30\\hmpc. An unbiased open universe CDM model (Omega h =0.2) and a biased CDM model with non-zero cosmological constant (Omega h =0.24, lambda_0 =0.6) match the observed power spectrum. The amplitude of the power spectrum depends on the luminosity of galaxies in the sample; bright (L>L(*) ) galaxies are more strongly clustered than faint galaxies. The paucity of bright galaxies in low-density regions may explain this dependence. To measure the topology of large-scale structure, we compute the genus of isodensity surfaces of the smoothed density field. On scales in the ``non-linear" regime, <= 10\\hmpc, the high- and low-density regions are multiply-connected over a broad range of density threshold, as in a filamentary net. On smoothing scales >10\\hmpc, the topology is consistent with statistics of a Gaussian random field. Simulations of CDM models fail to produce the observed coherence of structure on non-linear scales (>95% confidence level). The underdensity probability (the frequency of regions with density contrast delta rho //lineρ=-0.8) depends strongly on the luminosity of galaxies; underdense regions are significantly more common (>2sigma ) in bright (L>L(*) ) galaxy samples than in samples which include fainter galaxies.

  20. Management of large-scale multimedia conferencing

    NASA Astrophysics Data System (ADS)

    Cidon, Israel; Nachum, Youval

    1998-12-01

    The goal of this work is to explore management strategies and algorithms for large-scale multimedia conferencing over a communication network. Since the use of multimedia conferencing is still limited, the management of such systems has not yet been studied in depth. A well organized and human friendly multimedia conference management should utilize efficiently and fairly its limited resources as well as take into account the requirements of the conference participants. The ability of the management to enforce fair policies and to quickly take into account the participants preferences may even lead to a conference environment that is more pleasant and more effective than a similar face to face meeting. We suggest several principles for defining and solving resource sharing problems in this context. The conference resources which are addressed in this paper are the bandwidth (conference network capacity), time (participants' scheduling) and limitations of audio and visual equipment. The participants' requirements for these resources are defined and translated in terms of Quality of Service requirements and the fairness criteria.

  1. Large-scale wind turbine structures

    NASA Technical Reports Server (NTRS)

    Spera, David A.

    1988-01-01

    The purpose of this presentation is to show how structural technology was applied in the design of modern wind turbines, which were recently brought to an advanced stage of development as sources of renewable power. Wind turbine structures present many difficult problems because they are relatively slender and flexible; subject to vibration and aeroelastic instabilities; acted upon by loads which are often nondeterministic; operated continuously with little maintenance in all weather; and dominated by life-cycle cost considerations. Progress in horizontal-axis wind turbines (HAWT) development was paced by progress in the understanding of structural loads, modeling of structural dynamic response, and designing of innovative structural response. During the past 15 years a series of large HAWTs was developed. This has culminated in the recent completion of the world's largest operating wind turbine, the 3.2 MW Mod-5B power plane installed on the island of Oahu, Hawaii. Some of the applications of structures technology to wind turbine will be illustrated by referring to the Mod-5B design. First, a video overview will be presented to provide familiarization with the Mod-5B project and the important components of the wind turbine system. Next, the structural requirements for large-scale wind turbines will be discussed, emphasizing the difficult fatigue-life requirements. Finally, the procedures used to design the structure will be presented, including the use of the fracture mechanics approach for determining allowable fatigue stresses.

  2. Large-scale tides in general relativity

    NASA Astrophysics Data System (ADS)

    Ip, Hiu Yan; Schmidt, Fabian

    2017-02-01

    Density perturbations in cosmology, i.e. spherically symmetric adiabatic perturbations of a Friedmann-Lemaȋtre-Robertson-Walker (FLRW) spacetime, are locally exactly equivalent to a different FLRW solution, as long as their wavelength is much larger than the sound horizon of all fluid components. This fact is known as the "separate universe" paradigm. However, no such relation is known for anisotropic adiabatic perturbations, which correspond to an FLRW spacetime with large-scale tidal fields. Here, we provide a closed, fully relativistic set of evolutionary equations for the nonlinear evolution of such modes, based on the conformal Fermi (CFC) frame. We show explicitly that the tidal effects are encoded by the Weyl tensor, and are hence entirely different from an anisotropic Bianchi I spacetime, where the anisotropy is sourced by the Ricci tensor. In order to close the system, certain higher derivative terms have to be dropped. We show that this approximation is equivalent to the local tidal approximation of Hui and Bertschinger [1]. We also show that this very simple set of equations matches the exact evolution of the density field at second order, but fails at third and higher order. This provides a useful, easy-to-use framework for computing the fully relativistic growth of structure at second order.

  3. Large scale mechanical metamaterials as seismic shields

    NASA Astrophysics Data System (ADS)

    Miniaci, Marco; Krushynska, Anastasiia; Bosia, Federico; Pugno, Nicola M.

    2016-08-01

    Earthquakes represent one of the most catastrophic natural events affecting mankind. At present, a universally accepted risk mitigation strategy for seismic events remains to be proposed. Most approaches are based on vibration isolation of structures rather than on the remote shielding of incoming waves. In this work, we propose a novel approach to the problem and discuss the feasibility of a passive isolation strategy for seismic waves based on large-scale mechanical metamaterials, including for the first time numerical analysis of both surface and guided waves, soil dissipation effects, and adopting a full 3D simulations. The study focuses on realistic structures that can be effective in frequency ranges of interest for seismic waves, and optimal design criteria are provided, exploring different metamaterial configurations, combining phononic crystals and locally resonant structures and different ranges of mechanical properties. Dispersion analysis and full-scale 3D transient wave transmission simulations are carried out on finite size systems to assess the seismic wave amplitude attenuation in realistic conditions. Results reveal that both surface and bulk seismic waves can be considerably attenuated, making this strategy viable for the protection of civil structures against seismic risk. The proposed remote shielding approach could open up new perspectives in the field of seismology and in related areas of low-frequency vibration damping or blast protection.

  4. Food appropriation through large scale land acquisitions

    NASA Astrophysics Data System (ADS)

    Rulli, Maria Cristina; D'Odorico, Paolo

    2014-05-01

    The increasing demand for agricultural products and the uncertainty of international food markets has recently drawn the attention of governments and agribusiness firms toward investments in productive agricultural land, mostly in the developing world. The targeted countries are typically located in regions that have remained only marginally utilized because of lack of modern technology. It is expected that in the long run large scale land acquisitions (LSLAs) for commercial farming will bring the technology required to close the existing crops yield gaps. While the extent of the acquired land and the associated appropriation of freshwater resources have been investigated in detail, the amount of food this land can produce and the number of people it could feed still need to be quantified. Here we use a unique dataset of land deals to provide a global quantitative assessment of the rates of crop and food appropriation potentially associated with LSLAs. We show how up to 300-550 million people could be fed by crops grown in the acquired land, should these investments in agriculture improve crop production and close the yield gap. In contrast, about 190-370 million people could be supported by this land without closing of the yield gap. These numbers raise some concern because the food produced in the acquired land is typically exported to other regions, while the target countries exhibit high levels of malnourishment. Conversely, if used for domestic consumption, the crops harvested in the acquired land could ensure food security to the local populations.

  5. Large scale structure of the sun's corona

    NASA Astrophysics Data System (ADS)

    Kundu, Mukul R.

    Results concerning the large-scale structure of the solar corona obtained by observations at meter-decameter wavelengths are reviewed. Coronal holes observed on the disk at multiple frequencies show the radial and azimuthal geometry of the hole. At the base of the hole there is good correspondence to the chromospheric signature in He I 10,830 A, but at greater heights the hole may show departures from symmetry. Two-dimensional imaging of weak-type III bursts simultaneously with the HAO SMM coronagraph/polarimeter measurements indicate that these bursts occur along elongated features emanating from the quiet sun, corresponding in position angle to the bright coronal streamers. It is shown that the densest regions of streamers and the regions of maximum intensity of type II bursts coincide closely. Non-flare-associated type II/type IV bursts associated with coronal streamer disruption events are studied along with correlated type II burst emissions originating from distant centers on the sun.

  6. Large-scale carbon fiber tests

    NASA Technical Reports Server (NTRS)

    Pride, R. A.

    1980-01-01

    A realistic release of carbon fibers was established by burning a minimum of 45 kg of carbon fiber composite aircraft structural components in each of five large scale, outdoor aviation jet fuel fire tests. This release was quantified by several independent assessments with various instruments developed specifically for these tests. The most likely values for the mass of single carbon fibers released ranged from 0.2 percent of the initial mass of carbon fiber for the source tests (zero wind velocity) to a maximum of 0.6 percent of the initial carbon fiber mass for dissemination tests (5 to 6 m/s wind velocity). Mean fiber lengths for fibers greater than 1 mm in length ranged from 2.5 to 3.5 mm. Mean diameters ranged from 3.6 to 5.3 micrometers which was indicative of significant oxidation. Footprints of downwind dissemination of the fire released fibers were measured to 19.1 km from the fire.

  7. Large-scale clustering of cosmic voids

    NASA Astrophysics Data System (ADS)

    Chan, Kwan Chuen; Hamaus, Nico; Desjacques, Vincent

    2014-11-01

    We study the clustering of voids using N -body simulations and simple theoretical models. The excursion-set formalism describes fairly well the abundance of voids identified with the watershed algorithm, although the void formation threshold required is quite different from the spherical collapse value. The void cross bias bc is measured and its large-scale value is found to be consistent with the peak background split results. A simple fitting formula for bc is found. We model the void auto-power spectrum taking into account the void biasing and exclusion effect. A good fit to the simulation data is obtained for voids with radii ≳30 Mpc h-1 , especially when the void biasing model is extended to 1-loop order. However, the best-fit bias parameters do not agree well with the peak-background results. Being able to fit the void auto-power spectrum is particularly important not only because it is the direct observable in galaxy surveys, but also our method enables us to treat the bias parameters as nuisance parameters, which are sensitive to the techniques used to identify voids.

  8. Large-scale autostereoscopic outdoor display

    NASA Astrophysics Data System (ADS)

    Reitterer, Jörg; Fidler, Franz; Saint Julien-Wallsee, Ferdinand; Schmid, Gerhard; Gartner, Wolfgang; Leeb, Walter; Schmid, Ulrich

    2013-03-01

    State-of-the-art autostereoscopic displays are often limited in size, effective brightness, number of 3D viewing zones, and maximum 3D viewing distances, all of which are mandatory requirements for large-scale outdoor displays. Conventional autostereoscopic indoor concepts like lenticular lenses or parallax barriers cannot simply be adapted for these screens due to the inherent loss of effective resolution and brightness, which would reduce both image quality and sunlight readability. We have developed a modular autostereoscopic multi-view laser display concept with sunlight readable effective brightness, theoretically up to several thousand 3D viewing zones, and maximum 3D viewing distances of up to 60 meters. For proof-of-concept purposes a prototype display with two pixels was realized. Due to various manufacturing tolerances each individual pixel has slightly different optical properties, and hence the 3D image quality of the display has to be calculated stochastically. In this paper we present the corresponding stochastic model, we evaluate the simulation and measurement results of the prototype display, and we calculate the achievable autostereoscopic image quality to be expected for our concept.

  9. Large Scale EOF Analysis of Climate Data

    NASA Astrophysics Data System (ADS)

    Prabhat, M.; Gittens, A.; Kashinath, K.; Cavanaugh, N. R.; Mahoney, M.

    2016-12-01

    We present a distributed approach towards extracting EOFs from 3D climate data. We implement the method in Apache Spark, and process multi-TB sized datasets on O(1000-10,000) cores. We apply this method to latitude-weighted ocean temperature data from CSFR, a 2.2 terabyte-sized data set comprising ocean and subsurface reanalysis measurements collected at 41 levels in the ocean, at 6 hour intervals over 31 years. We extract the first 100 EOFs of this full data set and compare to the EOFs computed simply on the surface temperature field. Our analyses provide evidence of Kelvin and Rossy waves and components of large-scale modes of oscillation including the ENSO and PDO that are not visible in the usual SST EOFs. Further, they provide information on the the most influential parts of the ocean, such as the thermocline, that exist below the surface. Work is ongoing to understand the factors determining the depth-varying spatial patterns observed in the EOFs. We will experiment with weighting schemes to appropriately account for the differing depths of the observations. We also plan to apply the same distributed approach to analysis of analysis of 3D atmospheric climatic data sets, including multiple variables. Because the atmosphere changes on a quicker time-scale than the ocean, we expect that the results will demonstrate an even greater advantage to computing 3D EOFs in lieu of 2D EOFs.

  10. Numerical Modeling for Large Scale Hydrothermal System

    NASA Astrophysics Data System (ADS)

    Sohrabi, Reza; Jansen, Gunnar; Malvoisin, Benjamin; Mazzini, Adriano; Miller, Stephen A.

    2017-04-01

    Moderate-to-high enthalpy systems are driven by multiphase and multicomponent processes, fluid and rock mechanics, and heat transport processes, all of which present challenges in developing realistic numerical models of the underlying physics. The objective of this work is to present an approach, and some initial results, for modeling and understanding dynamics of the birth of large scale hydrothermal systems. Numerical modeling of such complex systems must take into account a variety of coupled thermal, hydraulic, mechanical and chemical processes, which is numerically challenging. To provide first estimates of the behavior of this deep complex systems, geological structures must be constrained, and the fluid dynamics, mechanics and the heat transport need to be investigated in three dimensions. Modeling these processes numerically at adequate resolution and reasonable computation times requires a suite of tools that we are developing and/or utilizing to investigate such systems. Our long-term goal is to develop 3D numerical models, based on a geological models, which couples mechanics with the hydraulics and thermal processes driving hydrothermal system. Our first results from the Lusi hydrothermal system in East Java, Indonesia provide a basis for more sophisticated studies, eventually in 3D, and we introduce a workflow necessary to achieve these objectives. Future work focuses with the aim and parallelization suitable for High Performance Computing (HPC). Such developments are necessary to achieve high-resolution simulations to more fully understand the complex dynamics of hydrothermal systems.

  11. Large scale digital atlases in neuroscience

    NASA Astrophysics Data System (ADS)

    Hawrylycz, M.; Feng, D.; Lau, C.; Kuan, C.; Miller, J.; Dang, C.; Ng, L.

    2014-03-01

    Imaging in neuroscience has revolutionized our current understanding of brain structure, architecture and increasingly its function. Many characteristics of morphology, cell type, and neuronal circuitry have been elucidated through methods of neuroimaging. Combining this data in a meaningful, standardized, and accessible manner is the scope and goal of the digital brain atlas. Digital brain atlases are used today in neuroscience to characterize the spatial organization of neuronal structures, for planning and guidance during neurosurgery, and as a reference for interpreting other data modalities such as gene expression and connectivity data. The field of digital atlases is extensive and in addition to atlases of the human includes high quality brain atlases of the mouse, rat, rhesus macaque, and other model organisms. Using techniques based on histology, structural and functional magnetic resonance imaging as well as gene expression data, modern digital atlases use probabilistic and multimodal techniques, as well as sophisticated visualization software to form an integrated product. Toward this goal, brain atlases form a common coordinate framework for summarizing, accessing, and organizing this knowledge and will undoubtedly remain a key technology in neuroscience in the future. Since the development of its flagship project of a genome wide image-based atlas of the mouse brain, the Allen Institute for Brain Science has used imaging as a primary data modality for many of its large scale atlas projects. We present an overview of Allen Institute digital atlases in neuroscience, with a focus on the challenges and opportunities for image processing and computation.

  12. Sensitivity technologies for large scale simulation.

    SciTech Connect

    Collis, Samuel Scott; Bartlett, Roscoe Ainsworth; Smith, Thomas Michael; Heinkenschloss, Matthias; Wilcox, Lucas C.; Hill, Judith C.; Ghattas, Omar; Berggren, Martin Olof; Akcelik, Volkan; Ober, Curtis Curry; van Bloemen Waanders, Bart Gustaaf; Keiter, Eric Richard

    2005-01-01

    Sensitivity analysis is critically important to numerous analysis algorithms, including large scale optimization, uncertainty quantification,reduced order modeling, and error estimation. Our research focused on developing tools, algorithms and standard interfaces to facilitate the implementation of sensitivity type analysis into existing code and equally important, the work was focused on ways to increase the visibility of sensitivity analysis. We attempt to accomplish the first objective through the development of hybrid automatic differentiation tools, standard linear algebra interfaces for numerical algorithms, time domain decomposition algorithms and two level Newton methods. We attempt to accomplish the second goal by presenting the results of several case studies in which direct sensitivities and adjoint methods have been effectively applied, in addition to an investigation of h-p adaptivity using adjoint based a posteriori error estimation. A mathematical overview is provided of direct sensitivities and adjoint methods for both steady state and transient simulations. Two case studies are presented to demonstrate the utility of these methods. A direct sensitivity method is implemented to solve a source inversion problem for steady state internal flows subject to convection diffusion. Real time performance is achieved using novel decomposition into offline and online calculations. Adjoint methods are used to reconstruct initial conditions of a contamination event in an external flow. We demonstrate an adjoint based transient solution. In addition, we investigated time domain decomposition algorithms in an attempt to improve the efficiency of transient simulations. Because derivative calculations are at the root of sensitivity calculations, we have developed hybrid automatic differentiation methods and implemented this approach for shape optimization for gas dynamics using the Euler equations. The hybrid automatic differentiation method was applied to a first

  13. Large Scale Flame Spread Environmental Characterization Testing

    NASA Technical Reports Server (NTRS)

    Clayman, Lauren K.; Olson, Sandra L.; Gokoghi, Suleyman A.; Brooker, John E.; Ferkul, Paul V.; Kacher, Henry F.

    2013-01-01

    Under the Advanced Exploration Systems (AES) Spacecraft Fire Safety Demonstration Project (SFSDP), as a risk mitigation activity in support of the development of a large-scale fire demonstration experiment in microgravity, flame-spread tests were conducted in normal gravity on thin, cellulose-based fuels in a sealed chamber. The primary objective of the tests was to measure pressure rise in a chamber as sample material, burning direction (upward/downward), total heat release, heat release rate, and heat loss mechanisms were varied between tests. A Design of Experiments (DOE) method was imposed to produce an array of tests from a fixed set of constraints and a coupled response model was developed. Supplementary tests were run without experimental design to additionally vary select parameters such as initial chamber pressure. The starting chamber pressure for each test was set below atmospheric to prevent chamber overpressure. Bottom ignition, or upward propagating burns, produced rapid acceleratory turbulent flame spread. Pressure rise in the chamber increases as the amount of fuel burned increases mainly because of the larger amount of heat generation and, to a much smaller extent, due to the increase in gaseous number of moles. Top ignition, or downward propagating burns, produced a steady flame spread with a very small flat flame across the burning edge. Steady-state pressure is achieved during downward flame spread as the pressure rises and plateaus. This indicates that the heat generation by the flame matches the heat loss to surroundings during the longer, slower downward burns. One heat loss mechanism included mounting a heat exchanger directly above the burning sample in the path of the plume to act as a heat sink and more efficiently dissipate the heat due to the combustion event. This proved an effective means for chamber overpressure mitigation for those tests producing the most total heat release and thusly was determined to be a feasible mitigation

  14. Transposon tagging and the study of root development in Arabidopsis

    NASA Technical Reports Server (NTRS)

    Tsugeki, R.; Olson, M. L.; Fedoroff, N. V.

    1998-01-01

    The maize Ac-Ds transposable element family has been used as the basis of transposon mutagenesis systems that function in a variety of plants, including Arabidopsis. We have developed modified transposons and methods which simplify the detection, cloning and analysis of insertion mutations. We have identified and are analyzing two plant lines in which genes expressed either in the root cap cells or in the quiescent cells, cortex/endodermal initial cells and columella cells of the root cap have been tagged with a transposon carrying a reporter gene. A gene expressed in root cap cells tagged with an enhancer-trap Ds was isolated and its corresponding EST cDNA was identified. Nucleotide and deduced amino acid sequences of the gene show no significant similarity to other genes in the database. Genetic ablation experiments have been done by fusing a root cap-specific promoter to the diphtheria toxin A-chain gene and introducing the fusion construct into Arabidopsis plants. We find that in addition to eliminating gravitropism, root cap ablation inhibits elongation of roots by lowering root meristematic activities.

  15. Synchronization of coupled large-scale Boolean networks

    SciTech Connect

    Li, Fangfei

    2014-03-15

    This paper investigates the complete synchronization and partial synchronization of two large-scale Boolean networks. First, the aggregation algorithm towards large-scale Boolean network is reviewed. Second, the aggregation algorithm is applied to study the complete synchronization and partial synchronization of large-scale Boolean networks. Finally, an illustrative example is presented to show the efficiency of the proposed results.

  16. The School Principal's Role in Large-Scale Assessment

    ERIC Educational Resources Information Center

    Newton, Paul; Tunison, Scott; Viczko, Melody

    2010-01-01

    This paper reports on an interpretive study in which 25 elementary principals were asked about their assessment knowledge, the use of large-scale assessments in their schools, and principals' perceptions on their roles with respect to large-scale assessments. Principals in this study suggested that the current context of large-scale assessment and…

  17. Synchronization of coupled large-scale Boolean networks

    NASA Astrophysics Data System (ADS)

    Li, Fangfei

    2014-03-01

    This paper investigates the complete synchronization and partial synchronization of two large-scale Boolean networks. First, the aggregation algorithm towards large-scale Boolean network is reviewed. Second, the aggregation algorithm is applied to study the complete synchronization and partial synchronization of large-scale Boolean networks. Finally, an illustrative example is presented to show the efficiency of the proposed results.

  18. Modified mariner transposons for random inducible-expression insertions and transcriptional reporter fusion insertions in Bacillus subtilis.

    PubMed

    Pozsgai, Eric R; Blair, Kris M; Kearns, Daniel B

    2012-02-01

    Transposons are mobile genetic elements bounded by insertion sequences that are recognized by a specific mobilizing transposase enzyme. The transposase may mobilize not only the insertion sequences but also intervening DNA. mariner is a particularly efficient transposon for the random chromosomal integration of genes and insertional mutagenesis. Here, we modify an existing mariner transposon, TnYLB, such that it can easily be genetically manipulated and introduced into Bacillus subtilis. We generate a series of three new mariner derivatives that mobilize spectinomycin, chloramphenicol, and kanamycin antibiotic resistance cassettes. Furthermore, we generate a series of transposons with a strong, outward-oriented, optionally isopropyl-β-D-thiogalactopyranoside (IPTG)-inducible promoter for the random overexpression of neighboring genes and a series of transposons with a promoterless lacZ gene for the random generation of transcriptional reporter fusions. We note that the modification of the base transposon is not restricted to B. subtilis and should be applicable to any mariner-compatible host organism, provided that in vitro mutagenesis or an in vivo species-specific delivery vector is employed.

  19. Large scale dynamics of protoplanetary discs

    NASA Astrophysics Data System (ADS)

    Béthune, William

    2017-08-01

    Planets form in the gaseous and dusty disks orbiting young stars. These protoplanetary disks are dispersed in a few million years, being accreted onto the central star or evaporated into the interstellar medium. To explain the observed accretion rates, it is commonly assumed that matter is transported through the disk by turbulence, although the mechanism sustaining turbulence is uncertain. On the other side, irradiation by the central star could heat up the disk surface and trigger a photoevaporative wind, but thermal effects cannot account for the observed acceleration and collimation of the wind into a narrow jet perpendicular to the disk plane. Both issues can be solved if the disk is sensitive to magnetic fields. Weak fields lead to the magnetorotational instability, whose outcome is a state of sustained turbulence. Strong fields can slow down the disk, causing it to accrete while launching a collimated wind. However, the coupling between the disk and the neutral gas is done via electric charges, each of which is outnumbered by several billion neutral molecules. The imperfect coupling between the magnetic field and the neutral gas is described in terms of "non-ideal" effects, introducing new dynamical behaviors. This thesis is devoted to the transport processes happening inside weakly ionized and weakly magnetized accretion disks; the role of microphysical effects on the large-scale dynamics of the disk is of primary importance. As a first step, I exclude the wind and examine the impact of non-ideal effects on the turbulent properties near the disk midplane. I show that the flow can spontaneously organize itself if the ionization fraction is low enough; in this case, accretion is halted and the disk exhibits axisymmetric structures, with possible consequences on planetary formation. As a second step, I study the launching of disk winds via a global model of stratified disk embedded in a warm atmosphere. This model is the first to compute non-ideal effects from

  20. Large-Scale Spacecraft Fire Safety Tests

    NASA Technical Reports Server (NTRS)

    Urban, David; Ruff, Gary A.; Ferkul, Paul V.; Olson, Sandra; Fernandez-Pello, A. Carlos; T'ien, James S.; Torero, Jose L.; Cowlard, Adam J.; Rouvreau, Sebastien; Minster, Olivier; hide

    2014-01-01

    An international collaborative program is underway to address open issues in spacecraft fire safety. Because of limited access to long-term low-gravity conditions and the small volume generally allotted for these experiments, there have been relatively few experiments that directly study spacecraft fire safety under low-gravity conditions. Furthermore, none of these experiments have studied sample sizes and environment conditions typical of those expected in a spacecraft fire. The major constraint has been the size of the sample, with prior experiments limited to samples of the order of 10 cm in length and width or smaller. This lack of experimental data forces spacecraft designers to base their designs and safety precautions on 1-g understanding of flame spread, fire detection, and suppression. However, low-gravity combustion research has demonstrated substantial differences in flame behavior in low-gravity. This, combined with the differences caused by the confined spacecraft environment, necessitates practical scale spacecraft fire safety research to mitigate risks for future space missions. To address this issue, a large-scale spacecraft fire experiment is under development by NASA and an international team of investigators. This poster presents the objectives, status, and concept of this collaborative international project (Saffire). The project plan is to conduct fire safety experiments on three sequential flights of an unmanned ISS re-supply spacecraft (the Orbital Cygnus vehicle) after they have completed their delivery of cargo to the ISS and have begun their return journeys to earth. On two flights (Saffire-1 and Saffire-3), the experiment will consist of a flame spread test involving a meter-scale sample ignited in the pressurized volume of the spacecraft and allowed to burn to completion while measurements are made. On one of the flights (Saffire-2), 9 smaller (5 x 30 cm) samples will be tested to evaluate NASAs material flammability screening tests

  1. Large scale simulations of Brownian suspensions

    NASA Astrophysics Data System (ADS)

    Viera, Marc Nathaniel

    Particle suspensions occur in a wide variety of natural and engineering materials. Some examples are colloids, polymers, paints, and slurries. These materials exhibit complex behavior owing to the forces which act among the particles and are transmitted through the fluid medium. Depending on the application, particle sizes range from large macroscopic molecules of 100mum to smaller colloidal particles in the range of 10nm to 1mum. Particles of this size interact though interparticle forces such as electrostatic and van der Waals, as well as hydrodynamic forces transmitted through the fluid medium. Additionally, the particles are subjected to random thermal fluctuations in the fluid giving rise to Brownian motion. The central objective of our research is to develop efficient numerical algorithms for the large scale dynamic simulation of particle suspensions. While previous methods have incurred a computational cost of O(N3), where N is the number of particles, we have developed a novel algorithm capable of solving this problem in O(N ln N) operations. This has allowed us to perform dynamic simulations with up to 64,000 particles and Monte Carlo realizations of up to 1 million particles. Our algorithm follows a Stokesian dynamics formulation by evaluating many-body hydrodynamic interactions using a far-field multipole expansion combined with a near-field lubrication correction. The breakthrough O(N ln N) scaling is obtained by employing a Particle-Mesh-Ewald (PME) approach whereby near-field interactions are evaluated directly and far-field interactions are evaluated using a grid based velocity computed with FFT's. This approach is readily extended to include the effects of Brownian motion. For interacting particles, the fluctuation-dissipation theorem requires that the individual Brownian forces satisfy a correlation based on the N body resistance tensor R. The accurate modeling of these forces requires the computation of a matrix square root R 1/2 for matrices up

  2. Plant regeneration methods for rapid generation of a large scale Ds transposant population in rice.

    PubMed

    Xuan, Yuan Hu; Huang, Jin; Yi, Gihwan; Park, Dong-Soo; Park, Soo Kwon; Eun, Moo Young; Yun, Doh Won; Lee, Gang-Seob; Kim, Tae Ho; Han, Chang-deok

    2013-01-01

    To mutagenize rice genomes, a two-element system is utilized. This system comprises an immobile Ac element driven by the CaMV 35S promoter, and a gene trap Ds carrying a partial intron with alternative splice acceptors fused to the GUS coding region. Rapid, large-scale generation of a Ds transposant population was achieved using a plant regeneration procedure involving the tissue culture of seed-derived calli carrying Ac and Ds elements. During tissue cultures, Ds mobility accompanies changes in methylation patterns of a terminal region of Ds, where over 70% of plants contained independent Ds insertions. In the transposon population, around 12% of plants expressed GUS at the early seedling stage. A flanking-sequence-tag (FST) database has been established by cloning over 19,968 Ds insertion sites and the Ds map shows relatively uniform distribution across the rice chromosomes.

  3. Genomic landscape of human, bat, and ex vivo DNA transposon integrations.

    PubMed

    Campos-Sánchez, Rebeca; Kapusta, Aurélie; Feschotte, Cédric; Chiaromonte, Francesca; Makova, Kateryna D

    2014-07-01

    The integration and fixation preferences of DNA transposons, one of the major classes of eukaryotic transposable elements, have never been evaluated comprehensively on a genome-wide scale. Here, we present a detailed study of the distribution of DNA transposons in the human and bat genomes. We studied three groups of DNA transposons that integrated at different evolutionary times: 1) ancient (>40 My) and currently inactive human elements, 2) younger (<40 My) bat elements, and 3) ex vivo integrations of piggyBat and Sleeping Beauty elements in HeLa cells. Although the distribution of ex vivo elements reflected integration preferences, the distribution of human and (to a lesser extent) bat elements was also affected by selection. We used regression techniques (linear, negative binomial, and logistic regression models with multiple predictors) applied to 20-kb and 1-Mb windows to investigate how the genomic landscape in the vicinity of DNA transposons contributes to their integration and fixation. Our models indicate that genomic landscape explains 16-79% of variability in DNA transposon genome-wide distribution. Importantly, we not only confirmed previously identified predictors (e.g., DNA conformation and recombination hotspots) but also identified several novel predictors (e.g., signatures of double-strand breaks and telomere hexamer). Ex vivo integrations showed a bias toward actively transcribed regions. Older DNA transposons were located in genomic regions scarce in most conserved elements-likely reflecting purifying selection. Our study highlights how DNA transposons are integral to the evolution of bat and human genomes, and has implications for the development of DNA transposon assays for gene therapy and mutagenesis applications.

  4. Size matters: versatile use of PiggyBac transposons as a genetic manipulation tool.

    PubMed

    Kim, Adele; Pyykko, Ilmari

    2011-08-01

    Transposons have been promising elements for gene integration, and the Sleeping Beauty (SB) system has been the major one for many years, although there have been several other transposon systems available, for example, Tol2. However, recently another system known as PiggyBac (PB) has been introduced and developed for fulfilling the same purposes, for example, mutagenesis, transgenesis and gene therapy and in some cases with improved transposition efficiency and advantages over the Sleeping Beauty transposon system, although improved hyperactive transposase has highly increased the transposition efficacy for SB. The PB systems have been used in many different scientific research fields; therefore, the purpose of this review is to describe some of these versatile uses of the PiggyBac system to give readers an overview on the usage of PiggyBac system.

  5. Transposon tagging of disease resistance genes. Final report, May 1, 1988--April 30, 1993

    SciTech Connect

    Michelmore, R.

    1994-09-01

    The goal of this project was to develop a transposon mutagenesis system for lettuce and to clone and characterize disease resistance genes by transposon tagging. The majority of studies were conducted with the Ac/Ds System. Researchers made and tested several constructs as well as utilized constructions shown to be functional in other plant species. Researchers demonstrated movement of Ac and DS in lettuce; however, they transposed at much lower frequencies in lettuce than in other plant species. Therefore, further manipulation of the system, particularly for flower specific expression of transposase, is required before a routine transposon system is available for lettuce. Populations of lettuce were generated and screened to test for the stability of resistance genes and several spontaneous mutations were isolated. Researchers also identified a resistance gene mutant in plants transformed with a Ds element and chimeric transposase gene. This is currently being characterized in detail.

  6. Population generation for large-scale simulation

    NASA Astrophysics Data System (ADS)

    Hannon, Andrew C.; King, Gary; Morrison, Clayton; Galstyan, Aram; Cohen, Paul

    2005-05-01

    Computer simulation is used to research phenomena ranging from the structure of the space-time continuum to population genetics and future combat.1-3 Multi-agent simulations in particular are now commonplace in many fields.4, 5 By modeling populations whose complex behavior emerges from individual interactions, these simulations help to answer questions about effects where closed form solutions are difficult to solve or impossible to derive.6 To be useful, simulations must accurately model the relevant aspects of the underlying domain. In multi-agent simulation, this means that the modeling must include both the agents and their relationships. Typically, each agent can be modeled as a set of attributes drawn from various distributions (e.g., height, morale, intelligence and so forth). Though these can interact - for example, agent height is related to agent weight - they are usually independent. Modeling relations between agents, on the other hand, adds a new layer of complexity, and tools from graph theory and social network analysis are finding increasing application.7, 8 Recognizing the role and proper use of these techniques, however, remains the subject of ongoing research. We recently encountered these complexities while building large scale social simulations.9-11 One of these, the Hats Simulator, is designed to be a lightweight proxy for intelligence analysis problems. Hats models a "society in a box" consisting of many simple agents, called hats. Hats gets its name from the classic spaghetti western, in which the heroes and villains are known by the color of the hats they wear. The Hats society also has its heroes and villains, but the challenge is to identify which color hat they should be wearing based on how they behave. There are three types of hats: benign hats, known terrorists, and covert terrorists. Covert terrorists look just like benign hats but act like terrorists. Population structure can make covert hat identification significantly more

  7. Large-scale Fractal Motion of Clouds

    NASA Image and Video Library

    2017-09-27

    waters surrounding the island.) The “swallowed” gulps of clear island air get carried along within the vortices, but these are soon mixed into the surrounding clouds. Landsat is unique in its ability to image both the small-scale eddies that mix clear and cloudy air, down to the 30 meter pixel size of Landsat, but also having a wide enough field-of-view, 180 km, to reveal the connection of the turbulence to large-scale flows such as the subtropical oceanic gyres. Landsat 7, with its new onboard digital recorder, has extended this capability away from the few Landsat ground stations to remote areas such as Alejandro Island, and thus is gradually providing a global dynamic picture of evolving human-scale phenomena. For more details on von Karman vortices, refer to climate.gsfc.nasa.gov/~cahalan. Image and caption courtesy Bob Cahalan, NASA GSFC Instrument: Landsat 7 - ETM+ Credit: NASA/GSFC/Landsat NASA Goddard Space Flight Center enables NASA’s mission through four scientific endeavors: Earth Science, Heliophysics, Solar System Exploration, and Astrophysics. Goddard plays a leading role in NASA’s accomplishments by contributing compelling scientific knowledge to advance the Agency’s mission. Follow us on Twitter Join us on Facebook

  8. Large-scale assembly of colloidal particles

    NASA Astrophysics Data System (ADS)

    Yang, Hongta

    This study reports a simple, roll-to-roll compatible coating technology for producing three-dimensional highly ordered colloidal crystal-polymer composites, colloidal crystals, and macroporous polymer membranes. A vertically beveled doctor blade is utilized to shear align silica microsphere-monomer suspensions to form large-area composites in a single step. The polymer matrix and the silica microspheres can be selectively removed to create colloidal crystals and self-standing macroporous polymer membranes. The thickness of the shear-aligned crystal is correlated with the viscosity of the colloidal suspension and the coating speed, and the correlations can be qualitatively explained by adapting the mechanisms developed for conventional doctor blade coating. Five important research topics related to the application of large-scale three-dimensional highly ordered macroporous films by doctor blade coating are covered in this study. The first topic describes the invention in large area and low cost color reflective displays. This invention is inspired by the heat pipe technology. The self-standing macroporous polymer films exhibit brilliant colors which originate from the Bragg diffractive of visible light form the three-dimensional highly ordered air cavities. The colors can be easily changed by tuning the size of the air cavities to cover the whole visible spectrum. When the air cavities are filled with a solvent which has the same refractive index as that of the polymer, the macroporous polymer films become completely transparent due to the index matching. When the solvent trapped in the cavities is evaporated by in-situ heating, the sample color changes back to brilliant color. This process is highly reversible and reproducible for thousands of cycles. The second topic reports the achievement of rapid and reversible vapor detection by using 3-D macroporous photonic crystals. Capillary condensation of a condensable vapor in the interconnected macropores leads to the

  9. Generation of induced pluripotent stem cells from human foetal fibroblasts using the Sleeping Beauty transposon gene delivery system.

    PubMed

    Davis, Richard P; Nemes, Csilla; Varga, Eszter; Freund, Christian; Kosmidis, Georgios; Gkatzis, Konstantinos; de Jong, Danielle; Szuhai, Károly; Dinnyés, András; Mummery, Christine L

    2013-01-01

    Transposon gene delivery systems offer an alternative, non-viral-based approach to generate induced pluripotent stem cells (iPSCs). Here we used the Sleeping Beauty (SB) transposon to generate four human iPSC lines from foetal fibroblasts. In contrast to other gene delivery systems, the SB transposon does not exhibit an integration bias towards particular genetic elements, thereby reducing the risk of insertional mutagenesis. Furthermore, unlike the alternative transposon piggyBac, SB has no SB-like elements within the human genome, minimising the possibility of mobilising endogenous transposon elements. All iPSC lines exhibited the expected characteristics of pluripotent human cells, including the ability to differentiate to derivatives of all three germ layers in vitro. Re-expression of the SB transposase in the iPSCs after reprogramming resulted in the mobilisation of some of the transposons. These results indicate that the SB transposon system is a useful addition to methods for generating human iPSCs, both for basic and applied biomedical research, and in the context of future therapeutic application. © 2013 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  10. Multitree Algorithms for Large-Scale Astrostatistics

    NASA Astrophysics Data System (ADS)

    March, William B.; Ozakin, Arkadas; Lee, Dongryeol; Riegel, Ryan; Gray, Alexander G.

    2012-03-01

    this number every week, resulting in billions of objects. At such scales, even linear-time analysis operations present challenges, particularly since statistical analyses are inherently interactive processes, requiring that computations complete within some reasonable human attention span. The quadratic (or worse) runtimes of straightforward implementations become quickly unbearable. Examples of applications. These analysis subroutines occur ubiquitously in astrostatistical work. We list just a few examples. The need to cross-match objects across different catalogs has led to various algorithms, which at some point perform an AllNN computation. 2-point and higher-order spatial correlations for the basis of spatial statistics, and are utilized in astronomy to compare the spatial structures of two datasets, such as an observed sample and a theoretical sample, for example, forming the basis for two-sample hypothesis testing. Friends-of-friends clustering is often used to identify halos in data from astrophysical simulations. Minimum spanning tree properties have also been proposed as statistics of large-scale structure. Comparison of the distributions of different kinds of objects requires accurate density estimation, for which KDE is the overall statistical method of choice. The prediction of redshifts from optical data requires accurate regression, for which kernel regression is a powerful method. The identification of objects of various types in astronomy, such as stars versus galaxies, requires accurate classification, for which KDA is a powerful method. Overview. In this chapter, we will briefly sketch the main ideas behind recent fast algorithms which achieve, for example, linear runtimes for pairwise-distance problems, or similarly dramatic reductions in computational growth. In some cases, the runtime orders for these algorithms are mathematically provable statements, while in others we have only conjectures backed by experimental observations for the time being

  11. The expanding universe of transposon technologies for gene and cell engineering.

    PubMed

    Ivics, Zoltán; Izsvák, Zsuzsanna

    2010-12-07

    Transposable elements can be viewed as natural DNA transfer vehicles that, similar to integrating viruses, are capable of efficient genomic insertion. The mobility of class II transposable elements (DNA transposons) can be controlled by conditionally providing the transposase component of the transposition reaction. Thus, a DNA of interest (be it a fluorescent marker, a small hairpin (sh)RNA expression cassette, a mutagenic gene trap or a therapeutic gene construct) cloned between the inverted repeat sequences of a transposon-based vector can be used for stable genomic insertion in a regulated and highly efficient manner. This methodological paradigm opened up a number of avenues for genome manipulations in vertebrates, including transgenesis for the generation of transgenic cells in tissue culture, the production of germline transgenic animals for basic and applied research, forward genetic screens for functional gene annotation in model species, and therapy of genetic disorders in humans. Sleeping Beauty (SB) was the first transposon shown to be capable of gene transfer in vertebrate cells, and recent results confirm that SB supports a full spectrum of genetic engineering including transgenesis, insertional mutagenesis, and therapeutic somatic gene transfer both ex vivo and in vivo. The first clinical application of the SB system will help to validate both the safety and efficacy of this approach. In this review, we describe the major transposon systems currently available (with special emphasis on SB), discuss the various parameters and considerations pertinent to their experimental use, and highlight the state of the art in transposon technology in diverse genetic applications.

  12. Genome-scale metabolic network validation of Shewanella oneidensis using transposon insertion frequency analysis.

    PubMed

    Yang, Hong; Krumholz, Elias W; Brutinel, Evan D; Palani, Nagendra P; Sadowsky, Michael J; Odlyzko, Andrew M; Gralnick, Jeffrey A; Libourel, Igor G L

    2014-09-01

    Transposon mutagenesis, in combination with parallel sequencing, is becoming a powerful tool for en-masse mutant analysis. A probability generating function was used to explain observed miniHimar transposon insertion patterns, and gene essentiality calls were made by transposon insertion frequency analysis (TIFA). TIFA incorporated the observed genome and sequence motif bias of the miniHimar transposon. The gene essentiality calls were compared to: 1) previous genome-wide direct gene-essentiality assignments; and, 2) flux balance analysis (FBA) predictions from an existing genome-scale metabolic model of Shewanella oneidensis MR-1. A three-way comparison between FBA, TIFA, and the direct essentiality calls was made to validate the TIFA approach. The refinement in the interpretation of observed transposon insertions demonstrated that genes without insertions are not necessarily essential, and that genes that contain insertions are not always nonessential. The TIFA calls were in reasonable agreement with direct essentiality calls for S. oneidensis, but agreed more closely with E. coli essentiality calls for orthologs. The TIFA gene essentiality calls were in good agreement with the MR-1 FBA essentiality predictions, and the agreement between TIFA and FBA predictions was substantially better than between the FBA and the direct gene essentiality predictions.

  13. Genome-Scale Metabolic Network Validation of Shewanella oneidensis Using Transposon Insertion Frequency Analysis

    PubMed Central

    Yang, Hong; Krumholz, Elias W.; Brutinel, Evan D.; Palani, Nagendra P.; Sadowsky, Michael J.; Odlyzko, Andrew M.; Gralnick, Jeffrey A.; Libourel, Igor G. L.

    2014-01-01

    Transposon mutagenesis, in combination with parallel sequencing, is becoming a powerful tool for en-masse mutant analysis. A probability generating function was used to explain observed miniHimar transposon insertion patterns, and gene essentiality calls were made by transposon insertion frequency analysis (TIFA). TIFA incorporated the observed genome and sequence motif bias of the miniHimar transposon. The gene essentiality calls were compared to: 1) previous genome-wide direct gene-essentiality assignments; and, 2) flux balance analysis (FBA) predictions from an existing genome-scale metabolic model of Shewanella oneidensis MR-1. A three-way comparison between FBA, TIFA, and the direct essentiality calls was made to validate the TIFA approach. The refinement in the interpretation of observed transposon insertions demonstrated that genes without insertions are not necessarily essential, and that genes that contain insertions are not always nonessential. The TIFA calls were in reasonable agreement with direct essentiality calls for S. oneidensis, but agreed more closely with E. coli essentiality calls for orthologs. The TIFA gene essentiality calls were in good agreement with the MR-1 FBA essentiality predictions, and the agreement between TIFA and FBA predictions was substantially better than between the FBA and the direct gene essentiality predictions. PMID:25233219

  14. Signature tagged mutagenesis in the functional genetic analysis of gastrointestinal pathogens

    PubMed Central

    Cummins, Joanne; Gahan, Cormac G.M.

    2012-01-01

    Signature tagged mutagenesis is a genetic approach that was developed to identify novel bacterial virulence factors. It is a negative selection method in which unique identification tags allow analysis of pools of mutants in mixed populations. The approach is particularly well suited to functional genetic analysis of the gastrointestinal phase of infection in foodborne pathogens and has the capacity to guide the development of novel vaccines and therapeutics. In this review we outline the technical principles underpinning signature-tagged mutagenesis as well as novel sequencing-based approaches for transposon mutant identification such as TraDIS (transposon directed insertion-site sequencing). We also provide an analysis of screens that have been performed in gastrointestinal pathogens which are a global health concern (Escherichia coli, Listeria monocytogenes, Helicobacter pylori, Vibrio cholerae and Salmonella enterica). The identification of key virulence loci through the use of signature tagged mutagenesis in mice and relevant larger animal models is discussed. PMID:22555467

  15. Lethal mutagenesis of viruses.

    PubMed

    Perales, Celia; Martín, Verónica; Domingo, Esteban

    2011-11-01

    Lethal mutagenesis aims at extinguishing viruses by increased mutagenesis prompted by virus-specific mutagenic agents, mainly nucleoside analogues. It is derived from the error threshold relationship of quasispecies theory, and it is slowly finding its way towards a clinical application. We summarize the current situation of research in this field of antiviral therapy. Copyright © 2011 Elsevier B.V. All rights reserved.

  16. Using Web-Based Testing for Large-Scale Assessment.

    ERIC Educational Resources Information Center

    Hamilton, Laura S.; Klein, Stephen P.; Lorie, William

    This paper describes an approach to large-scale assessment that uses tests that are delivered to students over the Internet and that are tailored (adapted) to each student's own level of proficiency. A brief background on large-scale assessment is followed by a description of this new technology and an example. Issues that need to be investigated…

  17. Tyrosine Recombinase Retrotransposons and Transposons.

    PubMed

    Poulter, Russell T M; Butler, Margi I

    2015-04-01

    Retrotransposons carrying tyrosine recombinases (YR) are widespread in eukaryotes. The first described tyrosine recombinase mobile element, DIRS1, is a retroelement from the slime mold Dictyostelium discoideum. The YR elements are bordered by terminal repeats related to their replication via free circular dsDNA intermediates. Site-specific recombination is believed to integrate the circle without creating duplications of the target sites. Recently a large number of YR retrotransposons have been described, including elements from fungi (mucorales and basidiomycetes), plants (green algae) and a wide range of animals including nematodes, insects, sea urchins, fish, amphibia and reptiles. YR retrotransposons can be divided into three major groups: the DIRS elements, PAT-like and the Ngaro elements. The three groups form distinct clades on phylogenetic trees based on alignments of reverse transcriptase/ribonuclease H (RT/RH) and YR sequences, and also having some structural distinctions. A group of eukaryote DNA transposons, cryptons, also carry tyrosine recombinases. These DNA transposons do not encode a reverse transcriptase. They have been detected in several pathogenic fungi and oomycetes. Sequence comparisons suggest that the crypton YRs are related to those of the YR retrotransposons. We suggest that the YR retrotransposons arose from the combination of a crypton-like YR DNA transposon and the RT/RH encoding sequence of a retrotransposon. This acquisition must have occurred at a very early point in the evolution of eukaryotes.

  18. Transposon tagging of disease resistance genes. Progress report, May 1, 1988--1992

    SciTech Connect

    Michelmore, R.

    1994-06-01

    Our goal is to clone genes in lettuce determining resistance to downy mildew. One approach involves the mobilization of transposons into resistance genes to mutate and tag the target gene. Because transposons have yet to be isolated and characterized from lettuce, the majority of our experiments have involved Ac from corn as this is increasingly the best characterized transposon. Over the past several years, various labs have contributed to a detailed understanding of the biology of Ac in corn and heterologous plant species. We have collaborated closely with several of these labs, exchanged materials and incorporated their advances into our analysis of transposition in lettuce. The original proposal described the development of a transposon mutagenesis system for lettuce and its subsequent use to tag disease resistance genes. The development phase involved characterization and manipulation of Ac transposition, identification of suitable whole plant selectable markers for the construction of chimeric non-autonomous elements, and investigation of the stability of resistance genes. Investigation of Ac transposition in lettuce has received the majority of our attention. Initially, we made a simple construct with wildtype Ac and introduced it into lettuce. No transposition was observed; although other labs demonstrated that the same construct was functional in tomato. We then focused on assaying for Ac transposition with constructs of increasing sophistication that had been demonstrated by others to be functional in other species. The latest constructs for transposon mutagenesis clearly demonstrated transposition in lettuce. This allowed us to generate seed stocks that we will start to screen for insertional inactivation of resistance genes this year.

  19. Signature-tagged mutagenesis of Vibrio vulnificus

    PubMed Central

    YAMAMOTO, Mai; KASHIMOTO, Takashige; TONG, Ping; XIAO, Jianbo; SUGIYAMA, Michiko; INOUE, Miyuki; MATSUNAGA, Rie; HOSOHARA, Kohei; NAKATA, Kazue; YOKOTA, Kenji; OGUMA, Keiji; YAMAMOTO, Koichiro

    2015-01-01

    Vibrio vulnificus is the causative agent of primary septicemia, wound infection and gastroenteritis in immunocompromised people. In this study, signature-tagged mutagenesis (STM) was applied to identify the virulence genes of V. vulnificus. Using STM, 6,480 mutants in total were constructed and divided into 81 sets (INPUT pools); each mutant in a set was assigned a different tag. Each INPUT pool was intraperitoneally injected into iron-overloaded mice, and in vivo surviving mutants were collected from blood samples from the heart (OUTPUT pools). From the genomic DNA of mixed INPUT or OUTPUT pools, digoxigenin-labeled DNA probes against the tagged region were prepared and used for dot hybridization. Thirty tentatively attenuated mutants, which were hybridized clearly with INPUT probes but barely with OUTPUT probes, were negatively selected. Lethal doses of 11 of the 30 mutants were reduced to more than 1/100; of these, the lethal doses of 2 were reduced to as low as 1/100,000. Transposon-inserted genes in the 11 attenuated mutants were those for IMP dehydrogenase, UDP-N-acetylglucosamine-2-epimerase, aspartokinase, phosphoribosylformylglycinamidine cyclo-ligase, malate Na (+) symporter and hypothetical protein. When mice were immunized with an attenuated mutant strain into which IMP dehydrogenase had been inserted with a transposon, they were protected against V. vulnificus infection. In this study, we demonstrated that the STM method can be used to search for the virulence genes of V. vulnificus. PMID:25755021

  20. Distribution probability of large-scale landslides in central Nepal

    NASA Astrophysics Data System (ADS)

    Timilsina, Manita; Bhandary, Netra P.; Dahal, Ranjan Kumar; Yatabe, Ryuichi

    2014-12-01

    Large-scale landslides in the Himalaya are defined as huge, deep-seated landslide masses that occurred in the geological past. They are widely distributed in the Nepal Himalaya. The steep topography and high local relief provide high potential for such failures, whereas the dynamic geology and adverse climatic conditions play a key role in the occurrence and reactivation of such landslides. The major geoscientific problems related with such large-scale landslides are 1) difficulties in their identification and delineation, 2) sources of small-scale failures, and 3) reactivation. Only a few scientific publications have been published concerning large-scale landslides in Nepal. In this context, the identification and quantification of large-scale landslides and their potential distribution are crucial. Therefore, this study explores the distribution of large-scale landslides in the Lesser Himalaya. It provides simple guidelines to identify large-scale landslides based on their typical characteristics and using a 3D schematic diagram. Based on the spatial distribution of landslides, geomorphological/geological parameters and logistic regression, an equation of large-scale landslide distribution is also derived. The equation is validated by applying it to another area. For the new area, the area under the receiver operating curve of the landslide distribution probability in the new area is 0.699, and a distribution probability value could explain > 65% of existing landslides. Therefore, the regression equation can be applied to areas of the Lesser Himalaya of central Nepal with similar geological and geomorphological conditions.

  1. Organised convection embedded in a large-scale flow

    NASA Astrophysics Data System (ADS)

    Naumann, Ann Kristin; Stevens, Bjorn; Hohenegger, Cathy

    2017-04-01

    In idealised simulations of radiative convective equilibrium, convection aggregates spontaneously from randomly distributed convective cells into organized mesoscale convection despite homogeneous boundary conditions. Although these simulations apply very idealised setups, the process of self-aggregation is thought to be relevant for the development of tropical convective systems. One feature that idealised simulations usually neglect is the occurrence of a large-scale background flow. In the tropics, organised convection is embedded in a large-scale circulation system, which advects convection in along-wind direction and alters near surface convergence in the convective areas. A large-scale flow also modifies the surface fluxes, which are expected to be enhanced upwind of the convective area if a large-scale flow is applied. Convective clusters that are embedded in a large-scale flow therefore experience an asymmetric component of the surface fluxes, which influences the development and the pathway of a convective cluster. In this study, we use numerical simulations with explicit convection and add a large-scale flow to the established setup of radiative convective equilibrium. We then analyse how aggregated convection evolves when being exposed to wind forcing. The simulations suggest that convective line structures are more prevalent if a large-scale flow is present and that convective clusters move considerably slower than advection by the large-scale flow would suggest. We also study the asymmetric component of convective aggregation due to enhanced surface fluxes, and discuss the pathway and speed of convective clusters as a function of the large-scale wind speed.

  2. Maize-targeted mutagenesis: A knockout resource for maize.

    PubMed

    May, Bruce P; Liu, Hong; Vollbrecht, Erik; Senior, Lynn; Rabinowicz, Pablo D; Roh, Donna; Pan, Xiaokang; Stein, Lincoln; Freeling, Mike; Alexander, Danny; Martienssen, Rob

    2003-09-30

    We describe an efficient system for site-selected transposon mutagenesis in maize. A total of 43,776 F1 plants were generated by using Robertson's Mutator (Mu) pollen parents and self-pollinated to establish a library of transposon-mutagenized seed. The frequency of new seed mutants was between 10-4 and 10-5 per F1 plant. As a service to the maize community, maize-targeted mutagenesis selects insertions in genes of interest from this library by using the PCR. Pedigree, knockout, sequence, phenotype, and other information is stored in a powerful interactive database (maize-targeted mutagenesis database) that enables analysis of the entire population and the handling of knockout requests. By inhibiting Mu activity in most F1 plants, we sought to reduce somatic insertions that may cause false positives selected from pooled tissue. By monitoring the remaining Mu activity in the F2, however, we demonstrate that seed phenotypes depend on it, and false positives occur in lines that appear to lack it. We conclude that more than half of all mutations arising in this population are suppressed on losing Mu activity. These results have implications for epigenetic models of inbreeding and for functional genomics.

  3. Massive parallel insertion site sequencing of an arrayed Sinorhizobium meliloti signature-tagged mini-Tn 5 transposon mutant library.

    PubMed

    Serrania, Javier; Johner, Tobias; Rupp, Oliver; Goesmann, Alexander; Becker, Anke

    2017-02-21

    Transposon mutagenesis in conjunction with identification of genomic transposon insertion sites is a powerful tool for gene function studies. We have implemented a protocol for parallel determination of transposon insertion sites by Illumina sequencing involving a hierarchical barcoding method that allowed for tracking back insertion sites to individual clones of an arrayed signature-tagged transposon mutant library. This protocol was applied to further characterize a signature-tagged mini-Tn 5 mutant library comprising about 12,000 mutants of the symbiotic nitrogen-fixing alphaproteobacterium Sinorhizobium meliloti (Pobigaylo et al., 2006; Appl. Environ. Microbiol. 72, 4329-4337). Previously, insertion sites have been determined for 5000 mutants of this library. Combining an adapter-free, inverse PCR method for sequencing library preparation with next generation sequencing, we identified 4473 novel insertion sites, increasing the total number of transposon mutants with known insertion site to 9562. The number of protein-coding genes that were hit at least once by a transposon increased by 1231 to a total number of 3673 disrupted genes, which represents 59% of the predicted protein-coding genes in S. meliloti.

  4. Enhancer trapping in zebrafish using the Sleeping Beauty transposon

    PubMed Central

    Balciunas, Darius; Davidson, Ann E; Sivasubbu, Sridhar; Hermanson, Spencer B; Welle, Zachary; Ekker, Stephen C

    2004-01-01

    Background Among functional elements of a metazoan gene, enhancers are particularly difficult to find and annotate. Pioneering experiments in Drosophila have demonstrated the value of enhancer "trapping" using an invertebrate to address this functional genomics problem. Results We modulated a Sleeping Beauty transposon-based transgenesis cassette to establish an enhancer trapping technique for use in a vertebrate model system, zebrafish Danio rerio. We established 9 lines of zebrafish with distinct tissue- or organ-specific GFP expression patterns from 90 founders that produced GFP-expressing progeny. We have molecularly characterized these lines and show that in each line, a specific GFP expression pattern is due to a single transposition event. Many of the insertions are into introns of zebrafish genes predicted in the current genome assembly. We have identified both previously characterized as well as novel expression patterns from this screen. For example, the ET7 line harbors a transposon insertion near the mkp3 locus and expresses GFP in the midbrain-hindbrain boundary, forebrain and the ventricle, matching a subset of the known FGF8-dependent mkp3 expression domain. The ET2 line, in contrast, expresses GFP specifically in caudal primary motoneurons due to an insertion into the poly(ADP-ribose) glycohydrolase (PARG) locus. This surprising expression pattern was confirmed using in situ hybridization techniques for the endogenous PARG mRNA, indicating the enhancer trap has replicated this unexpected and highly localized PARG expression with good fidelity. Finally, we show that it is possible to excise a Sleeping Beauty transposon from a genomic location in the zebrafish germline. Conclusions This genomics tool offers the opportunity for large-scale biological approaches combining both expression and genomic-level sequence analysis using as a template an entire vertebrate genome. PMID:15347431

  5. Needs, opportunities, and options for large scale systems research

    SciTech Connect

    Thompson, G.L.

    1984-10-01

    The Office of Energy Research was recently asked to perform a study of Large Scale Systems in order to facilitate the development of a true large systems theory. It was decided to ask experts in the fields of electrical engineering, chemical engineering and manufacturing/operations research for their ideas concerning large scale systems research. The author was asked to distribute a questionnaire among these experts to find out their opinions concerning recent accomplishments and future research directions in large scale systems research. He was also requested to convene a conference which included three experts in each area as panel members to discuss the general area of large scale systems research. The conference was held on March 26--27, 1984 in Pittsburgh with nine panel members, and 15 other attendees. The present report is a summary of the ideas presented and the recommendations proposed by the attendees.

  6. Modified gravity and large scale flows, a review

    NASA Astrophysics Data System (ADS)

    Mould, Jeremy

    2017-02-01

    Large scale flows have been a challenging feature of cosmography ever since galaxy scaling relations came on the scene 40 years ago. The next generation of surveys will offer a serious test of the standard cosmology.

  7. Learning networks for sustainable, large-scale improvement.

    PubMed

    McCannon, C Joseph; Perla, Rocco J

    2009-05-01

    Large-scale improvement efforts known as improvement networks offer structured opportunities for exchange of information and insights into the adaptation of clinical protocols to a variety of settings.

  8. Amplification of large-scale magnetic field in nonhelical magnetohydrodynamics

    NASA Astrophysics Data System (ADS)

    Kumar, Rohit; Verma, Mahendra K.

    2017-09-01

    It is typically assumed that the kinetic and magnetic helicities play a crucial role in the growth of large-scale dynamo. In this paper, we demonstrate that helicity is not essential for the amplification of large-scale magnetic field. For this purpose, we perform nonhelical magnetohydrodynamic (MHD) simulation, and show that the large-scale magnetic field can grow in nonhelical MHD when random external forcing is employed at scale 1/10 the box size. The energy fluxes and shell-to-shell transfer rates computed using the numerical data show that the large-scale magnetic energy grows due to the energy transfers from the velocity field at the forcing scales.

  9. An Adaptive Multiscale Finite Element Method for Large Scale Simulations

    DTIC Science & Technology

    2015-09-28

    the method . Using the above definitions , the weak statement of the non-linear local problem at the kth 4 DISTRIBUTION A: Distribution approved for...AFRL-AFOSR-VA-TR-2015-0305 An Adaptive Multiscale Finite Element Method for Large Scale Simulations Carlos Duarte UNIVERSITY OF ILLINOIS CHAMPAIGN...14-07-2015 4. TITLE AND SUBTITLE An Adaptive Multiscale Generalized Finite Element Method for Large Scale Simulations 5a.  CONTRACT NUMBER 5b

  10. Large-scale studies of marked birds in North America

    USGS Publications Warehouse

    Tautin, J.; Metras, L.; Smith, G.

    1999-01-01

    The first large-scale, co-operative, studies of marked birds in North America were attempted in the 1950s. Operation Recovery, which linked numerous ringing stations along the east coast in a study of autumn migration of passerines, and the Preseason Duck Ringing Programme in prairie states and provinces, conclusively demonstrated the feasibility of large-scale projects. The subsequent development of powerful analytical models and computing capabilities expanded the quantitative potential for further large-scale projects. Monitoring Avian Productivity and Survivorship, and Adaptive Harvest Management are current examples of truly large-scale programmes. Their exemplary success and the availability of versatile analytical tools are driving changes in the North American bird ringing programme. Both the US and Canadian ringing offices are modifying operations to collect more and better data to facilitate large-scale studies and promote a more project-oriented ringing programme. New large-scale programmes such as the Cornell Nest Box Network are on the horizon.

  11. A study of MLFMA for large-scale scattering problems

    NASA Astrophysics Data System (ADS)

    Hastriter, Michael Larkin

    This research is centered in computational electromagnetics with a focus on solving large-scale problems accurately in a timely fashion using first principle physics. Error control of the translation operator in 3-D is shown. A parallel implementation of the multilevel fast multipole algorithm (MLFMA) was studied as far as parallel efficiency and scaling. The large-scale scattering program (LSSP), based on the ScaleME library, was used to solve ultra-large-scale problems including a 200lambda sphere with 20 million unknowns. As these large-scale problems were solved, techniques were developed to accurately estimate the memory requirements. Careful memory management is needed in order to solve these massive problems. The study of MLFMA in large-scale problems revealed significant errors that stemmed from inconsistencies in constants used by different parts of the algorithm. These were fixed to produce the most accurate data possible for large-scale surface scattering problems. Data was calculated on a missile-like target using both high frequency methods and MLFMA. This data was compared and analyzed to determine possible strategies to increase data acquisition speed and accuracy through multiple computation method hybridization.

  12. Random mutagenesis of the hyperthermophilic archaeon Pyrococcus furiosus using in vitro mariner transposition and natural transformation

    PubMed Central

    Guschinskaya, Natalia; Brunel, Romain; Tourte, Maxime; Lipscomb, Gina L.; Adams, Michael W. W.; Oger, Philippe; Charpentier, Xavier

    2016-01-01

    Transposition mutagenesis is a powerful tool to identify the function of genes, reveal essential genes and generally to unravel the genetic basis of living organisms. However, transposon-mediated mutagenesis has only been successfully applied to a limited number of archaeal species and has never been reported in Thermococcales. Here, we report random insertion mutagenesis in the hyperthermophilic archaeon Pyrococcus furiosus. The strategy takes advantage of the natural transformability of derivatives of the P. furiosus COM1 strain and of in vitro Mariner-based transposition. A transposon bearing a genetic marker is randomly transposed in vitro in genomic DNA that is then used for natural transformation of P. furiosus. A small-scale transposition reaction routinely generates several hundred and up to two thousands transformants. Southern analysis and sequencing showed that the obtained mutants contain a single and random genomic insertion. Polyploidy has been reported in Thermococcales and P. furiosus is suspected of being polyploid. Yet, about half of the mutants obtained on the first selection are homozygous for the transposon insertion. Two rounds of isolation on selective medium were sufficient to obtain gene conversion in initially heterozygous mutants. This transposition mutagenesis strategy will greatly facilitate functional exploration of the Thermococcales genomes. PMID:27824140

  13. First Streptococcus pyogenes signature-tagged mutagenesis screen identifies novel virulence determinants.

    PubMed

    Kizy, Anne E; Neely, Melody N

    2009-05-01

    The virulence of bacterial pathogens is a complex process that requires the dynamic expression of many genes for the pathogens to invade and circumvent host defenses, as well as to proliferate in vivo. In this study, we employed a large-scale screen, signature-tagged mutagenesis (STM), to identify Streptococcus pyogenes virulence genes important for pathogenesis within the host. Approximately 1,200 STM mutants were created and screened using the zebrafish infectious disease model. The transposon insertion site was identified for 29 of the 150 mutants that were considered attenuated for virulence. Previously reported streptococcal virulence genes, such as mga, hasA, amrA, smeZ, and two genes in the sil locus, were identified, confirming the utility of the model for revealing genes important for virulence. Multiple genes not previously implicated in virulence were also identified, including genes encoding putative transporters, hypothetical cytosolic proteins, and macrolide efflux pumps. The STM mutant strains display various levels of attenuation, and multiple separate insertions were identified in either the same gene or the same locus, suggesting that these factors are important for this type of acute, invasive infection. We further examined two such genes, silB and silC of a putative quorum-sensing regulon, and determined that they are significant virulence factors in our model of necrotizing fasciitis. sil locus promoter expression was examined under various in vitro conditions, as well as in zebrafish tissues, and was found to be differentially induced. This study was a unique investigation of S. pyogenes factors required for successful invasive infection.

  14. Generation of an inducible and optimized piggyBac transposon system†

    PubMed Central

    Cadiñanos, Juan; Bradley, Allan

    2007-01-01

    Genomic studies in the mouse have been slowed by the lack of transposon-mediated mutagenesis. However, since the resurrection of Sleeping Beauty (SB), the possibility of performing forward genetics in mice has been reinforced. Recently, piggyBac (PB), a functional transposon from insects, was also described to work in mammals. As the activity of PB is higher than that of SB11 and SB12, two hyperactive SB transposases, we have characterized and improved the PB system in mouse ES cells. We have generated a mouse codon-optimized version of the PB transposase coding sequence (CDS) which provides transposition levels greater than the original. We have also found that the promoter sequence predicted in the 5′-terminal repeat of the PB transposon is active in the mammalian context. Finally, we have engineered inducible versions of the optimized piggyBac transposase fused with ERT2. One of them, when induced, provides higher levels of transposition than the native piggyBac CDS, whereas in the absence of induction its activity is indistinguishable from background. We expect that these tools, adaptable to perform mouse-germline mutagenesis, will facilitate the identification of genes involved in pathological and physiological processes, such as cancer or ES cell differentiation. PMID:17576687

  15. Chemical mutagens, transposons, and transgenes to interrogate gene function in Drosophila melanogaster.

    PubMed

    Venken, Koen J T; Bellen, Hugo J

    2014-06-15

    The study of genetics, genes, and chromosomal inheritance was initiated by Thomas Morgan in 1910, when the first visible mutations were identified in fruit flies. The field expanded upon the work initiated by Herman Muller in 1926 when he used X-rays to develop the first balancer chromosomes. Today, balancers are still invaluable to maintain mutations and transgenes but the arsenal of tools has expanded vastly and numerous new methods have been developed, many relying on the availability of the genome sequence and transposable elements. Forward genetic screens based on chemical mutagenesis or transposable elements have resulted in the unbiased identification of many novel players involved in processes probed by specific phenotypic assays. Reverse genetic approaches have relied on the availability of a carefully selected set of transposon insertions spread throughout the genome to allow the manipulation of the region in the vicinity of each insertion. Lastly, the ability to transform Drosophila with single copy transgenes using transposons or site-specific integration using the ΦC31 integrase has allowed numerous manipulations, including the ability to create and integrate genomic rescue constructs, generate duplications, RNAi knock-out technology, binary expression systems like the GAL4/UAS system as well as other methods. Here, we will discuss the most useful methodologies to interrogate the fruit fly genome in vivo focusing on chemical mutagenesis, transposons and transgenes. Genome engineering approaches based on nucleases and RNAi technology are discussed in following chapters.

  16. Chemical Mutagens, Transposons, and Transgenes to Interrogate Gene Function in Drosophila melanogaster

    PubMed Central

    Venken, Koen J.T.; Bellen, Hugo J.

    2014-01-01

    The study of genetics, genes, and chromosomal inheritance was initiated by Thomas Morgan in when the first visible mutations were identified in fruit flies. The field expanded upon the work initiated by Herman Muller in 1926 when he used X-rays to develop the first balancer chromosomes. Today, balancers are still invaluable to maintain mutations and transgenes but the arsenal of tools has expanded vastly and numerous new methods have been developed, many relying on the availability of the genome sequence and transposable elements. Forward genetic screens based on chemical mutagenesis or transposable elements have resulted in the unbiased identification of many novel players involved in processes probed by specific phenotypic assays. Reverse genetic approaches have relied on the availability of a carefully selected set of transposon insertions spread throughout the genome to allow the manipulation of the region in the vicinity of each insertion. Lastly, the ability to transform Drosophila with single copy transgenes using transposons or site-specific integration using the ΦC31 integrase has allowed numerous manipulations, including the ability to create and integrate genomic rescue constructs, generate duplications, RNAi knock-out technology, binary expression systems like the GAL4/UAS system as well as other methods. Here, we will discuss the most useful methodologies to interrogate the fruit fly genome in vivo focusing on chemical mutagenesis, transposons and transgenes. Genome engineering approaches based on nucleases and RNAi technology are discussed in following chapters. PMID:24583113

  17. Large-scale filament formation inhibits the activity of CTP synthetase

    PubMed Central

    Barry, Rachael M; Bitbol, Anne-Florence; Lorestani, Alexander; Charles, Emeric J; Habrian, Chris H; Hansen, Jesse M; Li, Hsin-Jung; Baldwin, Enoch P; Wingreen, Ned S; Kollman, Justin M; Gitai, Zemer

    2014-01-01

    CTP Synthetase (CtpS) is a universally conserved and essential metabolic enzyme. While many enzymes form small oligomers, CtpS forms large-scale filamentous structures of unknown function in prokaryotes and eukaryotes. By simultaneously monitoring CtpS polymerization and enzymatic activity, we show that polymerization inhibits activity, and CtpS's product, CTP, induces assembly. To understand how assembly inhibits activity, we used electron microscopy to define the structure of CtpS polymers. This structure suggests that polymerization sterically hinders a conformational change necessary for CtpS activity. Structure-guided mutagenesis and mathematical modeling further indicate that coupling activity to polymerization promotes cooperative catalytic regulation. This previously uncharacterized regulatory mechanism is important for cellular function since a mutant that disrupts CtpS polymerization disrupts E. coli growth and metabolic regulation without reducing CTP levels. We propose that regulation by large-scale polymerization enables ultrasensitive control of enzymatic activity while storing an enzyme subpopulation in a conformationally restricted form that is readily activatable. DOI: http://dx.doi.org/10.7554/eLife.03638.001 PMID:25030911

  18. Large-scale filament formation inhibits the activity of CTP synthetase.

    PubMed

    Barry, Rachael M; Bitbol, Anne-Florence; Lorestani, Alexander; Charles, Emeric J; Habrian, Chris H; Hansen, Jesse M; Li, Hsin-Jung; Baldwin, Enoch P; Wingreen, Ned S; Kollman, Justin M; Gitai, Zemer

    2014-07-16

    CTP Synthetase (CtpS) is a universally conserved and essential metabolic enzyme. While many enzymes form small oligomers, CtpS forms large-scale filamentous structures of unknown function in prokaryotes and eukaryotes. By simultaneously monitoring CtpS polymerization and enzymatic activity, we show that polymerization inhibits activity, and CtpS's product, CTP, induces assembly. To understand how assembly inhibits activity, we used electron microscopy to define the structure of CtpS polymers. This structure suggests that polymerization sterically hinders a conformational change necessary for CtpS activity. Structure-guided mutagenesis and mathematical modeling further indicate that coupling activity to polymerization promotes cooperative catalytic regulation. This previously uncharacterized regulatory mechanism is important for cellular function since a mutant that disrupts CtpS polymerization disrupts E. coli growth and metabolic regulation without reducing CTP levels. We propose that regulation by large-scale polymerization enables ultrasensitive control of enzymatic activity while storing an enzyme subpopulation in a conformationally restricted form that is readily activatable.

  19. Fungicide-induced transposon movement in Monilinia fructicola.

    PubMed

    Chen, Fengping; Everhart, Sydney E; Bryson, P Karen; Luo, Chaoxi; Song, Xi; Liu, Xili; Schnabel, Guido

    2015-12-01

    Repeated applications of fungicides with a single mode of action are believed to select for pre-existing resistant strains in a pathogen population, while the impact of sub-lethal doses of such fungicides on sensitive members of the population is unknown. In this study, in vitro evidence is presented that continuous exposure of Monilinia fructicola mycelium to some fungicides can induce genetic change in form of transposon transposition. Three fungicide-sensitive M. fructicola isolates were exposed in 12 weekly transfers of mycelia to a dose gradient of demethylation inhibitor fungicide (DMI) SYP-Z048 and quinone outside inhibitor fungicide (QoI) azoxystrobin in solo or mixture treatments. Evidence of mutagenesis was assessed by monitoring Mftc1, a multicopy transposable element of M. fructicola, by PCR and Southern blot analysis. Movement of Mftc1 was observed following azoxystrobin and azoxystrobin plus SYP-Z048 treatments in two of the three isolates, but not in the non-fungicide-treated controls. Interestingly, the upstream promoter region of MfCYP51 was a prime target for Mftc1 transposition in these isolates. Transposition of Mftc1 was verified by Southern blot in two of three isolates from another, similar experiment following prolonged, sublethal azoxystrobin exposure, although in these isolates movement of Mftc1 in the upstream MfCYP51 promoter region was not observed. More research is warranted to determine whether fungicide-induced mutagenesis may also happen under field conditions. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Generation of non-viral, transgene-free hepatocyte like cells with piggyBac transposon

    PubMed Central

    Katayama, Hokahiro; Yasuchika, Kentaro; Miyauchi, Yuya; Kojima, Hidenobu; Yamaoka, Ryoya; Kawai, Takayuki; Yukie Yoshitoshi, Elena; Ogiso, Satoshi; Kita, Sadahiko; Yasuda, Katsutaro; Sasaki, Naoya; Fukumitsu, Ken; Komori, Junji; Ishii, Takamichi; Uemoto, Shinji

    2017-01-01

    Somatic cells can be reprogrammed to induced hepatocyte-like cells (iHeps) by overexpressing certain defined factors in direct reprogramming techniques. Of the various methods to deliver genes into cells, typically used genome-integrating viral vectors are associated with integration-related adverse events such as mutagenesis, whereas non-integrating viral vectors have low efficiency, making viral vectors unsuitable for clinical application. Therefore, we focused on developing a transposon system to establish a non-viral reprogramming method. Transposons are unique DNA elements that can be integrated into and removed from chromosomes. PiggyBac, a type of transposon, has high transduction efficiency and cargo capacity, and the integrated transgene can be precisely excised in the presence of transposase. This feature enables the piggyBac vector to achieve efficient transgene expression and a transgene-free state, thus making it a promising method for cell reprogramming. Here, we attempted to utilize the piggyBac transposon system to generate iHeps by integrating a transgene consisting of Hnf4a and Foxa3, and successfully obtained functional iHeps. We then demonstrated removal of the transgene to obtain transgene-free iHeps, which still maintained hepatocyte functions. This non-viral, transgene-free reprogramming method using the piggyBac vector may facilitate clinical applications of iHeps in upcoming cell therapy. PMID:28295042

  1. EINSTEIN'S SIGNATURE IN COSMOLOGICAL LARGE-SCALE STRUCTURE

    SciTech Connect

    Bruni, Marco; Hidalgo, Juan Carlos; Wands, David

    2014-10-10

    We show how the nonlinearity of general relativity generates a characteristic nonGaussian signal in cosmological large-scale structure that we calculate at all perturbative orders in a large-scale limit. Newtonian gravity and general relativity provide complementary theoretical frameworks for modeling large-scale structure in ΛCDM cosmology; a relativistic approach is essential to determine initial conditions, which can then be used in Newtonian simulations studying the nonlinear evolution of the matter density. Most inflationary models in the very early universe predict an almost Gaussian distribution for the primordial metric perturbation, ζ. However, we argue that it is the Ricci curvature of comoving-orthogonal spatial hypersurfaces, R, that drives structure formation at large scales. We show how the nonlinear relation between the spatial curvature, R, and the metric perturbation, ζ, translates into a specific nonGaussian contribution to the initial comoving matter density that we calculate for the simple case of an initially Gaussian ζ. Our analysis shows the nonlinear signature of Einstein's gravity in large-scale structure.

  2. Recursive architecture for large-scale adaptive system

    NASA Astrophysics Data System (ADS)

    Hanahara, Kazuyuki; Sugiyama, Yoshihiko

    1994-09-01

    'Large scale' is one of major trends in the research and development of recent engineering, especially in the field of aerospace structural system. This term expresses the large scale of an artifact in general, however, it also implies the large number of the components which make up the artifact in usual. Considering a large scale system which is especially used in remote space or deep-sea, such a system should be adaptive as well as robust by itself, because its control as well as maintenance by human operators are not easy due to the remoteness. An approach to realizing this large scale, adaptive and robust system is to build the system as an assemblage of components which are respectively adaptive by themselves. In this case, the robustness of the system can be achieved by using a large number of such components and suitable adaptation as well as maintenance strategies. Such a system gathers many research's interest and their studies such as decentralized motion control, configurating algorithm and characteristics of structural elements are reported. In this article, a recursive architecture concept is developed and discussed towards the realization of large scale system which consists of a number of uniform adaptive components. We propose an adaptation strategy based on the architecture and its implementation by means of hierarchically connected processing units. The robustness and the restoration from degeneration of the processing unit are also discussed. Two- and three-dimensional adaptive truss structures are conceptually designed based on the recursive architecture.

  3. The Influence of Large-scale Environments on Galaxy Properties

    NASA Astrophysics Data System (ADS)

    Wei, Yu-qing; Wang, Lei; Dai, Cai-ping

    2017-07-01

    The star formation properties of galaxies and their dependence on environments play an important role for understanding the formation and evolution of galaxies. Using the galaxy sample of the Sloan Digital Sky Survey (SDSS), different research groups have studied the physical properties of galaxies and their large-scale environments. Here, using the filament catalog from Tempel et al. and the galaxy catalog of large-scale structure classification from Wang et al., and taking the influence of the galaxy morphology, high/low local density environment, and central (satellite) galaxy into consideration, we have found that the properties of galaxies are correlated with their residential large-scale environments: the SSFR (specific star formation rate) and SFR (star formation rate) strongly depend on the large-scale environment for spiral galaxies and satellite galaxies, but this dependence is very weak for elliptical galaxies and central galaxies, and the influence of large-scale environments on galaxies in low density region is more sensitive than that in high density region. The above conclusions remain valid even for the galaxies with the same mass. In addition, the SSFR distributions derived from the catalogs of Tempel et al. and Wang et al. are not entirely consistent.

  4. High-Throughput Parallel Sequencing to Measure Fitness of Leptospira interrogans Transposon Insertion Mutants during Acute Infection

    PubMed Central

    Matsunaga, James; Haake, David A.

    2016-01-01

    Pathogenic species of Leptospira are the causative agents of leptospirosis, a zoonotic disease that causes mortality and morbidity worldwide. The understanding of the virulence mechanisms of Leptospira spp is still at an early stage due to the limited number of genetic tools available for this microorganism. The development of random transposon mutagenesis in pathogenic strains a decade ago has contributed to the identification of several virulence factors. In this study, we used the transposon sequencing (Tn-Seq) technique, which combines transposon mutagenesis with massive parallel sequencing, to study the in vivo fitness of a pool of Leptospira interrogans mutants. We infected hamsters with a pool of 42 mutants (input pool), which included control mutants with insertions in four genes previously analyzed by virulence testing (loa22, ligB, flaA1, and lic20111) and 23 mutants with disrupted signal transduction genes. We quantified the mutants in different tissues (blood, kidney and liver) at 4 days post-challenge by high-throughput sequencing and compared the frequencies of mutants recovered from tissues to their frequencies in the input pool. Control mutants that were less fit in the Tn-Seq experiment were attenuated for virulence when tested separately in the hamster model of lethal leptospirosis. Control mutants with unaltered fitness were as virulent as the wild-type strain. We identified two mutants with the transposon inserted in the same putative adenylate/guanylate cyclase gene (lic12327) that had reduced in vivo fitness in blood, kidney and liver. Both lic12327 mutants were attenuated for virulence when tested individually in hamsters. Growth of the control mutants and lic12327 mutants in culture medium were similar to that of the wild-type strain. These results demonstrate the feasibility of screening large pools of L. interrogans transposon mutants for those with altered fitness, and potentially attenuated virulence, by transposon sequencing. PMID

  5. High-Throughput Parallel Sequencing to Measure Fitness of Leptospira interrogans Transposon Insertion Mutants during Acute Infection.

    PubMed

    Lourdault, Kristel; Matsunaga, James; Haake, David A

    2016-11-01

    Pathogenic species of Leptospira are the causative agents of leptospirosis, a zoonotic disease that causes mortality and morbidity worldwide. The understanding of the virulence mechanisms of Leptospira spp is still at an early stage due to the limited number of genetic tools available for this microorganism. The development of random transposon mutagenesis in pathogenic strains a decade ago has contributed to the identification of several virulence factors. In this study, we used the transposon sequencing (Tn-Seq) technique, which combines transposon mutagenesis with massive parallel sequencing, to study the in vivo fitness of a pool of Leptospira interrogans mutants. We infected hamsters with a pool of 42 mutants (input pool), which included control mutants with insertions in four genes previously analyzed by virulence testing (loa22, ligB, flaA1, and lic20111) and 23 mutants with disrupted signal transduction genes. We quantified the mutants in different tissues (blood, kidney and liver) at 4 days post-challenge by high-throughput sequencing and compared the frequencies of mutants recovered from tissues to their frequencies in the input pool. Control mutants that were less fit in the Tn-Seq experiment were attenuated for virulence when tested separately in the hamster model of lethal leptospirosis. Control mutants with unaltered fitness were as virulent as the wild-type strain. We identified two mutants with the transposon inserted in the same putative adenylate/guanylate cyclase gene (lic12327) that had reduced in vivo fitness in blood, kidney and liver. Both lic12327 mutants were attenuated for virulence when tested individually in hamsters. Growth of the control mutants and lic12327 mutants in culture medium were similar to that of the wild-type strain. These results demonstrate the feasibility of screening large pools of L. interrogans transposon mutants for those with altered fitness, and potentially attenuated virulence, by transposon sequencing.

  6. Global analysis of candidate genes important for fitness in a competitive biofilm using DNA-array-based transposon mapping.

    PubMed

    Junker, Lauren M; Peters, Joseph E; Hay, Anthony G

    2006-08-01

    Escherichia coli strain PHL628 was subjected to saturating Tn5 transposon mutagenesis and then grown under competitive planktonic or biofilm conditions. The locations of transposon insertions from the remaining cells were then mapped on a gene array. The results from the array mapping indicated that 4.5 % of the E. coli genome was important under these conditions. Specifically, 114 genes were identified as important for the biofilm lifestyle, whereas 80 genes were important for the planktonic lifestyle. Four broad functional categories were identified as biofilm-important. These included genes encoding cell structures, small-molecule transport, energy metabolism and regulatory functions. For one of these genes, arcA, an insertion mutant was generated and its biofilm-related phenotype was examined. Results from both the transposon array and insertion mutagenesis indicated that arcA, which is known to be a negative response regulator of genes in aerobic pathways, was important for competitiveness in E. coli PHL628 biofilms. This work also demonstrated that ligation-mediated PCR, coupled with array-based transposon mapping, was an effective tool for identifying a large variety of candidate genes that are important for biofilm fitness.

  7. Application of an inducible transposon with anther culture in generation of di-haploid homologous mutants.

    PubMed

    Yang, Hsiu-Chun; Charng, Yuh-Chyang

    2014-12-01

    Insertional mutagenesis represents one of the most effective ways to acquire information about a plant gene's function. However, it is hindered by the autosomal genome being diploid and therefore, most mutations being recessive. The problem is addressed by inducing the transposition during anther culture so that selected mutations can be transmitted and then regenerated to a homozygous state. To this end, we treated transgenic rice floral tissues containing the inducible transposon with an inducer, salicylic acid. Excision events were detected in regenerated calli and subsequent plantlets. DNA blot and PCR assay were used to determine the homogeneity of knockout mutants. About 5% of the mutants containing transposition events were homozygous. Furthermore, the inducible transposon was active during calli regeneration. This strategy could be applicable to improve transposition efficiency in microspore development stages to create stable di-haploid mutants in plants.

  8. Toward Improved Support for Loosely Coupled Large Scale Simulation Workflows

    SciTech Connect

    Boehm, Swen; Elwasif, Wael R; Naughton, III, Thomas J; Vallee, Geoffroy R

    2014-01-01

    High-performance computing (HPC) workloads are increasingly leveraging loosely coupled large scale simula- tions. Unfortunately, most large-scale HPC platforms, including Cray/ALPS environments, are designed for the execution of long-running jobs based on coarse-grained launch capabilities (e.g., one MPI rank per core on all allocated compute nodes). This assumption limits capability-class workload campaigns that require large numbers of discrete or loosely coupled simulations, and where time-to-solution is an untenable pacing issue. This paper describes the challenges related to the support of fine-grained launch capabilities that are necessary for the execution of loosely coupled large scale simulations on Cray/ALPS platforms. More precisely, we present the details of an enhanced runtime system to support this use case, and report on initial results from early testing on systems at Oak Ridge National Laboratory.

  9. Seismic safety in conducting large-scale blasts

    NASA Astrophysics Data System (ADS)

    Mashukov, I. V.; Chaplygin, V. V.; Domanov, V. P.; Semin, A. A.; Klimkin, M. A.

    2017-09-01

    In mining enterprises to prepare hard rocks for excavation a drilling and blasting method is used. With the approach of mining operations to settlements the negative effect of large-scale blasts increases. To assess the level of seismic impact of large-scale blasts the scientific staff of Siberian State Industrial University carried out expertise for coal mines and iron ore enterprises. Determination of the magnitude of surface seismic vibrations caused by mass explosions was performed using seismic receivers, an analog-digital converter with recording on a laptop. The registration results of surface seismic vibrations during production of more than 280 large-scale blasts at 17 mining enterprises in 22 settlements are presented. The maximum velocity values of the Earth’s surface vibrations are determined. The safety evaluation of seismic effect was carried out according to the permissible value of vibration velocity. For cases with exceedance of permissible values recommendations were developed to reduce the level of seismic impact.

  10. PKI security in large-scale healthcare networks.

    PubMed

    Mantas, Georgios; Lymberopoulos, Dimitrios; Komninos, Nikos

    2012-06-01

    During the past few years a lot of PKI (Public Key Infrastructures) infrastructures have been proposed for healthcare networks in order to ensure secure communication services and exchange of data among healthcare professionals. However, there is a plethora of challenges in these healthcare PKI infrastructures. Especially, there are a lot of challenges for PKI infrastructures deployed over large-scale healthcare networks. In this paper, we propose a PKI infrastructure to ensure security in a large-scale Internet-based healthcare network connecting a wide spectrum of healthcare units geographically distributed within a wide region. Furthermore, the proposed PKI infrastructure facilitates the trust issues that arise in a large-scale healthcare network including multi-domain PKI infrastructures.

  11. Acoustic Studies of the Large Scale Ocean Circulation

    NASA Technical Reports Server (NTRS)

    Menemenlis, Dimitris

    1999-01-01

    Detailed knowledge of ocean circulation and its transport properties is prerequisite to an understanding of the earth's climate and of important biological and chemical cycles. Results from two recent experiments, THETIS-2 in the Western Mediterranean and ATOC in the North Pacific, illustrate the use of ocean acoustic tomography for studies of the large scale circulation. The attraction of acoustic tomography is its ability to sample and average the large-scale oceanic thermal structure, synoptically, along several sections, and at regular intervals. In both studies, the acoustic data are compared to, and then combined with, general circulation models, meteorological analyses, satellite altimetry, and direct measurements from ships. Both studies provide complete regional descriptions of the time-evolving, three-dimensional, large scale circulation, albeit with large uncertainties. The studies raise serious issues about existing ocean observing capability and provide guidelines for future efforts.

  12. Large-scale velocity structures in turbulent thermal convection.

    PubMed

    Qiu, X L; Tong, P

    2001-09-01

    A systematic study of large-scale velocity structures in turbulent thermal convection is carried out in three different aspect-ratio cells filled with water. Laser Doppler velocimetry is used to measure the velocity profiles and statistics over varying Rayleigh numbers Ra and at various spatial positions across the whole convection cell. Large velocity fluctuations are found both in the central region and near the cell boundary. Despite the large velocity fluctuations, the flow field still maintains a large-scale quasi-two-dimensional structure, which rotates in a coherent manner. This coherent single-roll structure scales with Ra and can be divided into three regions in the rotation plane: (1) a thin viscous boundary layer, (2) a fully mixed central core region with a constant mean velocity gradient, and (3) an intermediate plume-dominated buffer region. The experiment reveals a unique driving mechanism for the large-scale coherent rotation in turbulent convection.

  13. Large-scale simulations of complex physical systems

    NASA Astrophysics Data System (ADS)

    Belić, A.

    2007-04-01

    Scientific computing has become a tool as vital as experimentation and theory for dealing with scientific challenges of the twenty-first century. Large scale simulations and modelling serve as heuristic tools in a broad problem-solving process. High-performance computing facilities make possible the first step in this process - a view of new and previously inaccessible domains in science and the building up of intuition regarding the new phenomenology. The final goal of this process is to translate this newly found intuition into better algorithms and new analytical results. In this presentation we give an outline of the research themes pursued at the Scientific Computing Laboratory of the Institute of Physics in Belgrade regarding large-scale simulations of complex classical and quantum physical systems, and present recent results obtained in the large-scale simulations of granular materials and path integrals.

  14. Large-scale simulations of complex physical systems

    SciTech Connect

    Belic, A.

    2007-04-23

    Scientific computing has become a tool as vital as experimentation and theory for dealing with scientific challenges of the twenty-first century. Large scale simulations and modelling serve as heuristic tools in a broad problem-solving process. High-performance computing facilities make possible the first step in this process - a view of new and previously inaccessible domains in science and the building up of intuition regarding the new phenomenology. The final goal of this process is to translate this newly found intuition into better algorithms and new analytical results.In this presentation we give an outline of the research themes pursued at the Scientific Computing Laboratory of the Institute of Physics in Belgrade regarding large-scale simulations of complex classical and quantum physical systems, and present recent results obtained in the large-scale simulations of granular materials and path integrals.

  15. A relativistic signature in large-scale structure

    NASA Astrophysics Data System (ADS)

    Bartolo, Nicola; Bertacca, Daniele; Bruni, Marco; Koyama, Kazuya; Maartens, Roy; Matarrese, Sabino; Sasaki, Misao; Verde, Licia; Wands, David

    2016-09-01

    In General Relativity, the constraint equation relating metric and density perturbations is inherently nonlinear, leading to an effective non-Gaussianity in the dark matter density field on large scales-even if the primordial metric perturbation is Gaussian. Intrinsic non-Gaussianity in the large-scale dark matter overdensity in GR is real and physical. However, the variance smoothed on a local physical scale is not correlated with the large-scale curvature perturbation, so that there is no relativistic signature in the galaxy bias when using the simplest model of bias. It is an open question whether the observable mass proxies such as luminosity or weak lensing correspond directly to the physical mass in the simple halo bias model. If not, there may be observables that encode this relativistic signature.

  16. Magnetic Helicity and Large Scale Magnetic Fields: A Primer

    NASA Astrophysics Data System (ADS)

    Blackman, Eric G.

    2015-05-01

    Magnetic fields of laboratory, planetary, stellar, and galactic plasmas commonly exhibit significant order on large temporal or spatial scales compared to the otherwise random motions within the hosting system. Such ordered fields can be measured in the case of planets, stars, and galaxies, or inferred indirectly by the action of their dynamical influence, such as jets. Whether large scale fields are amplified in situ or a remnant from previous stages of an object's history is often debated for objects without a definitive magnetic activity cycle. Magnetic helicity, a measure of twist and linkage of magnetic field lines, is a unifying tool for understanding large scale field evolution for both mechanisms of origin. Its importance stems from its two basic properties: (1) magnetic helicity is typically better conserved than magnetic energy; and (2) the magnetic energy associated with a fixed amount of magnetic helicity is minimized when the system relaxes this helical structure to the largest scale available. Here I discuss how magnetic helicity has come to help us understand the saturation of and sustenance of large scale dynamos, the need for either local or global helicity fluxes to avoid dynamo quenching, and the associated observational consequences. I also discuss how magnetic helicity acts as a hindrance to turbulent diffusion of large scale fields, and thus a helper for fossil remnant large scale field origin models in some contexts. I briefly discuss the connection between large scale fields and accretion disk theory as well. The goal here is to provide a conceptual primer to help the reader efficiently penetrate the literature.

  17. Large Scale Processes and Extreme Floods in Brazil

    NASA Astrophysics Data System (ADS)

    Ribeiro Lima, C. H.; AghaKouchak, A.; Lall, U.

    2016-12-01

    Persistent large scale anomalies in the atmospheric circulation and ocean state have been associated with heavy rainfall and extreme floods in water basins of different sizes across the world. Such studies have emerged in the last years as a new tool to improve the traditional, stationary based approach in flood frequency analysis and flood prediction. Here we seek to advance previous studies by evaluating the dominance of large scale processes (e.g. atmospheric rivers/moisture transport) over local processes (e.g. local convection) in producing floods. We consider flood-prone regions in Brazil as case studies and the role of large scale climate processes in generating extreme floods in such regions is explored by means of observed streamflow, reanalysis data and machine learning methods. The dynamics of the large scale atmospheric circulation in the days prior to the flood events are evaluated based on the vertically integrated moisture flux and its divergence field, which are interpreted in a low-dimensional space as obtained by machine learning techniques, particularly supervised kernel principal component analysis. In such reduced dimensional space, clusters are obtained in order to better understand the role of regional moisture recycling or teleconnected moisture in producing floods of a given magnitude. The convective available potential energy (CAPE) is also used as a measure of local convection activities. We investigate for individual sites the exceedance probability in which large scale atmospheric fluxes dominate the flood process. Finally, we analyze regional patterns of floods and how the scaling law of floods with drainage area responds to changes in the climate forcing mechanisms (e.g. local vs large scale).

  18. [Issues of large scale tissue culture of medicinal plant].

    PubMed

    Lv, Dong-Mei; Yuan, Yuan; Zhan, Zhi-Lai

    2014-09-01

    In order to increase the yield and quality of the medicinal plant and enhance the competitive power of industry of medicinal plant in our country, this paper analyzed the status, problem and countermeasure of the tissue culture of medicinal plant on large scale. Although the biotechnology is one of the most efficient and promising means in production of medicinal plant, it still has problems such as stability of the material, safety of the transgenic medicinal plant and optimization of cultured condition. Establishing perfect evaluation system according to the characteristic of the medicinal plant is the key measures to assure the sustainable development of the tissue culture of medicinal plant on large scale.

  19. The CLASSgal code for relativistic cosmological large scale structure

    NASA Astrophysics Data System (ADS)

    Di Dio, Enea; Montanari, Francesco; Lesgourgues, Julien; Durrer, Ruth

    2013-11-01

    We present accurate and efficient computations of large scale structure observables, obtained with a modified version of the CLASS code which is made publicly available. This code includes all relativistic corrections and computes both the power spectrum Cl(z1,z2) and the corresponding correlation function ξ(θ,z1,z2) of the matter density and the galaxy number fluctuations in linear perturbation theory. For Gaussian initial perturbations, these quantities contain the full information encoded in the large scale matter distribution at the level of linear perturbation theory. We illustrate the usefulness of our code for cosmological parameter estimation through a few simple examples.

  20. Corridors Increase Plant Species Richness at Large Scales

    SciTech Connect

    Damschen, Ellen I.; Haddad, Nick M.; Orrock,John L.; Tewksbury, Joshua J.; Levey, Douglas J.

    2006-09-01

    Habitat fragmentation is one of the largest threats to biodiversity. Landscape corridors, which are hypothesized to reduce the negative consequences of fragmentation, have become common features of ecological management plans worldwide. Despite their popularity, there is little evidence documenting the effectiveness of corridors in preserving biodiversity at large scales. Using a large-scale replicated experiment, we showed that habitat patches connected by corridors retain more native plant species than do isolated patches, that this difference increases over time, and that corridors do not promote invasion by exotic species. Our results support the use of corridors in biodiversity conservation.

  1. Large-Scale Graph Processing Analysis using Supercomputer Cluster

    NASA Astrophysics Data System (ADS)

    Vildario, Alfrido; Fitriyani; Nugraha Nurkahfi, Galih

    2017-01-01

    Graph implementation is widely use in various sector such as automotive, traffic, image processing and many more. They produce graph in large-scale dimension, cause the processing need long computational time and high specification resources. This research addressed the analysis of implementation large-scale graph using supercomputer cluster. We impelemented graph processing by using Breadth-First Search (BFS) algorithm with single destination shortest path problem. Parallel BFS implementation with Message Passing Interface (MPI) used supercomputer cluster at High Performance Computing Laboratory Computational Science Telkom University and Stanford Large Network Dataset Collection. The result showed that the implementation give the speed up averages more than 30 times and eficiency almost 90%.

  2. Survey of decentralized control methods. [for large scale dynamic systems

    NASA Technical Reports Server (NTRS)

    Athans, M.

    1975-01-01

    An overview is presented of the types of problems that are being considered by control theorists in the area of dynamic large scale systems with emphasis on decentralized control strategies. Approaches that deal directly with decentralized decision making for large scale systems are discussed. It is shown that future advances in decentralized system theory are intimately connected with advances in the stochastic control problem with nonclassical information pattern. The basic assumptions and mathematical tools associated with the latter are summarized, and recommendations concerning future research are presented.

  3. Clearing and Labeling Techniques for Large-Scale Biological Tissues

    PubMed Central

    Seo, Jinyoung; Choe, Minjin; Kim, Sung-Yon

    2016-01-01

    Clearing and labeling techniques for large-scale biological tissues enable simultaneous extraction of molecular and structural information with minimal disassembly of the sample, facilitating the integration of molecular, cellular and systems biology across different scales. Recent years have witnessed an explosive increase in the number of such methods and their applications, reflecting heightened interest in organ-wide clearing and labeling across many fields of biology and medicine. In this review, we provide an overview and comparison of existing clearing and labeling techniques and discuss challenges and opportunities in the investigations of large-scale biological systems. PMID:27239813

  4. The Evolution of Baryons in Cosmic Large Scale Structure

    NASA Astrophysics Data System (ADS)

    Snedden, Ali; Arielle Phillips, Lara; Mathews, Grant James; Coughlin, Jared; Suh, In-Saeng; Bhattacharya, Aparna

    2015-01-01

    The environments of galaxies play a critical role in their formation and evolution. We study these environments using cosmological simulations with star formation and supernova feedback included. From these simulations, we parse the large scale structure into clusters, filaments and voids using a segmentation algorithm adapted from medical imaging. We trace the star formation history, gas phase and metal evolution of the baryons in the intergalactic medium as function of structure. We find that our algorithm reproduces the baryon fraction in the intracluster medium and that the majority of star formation occurs in cold, dense filaments. We present the consequences this large scale environment has for galactic halos and galaxy evolution.

  5. Large scale purification of RNA nanoparticles by preparative ultracentrifugation.

    PubMed

    Jasinski, Daniel L; Schwartz, Chad T; Haque, Farzin; Guo, Peixuan

    2015-01-01

    Purification of large quantities of supramolecular RNA complexes is of paramount importance due to the large quantities of RNA needed and the purity requirements for in vitro and in vivo assays. Purification is generally carried out by liquid chromatography (HPLC), polyacrylamide gel electrophoresis (PAGE), or agarose gel electrophoresis (AGE). Here, we describe an efficient method for the large-scale purification of RNA prepared by in vitro transcription using T7 RNA polymerase by cesium chloride (CsCl) equilibrium density gradient ultracentrifugation and the large-scale purification of RNA nanoparticles by sucrose gradient rate-zonal ultracentrifugation or cushioned sucrose gradient rate-zonal ultracentrifugation.

  6. p53 in the game of transposons.

    PubMed

    Wylie, Annika; Jones, Amanda E; Abrams, John M

    2016-11-01

    Throughout the animal kingdom, p53 genes function to restrain mobile elements and recent observations indicate that transposons become derepressed in human cancers. Together, these emerging lines of evidence suggest that cancers driven by p53 mutations could represent "transpospoathies," i.e. disease states linked to eruptions of mobile elements. The transposopathy hypothesis predicts that p53 acts through conserved mechanisms to contain transposon movement, and in this way, prevents tumor formation. How transposon eruptions provoke neoplasias is not well understood but, from a broader perspective, this hypothesis also provides an attractive framework to explore unrestrained mobile elements as inciters of late-onset idiopathic disease. Also see the video abstract here.

  7. Generation of a Tn5 transposon library in Haemophilus parasuis and analysis by transposon-directed insertion-site sequencing (TraDIS).

    PubMed

    Luan, Shi-Lu; Chaudhuri, Roy R; Peters, Sarah E; Mayho, Matthew; Weinert, Lucy A; Crowther, Sarah A; Wang, Jinhong; Langford, Paul R; Rycroft, Andrew; Wren, Brendan W; Tucker, Alexander W; Maskell, Duncan J

    2013-10-25

    Haemophilus parasuis is an important respiratory tract pathogen of swine and the etiological agent of Glässer's disease. The molecular pathogenesis of H. parasuis is not well studied, mainly due to the lack of efficient tools for genetic manipulation of this bacterium. In this study we describe a Tn5-based random mutagenesis method for use in H. parasuis. A novel chloramphenicol-resistant Tn5 transposome was electroporated into the virulent H. parasuis serovar 5 strain 29755. High transposition efficiency of Tn5, up to 10(4) transformants/μg of transposon DNA, was obtained by modification of the Tn5 DNA in the H. parasuis strain HS071 and establishment of optimal electrotransformation conditions, and a library of approximately 10,500 mutants was constructed. Analysis of the library using transposon-directed insertion-site sequencing (TraDIS) revealed that the insertion of Tn5 was evenly distributed throughout the genome. 10,001 individual mutants were identified, with 1561 genes being disrupted (69.4% of the genome). This newly-developed, efficient mutagenesis approach will be a powerful tool for genetic manipulation of H. parasuis in order to study its physiology and pathogenesis.

  8. The Large-Scale Structure of Scientific Method

    ERIC Educational Resources Information Center

    Kosso, Peter

    2009-01-01

    The standard textbook description of the nature of science describes the proposal, testing, and acceptance of a theoretical idea almost entirely in isolation from other theories. The resulting model of science is a kind of piecemeal empiricism that misses the important network structure of scientific knowledge. Only the large-scale description of…

  9. A bibliographical surveys of large-scale systems

    NASA Technical Reports Server (NTRS)

    Corliss, W. R.

    1970-01-01

    A limited, partly annotated bibliography was prepared on the subject of large-scale system control. Approximately 400 references are divided into thirteen application areas, such as large societal systems and large communication systems. A first-author index is provided.

  10. Mixing Metaphors: Building Infrastructure for Large Scale School Turnaround

    ERIC Educational Resources Information Center

    Peurach, Donald J.; Neumerski, Christine M.

    2015-01-01

    The purpose of this analysis is to increase understanding of the possibilities and challenges of building educational infrastructure--the basic, foundational structures, systems, and resources--to support large-scale school turnaround. Building educational infrastructure often exceeds the capacity of schools, districts, and state education…

  11. Firebrands and spotting ignition in large-scale fires

    Treesearch

    Eunmo Koo; Patrick J. Pagni; David R. Weise; John P. Woycheese

    2010-01-01

    Spotting ignition by lofted firebrands is a significant mechanism of fire spread, as observed in many largescale fires. The role of firebrands in fire propagation and the important parameters involved in spot fire development are studied. Historical large-scale fires, including wind-driven urban and wildland conflagrations and post-earthquake fires are given as...

  12. Large Scale Survey Data in Career Development Research

    ERIC Educational Resources Information Center

    Diemer, Matthew A.

    2008-01-01

    Large scale survey datasets have been underutilized but offer numerous advantages for career development scholars, as they contain numerous career development constructs with large and diverse samples that are followed longitudinally. Constructs such as work salience, vocational expectations, educational expectations, work satisfaction, and…

  13. Measurement, Sampling, and Equating Errors in Large-Scale Assessments

    ERIC Educational Resources Information Center

    Wu, Margaret

    2010-01-01

    In large-scale assessments, such as state-wide testing programs, national sample-based assessments, and international comparative studies, there are many steps involved in the measurement and reporting of student achievement. There are always sources of inaccuracies in each of the steps. It is of interest to identify the source and magnitude of…

  14. US National Large-scale City Orthoimage Standard Initiative

    USGS Publications Warehouse

    Zhou, G.; Song, C.; Benjamin, S.; Schickler, W.

    2003-01-01

    The early procedures and algorithms for National digital orthophoto generation in National Digital Orthophoto Program (NDOP) were based on earlier USGS mapping operations, such as field control, aerotriangulation (derived in the early 1920's), the quarter-quadrangle-centered (3.75 minutes of longitude and latitude in geographic extent), 1:40,000 aerial photographs, and 2.5 D digital elevation models. However, large-scale city orthophotos using early procedures have disclosed many shortcomings, e.g., ghost image, occlusion, shadow. Thus, to provide the technical base (algorithms, procedure) and experience needed for city large-scale digital orthophoto creation is essential for the near future national large-scale digital orthophoto deployment and the revision of the Standards for National Large-scale City Digital Orthophoto in National Digital Orthophoto Program (NDOP). This paper will report our initial research results as follows: (1) High-precision 3D city DSM generation through LIDAR data processing, (2) Spatial objects/features extraction through surface material information and high-accuracy 3D DSM data, (3) 3D city model development, (4) Algorithm development for generation of DTM-based orthophoto, and DBM-based orthophoto, (5) True orthophoto generation by merging DBM-based orthophoto and DTM-based orthophoto, and (6) Automatic mosaic by optimizing and combining imagery from many perspectives.

  15. DESIGN OF LARGE-SCALE AIR MONITORING NETWORKS

    EPA Science Inventory

    The potential effects of air pollution on human health have received much attention in recent years. In the U.S. and other countries, there are extensive large-scale monitoring networks designed to collect data to inform the public of exposure risks to air pollution. A major crit...

  16. Large-Scale Environmental Influences on Aquatic Animal Health

    EPA Science Inventory

    In the latter portion of the 20th century, North America experienced numerous large-scale mortality events affecting a broad diversity of aquatic animals. Short-term forensic investigations of these events have sometimes characterized a causative agent or condition, but have rare...

  17. DESIGN OF LARGE-SCALE AIR MONITORING NETWORKS

    EPA Science Inventory

    The potential effects of air pollution on human health have received much attention in recent years. In the U.S. and other countries, there are extensive large-scale monitoring networks designed to collect data to inform the public of exposure risks to air pollution. A major crit...

  18. Developing and Understanding Methods for Large-Scale Nonlinear Optimization

    DTIC Science & Technology

    2006-07-24

    algorithms for large-scale uncon- strained and constrained optimization problems, including limited-memory methods for problems with -2- many thousands...34Published in peer-reviewed journals" E. Eskow, B. Bader, R. Byrd, S. Crivelli, T. Head-Gordon, V. Lamberti and R. Schnabel, "An optimization approach to the

  19. Probabilistic Cuing in Large-Scale Environmental Search

    ERIC Educational Resources Information Center

    Smith, Alastair D.; Hood, Bruce M.; Gilchrist, Iain D.

    2010-01-01

    Finding an object in our environment is an important human ability that also represents a critical component of human foraging behavior. One type of information that aids efficient large-scale search is the likelihood of the object being in one location over another. In this study we investigated the conditions under which individuals respond to…

  20. Feasibility of large-scale aquatic microcosms. Final report

    SciTech Connect

    Pease, T.; Wyman, R.L.; Logan, D.T.; Logan, C.M.; Lispi, D.R.

    1982-02-01

    Microcosms have been used to study a number of fundamental ecological principles and more recently to investigate the effects of man-made perturbations on ecosystems. In this report the feasibility of using large-scale microcosms to access aquatic impacts of power generating facilities is evaluated. Aquatic problems of concern to utilities are outlined, and various research approaches, including large and small microcosms, bioassays, and other laboratory experiments, are discussed. An extensive critical review and synthesis of the literature on recent microcosm research, which includes a comparison of the factors influencing physical, chemical, and biological processes in small vs large microcosms and in microcosms vs nature, led the authors to conclude that large-scale microcosms offer several advantages over other study techniques for particular types of problems. A hypothetical large-scale facility simulating a lake ecosystem is presented to illustrate the size, cost, and complexity of such facilities. The rationale for designing a lake-simulating large-scale microcosm is presented.

  1. Assuring Quality in Large-Scale Online Course Development

    ERIC Educational Resources Information Center

    Parscal, Tina; Riemer, Deborah

    2010-01-01

    Student demand for online education requires colleges and universities to rapidly expand the number of courses and programs offered online while maintaining high quality. This paper outlines two universities respective processes to assure quality in large-scale online programs that integrate instructional design, eBook custom publishing, Quality…

  2. Improving the Utility of Large-Scale Assessments in Canada

    ERIC Educational Resources Information Center

    Rogers, W. Todd

    2014-01-01

    Principals and teachers do not use large-scale assessment results because the lack of distinct and reliable subtests prevents identifying strengths and weaknesses of students and instruction, the results arrive too late to be used, and principals and teachers need assistance to use the results to improve instruction so as to improve student…

  3. Research directions in large scale systems and decentralized control

    NASA Technical Reports Server (NTRS)

    Tenney, R. R.

    1980-01-01

    Control theory provides a well established framework for dealing with automatic decision problems and a set of techniques for automatic decision making which exploit special structure, but it does not deal well with complexity. The potential exists for combining control theoretic and knowledge based concepts into a unified approach. The elements of control theory are diagrammed, including modern control and large scale systems.

  4. Efficient On-Demand Operations in Large-Scale Infrastructures

    ERIC Educational Resources Information Center

    Ko, Steven Y.

    2009-01-01

    In large-scale distributed infrastructures such as clouds, Grids, peer-to-peer systems, and wide-area testbeds, users and administrators typically desire to perform "on-demand operations" that deal with the most up-to-date state of the infrastructure. However, the scale and dynamism present in the operating environment make it challenging to…

  5. Ecosystem resilience despite large-scale altered hydro climatic conditions

    USDA-ARS?s Scientific Manuscript database

    Climate change is predicted to increase both drought frequency and duration, and when coupled with substantial warming, will establish a new hydroclimatological paradigm for many regions. Large-scale, warm droughts have recently impacted North America, Africa, Europe, Amazonia, and Australia result...

  6. The Large-Scale Structure of Scientific Method

    ERIC Educational Resources Information Center

    Kosso, Peter

    2009-01-01

    The standard textbook description of the nature of science describes the proposal, testing, and acceptance of a theoretical idea almost entirely in isolation from other theories. The resulting model of science is a kind of piecemeal empiricism that misses the important network structure of scientific knowledge. Only the large-scale description of…

  7. Large-Scale Assessments and Educational Policies in Italy

    ERIC Educational Resources Information Center

    Damiani, Valeria

    2016-01-01

    Despite Italy's extensive participation in most large-scale assessments, their actual influence on Italian educational policies is less easy to identify. The present contribution aims at highlighting and explaining reasons for the weak and often inconsistent relationship between international surveys and policy-making processes in Italy.…

  8. Large-Scale Innovation and Change in UK Higher Education

    ERIC Educational Resources Information Center

    Brown, Stephen

    2013-01-01

    This paper reflects on challenges universities face as they respond to change. It reviews current theories and models of change management, discusses why universities are particularly difficult environments in which to achieve large scale, lasting change and reports on a recent attempt by the UK JISC to enable a range of UK universities to employ…

  9. Current Scientific Issues in Large Scale Atmospheric Dynamics

    NASA Technical Reports Server (NTRS)

    Miller, T. L. (Compiler)

    1986-01-01

    Topics in large scale atmospheric dynamics are discussed. Aspects of atmospheric blocking, the influence of transient baroclinic eddies on planetary-scale waves, cyclogenesis, the effects of orography on planetary scale flow, small scale frontal structure, and simulations of gravity waves in frontal zones are discussed.

  10. Large-Scale Assessments and Educational Policies in Italy

    ERIC Educational Resources Information Center

    Damiani, Valeria

    2016-01-01

    Despite Italy's extensive participation in most large-scale assessments, their actual influence on Italian educational policies is less easy to identify. The present contribution aims at highlighting and explaining reasons for the weak and often inconsistent relationship between international surveys and policy-making processes in Italy.…

  11. Large scale fire whirls: Can their formation be predicted?

    Treesearch

    J. Forthofer; Bret Butler

    2010-01-01

    Large scale fire whirls have not traditionally been recognized as a frequent phenomenon on wildland fires. However, there are anecdotal data suggesting that they can and do occur with some regularity. This paper presents a brief summary of this information and an analysis of the causal factors leading to their formation.

  12. Large-Scale Environmental Influences on Aquatic Animal Health

    EPA Science Inventory

    In the latter portion of the 20th century, North America experienced numerous large-scale mortality events affecting a broad diversity of aquatic animals. Short-term forensic investigations of these events have sometimes characterized a causative agent or condition, but have rare...

  13. International Large-Scale Assessments: What Uses, What Consequences?

    ERIC Educational Resources Information Center

    Johansson, Stefan

    2016-01-01

    Background: International large-scale assessments (ILSAs) are a much-debated phenomenon in education. Increasingly, their outcomes attract considerable media attention and influence educational policies in many jurisdictions worldwide. The relevance, uses and consequences of these assessments are often the focus of research scrutiny. Whilst some…

  14. Extracting Useful Semantic Information from Large Scale Corpora of Text

    ERIC Educational Resources Information Center

    Mendoza, Ray Padilla, Jr.

    2012-01-01

    Extracting and representing semantic information from large scale corpora is at the crux of computer-assisted knowledge generation. Semantic information depends on collocation extraction methods, mathematical models used to represent distributional information, and weighting functions which transform the space. This dissertation provides a…

  15. Large-Scale Innovation and Change in UK Higher Education

    ERIC Educational Resources Information Center

    Brown, Stephen

    2013-01-01

    This paper reflects on challenges universities face as they respond to change. It reviews current theories and models of change management, discusses why universities are particularly difficult environments in which to achieve large scale, lasting change and reports on a recent attempt by the UK JISC to enable a range of UK universities to employ…

  16. Mixing Metaphors: Building Infrastructure for Large Scale School Turnaround

    ERIC Educational Resources Information Center

    Peurach, Donald J.; Neumerski, Christine M.

    2015-01-01

    The purpose of this analysis is to increase understanding of the possibilities and challenges of building educational infrastructure--the basic, foundational structures, systems, and resources--to support large-scale school turnaround. Building educational infrastructure often exceeds the capacity of schools, districts, and state education…

  17. Individual Skill Differences and Large-Scale Environmental Learning

    ERIC Educational Resources Information Center

    Fields, Alexa W.; Shelton, Amy L.

    2006-01-01

    Spatial skills are known to vary widely among normal individuals. This project was designed to address whether these individual differences are differentially related to large-scale environmental learning from route (ground-level) and survey (aerial) perspectives. Participants learned two virtual environments (route and survey) with limited…

  18. Newton Methods for Large Scale Problems in Machine Learning

    ERIC Educational Resources Information Center

    Hansen, Samantha Leigh

    2014-01-01

    The focus of this thesis is on practical ways of designing optimization algorithms for minimizing large-scale nonlinear functions with applications in machine learning. Chapter 1 introduces the overarching ideas in the thesis. Chapters 2 and 3 are geared towards supervised machine learning applications that involve minimizing a sum of loss…

  19. Large-Scale Machine Learning for Classification and Search

    ERIC Educational Resources Information Center

    Liu, Wei

    2012-01-01

    With the rapid development of the Internet, nowadays tremendous amounts of data including images and videos, up to millions or billions, can be collected for training machine learning models. Inspired by this trend, this thesis is dedicated to developing large-scale machine learning techniques for the purpose of making classification and nearest…

  20. Global smoothing and continuation for large-scale molecular optimization

    SciTech Connect

    More, J.J.; Wu, Zhijun

    1995-10-01

    We discuss the formulation of optimization problems that arise in the study of distance geometry, ionic systems, and molecular clusters. We show that continuation techniques based on global smoothing are applicable to these molecular optimization problems, and we outline the issues that must be resolved in the solution of large-scale molecular optimization problems.

  1. Large-scale Eucalyptus energy farms and power cogeneration

    Treesearch

    Robert C. Noroña

    1983-01-01

    A thorough evaluation of all factors possibly affecting a large-scale planting of eucalyptus is foremost in determining the cost effectiveness of the planned operation. Seven basic areas of concern must be analyzed:1. Species Selection 2. Site Preparation 3. Planting 4. Weed Control 5....

  2. Probabilistic Cuing in Large-Scale Environmental Search

    ERIC Educational Resources Information Center

    Smith, Alastair D.; Hood, Bruce M.; Gilchrist, Iain D.

    2010-01-01

    Finding an object in our environment is an important human ability that also represents a critical component of human foraging behavior. One type of information that aids efficient large-scale search is the likelihood of the object being in one location over another. In this study we investigated the conditions under which individuals respond to…

  3. Lessons from Large-Scale Renewable Energy Integration Studies: Preprint

    SciTech Connect

    Bird, L.; Milligan, M.

    2012-06-01

    In general, large-scale integration studies in Europe and the United States find that high penetrations of renewable generation are technically feasible with operational changes and increased access to transmission. This paper describes other key findings such as the need for fast markets, large balancing areas, system flexibility, and the use of advanced forecasting.

  4. The large scale microwave background anisotropy in decaying particle cosmology

    SciTech Connect

    Panek, M.

    1987-06-01

    We investigate the large-scale anisotropy of the microwave background radiation in cosmological models with decaying particles. The observed value of the quadrupole moment combined with other constraints gives an upper limit on the redshift of the decay z/sub d/ < 3-5. 12 refs., 2 figs.

  5. Large-scale search for dark-matter axions

    SciTech Connect

    Kinion, D; van Bibber, K

    2000-08-30

    We review the status of two ongoing large-scale searches for axions which may constitute the dark matter of our Milky Way halo. The experiments are based on the microwave cavity technique proposed by Sikivie, and marks a ''second-generation'' to the original experiments performed by the Rochester-Brookhaven-Fermilab collaboration, and the University of Florida group.

  6. Resilience of Florida Keys coral communities following large scale disturbances

    EPA Science Inventory

    The decline of coral reefs in the Caribbean over the last 40 years has been attributed to multiple chronic stressors and episodic large-scale disturbances. This study assessed the resilience of coral communities in two different regions of the Florida Keys reef system between 199...

  7. Large Scale Survey Data in Career Development Research

    ERIC Educational Resources Information Center

    Diemer, Matthew A.

    2008-01-01

    Large scale survey datasets have been underutilized but offer numerous advantages for career development scholars, as they contain numerous career development constructs with large and diverse samples that are followed longitudinally. Constructs such as work salience, vocational expectations, educational expectations, work satisfaction, and…

  8. The Role of Plausible Values in Large-Scale Surveys

    ERIC Educational Resources Information Center

    Wu, Margaret

    2005-01-01

    In large-scale assessment programs such as NAEP, TIMSS and PISA, students' achievement data sets provided for secondary analysts contain so-called "plausible values." Plausible values are multiple imputations of the unobservable latent achievement for each student. In this article it has been shown how plausible values are used to: (1)…

  9. Large-scale silicon optical switches for optical interconnection

    NASA Astrophysics Data System (ADS)

    Qiao, Lei; Tang, Weijie; Chu, Tao

    2016-11-01

    Large-scale optical switches are greatly demanded in building optical interconnections in data centers and high performance computers (HPCs). Silicon optical switches have advantages of being compact and CMOS process compatible, which can be easily monolithically integrated. However, there are difficulties to construct large ports silicon optical switches. One of them is the non-uniformity of the switch units in large scale silicon optical switches, which arises from the fabrication error and causes confusion in finding the unit optimum operation points. In this paper, we proposed a method to detect the optimum operating point in large scale switch with limited build-in power monitors. We also propose methods for improving the unbalanced crosstalk of cross/bar states in silicon electro-optical MZI switches and insertion losses. Our recent progress in large scale silicon optical switches, including 64 × 64 thermal-optical and 32 × 32 electro-optical switches will be introduced. To the best our knowledge, both of them are the largest scale silicon optical switches in their sections, respectively. The switches were fabricated on 340-nm SOI substrates with CMOS 180- nm processes. The crosstalk of the 32 × 32 electro-optic switch was -19.2dB to -25.1 dB, while the value of the 64 × 64 thermal-optic switch was -30 dB to -48.3 dB.

  10. Assuring Quality in Large-Scale Online Course Development

    ERIC Educational Resources Information Center

    Parscal, Tina; Riemer, Deborah

    2010-01-01

    Student demand for online education requires colleges and universities to rapidly expand the number of courses and programs offered online while maintaining high quality. This paper outlines two universities respective processes to assure quality in large-scale online programs that integrate instructional design, eBook custom publishing, Quality…

  11. Computational Complexity, Efficiency and Accountability in Large Scale Teleprocessing Systems.

    DTIC Science & Technology

    1980-12-01

    COMPLEXITY, EFFICIENCY AND ACCOUNTABILITY IN LARGE SCALE TELEPROCESSING SYSTEMS DAAG29-78-C-0036 STANFORD UNIVERSITY JOHN T. GILL MARTIN E. BELLMAN...solve but easy to check. Ve have also suggested howy sucb random tapes can be simulated by determin- istically generating "pseudorandom" numbers by a

  12. Large-Scale Assessment and English Language Learners with Disabilities

    ERIC Educational Resources Information Center

    Liu, Kristin K.; Ward, Jenna M.; Thurlow, Martha L.; Christensen, Laurene L.

    2017-01-01

    This article highlights a set of principles and guidelines, developed by a diverse group of specialists in the field, for appropriately including English language learners (ELLs) with disabilities in large-scale assessments. ELLs with disabilities make up roughly 9% of the rapidly increasing ELL population nationwide. In spite of the small overall…

  13. Large-scale silviculture experiments of western Oregon and Washington.

    Treesearch

    Nathan J. Poage; Paul D. Anderson

    2007-01-01

    We review 12 large-scale silviculture experiments (LSSEs) in western Washington and Oregon with which the Pacific Northwest Research Station of the USDA Forest Service is substantially involved. We compiled and arrayed information about the LSSEs as a series of matrices in a relational database, which is included on the compact disc published with this report and...

  14. Newton Methods for Large Scale Problems in Machine Learning

    ERIC Educational Resources Information Center

    Hansen, Samantha Leigh

    2014-01-01

    The focus of this thesis is on practical ways of designing optimization algorithms for minimizing large-scale nonlinear functions with applications in machine learning. Chapter 1 introduces the overarching ideas in the thesis. Chapters 2 and 3 are geared towards supervised machine learning applications that involve minimizing a sum of loss…

  15. Efficient On-Demand Operations in Large-Scale Infrastructures

    ERIC Educational Resources Information Center

    Ko, Steven Y.

    2009-01-01

    In large-scale distributed infrastructures such as clouds, Grids, peer-to-peer systems, and wide-area testbeds, users and administrators typically desire to perform "on-demand operations" that deal with the most up-to-date state of the infrastructure. However, the scale and dynamism present in the operating environment make it challenging to…

  16. Large-Scale Machine Learning for Classification and Search

    ERIC Educational Resources Information Center

    Liu, Wei

    2012-01-01

    With the rapid development of the Internet, nowadays tremendous amounts of data including images and videos, up to millions or billions, can be collected for training machine learning models. Inspired by this trend, this thesis is dedicated to developing large-scale machine learning techniques for the purpose of making classification and nearest…

  17. Moon-based Earth Observation for Large Scale Geoscience Phenomena

    NASA Astrophysics Data System (ADS)

    Guo, Huadong; Liu, Guang; Ding, Yixing

    2016-07-01

    The capability of Earth observation for large-global-scale natural phenomena needs to be improved and new observing platform are expected. We have studied the concept of Moon as an Earth observation in these years. Comparing with manmade satellite platform, Moon-based Earth observation can obtain multi-spherical, full-band, active and passive information,which is of following advantages: large observation range, variable view angle, long-term continuous observation, extra-long life cycle, with the characteristics of longevity ,consistency, integrity, stability and uniqueness. Moon-based Earth observation is suitable for monitoring the large scale geoscience phenomena including large scale atmosphere change, large scale ocean change,large scale land surface dynamic change,solid earth dynamic change,etc. For the purpose of establishing a Moon-based Earth observation platform, we already have a plan to study the five aspects as follows: mechanism and models of moon-based observing earth sciences macroscopic phenomena; sensors' parameters optimization and methods of moon-based Earth observation; site selection and environment of moon-based Earth observation; Moon-based Earth observation platform; and Moon-based Earth observation fundamental scientific framework.

  18. Large-scale societal changes and intentionality - an uneasy marriage.

    PubMed

    Bodor, Péter; Fokas, Nikos

    2014-08-01

    Our commentary focuses on juxtaposing the proposed science of intentional change with facts and concepts pertaining to the level of large populations or changes on a worldwide scale. Although we find a unified evolutionary theory promising, we think that long-term and large-scale, scientifically guided - that is, intentional - social change is not only impossible, but also undesirable.

  19. Large-scale screening by the automated Wassermann reaction

    PubMed Central

    Wagstaff, W.; Firth, R.; Booth, J. R.; Bowley, C. C.

    1969-01-01

    In view of the drawbacks in the use of the Kahn test for large-scale screening of blood donors, mainly those of human error through work overload and fatiguability, an attempt was made to adapt an existing automated complement-fixation technique for this purpose. This paper reports the successful results of that adaptation. PMID:5776559

  20. International Large-Scale Assessments: What Uses, What Consequences?

    ERIC Educational Resources Information Center

    Johansson, Stefan

    2016-01-01

    Background: International large-scale assessments (ILSAs) are a much-debated phenomenon in education. Increasingly, their outcomes attract considerable media attention and influence educational policies in many jurisdictions worldwide. The relevance, uses and consequences of these assessments are often the focus of research scrutiny. Whilst some…

  1. Cosmic strings and the large-scale structure

    NASA Technical Reports Server (NTRS)

    Stebbins, Albert

    1988-01-01

    A possible problem for cosmic string models of galaxy formation is presented. If very large voids are common and if loop fragmentation is not much more efficient than presently believed, then it may be impossible for string scenarios to produce the observed large-scale structure with Omega sub 0 = 1 and without strong environmental biasing.

  2. Extracting Useful Semantic Information from Large Scale Corpora of Text

    ERIC Educational Resources Information Center

    Mendoza, Ray Padilla, Jr.

    2012-01-01

    Extracting and representing semantic information from large scale corpora is at the crux of computer-assisted knowledge generation. Semantic information depends on collocation extraction methods, mathematical models used to represent distributional information, and weighting functions which transform the space. This dissertation provides a…

  3. Large scale structure of the sun's radio corona

    NASA Technical Reports Server (NTRS)

    Kundu, M. R.

    1986-01-01

    Results of studies of large scale structures of the corona at long radio wavelengths are presented, using data obtained with the multifrequency radioheliograph of the Clark Lake Radio Observatory. It is shown that features corresponding to coronal streamers and coronal holes are readily apparent in the Clark Lake maps.

  4. Resilience of Florida Keys coral communities following large scale disturbances

    EPA Science Inventory

    The decline of coral reefs in the Caribbean over the last 40 years has been attributed to multiple chronic stressors and episodic large-scale disturbances. This study assessed the resilience of coral communities in two different regions of the Florida Keys reef system between 199...

  5. Rapid quantification of mutant fitness in diverse bacteria by sequencing randomly bar-coded transposons

    DOE PAGES

    Wetmore, Kelly M.; Price, Morgan N.; Waters, Robert J.; ...

    2015-05-12

    Transposon mutagenesis with next-generation sequencing (TnSeq) is a powerful approach to annotate gene function in bacteria, but existing protocols for TnSeq require laborious preparation of every sample before sequencing. Thus, the existing protocols are not amenable to the throughput necessary to identify phenotypes and functions for the majority of genes in diverse bacteria. Here, we present a method, random bar code transposon-site sequencing (RB-TnSeq), which increases the throughput of mutant fitness profiling by incorporating random DNA bar codes into Tn5 and mariner transposons and by using bar code sequencing (BarSeq) to assay mutant fitness. RB-TnSeq can be used with anymore » transposon, and TnSeq is performed once per organism instead of once per sample. Each BarSeq assay requires only a simple PCR, and 48 to 96 samples can be sequenced on one lane of an Illumina HiSeq system. We demonstrate the reproducibility and biological significance of RB-TnSeq with Escherichia coli, Phaeobacter inhibens, Pseudomonas stutzeri, Shewanella amazonensis, and Shewanella oneidensis. To demonstrate the increased throughput of RB-TnSeq, we performed 387 successful genome-wide mutant fitness assays representing 130 different bacterium-carbon source combinations and identified 5,196 genes with significant phenotypes across the five bacteria. In P. inhibens, we used our mutant fitness data to identify genes important for the utilization of diverse carbon substrates, including a putative D-mannose isomerase that is required for mannitol catabolism. RB-TnSeq will enable the cost-effective functional annotation of diverse bacteria using mutant fitness profiling. A large challenge in microbiology is the functional assessment of the millions of uncharacterized genes identified by genome sequencing. Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach to assign phenotypes and functions to genes. However, the current strategies for TnSeq are

  6. Rapid quantification of mutant fitness in diverse bacteria by sequencing randomly bar-coded transposons

    SciTech Connect

    Wetmore, Kelly M.; Price, Morgan N.; Waters, Robert J.; Lamson, Jacob S.; He, Jennifer; Hoover, Cindi A.; Blow, Matthew J.; Bristow, James; Butland, Gareth; Arkin, Adam P.; Deutschbauer, Adam

    2015-05-12

    Transposon mutagenesis with next-generation sequencing (TnSeq) is a powerful approach to annotate gene function in bacteria, but existing protocols for TnSeq require laborious preparation of every sample before sequencing. Thus, the existing protocols are not amenable to the throughput necessary to identify phenotypes and functions for the majority of genes in diverse bacteria. Here, we present a method, random bar code transposon-site sequencing (RB-TnSeq), which increases the throughput of mutant fitness profiling by incorporating random DNA bar codes into Tn5 and mariner transposons and by using bar code sequencing (BarSeq) to assay mutant fitness. RB-TnSeq can be used with any transposon, and TnSeq is performed once per organism instead of once per sample. Each BarSeq assay requires only a simple PCR, and 48 to 96 samples can be sequenced on one lane of an Illumina HiSeq system. We demonstrate the reproducibility and biological significance of RB-TnSeq with Escherichia coli, Phaeobacter inhibens, Pseudomonas stutzeri, Shewanella amazonensis, and Shewanella oneidensis. To demonstrate the increased throughput of RB-TnSeq, we performed 387 successful genome-wide mutant fitness assays representing 130 different bacterium-carbon source combinations and identified 5,196 genes with significant phenotypes across the five bacteria. In P. inhibens, we used our mutant fitness data to identify genes important for the utilization of diverse carbon substrates, including a putative D-mannose isomerase that is required for mannitol catabolism. RB-TnSeq will enable the cost-effective functional annotation of diverse bacteria using mutant fitness profiling. A large challenge in microbiology is the functional assessment of the millions of uncharacterized genes identified by genome sequencing. Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach to assign phenotypes and functions to genes

  7. 2004 Mutagenesis Gordon Conference

    SciTech Connect

    Dr. Sue Jinks-Robertson

    2005-09-16

    Mutations are genetic alterations that drive biological evolution and cause many, if not all, human diseases. Mutation originates via two distinct mechanisms: ''vertical'' variation is de novo change of one or few bases, whereas ''horizontal'' variation occurs by genetic recombination, which creates new mosaics of pre-existing sequences. The Mutagenesis Conference has traditionally focused on the generation of mutagenic intermediates during normal DNA synthesis or in response to environmental insults, as well as the diverse repair mechanisms that prevent the fixation of such intermediates as permanent mutations. While the 2004 Conference will continue to focus on the molecular mechanisms of mutagenesis, there will be increased emphasis on the biological consequences of mutations, both in terms of evolutionary processes and in terms of human disease. The meeting will open with two historical accounts of mutation research that recapitulate the intellectual framework of this field and thereby place the current research paradigms into perspective. The two introductory keynote lectures will be followed by sessions on: (1) mutagenic systems, (2) hypermutable sequences, (3) mechanisms of mutation, (4) mutation avoidance systems, (5) mutation in human hereditary and infectious diseases, (6) mutation rates in evolution and genotype-phenotype relationships, (7) ecology, mutagenesis and the modeling of evolution and (8) genetic diversity of the human population and models for human mutagenesis. The Conference will end with a synthesis of the meeting as the keynote closing lecture.

  8. Computer Simulation of Mutagenesis.

    ERIC Educational Resources Information Center

    North, J. C.; Dent, M. T.

    1978-01-01

    A FORTRAN program is described which simulates point-substitution mutations in the DNA strands of typical organisms. Its objective is to help students to understand the significance and structure of the genetic code, and the mechanisms and effect of mutagenesis. (Author/BB)

  9. Computer Simulation of Mutagenesis.

    ERIC Educational Resources Information Center

    North, J. C.; Dent, M. T.

    1978-01-01

    A FORTRAN program is described which simulates point-substitution mutations in the DNA strands of typical organisms. Its objective is to help students to understand the significance and structure of the genetic code, and the mechanisms and effect of mutagenesis. (Author/BB)

  10. Mechanism of proflavin mutagenesis.

    PubMed

    Sarabhai, A; Lamfrom, H

    1969-08-01

    The mutagenic action of proflavin on bacteriophage T4 is greater in the presence of defective T4 ligase than in the presence of normal T4 ligase. This suggests that the persistence of single-strand breaks in DNA enhances proflavin mutagenesis.

  11. Electron drift in a large scale solid xenon

    SciTech Connect

    Yoo, J.; Jaskierny, W. F.

    2015-08-21

    A study of charge drift in a large scale optically transparent solid xenon is reported. A pulsed high power xenon light source is used to liberate electrons from a photocathode. The drift speeds of the electrons are measured using a 8.7 cm long electrode in both the liquid and solid phase of xenon. In the liquid phase (163 K), the drift speed is 0.193 ± 0.003 cm/μs while the drift speed in the solid phase (157 K) is 0.397 ± 0.006 cm/μs at 900 V/cm over 8.0 cm of uniform electric fields. Furthermore, it is demonstrated that a factor two faster electron drift speed in solid phase xenon compared to that in liquid in a large scale solid xenon.

  12. Electron drift in a large scale solid xenon

    DOE PAGES

    Yoo, J.; Jaskierny, W. F.

    2015-08-21

    A study of charge drift in a large scale optically transparent solid xenon is reported. A pulsed high power xenon light source is used to liberate electrons from a photocathode. The drift speeds of the electrons are measured using a 8.7 cm long electrode in both the liquid and solid phase of xenon. In the liquid phase (163 K), the drift speed is 0.193 ± 0.003 cm/μs while the drift speed in the solid phase (157 K) is 0.397 ± 0.006 cm/μs at 900 V/cm over 8.0 cm of uniform electric fields. Furthermore, it is demonstrated that a factor twomore » faster electron drift speed in solid phase xenon compared to that in liquid in a large scale solid xenon.« less

  13. GAIA: A WINDOW TO LARGE-SCALE MOTIONS

    SciTech Connect

    Nusser, Adi; Branchini, Enzo; Davis, Marc E-mail: branchin@fis.uniroma3.it

    2012-08-10

    Using redshifts as a proxy for galaxy distances, estimates of the two-dimensional (2D) transverse peculiar velocities of distant galaxies could be obtained from future measurements of proper motions. We provide the mathematical framework for analyzing 2D transverse motions and show that they offer several advantages over traditional probes of large-scale motions. They are completely independent of any intrinsic relations between galaxy properties; hence, they are essentially free of selection biases. They are free from homogeneous and inhomogeneous Malmquist biases that typically plague distance indicator catalogs. They provide additional information to traditional probes that yield line-of-sight peculiar velocities only. Further, because of their 2D nature, fundamental questions regarding vorticity of large-scale flows can be addressed. Gaia, for example, is expected to provide proper motions of at least bright galaxies with high central surface brightness, making proper motions a likely contender for traditional probes based on current and future distance indicator measurements.

  14. Large scale meteorological influence during the Geysers 1979 field experiment

    SciTech Connect

    Barr, S.

    1980-01-01

    A series of meteorological field measurements conducted during July 1979 near Cobb Mountain in Northern California reveals evidence of several scales of atmospheric circulation consistent with the climatic pattern of the area. The scales of influence are reflected in the structure of wind and temperature in vertically stratified layers at a given observation site. Large scale synoptic gradient flow dominates the wind field above about twice the height of the topographic ridge. Below that there is a mixture of effects with evidence of a diurnal sea breeze influence and a sublayer of katabatic winds. The July observations demonstrate that weak migratory circulations in the large scale synoptic meteorological pattern have a significant influence on the day-to-day gradient winds and must be accounted for in planning meteorological programs including tracer experiments.

  15. The Large Scale Synthesis of Aligned Plate Nanostructures

    PubMed Central

    Zhou, Yang; Nash, Philip; Liu, Tian; Zhao, Naiqin; Zhu, Shengli

    2016-01-01

    We propose a novel technique for the large-scale synthesis of aligned-plate nanostructures that are self-assembled and self-supporting. The synthesis technique involves developing nanoscale two-phase microstructures through discontinuous precipitation followed by selective etching to remove one of the phases. The method may be applied to any alloy system in which the discontinuous precipitation transformation goes to completion. The resulting structure may have many applications in catalysis, filtering and thermal management depending on the phase selection and added functionality through chemical reaction with the retained phase. The synthesis technique is demonstrated using the discontinuous precipitation of a γ′ phase, (Ni, Co)3Al, followed by selective dissolution of the γ matrix phase. The production of the nanostructure requires heat treatments on the order of minutes and can be performed on a large scale making this synthesis technique of great economic potential. PMID:27439672

  16. Lagrangian space consistency relation for large scale structure

    SciTech Connect

    Horn, Bart; Hui, Lam; Xiao, Xiao E-mail: lh399@columbia.edu

    2015-09-01

    Consistency relations, which relate the squeezed limit of an (N+1)-point correlation function to an N-point function, are non-perturbative symmetry statements that hold even if the associated high momentum modes are deep in the nonlinear regime and astrophysically complex. Recently, Kehagias and Riotto and Peloso and Pietroni discovered a consistency relation applicable to large scale structure. We show that this can be recast into a simple physical statement in Lagrangian space: that the squeezed correlation function (suitably normalized) vanishes. This holds regardless of whether the correlation observables are at the same time or not, and regardless of whether multiple-streaming is present. The simplicity of this statement suggests that an analytic understanding of large scale structure in the nonlinear regime may be particularly promising in Lagrangian space.

  17. The workshop on iterative methods for large scale nonlinear problems

    SciTech Connect

    Walker, H.F.; Pernice, M.

    1995-12-01

    The aim of the workshop was to bring together researchers working on large scale applications with numerical specialists of various kinds. Applications that were addressed included reactive flows (combustion and other chemically reacting flows, tokamak modeling), porous media flows, cardiac modeling, chemical vapor deposition, image restoration, macromolecular modeling, and population dynamics. Numerical areas included Newton iterative (truncated Newton) methods, Krylov subspace methods, domain decomposition and other preconditioning methods, large scale optimization and optimal control, and parallel implementations and software. This report offers a brief summary of workshop activities and information about the participants. Interested readers are encouraged to look into an online proceedings available at http://www.usi.utah.edu/logan.proceedings. In this, the material offered here is augmented with hypertext abstracts that include links to locations such as speakers` home pages, PostScript copies of talks and papers, cross-references to related talks, and other information about topics addresses at the workshop.

  18. Large Scale Deformation of the Western US Cordillera

    NASA Technical Reports Server (NTRS)

    Bennett, Richard A.

    2001-01-01

    Destructive earthquakes occur throughout the western US Cordillera (WUSC), not just within the San Andreas fault zone. But because we do not understand the present-day large-scale deformations of the crust throughout the WUSC, our ability to assess the potential for seismic hazards in this region remains severely limited. To address this problem, we are using a large collection of Global Positioning System (GPS) networks which spans the WUSC to precisely quantify present-day large-scale crustal deformations in a single uniform reference frame. Our work can roughly be divided into an analysis of the GPS observations to infer the deformation field across and within the entire plate boundary zone and an investigation of the implications of this deformation field regarding plate boundary dynamics.

  19. Large-scale linear nonparallel support vector machine solver.

    PubMed

    Tian, Yingjie; Ping, Yuan

    2014-02-01

    Twin support vector machines (TWSVMs), as the representative nonparallel hyperplane classifiers, have shown the effectiveness over standard SVMs from some aspects. However, they still have some serious defects restricting their further study and real applications: (1) They have to compute and store the inverse matrices before training, it is intractable for many applications where data appear with a huge number of instances as well as features; (2) TWSVMs lost the sparseness by using a quadratic loss function making the proximal hyperplane close enough to the class itself. This paper proposes a Sparse Linear Nonparallel Support Vector Machine, termed as L1-NPSVM, to deal with large-scale data based on an efficient solver-dual coordinate descent (DCD) method. Both theoretical analysis and experiments indicate that our method is not only suitable for large scale problems, but also performs as good as TWSVMs and SVMs.

  20. Instrumentation Development for Large Scale Hypersonic Inflatable Aerodynamic Decelerator Characterization

    NASA Technical Reports Server (NTRS)

    Swanson, Gregory T.; Cassell, Alan M.

    2011-01-01

    Hypersonic Inflatable Aerodynamic Decelerator (HIAD) technology is currently being considered for multiple atmospheric entry applications as the limitations of traditional entry vehicles have been reached. The Inflatable Re-entry Vehicle Experiment (IRVE) has successfully demonstrated this technology as a viable candidate with a 3.0 m diameter vehicle sub-orbital flight. To further this technology, large scale HIADs (6.0 8.5 m) must be developed and tested. To characterize the performance of large scale HIAD technology new instrumentation concepts must be developed to accommodate the flexible nature inflatable aeroshell. Many of the concepts that are under consideration for the HIAD FY12 subsonic wind tunnel test series are discussed below.

  1. Long gradient mode and large-scale structure observables

    NASA Astrophysics Data System (ADS)

    Allahyari, Alireza; Firouzjaee, Javad T.

    2017-03-01

    We extend the study of long-mode perturbations to other large-scale observables such as cosmic rulers, galaxy-number counts, and halo bias. The long mode is a pure gradient mode that is still outside an observer's horizon. We insist that gradient-mode effects on observables vanish. It is also crucial that the expressions for observables are relativistic. This allows us to show that the effects of a gradient mode on the large-scale observables vanish identically in a relativistic framework. To study the potential modulation effect of the gradient mode on halo bias, we derive a consistency condition to the first order in gradient expansion. We find that the matter variance at a fixed physical scale is not modulated by the long gradient mode perturbations when the consistency condition holds. This shows that the contribution of long gradient modes to bias vanishes in this framework.

  2. LARGE SCALE PURIFICATION OF PROTEINASES FROM CLOSTRIDIUM HISTOLYTICUM FILTRATES

    PubMed Central

    Conklin, David A.; Webster, Marion E.; Altieri, Patricia L.; Berman, Sanford; Lowenthal, Joseph P.; Gochenour, Raymond B.

    1961-01-01

    Conklin, David A. (Walter Reed Army Institute of Research, Washington, D. C.), Marion E. Webster, Patricia L. Altieri, Sanford Berman, Joseph P. Lowenthal, and Raymond B. Gochenour. Large scale purification of proteinases from Clostridium histolyticum filtrates. J. Bacteriol. 82:589–594. 1961.—A method for the large scale preparation and partial purification of Clostridium histolyticum proteinases by fractional precipitation with ammonium sulfate is described. Conditions for adequate separation and purification of the δ-proteinase and the gelatinase were obtained. Collagenase, on the other hand, was found distributed in four to five fractions and little increase in purity was achieved as compared to the crude ammonium sulfate precipitates. PMID:13880849

  3. Prototype Vector Machine for Large Scale Semi-Supervised Learning

    SciTech Connect

    Zhang, Kai; Kwok, James T.; Parvin, Bahram

    2009-04-29

    Practicaldataminingrarelyfalls exactlyinto the supervisedlearning scenario. Rather, the growing amount of unlabeled data poses a big challenge to large-scale semi-supervised learning (SSL). We note that the computationalintensivenessofgraph-based SSLarises largely from the manifold or graph regularization, which in turn lead to large models that are dificult to handle. To alleviate this, we proposed the prototype vector machine (PVM), a highlyscalable,graph-based algorithm for large-scale SSL. Our key innovation is the use of"prototypes vectors" for effcient approximation on both the graph-based regularizer and model representation. The choice of prototypes are grounded upon two important criteria: they not only perform effective low-rank approximation of the kernel matrix, but also span a model suffering the minimum information loss compared with the complete model. We demonstrate encouraging performance and appealing scaling properties of the PVM on a number of machine learning benchmark data sets.

  4. The Large Scale Synthesis of Aligned Plate Nanostructures

    NASA Astrophysics Data System (ADS)

    Zhou, Yang; Nash, Philip; Liu, Tian; Zhao, Naiqin; Zhu, Shengli

    2016-07-01

    We propose a novel technique for the large-scale synthesis of aligned-plate nanostructures that are self-assembled and self-supporting. The synthesis technique involves developing nanoscale two-phase microstructures through discontinuous precipitation followed by selective etching to remove one of the phases. The method may be applied to any alloy system in which the discontinuous precipitation transformation goes to completion. The resulting structure may have many applications in catalysis, filtering and thermal management depending on the phase selection and added functionality through chemical reaction with the retained phase. The synthesis technique is demonstrated using the discontinuous precipitation of a γ‧ phase, (Ni, Co)3Al, followed by selective dissolution of the γ matrix phase. The production of the nanostructure requires heat treatments on the order of minutes and can be performed on a large scale making this synthesis technique of great economic potential.

  5. Comparative study of large-scale nonlinear optimization methods

    SciTech Connect

    Alemzadeh, S.A.

    1987-01-01

    Solving large-scale nonlinear optimization problems has been one of the active research areas for the last twenty years. Several heuristic algorithms with codes have been developed and implemented since 1966. This study explores the motivation and basic mathematical ideas leading to the development of MINOS-1.0, GRG-2,and MINOS-5.0 algorithms and their codes. The reliability, accuracy, and complexity of the algorithms and software depend upon their use of the gradient, Jacobian, and the Hessian. MINOS-1.0 and GRG-2 incorporate all of the input and output features, while MINOS-1.0 is not able to handle the nonlinearly constrained problems, and GRG-2 is not able to handle large-scale problems, MINOS-5.0 is a robust and an efficient software that incorporates all input, output features.

  6. LARGE-SCALE MOTIONS IN THE PERSEUS GALAXY CLUSTER

    SciTech Connect

    Simionescu, A.; Werner, N.; Urban, O.; Allen, S. W.; Fabian, A. C.; Sanders, J. S.; Mantz, A.; Nulsen, P. E. J.; Takei, Y.

    2012-10-01

    By combining large-scale mosaics of ROSAT PSPC, XMM-Newton, and Suzaku X-ray observations, we present evidence for large-scale motions in the intracluster medium of the nearby, X-ray bright Perseus Cluster. These motions are suggested by several alternating and interleaved X-ray bright, low-temperature, low-entropy arcs located along the east-west axis, at radii ranging from {approx}10 kpc to over a Mpc. Thermodynamic features qualitatively similar to these have previously been observed in the centers of cool-core clusters, and were successfully modeled as a consequence of the gas sloshing/swirling motions induced by minor mergers. Our observations indicate that such sloshing/swirling can extend out to larger radii than previously thought, on scales approaching the virial radius.

  7. The CLASSgal code for relativistic cosmological large scale structure

    SciTech Connect

    Dio, Enea Di; Montanari, Francesco; Durrer, Ruth; Lesgourgues, Julien E-mail: Francesco.Montanari@unige.ch E-mail: Ruth.Durrer@unige.ch

    2013-11-01

    We present accurate and efficient computations of large scale structure observables, obtained with a modified version of the CLASS code which is made publicly available. This code includes all relativistic corrections and computes both the power spectrum C{sub ℓ}(z{sub 1},z{sub 2}) and the corresponding correlation function ξ(θ,z{sub 1},z{sub 2}) of the matter density and the galaxy number fluctuations in linear perturbation theory. For Gaussian initial perturbations, these quantities contain the full information encoded in the large scale matter distribution at the level of linear perturbation theory. We illustrate the usefulness of our code for cosmological parameter estimation through a few simple examples.

  8. Large-Scale Optimization for Bayesian Inference in Complex Systems

    SciTech Connect

    Willcox, Karen; Marzouk, Youssef

    2013-11-12

    The SAGUARO (Scalable Algorithms for Groundwater Uncertainty Analysis and Robust Optimization) Project focused on the development of scalable numerical algorithms for large-scale Bayesian inversion in complex systems that capitalize on advances in large-scale simulation-based optimization and inversion methods. The project was a collaborative effort among MIT, the University of Texas at Austin, Georgia Institute of Technology, and Sandia National Laboratories. The research was directed in three complementary areas: efficient approximations of the Hessian operator, reductions in complexity of forward simulations via stochastic spectral approximations and model reduction, and employing large-scale optimization concepts to accelerate sampling. The MIT--Sandia component of the SAGUARO Project addressed the intractability of conventional sampling methods for large-scale statistical inverse problems by devising reduced-order models that are faithful to the full-order model over a wide range of parameter values; sampling then employs the reduced model rather than the full model, resulting in very large computational savings. Results indicate little effect on the computed posterior distribution. On the other hand, in the Texas--Georgia Tech component of the project, we retain the full-order model, but exploit inverse problem structure (adjoint-based gradients and partial Hessian information of the parameter-to-observation map) to implicitly extract lower dimensional information on the posterior distribution; this greatly speeds up sampling methods, so that fewer sampling points are needed. We can think of these two approaches as ``reduce then sample'' and ``sample then reduce.'' In fact, these two approaches are complementary, and can be used in conjunction with each other. Moreover, they both exploit deterministic inverse problem structure, in the form of adjoint-based gradient and Hessian information of the underlying parameter-to-observation map, to achieve their

  9. Turbulent amplification of large-scale magnetic fields

    NASA Technical Reports Server (NTRS)

    Montgomery, D.; Chen, H.

    1984-01-01

    Previously-introduced methods for analytically estimating the effects of small-scale turbulent fluctuations on large-scale dynamics are extended to fully three-dimensional magnetohydrodynamics. The problem becomes algebraically tractable in the presence of sufficiently large spectral gaps. The calculation generalizes 'alpha dynamo' calculations, except that the velocity fluctuations and magnetic fluctuations are treated on an independent and equal footing. Earlier expressions for the 'alpha coefficients' of turbulent magnetic field amplification are recovered as a special case.

  10. Concurrent Programming Using Actors: Exploiting Large-Scale Parallelism,

    DTIC Science & Technology

    1985-10-07

    ORGANIZATION NAME AND ADDRESS 10. PROGRAM ELEMENT. PROJECT. TASK* Artificial Inteligence Laboratory AREA Is WORK UNIT NUMBERS 545 Technology Square...D-R162 422 CONCURRENT PROGRMMIZNG USING f"OS XL?ITP TEH l’ LARGE-SCALE PARALLELISH(U) NASI AC E Al CAMBRIDGE ARTIFICIAL INTELLIGENCE L. G AGHA ET AL...RESOLUTION TEST CHART N~ATIONAL BUREAU OF STANDA.RDS - -96 A -E. __ _ __ __’ .,*- - -- •. - MASSACHUSETTS INSTITUTE OF TECHNOLOGY ARTIFICIAL

  11. Host Immunity via Mutable Virtualized Large-Scale Network Containers

    DTIC Science & Technology

    2016-07-25

    migrate to different IP addresses multiple 6mes. We implement a virtual machine based system prototype and evaluate it using state-of-the-a1t scanning...entire !Pv4 address space within 5 Host Immunity via Mutable Virtualized Large-Scale Network Containers 45 minutes from a single machine . Second, when...that the attacker will be trapped into one decoy instead of the real server. We implement a virtual machine (VM)-based prototype that integrates

  12. Developing and Understanding Methods for Large Scale Nonlinear Optimization

    DTIC Science & Technology

    2001-12-01

    development of new algorithms for large-scale uncon- strained and constrained optimization problems, including limited-memory methods for problems with...analysis of tensor and SQP methods for singular con- strained optimization", to appear in SIAM Journal on Optimization. Published in peer-reviewed...Mathematica, Vol III, Journal der Deutschen Mathematiker-Vereinigung, 1998. S. Crivelli, B. Bader, R. Byrd, E. Eskow, V. Lamberti , R.Schnabel and T

  13. Wiggly cosmic strings, neutrinos and large-scale structure

    NASA Astrophysics Data System (ADS)

    Vachaspati, Tanmay

    1993-04-01

    We discuss the cosmic string scenario of large-scale structure formation in light of the result that the strings are not smooth but instead have a lot of sub-structure or wiggles on them. It appears from the results of Albrecht and Stebbins that the scenario works best if the universe is dominated by massive neutrinos or some other form of hot dark matter. Some unique features of the scenario, such as the generation of primordial magnetic fields, are also described.

  14. Analysis plan for 1985 large-scale tests. Technical report

    SciTech Connect

    McMullan, F.W.

    1983-01-01

    The purpose of this effort is to assist DNA in planning for large-scale (upwards of 5000 tons) detonations of conventional explosives in the 1985 and beyond time frame. Primary research objectives were to investigate potential means to increase blast duration and peak pressures. This report identifies and analyzes several candidate explosives. It examines several charge designs and identifies advantages and disadvantages of each. Other factors including terrain and multiburst techniques are addressed as are test site considerations.

  15. Multimodel Design of Large Scale Systems with Multiple Decision Makers.

    DTIC Science & Technology

    1982-08-01

    virtue. 5- , Lead me from darkneu to light. - Lead me from death to eternal Life. ( Vedic Payer) p. I, MULTIMODEL DESIGN OF LARGE SCALE SYSTEMS WITH...guidance during the course of *: this research . He would also like to thank Professors W. R. Perkins, P. V. Kokotovic, T. Basar, and T. N. Trick for...thesis concludes with Chapter 7 where we summarize the results obtained, outline the main contributions, and indicate directions for future research . 7- I

  16. Critical Problems in Very Large Scale Computer Systems

    DTIC Science & Technology

    1990-03-31

    MAY I i9cu( CRITICAL PROBLEMS IN VERY LARGE SCALE COMPUTER SYSTEMS Semiannual Technical Report for the Period October 1, 1989 to...suitability for supporting popular models of parallel computation . During the reporting period they have developed an interface definition. A simulator has...queries in computational geometry . Range queries are a fundamental problem in computational geometry with applications to computer graphics and

  17. Supporting large scale applications on networks of workstations

    NASA Technical Reports Server (NTRS)

    Cooper, Robert; Birman, Kenneth P.

    1989-01-01

    Distributed applications on networks of workstations are an increasingly common way to satisfy computing needs. However, existing mechanisms for distributed programming exhibit poor performance and reliability as application size increases. Extension of the ISIS distributed programming system to support large scale distributed applications by providing hierarchical process groups is discussed. Incorporation of hierarchy in the program structure and exploitation of this to limit the communication and storage required in any one component of the distributed system is examined.

  18. Large Scale Airflow Perturbations and Resultant Dune Dynamics

    NASA Astrophysics Data System (ADS)

    Smith, Alexander B.; Jackson, Derek W. T.; Cooper, J. Andrew G.; Beyers, Meiring

    2017-04-01

    Large-scale atmospheric turbulence can have a large impact on the regional wind regime effecting dune environments. Depending on the incident angle of mesoscale airflow, local topographic steering can also alter wind conditions and subsequent aeolian dynamics. This research analyses the influence of large-scale airflow perturbations occurring at the Maspalomas dunefield located on the southern coast of Gran Canaria, Spain. These perturbations in turn significantly influence the morphometry and migration rates of barchan dunes, monitored at the study site through time. The main meteorological station on Gran Canaria records highly uni-modal NNE wind conditions; however, simultaneously measured winds are highly variable around the island, showing a high degree of steering. Large Eddy Simulations (LES) were used to identify large-scale airflow perturbations around the island of Gran Canaria during NNE, N, and NNW incident flow directions. Results indicate that approaching surface airflow bifurcates around the island's coastline before converging at the lee coast. Winds in areas located around the islands lateral coast are controlled by these diverging flow patterns, whereas lee-side areas are influenced primarily by the islands upwind canyon topography leading to highly turbulent flow. Characteristic turbulent eddies show a complex wind environment at Maspalomas with winds diverging-converging up to 180° between the eastern and western sections of the dunefield. Multi-directional flow conditions lead to highly altered dune dynamics including the production of temporary slip faces on the stoss slopes, rapid reduction in crest height and slope length, and development of bi-crested dunes. This indicates a distinct bi-modality of airflow conditions that control the geomorphic evolution of the dunefield. Variability in wind conditions is not evident in the long-term meteorological records on the island, indicating the significance of large scale atmospheric steering on

  19. A Holistic Management Architecture for Large-Scale Adaptive Networks

    DTIC Science & Technology

    2007-09-01

    MANAGEMENT ARCHITECTURE FOR LARGE-SCALE ADAPTIVE NETWORKS by Michael R. Clement September 2007 Thesis Advisor: Alex Bordetsky Second Reader...TECHNOLOGY MANAGEMENT from the NAVAL POSTGRADUATE SCHOOL September 2007 Author: Michael R. Clement Approved by: Dr. Alex ...achieve in life is by His will. Ad Majorem Dei Gloriam. To my parents, my family, and Caitlin: For supporting me, listening to me when I got

  20. A Cloud Computing Platform for Large-Scale Forensic Computing

    NASA Astrophysics Data System (ADS)

    Roussev, Vassil; Wang, Liqiang; Richard, Golden; Marziale, Lodovico

    The timely processing of massive digital forensic collections demands the use of large-scale distributed computing resources and the flexibility to customize the processing performed on the collections. This paper describes MPI MapReduce (MMR), an open implementation of the MapReduce processing model that outperforms traditional forensic computing techniques. MMR provides linear scaling for CPU-intensive processing and super-linear scaling for indexing-related workloads.

  1. Large-Scale Weather Disturbances in Mars’ Southern Extratropics

    NASA Astrophysics Data System (ADS)

    Hollingsworth, Jeffery L.; Kahre, Melinda A.

    2015-11-01

    Between late autumn and early spring, Mars’ middle and high latitudes within its atmosphere support strong mean thermal gradients between the tropics and poles. Observations from both the Mars Global Surveyor (MGS) and Mars Reconnaissance Orbiter (MRO) indicate that this strong baroclinicity supports intense, large-scale eastward traveling weather systems (i.e., transient synoptic-period waves). These extratropical weather disturbances are key components of the global circulation. Such wave-like disturbances act as agents in the transport of heat and momentum, and generalized scalar/tracer quantities (e.g., atmospheric dust, water-vapor and ice clouds). The character of large-scale, traveling extratropical synoptic-period disturbances in Mars' southern hemisphere during late winter through early spring is investigated using a moderately high-resolution Mars global climate model (Mars GCM). This Mars GCM imposes interactively lifted and radiatively active dust based on a threshold value of the surface stress. The model exhibits a reasonable "dust cycle" (i.e., globally averaged, a dustier atmosphere during southern spring and summer occurs). Compared to their northern-hemisphere counterparts, southern synoptic-period weather disturbances and accompanying frontal waves have smaller meridional and zonal scales, and are far less intense. Influences of the zonally asymmetric (i.e., east-west varying) topography on southern large-scale weather are examined. Simulations that adapt Mars’ full topography compared to simulations that utilize synthetic topographies emulating key large-scale features of the southern middle latitudes indicate that Mars’ transient barotropic/baroclinic eddies are highly influenced by the great impact basins of this hemisphere (e.g., Argyre and Hellas). The occurrence of a southern storm zone in late winter and early spring appears to be anchored to the western hemisphere via orographic influences from the Tharsis highlands, and the Argyre

  2. The large-scale anisotropy with the PAMELA calorimeter

    NASA Astrophysics Data System (ADS)

    Karelin, A.; Adriani, O.; Barbarino, G.; Bazilevskaya, G.; Bellotti, R.; Boezio, M.; Bogomolov, E.; Bongi, M.; Bonvicini, V.; Bottai, S.; Bruno, A.; Cafagna, F.; Campana, D.; Carbone, R.; Carlson, P.; Casolino, M.; Castellini, G.; De Donato, C.; De Santis, C.; De Simone, N.; Di Felice, V.; Formato, V.; Galper, A.; Koldashov, S.; Koldobskiy, S.; Krut'kov, S.; Kvashnin, A.; Leonov, A.; Malakhov, V.; Marcelli, L.; Martucci, M.; Mayorov, A.; Menn, W.; Mergé, M.; Mikhailov, V.; Mocchiutti, E.; Monaco, A.; Mori, N.; Munini, R.; Osteria, G.; Palma, F.; Panico, B.; Papini, P.; Pearce, M.; Picozza, P.; Ricci, M.; Ricciarini, S.; Sarkar, R.; Simon, M.; Scotti, V.; Sparvoli, R.; Spillantini, P.; Stozhkov, Y.; Vacchi, A.; Vannuccini, E.; Vasilyev, G.; Voronov, S.; Yurkin, Y.; Zampa, G.; Zampa, N.

    2015-10-01

    The large-scale anisotropy (or the so-called star-diurnal wave) has been studied using the calorimeter of the space-born experiment PAMELA. The cosmic ray anisotropy has been obtained for the Southern and Northern hemispheres simultaneously in the equatorial coordinate system for the time period 2006-2014. The dipole amplitude and phase have been measured for energies 1-20 TeV n-1.

  3. Space transportation booster engine thrust chamber technology, large scale injector

    NASA Technical Reports Server (NTRS)

    Schneider, J. A.

    1993-01-01

    The objective of the Large Scale Injector (LSI) program was to deliver a 21 inch diameter, 600,000 lbf thrust class injector to NASA/MSFC for hot fire testing. The hot fire test program would demonstrate the feasibility and integrity of the full scale injector, including combustion stability, chamber wall compatibility (thermal management), and injector performance. The 21 inch diameter injector was delivered in September of 1991.

  4. Large Scale Density Estimation of Blue and Fin Whales (LSD)

    DTIC Science & Technology

    2014-09-30

    1 DISTRIBUTION STATEMENT A. Approved for public release; distribution is unlimited. Large Scale Density Estimation of Blue and Fin Whales ...estimating blue and fin whale density that is effective over large spatial scales and is designed to cope with spatial variation in animal density utilizing...a density estimation methodology for quantifying blue and fin whale abundance from passive acoustic data recorded on sparse hydrophone arrays in the

  5. Relic vector field and CMB large scale anomalies

    SciTech Connect

    Chen, Xingang; Wang, Yi E-mail: yw366@cam.ac.uk

    2014-10-01

    We study the most general effects of relic vector fields on the inflationary background and density perturbations. Such effects are observable if the number of inflationary e-folds is close to the minimum requirement to solve the horizon problem. We show that this can potentially explain two CMB large scale anomalies: the quadrupole-octopole alignment and the quadrupole power suppression. We discuss its effect on the parity anomaly. We also provide analytical template for more detailed data comparison.

  6. Large-scale controls on convective extreme precipitation

    NASA Astrophysics Data System (ADS)

    Loriaux, Jessica M.; Lenderink, Geert; Pier Siebesma, A.

    2017-04-01

    The influence of large-scale conditions on extreme precipitation is not yet understood well enough. We will present the results of Loriaux et al. (2017), in which we investigate the role of large-scale dynamics and environmental conditions on precipitation and on the precipitation response to climate change. To this end, we have set up a composite LES case for convective precipitation using strong large-scale forcing based on idealized profiles for the highest 10 percentiles of peak intensities over the Netherlands, as described by Loriaux et al. (2016). In this setting, we have performed sensitivity analyses for atmospheric stability, large-scale moisture convergence, and relative humidity, and compared present-day climate to a warmer future climate. The results suggest that amplification of the moisture convergence and destabilization of the atmosphere both lead to an increase in precipitation, but due to different effects; Atmospheric stability mainly influences the precipitation intensity, while the moisture convergence mainly controls the precipitation area fraction. Extreme precipitation intensities show qualitatively similar sensitivities to atmospheric stability and moisture convergence. Precipitation increases with RH due to an increase in area fraction, despite a decrease in intensity. The precipitation response to the climate perturbation shows a stronger response for the precipitation intensity than the overall precipitation, with no clear dependency of changes in atmospheric stability, moisture convergence and relative humidity. The difference in response between the precipitation intensity and overall precipitation is caused by a decrease in the precipitation area fraction from present-day to future climate. In other words, our climate perturbation indicates that with warming, it will rain more intensely but in less places. Loriaux, J.M., G. Lenderink, and A.P. Siebesma, 2016, doi: 10.1002/2015JD024274 Loriaux, J.M., G. Lenderink, and A.P. Siebesma

  7. On a Game of Large-Scale Projects Competition

    NASA Astrophysics Data System (ADS)

    Nikonov, Oleg I.; Medvedeva, Marina A.

    2009-09-01

    The paper is devoted to game-theoretical control problems motivated by economic decision making situations arising in realization of large-scale projects, such as designing and putting into operations the new gas or oil pipelines. A non-cooperative two player game is considered with payoff functions of special type for which standard existence theorems and algorithms for searching Nash equilibrium solutions are not applicable. The paper is based on and develops the results obtained in [1]-[5].

  8. Measuring large scale space perception in literary texts

    NASA Astrophysics Data System (ADS)

    Rossi, Paolo

    2007-07-01

    A center and radius of “perception” (in the sense of environmental cognition) can be formally associated with a written text and operationally defined. Simple algorithms for their computation are presented, and indicators for anisotropy in large scale space perception are introduced. The relevance of these notions for the analysis of literary and historical records is briefly discussed and illustrated with an example taken from medieval historiography.

  9. Semantic Concept Discovery for Large Scale Zero Shot Event Detection

    DTIC Science & Technology

    2015-07-25

    NO. 0704-0188 3. DATES COVERED (From - To) - UU UU UU UU 18-08-2015 Approved for public release; distribution is unlimited. Semantic Concept Discovery ...Office P.O. Box 12211 Research Triangle Park, NC 27709-2211 zero shot event detection, semantic concept discovery REPORT DOCUMENTATION PAGE 11...Mellon University 5000 Forbes Avenue Pittsburgh, PA 15213 -3815 ABSTRACT Semantic Concept Discovery for Large-Scale Zero-Shot Event Detection Report

  10. Large-scale Alfvén vortices

    SciTech Connect

    Onishchenko, O. G.; Horton, W.; Scullion, E.; Fedun, V.

    2015-12-15

    The new type of large-scale vortex structures of dispersionless Alfvén waves in collisionless plasma is investigated. It is shown that Alfvén waves can propagate in the form of Alfvén vortices of finite characteristic radius and characterised by magnetic flux ropes carrying orbital angular momentum. The structure of the toroidal and radial velocity, fluid and magnetic field vorticity, the longitudinal electric current in the plane orthogonal to the external magnetic field are discussed.

  11. The Phoenix series large scale LNG pool fire experiments.

    SciTech Connect

    Simpson, Richard B.; Jensen, Richard Pearson; Demosthenous, Byron; Luketa, Anay Josephine; Ricks, Allen Joseph; Hightower, Marion Michael; Blanchat, Thomas K.; Helmick, Paul H.; Tieszen, Sheldon Robert; Deola, Regina Anne; Mercier, Jeffrey Alan; Suo-Anttila, Jill Marie; Miller, Timothy J.

    2010-12-01

    The increasing demand for natural gas could increase the number and frequency of Liquefied Natural Gas (LNG) tanker deliveries to ports across the United States. Because of the increasing number of shipments and the number of possible new facilities, concerns about the potential safety of the public and property from an accidental, and even more importantly intentional spills, have increased. While improvements have been made over the past decade in assessing hazards from LNG spills, the existing experimental data is much smaller in size and scale than many postulated large accidental and intentional spills. Since the physics and hazards from a fire change with fire size, there are concerns about the adequacy of current hazard prediction techniques for large LNG spills and fires. To address these concerns, Congress funded the Department of Energy (DOE) in 2008 to conduct a series of laboratory and large-scale LNG pool fire experiments at Sandia National Laboratories (Sandia) in Albuquerque, New Mexico. This report presents the test data and results of both sets of fire experiments. A series of five reduced-scale (gas burner) tests (yielding 27 sets of data) were conducted in 2007 and 2008 at Sandia's Thermal Test Complex (TTC) to assess flame height to fire diameter ratios as a function of nondimensional heat release rates for extrapolation to large-scale LNG fires. The large-scale LNG pool fire experiments were conducted in a 120 m diameter pond specially designed and constructed in Sandia's Area III large-scale test complex. Two fire tests of LNG spills of 21 and 81 m in diameter were conducted in 2009 to improve the understanding of flame height, smoke production, and burn rate and therefore the physics and hazards of large LNG spills and fires.

  12. Rapid Discovery of Continuous-Performance Compounds and Powernap Compounds Through Large-Scale Mutagenesis in Drosophila

    DTIC Science & Technology

    2008-04-22

    CL, Vyazovskiy VV, Southard TL, Chiu S-Y, Messing A, Tononi G, Cirelli C (2007). Sleep in Kcna2 knockout mice. BMC Biology, 5(1):42. Jones S... Kcna2 knockout mice. BMC Biol 5:42. Faraguna U, Douglas CL, Cirelli C, Tononi G (2006) Two-pore domain potassium channel TASK-1: effects on sleep and

  13. Ultra-high frequency ultrasound biomicroscopy and high throughput cardiovascular phenotyping in a large scale mouse mutagenesis screen

    NASA Astrophysics Data System (ADS)

    Liu, Xiaoqin; Francis, Richard; Tobita, Kimimasa; Kim, Andy; Leatherbury, Linda; Lo, Cecilia W.

    2013-02-01

    Ultrasound biomicroscopy (UBM) is ideally suited for phenotyping fetal mice for congenital heart disease (CHD), as imaging can be carried out noninvasively to provide both hemodynamic and structural information essential for CHD diagnosis. Using the UBM (Vevo 2100; 40Hz) in conjunction with the clinical ultrasound system (Acuson Sequioa C512; 15Hz), we developed a two-step screening protocol to scan thousands fetuses derived from ENU mutagenized pedigrees. A wide spectrum of CHD was detected by the UBM, which were subsequently confirmed with follow-up necropsy and histopathology examination with episcopic fluorescence image capture. CHD observed included outflow anomalies, left/right heart obstructive lesions, septal/valvular defects and cardiac situs anomalies. Meanwhile, various extracardiac defects were found, such as polydactyly, craniofacial defects, exencephaly, omphalocele-cleft palate, most of which were associated with cardiac defects. Our analyses showed the UBM was better at assessing cardiac structure and blood flow profiles, while conventional ultrasound allowed higher throughput low-resolution screening. Our study showed the integration of conventional clinical ultrasound imaging with the UBM for fetal mouse cardiovascular phenotyping can maximize the detection and recovery of CHD mutants.

  14. Large-scale quantization from local correlations in space plasmas

    NASA Astrophysics Data System (ADS)

    Livadiotis, George; McComas, David J.

    2014-05-01

    This study examines the large-scale quantization that can characterize the phase space of certain physical systems. Plasmas are such systems where large-scale quantization, ħ*, is caused by Debye shielding that structures correlations between particles. The value of ħ* is constant—some 12 orders of magnitude larger than the Planck constant—across a wide range of space plasmas, from the solar wind in the inner heliosphere to the distant plasma in the inner heliosheath and the local interstellar medium. This paper develops the foundation and advances the understanding of the concept of plasma quantization; in particular, we (i) show the analogy of plasma to Planck quantization, (ii) show the key points of plasma quantization, (iii) construct some basic quantum mechanical concepts for the large-scale plasma quantization, (iv) investigate the correlation between plasma parameters that implies plasma quantization, when it is approximated by a relation between the magnetosonic energy and the plasma frequency, (v) analyze typical space plasmas throughout the heliosphere and show the constancy of plasma quantization over many orders of magnitude in plasma parameters, (vi) analyze Advanced Composition Explorer (ACE) solar wind measurements to develop another measurement of the value of ħ*, and (vii) apply plasma quantization to derive unknown plasma parameters when some key observable is missing.

  15. Large-scale investigation of genomic markers for severe periodontitis.

    PubMed

    Suzuki, Asami; Ji, Guijin; Numabe, Yukihiro; Ishii, Keisuke; Muramatsu, Masaaki; Kamoi, Kyuichi

    2004-09-01

    The purpose of the present study was to investigate the genomic markers for periodontitis, using large-scale single-nucleotide polymorphism (SNP) association studies comparing healthy volunteers and patients with periodontitis. Genomic DNA was obtained from 19 healthy volunteers and 22 patients with severe periodontitis, all of whom were Japanese. The subjects were genotyped at 637 SNPs in 244 genes on a large scale, using the TaqMan polymerase chain reaction (PCR) system. Statistically significant differences in allele and genotype frequencies were analyzed with Fisher's exact test. We found statistically significant differences (P < 0.01) between the healthy volunteers and patients with severe periodontitis in the following genes; gonadotropin-releasing hormone 1 (GNRH1), phosphatidylinositol 3-kinase regulatory 1 (PIK3R1), dipeptidylpeptidase 4 (DPP4), fibrinogen-like 2 (FGL2), and calcitonin receptor (CALCR). These results suggest that SNPs in the GNRH1, PIK3R1, DPP4, FGL2, and CALCR genes are genomic markers for severe periodontitis. Our findings indicate the necessity of analyzing SNPs in genes on a large scale (i.e., genome-wide approach), to identify genomic markers for periodontitis.

  16. Geospatial Optimization of Siting Large-Scale Solar Projects

    SciTech Connect

    Macknick, J.; Quinby, T.; Caulfield, E.; Gerritsen, M.; Diffendorfer, J.; Haines, S.

    2014-03-01

    Recent policy and economic conditions have encouraged a renewed interest in developing large-scale solar projects in the U.S. Southwest. However, siting large-scale solar projects is complex. In addition to the quality of the solar resource, solar developers must take into consideration many environmental, social, and economic factors when evaluating a potential site. This report describes a proof-of-concept, Web-based Geographical Information Systems (GIS) tool that evaluates multiple user-defined criteria in an optimization algorithm to inform discussions and decisions regarding the locations of utility-scale solar projects. Existing siting recommendations for large-scale solar projects from governmental and non-governmental organizations are not consistent with each other, are often not transparent in methods, and do not take into consideration the differing priorities of stakeholders. The siting assistance GIS tool we have developed improves upon the existing siting guidelines by being user-driven, transparent, interactive, capable of incorporating multiple criteria, and flexible. This work provides the foundation for a dynamic siting assistance tool that can greatly facilitate siting decisions among multiple stakeholders.

  17. Large-scale data mining pilot project in human genome

    SciTech Connect

    Musick, R.; Fidelis, R.; Slezak, T.

    1997-05-01

    This whitepaper briefly describes a new, aggressive effort in large- scale data Livermore National Labs. The implications of `large- scale` will be clarified Section. In the short term, this effort will focus on several @ssion-critical questions of Genome project. We will adapt current data mining techniques to the Genome domain, to quantify the accuracy of inference results, and lay the groundwork for a more extensive effort in large-scale data mining. A major aspect of the approach is that we will be fully-staffed data warehousing effort in the human Genome area. The long term goal is strong applications- oriented research program in large-@e data mining. The tools, skill set gained will be directly applicable to a wide spectrum of tasks involving a for large spatial and multidimensional data. This includes applications in ensuring non-proliferation, stockpile stewardship, enabling Global Ecology (Materials Database Industrial Ecology), advancing the Biosciences (Human Genome Project), and supporting data for others (Battlefield Management, Health Care).

  18. A model of plasma heating by large-scale flow

    NASA Astrophysics Data System (ADS)

    Pongkitiwanichakul, P.; Cattaneo, F.; Boldyrev, S.; Mason, J.; Perez, J. C.

    2015-12-01

    In this work, we study the process of energy dissipation triggered by a slow large-scale motion of a magnetized conducting fluid. Our consideration is motivated by the problem of heating the solar corona, which is believed to be governed by fast reconnection events set off by the slow motion of magnetic field lines anchored in the photospheric plasma. To elucidate the physics governing the disruption of the imposed laminar motion and the energy transfer to small scales, we propose a simplified model where the large-scale motion of magnetic field lines is prescribed not at the footpoints but rather imposed volumetrically. As a result, the problem can be treated numerically with an efficient, highly accurate spectral method, allowing us to use a resolution and statistical ensemble exceeding those of the previous work. We find that, even though the large-scale deformations are slow, they eventually lead to reconnection events that drive a turbulent state at smaller scales. The small-scale turbulence displays many of the universal features of field-guided magnetohydrodynamic turbulence like a well-developed inertial range spectrum. Based on these observations, we construct a phenomenological model that gives the scalings of the amplitude of the fluctuations and the energy-dissipation rate as functions of the input parameters. We find good agreement between the numerical results and the predictions of the model.

  19. Large-scale biodiversity patterns in freshwater phytoplankton.

    PubMed

    Stomp, Maayke; Huisman, Jef; Mittelbach, Gary G; Litchman, Elena; Klausmeier, Christopher A

    2011-11-01

    Our planet shows striking gradients in the species richness of plants and animals, from high biodiversity in the tropics to low biodiversity in polar and high-mountain regions. Recently, similar patterns have been described for some groups of microorganisms, but the large-scale biogeographical distribution of freshwater phytoplankton diversity is still largely unknown. We examined the species diversity of freshwater phytoplankton sampled from 540 lakes and reservoirs distributed across the continental United States and found strong latitudinal, longitudinal, and altitudinal gradients in phytoplankton biodiversity, demonstrating that microorganisms can show substantial geographic variation in biodiversity. Detailed analysis using structural equation models indicated that these large-scale biodiversity gradients in freshwater phytoplankton diversity were mainly driven by local environmental factors, although there were residual direct effects of latitude, longitude, and altitude as well. Specifically, we found that phytoplankton species richness was an increasing saturating function of lake chlorophyll a concentration, increased with lake surface area and possibly increased with water temperature, resembling effects of productivity, habitat area, and temperature on diversity patterns commonly observed for macroorganisms. In turn, these local environmental factors varied along latitudinal, longitudinal, and altitudinal gradients. These results imply that changes in land use or climate that affect these local environmental factors are likely to have major impacts on large-scale biodiversity patterns of freshwater phytoplankton.

  20. Homogenization of Large-Scale Movement Models in Ecology

    USGS Publications Warehouse

    Garlick, M.J.; Powell, J.A.; Hooten, M.B.; McFarlane, L.R.

    2011-01-01

    A difficulty in using diffusion models to predict large scale animal population dispersal is that individuals move differently based on local information (as opposed to gradients) in differing habitat types. This can be accommodated by using ecological diffusion. However, real environments are often spatially complex, limiting application of a direct approach. Homogenization for partial differential equations has long been applied to Fickian diffusion (in which average individual movement is organized along gradients of habitat and population density). We derive a homogenization procedure for ecological diffusion and apply it to a simple model for chronic wasting disease in mule deer. Homogenization allows us to determine the impact of small scale (10-100 m) habitat variability on large scale (10-100 km) movement. The procedure generates asymptotic equations for solutions on the large scale with parameters defined by small-scale variation. The simplicity of this homogenization procedure is striking when compared to the multi-dimensional homogenization procedure for Fickian diffusion,and the method will be equally straightforward for more complex models. ?? 2010 Society for Mathematical Biology.

  1. Channel capacity of next generation large scale MIMO systems

    NASA Astrophysics Data System (ADS)

    Alshammari, A.; Albdran, S.; Matin, M.

    2016-09-01

    Information rate that can be transferred over a given bandwidth is limited by the information theory. Capacity depends on many factors such as the signal to noise ratio (SNR), channel state information (CSI) and the spatial correlation in the propagation environment. It is very important to increase spectral efficiency in order to meet the growing demand for wireless services. Thus, Multiple input multiple output (MIMO) technology has been developed and applied in most of the wireless standards and it has been very successful in increasing capacity and reliability. As the demand is still increasing, attention now is shifting towards large scale multiple input multiple output (MIMO) which has a potential of bringing orders of magnitude of improvement in spectral and energy efficiency. It has been shown that users channels decorrelate after increasing the number of antennas. As a result, inter-user interference can be avoided since energy can be focused on precise directions. This paper investigates the limits of channel capacity for large scale MIMO. We study the relation between spectral efficiency and the number of antenna N. We use time division duplex (TDD) system in order to obtain CSI using training sequence in the uplink. The same CSI is used for the downlink because the channel is reciprocal. Spectral efficiency is measured for channel model that account for small scale fading while ignoring the effect of large scale fading. It is shown the spectral efficiency can be improved significantly when compared to single antenna systems in ideal circumstances.

  2. Sparse approximation through boosting for learning large scale kernel machines.

    PubMed

    Sun, Ping; Yao, Xin

    2010-06-01

    Recently, sparse approximation has become a preferred method for learning large scale kernel machines. This technique attempts to represent the solution with only a subset of original data points also known as basis vectors, which are usually chosen one by one with a forward selection procedure based on some selection criteria. The computational complexity of several resultant algorithms scales as O(NM(2)) in time and O(NM) in memory, where N is the number of training points and M is the number of basis vectors as well as the steps of forward selection. For some large scale data sets, to obtain a better solution, we are sometimes required to include more basis vectors, which means that M is not trivial in this situation. However, the limited computational resource (e.g., memory) prevents us from including too many vectors. To handle this dilemma, we propose to add an ensemble of basis vectors instead of only one at each forward step. The proposed method, closely related to gradient boosting, could decrease the required number M of forward steps significantly and thus a large fraction of computational cost is saved. Numerical experiments on three large scale regression tasks and a classification problem demonstrate the effectiveness of the proposed approach.

  3. Dispersal Mutualism Incorporated into Large-Scale, Infrequent Disturbances.

    PubMed

    Parker, V Thomas

    2015-01-01

    Because of their influence on succession and other community interactions, large-scale, infrequent natural disturbances also should play a major role in mutualistic interactions. Using field data and experiments, I test whether mutualisms have been incorporated into large-scale wildfire by whether the outcomes of a mutualism depend on disturbance. In this study a seed dispersal mutualism is shown to depend on infrequent, large-scale disturbances. A dominant shrubland plant (Arctostaphylos species) produces seeds that make up a persistent soil seed bank and requires fire to germinate. In post-fire stands, I show that seedlings emerging from rodent caches dominate sites experiencing higher fire intensity. Field experiments show that rodents (Perimyscus californicus, P. boylii) do cache Arctostaphylos fruit and bury most seed caches to a sufficient depth to survive a killing heat pulse that a fire might drive into the soil. While the rodent dispersal and caching behavior itself has not changed compared to other habitats, the environmental transformation caused by wildfire converts the caching burial of seed from a dispersal process to a plant fire adaptive trait, and provides the context for stimulating subsequent life history evolution in the plant host.

  4. Line segment extraction for large scale unorganized point clouds

    NASA Astrophysics Data System (ADS)

    Lin, Yangbin; Wang, Cheng; Cheng, Jun; Chen, Bili; Jia, Fukai; Chen, Zhonggui; Li, Jonathan

    2015-04-01

    Line segment detection in images is already a well-investigated topic, although it has received considerably less attention in 3D point clouds. Benefiting from current LiDAR devices, large-scale point clouds are becoming increasingly common. Most human-made objects have flat surfaces. Line segments that occur where pairs of planes intersect give important information regarding the geometric content of point clouds, which is especially useful for automatic building reconstruction and segmentation. This paper proposes a novel method that is capable of accurately extracting plane intersection line segments from large-scale raw scan points. The 3D line-support region, namely, a point set near a straight linear structure, is extracted simultaneously. The 3D line-support region is fitted by our Line-Segment-Half-Planes (LSHP) structure, which provides a geometric constraint for a line segment, making the line segment more reliable and accurate. We demonstrate our method on the point clouds of large-scale, complex, real-world scenes acquired by LiDAR devices. We also demonstrate the application of 3D line-support regions and their LSHP structures on urban scene abstraction.

  5. [A large-scale accident in Alpine terrain].

    PubMed

    Wildner, M; Paal, P

    2015-02-01

    Due to the geographical conditions, large-scale accidents amounting to mass casualty incidents (MCI) in Alpine terrain regularly present rescue teams with huge challenges. Using an example incident, specific conditions and typical problems associated with such a situation are presented. The first rescue team members to arrive have the elementary tasks of qualified triage and communication to the control room, which is required to dispatch the necessary additional support. Only with a clear "concept", to which all have to adhere, can the subsequent chaos phase be limited. In this respect, a time factor confounded by adverse weather conditions or darkness represents enormous pressure. Additional hazards are frostbite and hypothermia. If priorities can be established in terms of urgency, then treatment and procedure algorithms have proven successful. For evacuation of causalities, a helicopter should be strived for. Due to the low density of hospitals in Alpine regions, it is often necessary to distribute the patients over a wide area. Rescue operations in Alpine terrain have to be performed according to the particular conditions and require rescue teams to have specific knowledge and expertise. The possibility of a large-scale accident should be considered when planning events. With respect to optimization of rescue measures, regular training and exercises are rational, as is the analysis of previous large-scale Alpine accidents.

  6. Large scale structure in universes dominated by cold dark matter

    NASA Technical Reports Server (NTRS)

    Bond, J. Richard

    1986-01-01

    The theory of Gaussian random density field peaks is applied to a numerical study of the large-scale structure developing from adiabatic fluctuations in models of biased galaxy formation in universes with Omega = 1, h = 0.5 dominated by cold dark matter (CDM). The angular anisotropy of the cross-correlation function demonstrates that the far-field regions of cluster-scale peaks are asymmetric, as recent observations indicate. These regions will generate pancakes or filaments upon collapse. One-dimensional singularities in the large-scale bulk flow should arise in these CDM models, appearing as pancakes in position space. They are too rare to explain the CfA bubble walls, but pancakes that are just turning around now are sufficiently abundant and would appear to be thin walls normal to the line of sight in redshift space. Large scale streaming velocities are significantly smaller than recent observations indicate. To explain the reported 700 km/s coherent motions, mass must be significantly more clustered than galaxies with a biasing factor of less than 0.4 and a nonlinear redshift at cluster scales greater than one for both massive neutrino and cold models.

  7. Learning Short Binary Codes for Large-scale Image Retrieval.

    PubMed

    Liu, Li; Yu, Mengyang; Shao, Ling

    2017-03-01

    Large-scale visual information retrieval has become an active research area in this big data era. Recently, hashing/binary coding algorithms prove to be effective for scalable retrieval applications. Most existing hashing methods require relatively long binary codes (i.e., over hundreds of bits, sometimes even thousands of bits) to achieve reasonable retrieval accuracies. However, for some realistic and unique applications, such as on wearable or mobile devices, only short binary codes can be used for efficient image retrieval due to the limitation of computational resources or bandwidth on these devices. In this paper, we propose a novel unsupervised hashing approach called min-cost ranking (MCR) specifically for learning powerful short binary codes (i.e., usually the code length shorter than 100 b) for scalable image retrieval tasks. By exploring the discriminative ability of each dimension of data, MCR can generate one bit binary code for each dimension and simultaneously rank the discriminative separability of each bit according to the proposed cost function. Only top-ranked bits with minimum cost-values are then selected and grouped together to compose the final salient binary codes. Extensive experimental results on large-scale retrieval demonstrate that MCR can achieve comparative performance as the state-of-the-art hashing algorithms but with significantly shorter codes, leading to much faster large-scale retrieval.

  8. Robust regression for large-scale neuroimaging studies.

    PubMed

    Fritsch, Virgile; Da Mota, Benoit; Loth, Eva; Varoquaux, Gaël; Banaschewski, Tobias; Barker, Gareth J; Bokde, Arun L W; Brühl, Rüdiger; Butzek, Brigitte; Conrod, Patricia; Flor, Herta; Garavan, Hugh; Lemaitre, Hervé; Mann, Karl; Nees, Frauke; Paus, Tomas; Schad, Daniel J; Schümann, Gunter; Frouin, Vincent; Poline, Jean-Baptiste; Thirion, Bertrand

    2015-05-01

    Multi-subject datasets used in neuroimaging group studies have a complex structure, as they exhibit non-stationary statistical properties across regions and display various artifacts. While studies with small sample sizes can rarely be shown to deviate from standard hypotheses (such as the normality of the residuals) due to the poor sensitivity of normality tests with low degrees of freedom, large-scale studies (e.g. >100 subjects) exhibit more obvious deviations from these hypotheses and call for more refined models for statistical inference. Here, we demonstrate the benefits of robust regression as a tool for analyzing large neuroimaging cohorts. First, we use an analytic test based on robust parameter estimates; based on simulations, this procedure is shown to provide an accurate statistical control without resorting to permutations. Second, we show that robust regression yields more detections than standard algorithms using as an example an imaging genetics study with 392 subjects. Third, we show that robust regression can avoid false positives in a large-scale analysis of brain-behavior relationships with over 1500 subjects. Finally we embed robust regression in the Randomized Parcellation Based Inference (RPBI) method and demonstrate that this combination further improves the sensitivity of tests carried out across the whole brain. Altogether, our results show that robust procedures provide important advantages in large-scale neuroimaging group studies. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Reliability assessment for components of large scale photovoltaic systems

    NASA Astrophysics Data System (ADS)

    Ahadi, Amir; Ghadimi, Noradin; Mirabbasi, Davar

    2014-10-01

    Photovoltaic (PV) systems have significantly shifted from independent power generation systems to a large-scale grid-connected generation systems in recent years. The power output of PV systems is affected by the reliability of various components in the system. This study proposes an analytical approach to evaluate the reliability of large-scale, grid-connected PV systems. The fault tree method with an exponential probability distribution function is used to analyze the components of large-scale PV systems. The system is considered in the various sequential and parallel fault combinations in order to find all realistic ways in which the top or undesired events can occur. Additionally, it can identify areas that the planned maintenance should focus on. By monitoring the critical components of a PV system, it is possible not only to improve the reliability of the system, but also to optimize the maintenance costs. The latter is achieved by informing the operators about the system component's status. This approach can be used to ensure secure operation of the system by its flexibility in monitoring system applications. The implementation demonstrates that the proposed method is effective and efficient and can conveniently incorporate more system maintenance plans and diagnostic strategies.

  10. Impact of Large-scale Geological Architectures On Recharge

    NASA Astrophysics Data System (ADS)

    Troldborg, L.; Refsgaard, J. C.; Engesgaard, P.; Jensen, K. H.

    Geological and hydrogeological data constitutes the basis for assessment of ground- water flow pattern and recharge zones. The accessibility and applicability of hard ge- ological data is often a major obstacle in deriving plausible conceptual models. Nev- ertheless focus is often on parameter uncertainty caused by the effect of geological heterogeneity due to lack of hard geological data, thus neglecting the possibility of alternative conceptualizations of the large-scale geological architecture. For a catchment in the eastern part of Denmark we have constructed different geologi- cal models based on different conceptualization of the major geological trends and fa- cies architecture. The geological models are equally plausible in a conceptually sense and they are all calibrated to well head and river flow measurements. Comparison of differences in recharge zones and subsequently well protection zones emphasize the importance of assessing large-scale geological architecture in hydrological modeling on regional scale in a non-deterministic way. Geostatistical modeling carried out in a transitional probability framework shows the possibility of assessing multiple re- alizations of large-scale geological architecture from a combination of soft and hard geological information.

  11. Alteration of Large-Scale Chromatin Structure by Estrogen Receptor

    PubMed Central

    Nye, Anne C.; Rajendran, Ramji R.; Stenoien, David L.; Mancini, Michael A.; Katzenellenbogen, Benita S.; Belmont, Andrew S.

    2002-01-01

    The estrogen receptor (ER), a member of the nuclear hormone receptor superfamily important in human physiology and disease, recruits coactivators which modify local chromatin structure. Here we describe effects of ER on large-scale chromatin structure as visualized in live cells. We targeted ER to gene-amplified chromosome arms containing large numbers of lac operator sites either directly, through a lac repressor-ER fusion protein (lac rep-ER), or indirectly, by fusing lac repressor with the ER interaction domain of the coactivator steroid receptor coactivator 1. Significant decondensation of large-scale chromatin structure, comparable to that produced by the ∼150-fold-stronger viral protein 16 (VP16) transcriptional activator, was produced by ER in the absence of estradiol using both approaches. Addition of estradiol induced a partial reversal of this unfolding by green fluorescent protein-lac rep-ER but not by wild-type ER recruited by a lac repressor-SRC570-780 fusion protein. The chromatin decondensation activity did not require transcriptional activation by ER nor did it require ligand-induced coactivator interactions, and unfolding did not correlate with histone hyperacetylation. Ligand-induced coactivator interactions with helix 12 of ER were necessary for the partial refolding of chromatin in response to estradiol using the lac rep-ER tethering system. This work demonstrates that when tethered or recruited to DNA, ER possesses a novel large-scale chromatin unfolding activity. PMID:11971975

  12. Multiresolution comparison of precipitation datasets for large-scale models

    NASA Astrophysics Data System (ADS)

    Chun, K. P.; Sapriza Azuri, G.; Davison, B.; DeBeer, C. M.; Wheater, H. S.

    2014-12-01

    Gridded precipitation datasets are crucial for driving large-scale models which are related to weather forecast and climate research. However, the quality of precipitation products is usually validated individually. Comparisons between gridded precipitation products along with ground observations provide another avenue for investigating how the precipitation uncertainty would affect the performance of large-scale models. In this study, using data from a set of precipitation gauges over British Columbia and Alberta, we evaluate several widely used North America gridded products including the Canadian Gridded Precipitation Anomalies (CANGRD), the National Center for Environmental Prediction (NCEP) reanalysis, the Water and Global Change (WATCH) project, the thin plate spline smoothing algorithms (ANUSPLIN) and Canadian Precipitation Analysis (CaPA). Based on verification criteria for various temporal and spatial scales, results provide an assessment of possible applications for various precipitation datasets. For long-term climate variation studies (~100 years), CANGRD, NCEP, WATCH and ANUSPLIN have different comparative advantages in terms of their resolution and accuracy. For synoptic and mesoscale precipitation patterns, CaPA provides appealing performance of spatial coherence. In addition to the products comparison, various downscaling methods are also surveyed to explore new verification and bias-reduction methods for improving gridded precipitation outputs for large-scale models.

  13. Equivalent common path method in large-scale laser comparator

    NASA Astrophysics Data System (ADS)

    He, Mingzhao; Li, Jianshuang; Miao, Dongjing

    2015-02-01

    Large-scale laser comparator is main standard device that providing accurate, reliable and traceable measurements for high precision large-scale line and 3D measurement instruments. It mainly composed of guide rail, motion control system, environmental parameters monitoring system and displacement measurement system. In the laser comparator, the main error sources are temperature distribution, straightness of guide rail and pitch and yaw of measuring carriage. To minimize the measurement uncertainty, an equivalent common optical path scheme is proposed and implemented. Three laser interferometers are adjusted to parallel with the guide rail. The displacement in an arbitrary virtual optical path is calculated using three displacements without the knowledge of carriage orientations at start and end positions. The orientation of air floating carriage is calculated with displacements of three optical path and position of three retroreflectors which are precisely measured by Laser Tracker. A 4th laser interferometer is used in the virtual optical path as reference to verify this compensation method. This paper analyzes the effect of rail straightness on the displacement measurement. The proposed method, through experimental verification, can improve the measurement uncertainty of large-scale laser comparator.

  14. Dispersal Mutualism Incorporated into Large-Scale, Infrequent Disturbances

    PubMed Central

    Parker, V. Thomas

    2015-01-01

    Because of their influence on succession and other community interactions, large-scale, infrequent natural disturbances also should play a major role in mutualistic interactions. Using field data and experiments, I test whether mutualisms have been incorporated into large-scale wildfire by whether the outcomes of a mutualism depend on disturbance. In this study a seed dispersal mutualism is shown to depend on infrequent, large-scale disturbances. A dominant shrubland plant (Arctostaphylos species) produces seeds that make up a persistent soil seed bank and requires fire to germinate. In post-fire stands, I show that seedlings emerging from rodent caches dominate sites experiencing higher fire intensity. Field experiments show that rodents (Perimyscus californicus, P. boylii) do cache Arctostaphylos fruit and bury most seed caches to a sufficient depth to survive a killing heat pulse that a fire might drive into the soil. While the rodent dispersal and caching behavior itself has not changed compared to other habitats, the environmental transformation caused by wildfire converts the caching burial of seed from a dispersal process to a plant fire adaptive trait, and provides the context for stimulating subsequent life history evolution in the plant host. PMID:26151560

  15. Large-scale flow generation by inhomogeneous helicity.

    PubMed

    Yokoi, N; Brandenburg, A

    2016-03-01

    The effect of kinetic helicity (velocity-vorticity correlation) on turbulent momentum transport is investigated. The turbulent kinetic helicity (pseudoscalar) enters the Reynolds stress (mirror-symmetric tensor) expression in the form of a helicity gradient as the coupling coefficient for the mean vorticity and/or the angular velocity (axial vector), which suggests the possibility of mean-flow generation in the presence of inhomogeneous helicity. This inhomogeneous helicity effect, which was previously confirmed at the level of a turbulence- or closure-model simulation, is examined with the aid of direct numerical simulations of rotating turbulence with nonuniform helicity sustained by an external forcing. The numerical simulations show that the spatial distribution of the Reynolds stress is in agreement with the helicity-related term coupled with the angular velocity, and that a large-scale flow is generated in the direction of angular velocity. Such a large-scale flow is not induced in the case of homogeneous turbulent helicity. This result confirms the validity of the inhomogeneous helicity effect in large-scale flow generation and suggests that a vortex dynamo is possible even in incompressible turbulence where there is no baroclinicity effect.

  16. Large-scale flow experiments for managing river systems

    USGS Publications Warehouse

    Konrad, Christopher P.; Olden, Julian D.; Lytle, David A.; Melis, Theodore S.; Schmidt, John C.; Bray, Erin N.; Freeman, Mary C.; Gido, Keith B.; Hemphill, Nina P.; Kennard, Mark J.; McMullen, Laura E.; Mims, Meryl C.; Pyron, Mark; Robinson, Christopher T.; Williams, John G.

    2011-01-01

    Experimental manipulations of streamflow have been used globally in recent decades to mitigate the impacts of dam operations on river systems. Rivers are challenging subjects for experimentation, because they are open systems that cannot be isolated from their social context. We identify principles to address the challenges of conducting effective large-scale flow experiments. Flow experiments have both scientific and social value when they help to resolve specific questions about the ecological action of flow with a clear nexus to water policies and decisions. Water managers must integrate new information into operating policies for large-scale experiments to be effective. Modeling and monitoring can be integrated with experiments to analyze long-term ecological responses. Experimental design should include spatially extensive observations and well-defined, repeated treatments. Large-scale flow manipulations are only a part of dam operations that affect river systems. Scientists can ensure that experimental manipulations continue to be a valuable approach for the scientifically based management of river systems.

  17. Foundational perspectives on causality in large-scale brain networks.

    PubMed

    Mannino, Michael; Bressler, Steven L

    2015-12-01

    A profusion of recent work in cognitive neuroscience has been concerned with the endeavor to uncover causal influences in large-scale brain networks. However, despite the fact that many papers give a nod to the important theoretical challenges posed by the concept of causality, this explosion of research has generally not been accompanied by a rigorous conceptual analysis of the nature of causality in the brain. This review provides both a descriptive and prescriptive account of the nature of causality as found within and between large-scale brain networks. In short, it seeks to clarify the concept of causality in large-scale brain networks both philosophically and scientifically. This is accomplished by briefly reviewing the rich philosophical history of work on causality, especially focusing on contributions by David Hume, Immanuel Kant, Bertrand Russell, and Christopher Hitchcock. We go on to discuss the impact that various interpretations of modern physics have had on our understanding of causality. Throughout all this, a central focus is the distinction between theories of deterministic causality (DC), whereby causes uniquely determine their effects, and probabilistic causality (PC), whereby causes change the probability of occurrence of their effects. We argue that, given the topological complexity of its large-scale connectivity, the brain should be considered as a complex system and its causal influences treated as probabilistic in nature. We conclude that PC is well suited for explaining causality in the brain for three reasons: (1) brain causality is often mutual; (2) connectional convergence dictates that only rarely is the activity of one neuronal population uniquely determined by another one; and (3) the causal influences exerted between neuronal populations may not have observable effects. A number of different techniques are currently available to characterize causal influence in the brain. Typically, these techniques quantify the statistical

  18. Robust large-scale parallel nonlinear solvers for simulations.

    SciTech Connect

    Bader, Brett William; Pawlowski, Roger Patrick; Kolda, Tamara Gibson

    2005-11-01

    This report documents research to develop robust and efficient solution techniques for solving large-scale systems of nonlinear equations. The most widely used method for solving systems of nonlinear equations is Newton's method. While much research has been devoted to augmenting Newton-based solvers (usually with globalization techniques), little has been devoted to exploring the application of different models. Our research has been directed at evaluating techniques using different models than Newton's method: a lower order model, Broyden's method, and a higher order model, the tensor method. We have developed large-scale versions of each of these models and have demonstrated their use in important applications at Sandia. Broyden's method replaces the Jacobian with an approximation, allowing codes that cannot evaluate a Jacobian or have an inaccurate Jacobian to converge to a solution. Limited-memory methods, which have been successful in optimization, allow us to extend this approach to large-scale problems. We compare the robustness and efficiency of Newton's method, modified Newton's method, Jacobian-free Newton-Krylov method, and our limited-memory Broyden method. Comparisons are carried out for large-scale applications of fluid flow simulations and electronic circuit simulations. Results show that, in cases where the Jacobian was inaccurate or could not be computed, Broyden's method converged in some cases where Newton's method failed to converge. We identify conditions where Broyden's method can be more efficient than Newton's method. We also present modifications to a large-scale tensor method, originally proposed by Bouaricha, for greater efficiency, better robustness, and wider applicability. Tensor methods are an alternative to Newton-based methods and are based on computing a step based on a local quadratic model rather than a linear model. The advantage of Bouaricha's method is that it can use any existing linear solver, which makes it simple to write

  19. New probes of Cosmic Microwave Background large-scale anomalies

    NASA Astrophysics Data System (ADS)

    Aiola, Simone

    Fifty years of Cosmic Microwave Background (CMB) data played a crucial role in constraining the parameters of the LambdaCDM model, where Dark Energy, Dark Matter, and Inflation are the three most important pillars not yet understood. Inflation prescribes an isotropic universe on large scales, and it generates spatially-correlated density fluctuations over the whole Hubble volume. CMB temperature fluctuations on scales bigger than a degree in the sky, affected by modes on super-horizon scale at the time of recombination, are a clean snapshot of the universe after inflation. In addition, the accelerated expansion of the universe, driven by Dark Energy, leaves a hardly detectable imprint in the large-scale temperature sky at late times. Such fundamental predictions have been tested with current CMB data and found to be in tension with what we expect from our simple LambdaCDM model. Is this tension just a random fluke or a fundamental issue with the present model? In this thesis, we present a new framework to probe the lack of large-scale correlations in the temperature sky using CMB polarization data. Our analysis shows that if a suppression in the CMB polarization correlations is detected, it will provide compelling evidence for new physics on super-horizon scale. To further analyze the statistical properties of the CMB temperature sky, we constrain the degree of statistical anisotropy of the CMB in the context of the observed large-scale dipole power asymmetry. We find evidence for a scale-dependent dipolar modulation at 2.5sigma. To isolate late-time signals from the primordial ones, we test the anomalously high Integrated Sachs-Wolfe effect signal generated by superstructures in the universe. We find that the detected signal is in tension with the expectations from LambdaCDM at the 2.5sigma level, which is somewhat smaller than what has been previously argued. To conclude, we describe the current status of CMB observations on small scales, highlighting the

  20. Foundational perspectives on causality in large-scale brain networks

    NASA Astrophysics Data System (ADS)

    Mannino, Michael; Bressler, Steven L.

    2015-12-01

    A profusion of recent work in cognitive neuroscience has been concerned with the endeavor to uncover causal influences in large-scale brain networks. However, despite the fact that many papers give a nod to the important theoretical challenges posed by the concept of causality, this explosion of research has generally not been accompanied by a rigorous conceptual analysis of the nature of causality in the brain. This review provides both a descriptive and prescriptive account of the nature of causality as found within and between large-scale brain networks. In short, it seeks to clarify the concept of causality in large-scale brain networks both philosophically and scientifically. This is accomplished by briefly reviewing the rich philosophical history of work on causality, especially focusing on contributions by David Hume, Immanuel Kant, Bertrand Russell, and Christopher Hitchcock. We go on to discuss the impact that various interpretations of modern physics have had on our understanding of causality. Throughout all this, a central focus is the distinction between theories of deterministic causality (DC), whereby causes uniquely determine their effects, and probabilistic causality (PC), whereby causes change the probability of occurrence of their effects. We argue that, given the topological complexity of its large-scale connectivity, the brain should be considered as a complex system and its causal influences treated as probabilistic in nature. We conclude that PC is well suited for explaining causality in the brain for three reasons: (1) brain causality is often mutual; (2) connectional convergence dictates that only rarely is the activity of one neuronal population uniquely determined by another one; and (3) the causal influences exerted between neuronal populations may not have observable effects. A number of different techniques are currently available to characterize causal influence in the brain. Typically, these techniques quantify the statistical

  1. Development of a gene synthesis platform for the efficient large scale production of small genes encoding animal toxins.

    PubMed

    Sequeira, Ana Filipa; Brás, Joana L A; Guerreiro, Catarina I P D; Vincentelli, Renaud; Fontes, Carlos M G A

    2016-12-01

    Gene synthesis is becoming an important tool in many fields of recombinant DNA technology, including recombinant protein production. De novo gene synthesis is quickly replacing the classical cloning and mutagenesis procedures and allows generating nucleic acids for which no template is available. In addition, when coupled with efficient gene design algorithms that optimize codon usage, it leads to high levels of recombinant protein expression. Here, we describe the development of an optimized gene synthesis platform that was applied to the large scale production of small genes encoding venom peptides. This improved gene synthesis method uses a PCR-based protocol to assemble synthetic DNA from pools of overlapping oligonucleotides and was developed to synthesise multiples genes simultaneously. This technology incorporates an accurate, automated and cost effective ligation independent cloning step to directly integrate the synthetic genes into an effective Escherichia coli expression vector. The robustness of this technology to generate large libraries of dozens to thousands of synthetic nucleic acids was demonstrated through the parallel and simultaneous synthesis of 96 genes encoding animal toxins. An automated platform was developed for the large-scale synthesis of small genes encoding eukaryotic toxins. Large scale recombinant expression of synthetic genes encoding eukaryotic toxins will allow exploring the extraordinary potency and pharmacological diversity of animal venoms, an increasingly valuable but unexplored source of lead molecules for drug discovery.

  2. High-resolution definition of the Vibrio cholerae essential gene set with hidden Markov model-based analyses of transposon-insertion sequencing data.

    PubMed

    Chao, Michael C; Pritchard, Justin R; Zhang, Yanjia J; Rubin, Eric J; Livny, Jonathan; Davis, Brigid M; Waldor, Matthew K

    2013-10-01

    The coupling of high-density transposon mutagenesis to high-throughput DNA sequencing (transposon-insertion sequencing) enables simultaneous and genome-wide assessment of the contributions of individual loci to bacterial growth and survival. We have refined analysis of transposon-insertion sequencing data by normalizing for the effect of DNA replication on sequencing output and using a hidden Markov model (HMM)-based filter to exploit heretofore unappreciated information inherent in all transposon-insertion sequencing data sets. The HMM can smooth variations in read abundance and thereby reduce the effects of read noise, as well as permit fine scale mapping that is independent of genomic annotation and enable classification of loci into several functional categories (e.g. essential, domain essential or 'sick'). We generated a high-resolution map of genomic loci (encompassing both intra- and intergenic sequences) that are required or beneficial for in vitro growth of the cholera pathogen, Vibrio cholerae. This work uncovered new metabolic and physiologic requirements for V. cholerae survival, and by combining transposon-insertion sequencing and transcriptomic data sets, we also identified several novel noncoding RNA species that contribute to V. cholerae growth. Our findings suggest that HMM-based approaches will enhance extraction of biological meaning from transposon-insertion sequencing genomic data.

  3. High-resolution definition of the Vibrio cholerae essential gene set with hidden Markov model–based analyses of transposon-insertion sequencing data

    PubMed Central

    Chao, Michael C.; Pritchard, Justin R.; Zhang, Yanjia J.; Rubin, Eric J.; Livny, Jonathan; Davis, Brigid M.; Waldor, Matthew K.

    2013-01-01

    The coupling of high-density transposon mutagenesis to high-throughput DNA sequencing (transposon-insertion sequencing) enables simultaneous and genome-wide assessment of the contributions of individual loci to bacterial growth and survival. We have refined analysis of transposon-insertion sequencing data by normalizing for the effect of DNA replication on sequencing output and using a hidden Markov model (HMM)-based filter to exploit heretofore unappreciated information inherent in all transposon-insertion sequencing data sets. The HMM can smooth variations in read abundance and thereby reduce the effects of read noise, as well as permit fine scale mapping that is independent of genomic annotation and enable classification of loci into several functional categories (e.g. essential, domain essential or ‘sick’). We generated a high-resolution map of genomic loci (encompassing both intra- and intergenic sequences) that are required or beneficial for in vitro growth of the cholera pathogen, Vibrio cholerae. This work uncovered new metabolic and physiologic requirements for V. cholerae survival, and by combining transposon-insertion sequencing and transcriptomic data sets, we also identified several novel noncoding RNA species that contribute to V. cholerae growth. Our findings suggest that HMM-based approaches will enhance extraction of biological meaning from transposon-insertion sequencing genomic data. PMID:23901011

  4. Indy gene variation in natural populations confers fitness advantage and life span extension through transposon insertion.

    PubMed

    Zhu, Chen-Tseh; Chang, Chengyi; Reenan, Robert A; Helfand, Stephen L

    2014-01-01

    Natural selection acts to maximize reproductive fitness. However, antagonism between life span and reproductive success frequently poses a dilemma pitting the cost of fecundity against longevity. Here, we show that natural populations of Drosophila melanogaster harbor a Hoppel transposon insertion variant in the longevity gene Indy (I'm not dead yet), which confers both increased reproduction and longevity through metabolic changes. Heterozygosity for this natural long-lived variant has been maintained in isolates despite long-term inbreeding under laboratory conditions and advantageously confers increased fecundity. DNA sequences of variant chromosome isolates show evidence of selective sweep acting on the advantageous allele, suggesting that natural selection acts to maintain this variant. The transposon insertion also regulates Indy expression level, which has experimentally been shown to affect life span and fecundity. Thus, in the wild, evolution reaffirms that the mechanism of heterozygote advantage has acted upon the Indy gene to assure increased reproductive fitness and, coincidentally, longer life span through regulatory transposon mutagenesis.

  5. Genetic signature of histiocytic sarcoma revealed by a sleeping beauty transposon genetic screen in mice.

    PubMed

    Been, Raha A; Linden, Michael A; Hager, Courtney J; DeCoursin, Krista J; Abrahante, Juan E; Landman, Sean R; Steinbach, Michael; Sarver, Aaron L; Largaespada, David A; Starr, Timothy K

    2014-01-01

    Histiocytic sarcoma is a rare, aggressive neoplasm that responds poorly to therapy. Histiocytic sarcoma is thought to arise from macrophage precursor cells via genetic changes that are largely undefined. To improve our understanding of the etiology of histiocytic sarcoma we conducted a forward genetic screen in mice using the Sleeping Beauty transposon as a mutagen to identify genetic drivers of histiocytic sarcoma. Sleeping Beauty mutagenesis was targeted to myeloid lineage cells using the Lysozyme2 promoter. Mice with activated Sleeping Beauty mutagenesis had significantly shortened lifespan and the majority of these mice developed tumors resembling human histiocytic sarcoma. Analysis of transposon insertions identified 27 common insertion sites containing 28 candidate cancer genes. Several of these genes are known drivers of hematological neoplasms, like Raf1, Fli1, and Mitf, while others are well-known cancer genes, including Nf1, Myc, Jak2, and Pten. Importantly, several new potential drivers of histiocytic sarcoma were identified and could serve as targets for therapy for histiocytic sarcoma patients.

  6. Genetic Signature of Histiocytic Sarcoma Revealed by a Sleeping Beauty Transposon Genetic Screen in Mice

    PubMed Central

    Been, Raha A.; Linden, Michael A.; Hager, Courtney J.; DeCoursin, Krista J.; Abrahante, Juan E.; Landman, Sean R.; Steinbach, Michael; Sarver, Aaron L.; Largaespada, David A.; Starr, Timothy K.

    2014-01-01

    Histiocytic sarcoma is a rare, aggressive neoplasm that responds poorly to therapy. Histiocytic sarcoma is thought to arise from macrophage precursor cells via genetic changes that are largely undefined. To improve our understanding of the etiology of histiocytic sarcoma we conducted a forward genetic screen in mice using the Sleeping Beauty transposon as a mutagen to identify genetic drivers of histiocytic sarcoma. Sleeping Beauty mutagenesis was targeted to myeloid lineage cells using the Lysozyme2 promoter. Mice with activated Sleeping Beauty mutagenesis had significantly shortened lifespan and the majority of these mice developed tumors resembling human histiocytic sarcoma. Analysis of transposon insertions identified 27 common insertion sites containing 28 candidate cancer genes. Several of these genes are known drivers of hematological neoplasms, like Raf1, Fli1, and Mitf, while others are well-known cancer genes, including Nf1, Myc, Jak2, and Pten. Importantly, several new potential drivers of histiocytic sarcoma were identified and could serve as targets for therapy for histiocytic sarcoma patients. PMID:24827933

  7. Large-Scale Hybrid Motor Testing. Chapter 10

    NASA Technical Reports Server (NTRS)

    Story, George

    2006-01-01

    Hybrid rocket motors can be successfully demonstrated at a small scale virtually anywhere. There have been many suitcase sized portable test stands assembled for demonstration of hybrids. They show the safety of hybrid rockets to the audiences. These small show motors and small laboratory scale motors can give comparative burn rate data for development of different fuel/oxidizer combinations, however questions that are always asked when hybrids are mentioned for large scale applications are - how do they scale and has it been shown in a large motor? To answer those questions, large scale motor testing is required to verify the hybrid motor at its true size. The necessity to conduct large-scale hybrid rocket motor tests to validate the burn rate from the small motors to application size has been documented in several place^'^^.^. Comparison of small scale hybrid data to that of larger scale data indicates that the fuel burn rate goes down with increasing port size, even with the same oxidizer flux. This trend holds for conventional hybrid motors with forward oxidizer injection and HTPB based fuels. While the reason this is occurring would make a great paper or study or thesis, it is not thoroughly understood at this time. Potential causes include the fact that since hybrid combustion is boundary layer driven, the larger port sizes reduce the interaction (radiation, mixing and heat transfer) from the core region of the port. This chapter focuses on some of the large, prototype sized testing of hybrid motors. The largest motors tested have been AMROC s 250K-lbf thrust motor at Edwards Air Force Base and the Hybrid Propulsion Demonstration Program s 250K-lbf thrust motor at Stennis Space Center. Numerous smaller tests were performed to support the burn rate, stability and scaling concepts that went into the development of those large motors.

  8. The Large-Scale Current System During Auroral Substorms

    NASA Astrophysics Data System (ADS)

    Gjerloev, Jesper

    2015-04-01

    The substorm process has been discussed for more than four decades and new empirical large-scale models continue to be published. The continued activity implies both the importance and the complexity of the problem. We recently published a new model of the large-scale substorm current system (Gjerloev and Hoffman, JGR, 2014). Based on data from >100 ground magnetometers (obtained from SuperMAG), 116 isolated substorms, global auroral images (obtained by the Polar VIS Earth Camera) and a careful normalization technique we derived an empirical model of the ionospheric equivalent current system. Our model yield some unexpected features that appear inconsistent with the classical single current wedge current system. One of these features is a distinct latitudinal shift of the westward electrojet (WEJ) current between the pre- and post-midnight region and we find evidence that these two WEJ regions are quasi disconnected. This, and other observational facts, led us to propose a modified 3D current system configuration that consists of 2 wedge type systems: a current wedge in the pre-midnight region (bulge current wedge), and another current wedge system in the post-midnight region (oval current wedge). The two wedge systems are shifted in latitude but overlap in local time in the midnight region. Our model is at considerable variance with previous global models and conceptual schematics of the large-scale substorm current system. We speculate that the data coverage, the methodologies and the techniques used in these previous global studies are the cause of the differences in solutions. In this presentation we present our model, compare with other published models and discuss possible causes for the differences.

  9. Improving Design Efficiency for Large-Scale Heterogeneous Circuits

    NASA Astrophysics Data System (ADS)

    Gregerson, Anthony

    Despite increases in logic density, many Big Data applications must still be partitioned across multiple computing devices in order to meet their strict performance requirements. Among the most demanding of these applications is high-energy physics (HEP), which uses complex computing systems consisting of thousands of FPGAs and ASICs to process the sensor data created by experiments at particles accelerators such as the Large Hadron Collider (LHC). Designing such computing systems is challenging due to the scale of the systems, the exceptionally high-throughput and low-latency performance constraints that necessitate application-specific hardware implementations, the requirement that algorithms are efficiently partitioned across many devices, and the possible need to update the implemented algorithms during the lifetime of the system. In this work, we describe our research to develop flexible architectures for implementing such large-scale circuits on FPGAs. In particular, this work is motivated by (but not limited in scope to) high-energy physics algorithms for the Compact Muon Solenoid (CMS) experiment at the LHC. To make efficient use of logic resources in multi-FPGA systems, we introduce Multi-Personality Partitioning, a novel form of the graph partitioning problem, and present partitioning algorithms that can significantly improve resource utilization on heterogeneous devices while also reducing inter-chip connections. To reduce the high communication costs of Big Data applications, we also introduce Information-Aware Partitioning, a partitioning method that analyzes the data content of application-specific circuits, characterizes their entropy, and selects circuit partitions that enable efficient compression of data between chips. We employ our information-aware partitioning method to improve the performance of the hardware validation platform for evaluating new algorithms for the CMS experiment. Together, these research efforts help to improve the efficiency

  10. Solving large scale structure in ten easy steps with COLA

    NASA Astrophysics Data System (ADS)

    Tassev, Svetlin; Zaldarriaga, Matias; Eisenstein, Daniel J.

    2013-06-01

    We present the COmoving Lagrangian Acceleration (COLA) method: an N-body method for solving for Large Scale Structure (LSS) in a frame that is comoving with observers following trajectories calculated in Lagrangian Perturbation Theory (LPT). Unlike standard N-body methods, the COLA method can straightforwardly trade accuracy at small-scales in order to gain computational speed without sacrificing accuracy at large scales. This is especially useful for cheaply generating large ensembles of accurate mock halo catalogs required to study galaxy clustering and weak lensing, as those catalogs are essential for performing detailed error analysis for ongoing and future surveys of LSS. As an illustration, we ran a COLA-based N-body code on a box of size 100 Mpc/h with particles of mass ≈ 5 × 109Msolar/h. Running the code with only 10 timesteps was sufficient to obtain an accurate description of halo statistics down to halo masses of at least 1011Msolar/h. This is only at a modest speed penalty when compared to mocks obtained with LPT. A standard detailed N-body run is orders of magnitude slower than our COLA-based code. The speed-up we obtain with COLA is due to the fact that we calculate the large-scale dynamics exactly using LPT, while letting the N-body code solve for the small scales, without requiring it to capture exactly the internal dynamics of halos. Achieving a similar level of accuracy in halo statistics without the COLA method requires at least 3 times more timesteps than when COLA is employed.

  11. Large-scale smart passive system for civil engineering applications

    NASA Astrophysics Data System (ADS)

    Jung, Hyung-Jo; Jang, Dong-Doo; Lee, Heon-Jae; Cho, Sang-Won

    2008-03-01

    The smart passive system consisting of a magnetorheological (MR) damper and an electromagnetic induction (EMI) part has been recently proposed. An EMI part can generate the input current for an MR damper from vibration of a structure according to Faraday's law of electromagnetic induction. The control performance of the smart passive system has been demonstrated mainly by numerical simulations. It was verified from the numerical results that the system could be effective to reduce the structural responses in the cases of civil engineering structures such as buildings and bridges. On the other hand, the experimental validation of the system is not sufficiently conducted yet. In this paper, the feasibility of the smart passive system to real-scale structures is investigated. To do this, the large-scale smart passive system is designed, manufactured, and tested. The system consists of the large-capacity MR damper, which has a maximum force level of approximately +/-10,000N, a maximum stroke level of +/-35mm and the maximum current level of 3 A, and the large-scale EMI part, which is designed to generate sufficient induced current for the damper. The applicability of the smart passive system to large real-scale structures is examined through a series of shaking table tests. The magnitudes of the induced current of the EMI part with various sinusoidal excitation inputs are measured. According to the test results, the large-scale EMI part shows the possibility that it could generate the sufficient current or power for changing the damping characteristics of the large-capacity MR damper.

  12. Infectious diseases in large-scale cat hoarding investigations.

    PubMed

    Polak, K C; Levy, J K; Crawford, P C; Leutenegger, C M; Moriello, K A

    2014-08-01

    Animal hoarders accumulate animals in over-crowded conditions without adequate nutrition, sanitation, and veterinary care. As a result, animals rescued from hoarding frequently have a variety of medical conditions including respiratory infections, gastrointestinal disease, parasitism, malnutrition, and other evidence of neglect. The purpose of this study was to characterize the infectious diseases carried by clinically affected cats and to determine the prevalence of retroviral infections among cats in large-scale cat hoarding investigations. Records were reviewed retrospectively from four large-scale seizures of cats from failed sanctuaries from November 2009 through March 2012. The number of cats seized in each case ranged from 387 to 697. Cats were screened for feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) in all four cases and for dermatophytosis in one case. A subset of cats exhibiting signs of upper respiratory disease or diarrhea had been tested for infections by PCR and fecal flotation for treatment planning. Mycoplasma felis (78%), calicivirus (78%), and Streptococcus equi subspecies zooepidemicus (55%) were the most common respiratory infections. Feline enteric coronavirus (88%), Giardia (56%), Clostridium perfringens (49%), and Tritrichomonas foetus (39%) were most common in cats with diarrhea. The seroprevalence of FeLV and FIV were 8% and 8%, respectively. In the one case in which cats with lesions suspicious for dermatophytosis were cultured for Microsporum canis, 69/76 lesional cats were culture-positive; of these, half were believed to be truly infected and half were believed to be fomite carriers. Cats from large-scale hoarding cases had high risk for enteric and respiratory infections, retroviruses, and dermatophytosis. Case responders should be prepared for mass treatment of infectious diseases and should implement protocols to prevent transmission of feline or zoonotic infections during the emergency response and when

  13. Solving large scale structure in ten easy steps with COLA

    SciTech Connect

    Tassev, Svetlin; Zaldarriaga, Matias; Eisenstein, Daniel J. E-mail: matiasz@ias.edu

    2013-06-01

    We present the COmoving Lagrangian Acceleration (COLA) method: an N-body method for solving for Large Scale Structure (LSS) in a frame that is comoving with observers following trajectories calculated in Lagrangian Perturbation Theory (LPT). Unlike standard N-body methods, the COLA method can straightforwardly trade accuracy at small-scales in order to gain computational speed without sacrificing accuracy at large scales. This is especially useful for cheaply generating large ensembles of accurate mock halo catalogs required to study galaxy clustering and weak lensing, as those catalogs are essential for performing detailed error analysis for ongoing and future surveys of LSS. As an illustration, we ran a COLA-based N-body code on a box of size 100 Mpc/h with particles of mass ≈ 5 × 10{sup 9}M{sub s}un/h. Running the code with only 10 timesteps was sufficient to obtain an accurate description of halo statistics down to halo masses of at least 10{sup 11}M{sub s}un/h. This is only at a modest speed penalty when compared to mocks obtained with LPT. A standard detailed N-body run is orders of magnitude slower than our COLA-based code. The speed-up we obtain with COLA is due to the fact that we calculate the large-scale dynamics exactly using LPT, while letting the N-body code solve for the small scales, without requiring it to capture exactly the internal dynamics of halos. Achieving a similar level of accuracy in halo statistics without the COLA method requires at least 3 times more timesteps than when COLA is employed.

  14. Statistical analysis of large-scale neuronal recording data

    PubMed Central

    Reed, Jamie L.; Kaas, Jon H.

    2010-01-01

    Relating stimulus properties to the response properties of individual neurons and neuronal networks is a major goal of sensory research. Many investigators implant electrode arrays in multiple brain areas and record from chronically implanted electrodes over time to answer a variety of questions. Technical challenges related to analyzing large-scale neuronal recording data are not trivial. Several analysis methods traditionally used by neurophysiologists do not account for dependencies in the data that are inherent in multi-electrode recordings. In addition, when neurophysiological data are not best modeled by the normal distribution and when the variables of interest may not be linearly related, extensions of the linear modeling techniques are recommended. A variety of methods exist to analyze correlated data, even when data are not normally distributed and the relationships are nonlinear. Here we review expansions of the Generalized Linear Model designed to address these data properties. Such methods are used in other research fields, and the application to large-scale neuronal recording data will enable investigators to determine the variable properties that convincingly contribute to the variances in the observed neuronal measures. Standard measures of neuron properties such as response magnitudes can be analyzed using these methods, and measures of neuronal network activity such as spike timing correlations can be analyzed as well. We have done just that in recordings from 100-electrode arrays implanted in the primary somatosensory cortex of owl monkeys. Here we illustrate how one example method, Generalized Estimating Equations analysis, is a useful method to apply to large-scale neuronal recordings. PMID:20472395

  15. LARGE-SCALE CO2 TRANSPORTATION AND DEEP OCEAN SEQUESTRATION

    SciTech Connect

    Hamid Sarv

    1999-03-01

    Technical and economical feasibility of large-scale CO{sub 2} transportation and ocean sequestration at depths of 3000 meters or grater was investigated. Two options were examined for transporting and disposing the captured CO{sub 2}. In one case, CO{sub 2} was pumped from a land-based collection center through long pipelines laid on the ocean floor. Another case considered oceanic tanker transport of liquid carbon dioxide to an offshore floating structure for vertical injection to the ocean floor. In the latter case, a novel concept based on subsurface towing of a 3000-meter pipe, and attaching it to the offshore structure was considered. Budgetary cost estimates indicate that for distances greater than 400 km, tanker transportation and offshore injection through a 3000-meter vertical pipe provides the best method for delivering liquid CO{sub 2} to deep ocean floor depressions. For shorter distances, CO{sub 2} delivery by parallel-laid, subsea pipelines is more cost-effective. Estimated costs for 500-km transport and storage at a depth of 3000 meters by subsea pipelines and tankers were 1.5 and 1.4 dollars per ton of stored CO{sub 2}, respectively. At these prices, economics of ocean disposal are highly favorable. Future work should focus on addressing technical issues that are critical to the deployment of a large-scale CO{sub 2} transportation and disposal system. Pipe corrosion, structural design of the transport pipe, and dispersion characteristics of sinking CO{sub 2} effluent plumes have been identified as areas that require further attention. Our planned activities in the next Phase include laboratory-scale corrosion testing, structural analysis of the pipeline, analytical and experimental simulations of CO{sub 2} discharge and dispersion, and the conceptual economic and engineering evaluation of large-scale implementation.

  16. Genome ARTIST: a robust, high-accuracy aligner tool for mapping transposon insertions and self-insertions.

    PubMed

    Ecovoiu, Alexandru Al; Ghionoiu, Iulian Constantin; Ciuca, Andrei Mihai; Ratiu, Attila Cristian

    2016-01-01

    A critical topic of insertional mutagenesis experiments performed on model organisms is mapping the hits of artificial transposons (ATs) at nucleotide level accuracy. Mapping errors may occur when sequencing artifacts or mutations as single nucleotide polymorphisms (SNPs) and small indels are present very close to the junction between a genomic sequence and a transposon inverted repeat (TIR). Another particular item of insertional mutagenesis is mapping of the transposon self-insertions and, to our best knowledge, there is no publicly available mapping tool designed to analyze such molecular events. We developed Genome ARTIST, a pairwise gapped aligner tool which works out both issues by means of an original, robust mapping strategy. Genome ARTIST is not designed to use next-generation sequencing (NGS) data but to analyze ATs insertions obtained in small to medium-scale mutagenesis experiments. Genome ARTIST employs a heuristic approach to find DNA sequence similarities and harnesses a multi-step implementation of a Smith-Waterman adapted algorithm to compute the mapping alignments. The experience is enhanced by easily customizable parameters and a user-friendly interface that describes the genomic landscape surrounding the insertion. Genome ARTIST is functional with many genomes of bacteria and eukaryotes available in Ensembl and GenBank repositories. Our tool specifically harnesses the sequence annotation data provided by FlyBase for Drosophila melanogaster (the fruit fly), which enables mapping of insertions relative to various genomic features such as natural transposons. Genome ARTIST was tested against other alignment tools using relevant query sequences derived from the D. melanogaster and Mus musculus (mouse) genomes. Real and simulated query sequences were also comparatively inquired, revealing that Genome ARTIST is a very robust solution for mapping transposon insertions. Genome ARTIST is a stand-alone user-friendly application, designed for high

  17. Solving Large-scale Eigenvalue Problems in SciDACApplications

    SciTech Connect

    Yang, Chao

    2005-06-29

    Large-scale eigenvalue problems arise in a number of DOE applications. This paper provides an overview of the recent development of eigenvalue computation in the context of two SciDAC applications. We emphasize the importance of Krylov subspace methods, and point out its limitations. We discuss the value of alternative approaches that are more amenable to the use of preconditioners, and report the progression using the multi-level algebraic sub-structuring techniques to speed up eigenvalue calculation. In addition to methods for linear eigenvalue problems, we also examine new approaches to solving two types of non-linear eigenvalue problems arising from SciDAC applications.

  18. Analysis Plan for 1985 Large-Scale Tests.

    DTIC Science & Technology

    1983-01-01

    KEY WORDS (Continue on reverse side it necessary mnd Identify by block number) Large-Scale Blasting Agents Multiburst ANFO S:,ock Waves 20. ABSTRACT...CONSIDERATIONS 6 1.5 MULTIBURST TECHNIQUES 6 1.6 TEST SITE CONSIDERATIONS 6 2 CANDIDATE EXPLOSIVES 8 2.1 INTRODUCTION 82.2 ANFO 8 2.2.1 Bulk (Loose) ANFO 11...2.2.2 Bagged ANFO 13 2.3 APEX 1360 15 2.4 NITRIC ACID AND NITROPROPANE 17 2.5 NITROPROPANENITRATE (NPN) 19 2.6 DBA - 22M 21 2.7 HARDENING EMULSION 22 2.8

  19. Large-Scale Patterns of Filament Channels and Filaments

    NASA Astrophysics Data System (ADS)

    Mackay, Duncan

    2016-07-01

    In this review the properties and large-scale patterns of filament channels and filaments will be considered. Initially, the global formation locations of filament channels and filaments are discussed, along with their hemispheric pattern. Next, observations of the formation of filament channels and filaments are described where two opposing views are considered. Finally, the wide range of models that have been constructed to consider the formation of filament channels and filaments over long time-scales are described, along with the origin of the hemispheric pattern of filaments.

  20. Design of a large-scale CFB boiler

    SciTech Connect

    Darling, S.; Li, S.

    1997-12-31

    Many CFB boilers sized 100--150 MWe are in operation, and several others sized 150--250 MWe are in operation or under construction. The next step for CFB technology is the 300--400 MWe size range. This paper will describe Foster Wheeler`s large-scale CFB boiler experience and the design for a 300 MWe CFB boiler. The authors will show how the design incorporates Foster Wheeler`s unique combination of extensive utility experience and CFB boiler experience. All the benefits of CFB technology which include low emissions, fuel flexibility, low maintenance and competitive cost are now available in the 300--400 MWe size range.

  1. An iterative decoupling solution method for large scale Lyapunov equations

    NASA Technical Reports Server (NTRS)

    Athay, T. M.; Sandell, N. R., Jr.

    1976-01-01

    A great deal of attention has been given to the numerical solution of the Lyapunov equation. A useful classification of the variety of solution techniques are the groupings of direct, transformation, and iterative methods. The paper summarizes those methods that are at least partly favorable numerically, giving special attention to two criteria: exploitation of a general sparse system matrix structure and efficiency in resolving the governing linear matrix equation for different matrices. An iterative decoupling solution method is proposed as a promising approach for solving large-scale Lyapunov equation when the system matrix exhibits a general sparse structure. A Fortran computer program that realizes the iterative decoupling algorithm is also discussed.

  2. Large-scale normal fluid circulation in helium superflows

    NASA Astrophysics Data System (ADS)

    Galantucci, Luca; Sciacca, Michele; Barenghi, Carlo F.

    2017-01-01

    We perform fully coupled numerical simulations of helium II pure superflows in a channel, with vortex-line density typical of experiments. Peculiar to our model is the computation of the back-reaction of the superfluid vortex motion on the normal fluid and the presence of solid boundaries. We recover the uniform vortex-line density experimentally measured employing second sound resonators and we show that pure superflow in helium II is associated with a large-scale circulation of the normal fluid which can be detected using existing particle-tracking visualization techniques.

  3. On decentralized control of large-scale systems

    NASA Technical Reports Server (NTRS)

    Siljak, D. D.

    1978-01-01

    A scheme is presented for decentralized control of large-scale linear systems which are composed of a number of interconnected subsystems. By ignoring the interconnections, local feedback controls are chosen to optimize each decoupled subsystem. Conditions are provided to establish compatibility of the individual local controllers and achieve stability of the overall system. Besides computational simplifications, the scheme is attractive because of its structural features and the fact that it produces a robust decentralized regulator for large dynamic systems, which can tolerate a wide range of nonlinearities and perturbations among the subsystems.

  4. Large-Scale periodic solar velocities: An observational study

    NASA Technical Reports Server (NTRS)

    Dittmer, P. H.

    1977-01-01

    Observations of large-scale solar velocities were made using the mean field telescope and Babcock magnetograph of the Stanford Solar Observatory. Observations were made in the magnetically insensitive ion line at 5124 A, with light from the center (limb) of the disk right (left) circularly polarized, so that the magnetograph measures the difference in wavelength between center and limb. Computer calculations are made of the wavelength difference produced by global pulsations for spherical harmonics up to second order and of the signal produced by displacing the solar image relative to polarizing optics or diffraction grating.

  5. Large-Scale Compton Imaging for Wide-Area Surveillance

    SciTech Connect

    Lange, D J; Manini, H A; Wright, D M

    2006-03-01

    We study the performance of a large-scale Compton imaging detector placed in a low-flying aircraft, used to search wide areas for rad/nuc threat sources. In this paper we investigate the performance potential of equipping aerial platforms with gamma-ray detectors that have photon sensitivity up to a few MeV. We simulate the detector performance, and present receiver operating characteristics (ROC) curves for a benchmark scenario using a {sup 137}Cs source. The analysis uses a realistic environmental background energy spectrum and includes air attenuation.

  6. Large-Scale Measurement of Absolute Protein Glycosylation Stoichiometry.

    PubMed

    Sun, Shisheng; Zhang, Hui

    2015-07-07

    Protein glycosylation is one of the most important protein modifications. Glycosylation site occupancy alteration has been implicated in human diseases and cancers. However, current glycoproteomic methods focus on the identification and quantification of glycosylated peptides and glycosylation sites but not glycosylation occupancy or glycoform stoichiometry. Here we describe a method for large-scale determination of the absolute glycosylation stoichiometry using three independent relative ratios. Using this method, we determined 117 absolute N-glycosylation occupancies in OVCAR-3 cells. Finally, we investigated the possible functions and the determinants for partial glycosylation.

  7. Enabling Large-Scale Biomedical Analysis in the Cloud

    PubMed Central

    Lin, Ying-Chih; Yu, Chin-Sheng; Lin, Yen-Jen

    2013-01-01

    Recent progress in high-throughput instrumentations has led to an astonishing growth in both volume and complexity of biomedical data collected from various sources. The planet-size data brings serious challenges to the storage and computing technologies. Cloud computing is an alternative to crack the nut because it gives concurrent consideration to enable storage and high-performance computing on large-scale data. This work briefly introduces the data intensive computing system and summarizes existing cloud-based resources in bioinformatics. These developments and applications would facilitate biomedical research to make the vast amount of diversification data meaningful and usable. PMID:24288665

  8. Large-scale intermittency in the atmospheric boundary layer.

    PubMed

    Kholmyansky, M; Moriconi, L; Tsinober, A

    2007-08-01

    We find actual evidence, relying upon vorticity time series taken in a high-Reynolds-number atmospheric experiment, that to a very good approximation the surface boundary layer flow may be described, in a statistical sense and under certain regimes, as an advected ensemble of homogeneous turbulent systems, characterized by a log-normal distribution of fluctuating intensities. Our analysis suggests that the usual direct numerical simulations of homogeneous and isotropic turbulence, performed at moderate Reynolds numbers, may play an important role in the study of turbulent boundary layer flows, if supplemented with appropriate statistical information concerned with the structure of large-scale fluctuations.

  9. Frequency domain multiplexing for large-scale bolometer arrays

    SciTech Connect

    Spieler, Helmuth

    2002-05-31

    The development of planar fabrication techniques for superconducting transition-edge sensors has brought large-scale arrays of 1000 pixels or more to the realm of practicality. This raises the problem of reading out a large number of sensors with a tractable number of connections. A possible solution is frequency-domain multiplexing. I summarize basic principles, present various circuit topologies, and discuss design trade-offs, noise performance, cross-talk and dynamic range. The design of a practical device and its readout system is described with a discussion of fabrication issues, practical limits and future prospects.

  10. A Modular Ring Architecture for Large Scale Neural Network Implementations

    NASA Astrophysics Data System (ADS)

    Jump, Lance B.; Ligomenides, Panos A.

    1989-11-01

    Constructing fully parallel, large scale, neural networks is complicated by the problems of providing for massive interconnectivity and of overcoming fan in/out limitations in area-efficient VLSI/WSI realizations. A modular, bus switched, neural ring architecture employing primitive ring (pRing) processors is proposed, which solves the fan in/out and connectivity problems by a dynamically reconfigurable communication ring that synchronously serves identical, radially connected, processing elements. It also allows cost versus performance trade-offs by the assignment of variable numbers of logical neurons to each physical processing element.

  11. Simplified DGS procedure for large-scale genome structural study.

    PubMed

    Jung, Yong-Chul; Xu, Jia; Chen, Jun; Kim, Yeong; Winchester, David; Wang, San Ming

    2009-11-01

    Ditag genome scanning (DGS) uses next-generation DNA sequencing to sequence the ends of ditag fragments produced by restriction enzymes. These sequences are compared to known genome sequences to determine their structure. In order to use DGS for large-scale genome structural studies, we have substantially revised the original protocol by replacing the in vivo genomic DNA cloning with in vitro adaptor ligation, eliminating the ditag concatemerization steps, and replacing the 454 sequencer with Solexa or SOLiD sequencers for ditag sequence collection. This revised protocol further increases genome coverage and resolution and allows DGS to be used to analyze multiple genomes simultaneously.

  12. UAV Data Processing for Large Scale Topographical Mapping

    NASA Astrophysics Data System (ADS)

    Tampubolon, W.; Reinhardt, W.

    2014-06-01

    Large scale topographical mapping in the third world countries is really a prominent challenge in geospatial industries nowadays. On one side the demand is significantly increasing while on the other hand it is constrained by limited budgets available for mapping projects. Since the advent of Act Nr.4/yr.2011 about Geospatial Information in Indonesia, large scale topographical mapping has been on high priority for supporting the nationwide development e.g. detail spatial planning. Usually large scale topographical mapping relies on conventional aerial survey campaigns in order to provide high resolution 3D geospatial data sources. Widely growing on a leisure hobby, aero models in form of the so-called Unmanned Aerial Vehicle (UAV) bring up alternative semi photogrammetric aerial data acquisition possibilities suitable for relatively small Area of Interest (AOI) i.e. <5,000 hectares. For detail spatial planning purposes in Indonesia this area size can be used as a mapping unit since it usually concentrates on the basis of sub district area (kecamatan) level. In this paper different camera and processing software systems will be further analyzed for identifying the best optimum UAV data acquisition campaign components in combination with the data processing scheme. The selected AOI is covering the cultural heritage of Borobudur Temple as one of the Seven Wonders of the World. A detailed accuracy assessment will be concentrated within the object feature of the temple at the first place. Feature compilation involving planimetric objects (2D) and digital terrain models (3D) will be integrated in order to provide Digital Elevation Models (DEM) as the main interest of the topographic mapping activity. By doing this research, incorporating the optimum amount of GCPs in the UAV photo data processing will increase the accuracy along with its high resolution in 5 cm Ground Sampling Distance (GSD). Finally this result will be used as the benchmark for alternative geospatial

  13. Large Scale Composite Manufacturing for Heavy Lift Launch Vehicles

    NASA Technical Reports Server (NTRS)

    Stavana, Jacob; Cohen, Leslie J.; Houseal, Keth; Pelham, Larry; Lort, Richard; Zimmerman, Thomas; Sutter, James; Western, Mike; Harper, Robert; Stuart, Michael

    2012-01-01

    Risk reduction for the large scale composite manufacturing is an important goal to produce light weight components for heavy lift launch vehicles. NASA and an industry team successfully employed a building block approach using low-cost Automated Tape Layup (ATL) of autoclave and Out-of-Autoclave (OoA) prepregs. Several large, curved sandwich panels were fabricated at HITCO Carbon Composites. The aluminum honeycomb core sandwich panels are segments of a 1/16th arc from a 10 meter cylindrical barrel. Lessons learned highlight the manufacturing challenges required to produce light weight composite structures such as fairings for heavy lift launch vehicles.

  14. Clusters as cornerstones of large-scale structure.

    NASA Astrophysics Data System (ADS)

    Gottlöber, S.; Retzlaff, J.; Turchaninov, V.

    1997-04-01

    Galaxy clusters are one of the best tracers of large-scale structure in the Universe on scales well above 100 Mpc. The authors investigate here the clustering properties of a redshift sample of Abell/ACO clusters and compare the observational sample with mock samples constructed from N-body simulations on the basis of four different cosmological models. The authors discuss the power spectrum, the Minkowski functionals and the void statistics of these samples and conclude, that the SCDM and TCDM models are ruled out whereas the ACDM and BSI models are in agreement with the observational data.

  15. Artificial intelligence and large scale computation: A physics perspective

    NASA Astrophysics Data System (ADS)

    Hogg, Tad; Huberman, B. A.

    1987-12-01

    We study the macroscopic behavior of computation and examine both emergent collective phenomena and dynamical aspects with an emphasis on software issues, which are at the core of large scale distributed computation and artificial intelligence systems. By considering large systems, we exhibit novel phenomena which cannot be foreseen from examination of their smaller counterparts. We review both the symbolic and connectionist views of artificial intelligence, provide a number of examples which display these phenomena, and resort to statistical mechanics, dynamical systems theory and the theory of random graphs to elicit the range of possible behaviors.

  16. Potential for geophysical experiments in large scale tests

    SciTech Connect

    Dieterich, J.H.

    1981-07-01

    Potential research applications for large-specimen geophysical experiments include measurements of scale dependence of physical parameters and examination of interactions with heterogeneities, especially flaws such as cracks. In addition, increased specimen size provides opportunities for improved recording resolution and greater control of experimental variables. Large-scale experiments using a special purpose low stress (<40 MPa) bi-axial apparatus demonstrate that a minimum fault length is required to generate confined shear instabilities along pre-existing faults. Experimental analysis of source interactions for simulated earthquakes consisting of confined shear instabilities on a fault with gouge appears to require large specimens (approx.1m) and high confining pressures (>100 MPa).

  17. Quantum computation for large-scale image classification

    NASA Astrophysics Data System (ADS)

    Ruan, Yue; Chen, Hanwu; Tan, Jianing; Li, Xi

    2016-10-01

    Due to the lack of an effective quantum feature extraction method, there is currently no effective way to perform quantum image classification or recognition. In this paper, for the first time, a global quantum feature extraction method based on Schmidt decomposition is proposed. A revised quantum learning algorithm is also proposed that will classify images by computing the Hamming distance of these features. From the experimental results derived from the benchmark database Caltech 101, and an analysis of the algorithm, an effective approach to large-scale image classification is derived and proposed against the background of big data.

  18. Large-scale genotoxicity assessments in the marine environment.

    PubMed

    Hose, J E

    1994-12-01

    There are a number of techniques for detecting genotoxicity in the marine environment, and many are applicable to large-scale field assessments. Certain tests can be used to evaluate responses in target organisms in situ while others utilize surrogate organisms exposed to field samples in short-term laboratory bioassays. Genotoxicity endpoints appear distinct from traditional toxicity endpoints, but some have chemical or ecotoxicologic correlates. One versatile end point, the frequency of anaphase aberrations, has been used in several large marine assessments to evaluate genotoxicity in the New York Bight, in sediment from San Francisco Bay, and following the Exxon Valdez oil spill.

  19. Large-scale sodium spray fire code validation (SOFICOV) test

    SciTech Connect

    Jeppson, D.W.; Muhlestein, L.D.

    1985-01-01

    A large-scale, sodium, spray fire code validation test was performed in the HEDL 850-m/sup 3/ Containment System Test Facility (CSTF) as part of the Sodium Spray Fire Code Validation (SOFICOV) program. Six hundred fifty eight kilograms of sodium spray was sprayed in an air atmosphere for a period of 2400 s. The sodium spray droplet sizes and spray pattern distribution were estimated. The containment atmosphere temperature and pressure response, containment wall temperature response and sodium reaction rate with oxygen were measured. These results are compared to post-test predictions using SPRAY and NACOM computer codes.

  20. Water-based scintillators for large-scale liquid calorimetry

    SciTech Connect

    Winn, D.R.; Raftery, D.

    1985-02-01

    We have investigated primary and secondary solvent intermediates in search of a recipe to create a bulk liquid scintillator with water as the bulk solvent and common fluors as the solutes. As we are not concerned with energy resolution below 1 MeV in large-scale experiments, light-output at the 10% level of high-quality organic solvent based scintillators is acceptable. We have found encouraging performance from industrial surfactants as primary solvents for PPO and POPOP. This technique may allow economical and environmentally safe bulk scintillator for kiloton-sized high energy calorimetry.