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Sample records for affymetrix mouse genome

  1. X:Map: annotation and visualization of genome structure for Affymetrix exon array analysis

    PubMed Central

    Yates, Tim; Okoniewski, Michał J.; Miller, Crispin J.

    2008-01-01

    Affymetrix exon arrays aim to target every known and predicted exon in the human, mouse or rat genomes, and have reporters that extend beyond protein coding regions to other areas of the transcribed genome. This combination of increased coverage and precision is important because a substantial proportion of protein coding genes are predicted to be alternatively spliced, and because many non-coding genes are known also to be of biological significance. In order to fully exploit these arrays, it is necessary to associate each reporter on the array with the features of the genome it is targeting, and to relate these to gene and genome structure. X:Map is a genome annotation database that provides this information. Data can be browsed using a novel Google-maps based interface, and analysed and further visualized through an associated BioConductor package. The database can be found at http://xmap.picr.man.ac.uk. PMID:17932061

  2. Mouse genome database 2016

    PubMed Central

    Bult, Carol J.; Eppig, Janan T.; Blake, Judith A.; Kadin, James A.; Richardson, Joel E.

    2016-01-01

    The Mouse Genome Database (MGD; http://www.informatics.jax.org) is the primary community model organism database for the laboratory mouse and serves as the source for key biological reference data related to mouse genes, gene functions, phenotypes and disease models with a strong emphasis on the relationship of these data to human biology and disease. As the cost of genome-scale sequencing continues to decrease and new technologies for genome editing become widely adopted, the laboratory mouse is more important than ever as a model system for understanding the biological significance of human genetic variation and for advancing the basic research needed to support the emergence of genome-guided precision medicine. Recent enhancements to MGD include new graphical summaries of biological annotations for mouse genes, support for mobile access to the database, tools to support the annotation and analysis of sets of genes, and expanded support for comparative biology through the expansion of homology data. PMID:26578600

  3. Mouse genome database 2016.

    PubMed

    Bult, Carol J; Eppig, Janan T; Blake, Judith A; Kadin, James A; Richardson, Joel E

    2016-01-04

    The Mouse Genome Database (MGD; http://www.informatics.jax.org) is the primary community model organism database for the laboratory mouse and serves as the source for key biological reference data related to mouse genes, gene functions, phenotypes and disease models with a strong emphasis on the relationship of these data to human biology and disease. As the cost of genome-scale sequencing continues to decrease and new technologies for genome editing become widely adopted, the laboratory mouse is more important than ever as a model system for understanding the biological significance of human genetic variation and for advancing the basic research needed to support the emergence of genome-guided precision medicine. Recent enhancements to MGD include new graphical summaries of biological annotations for mouse genes, support for mobile access to the database, tools to support the annotation and analysis of sets of genes, and expanded support for comparative biology through the expansion of homology data.

  4. FULL-GENOME ANALYSIS OF ALTERNATIVE SPLICING IN MOUSE LIVER AFTER HEPATOTOXICANT EXPOSURE

    EPA Science Inventory

    Alternative splicing plays a role in determining gene function and protein diversity. We have employed whole genome exon profiling using Affymetrix Mouse Exon 1.0 ST arrays to understand the significance of alternative splicing on a genome-wide scale in response to multiple toxic...

  5. RIKEN mouse genome encyclopedia.

    PubMed

    Hayashizaki, Yoshihide

    2003-01-01

    We have been working to establish the comprehensive mouse full-length cDNA collection and sequence database to cover as many genes as we can, named Riken mouse genome encyclopedia. Recently we are constructing higher-level annotation (Functional ANnoTation Of Mouse cDNA; FANTOM) not only with homology search based annotation but also with expression data profile, mapping information and protein-protein database. More than 1,000,000 clones prepared from 163 tissues were end-sequenced to classify into 159,789 clusters and 60,770 representative clones were fully sequenced. As a conclusion, the 60,770 sequences contained 33,409 unique. The next generation of life science is clearly based on all of the genome information and resources. Based on our cDNA clones we developed the additional system to explore gene function. We developed cDNA microarray system to print all of these cDNA clones, protein-protein interaction screening system, protein-DNA interaction screening system and so on. The integrated database of all the information is very useful not only for analysis of gene transcriptional network and for the connection of gene to phenotype to facilitate positional candidate approach. In this talk, the prospect of the application of these genome resourced should be discussed. More information is available at the web page: http://genome.gsc.riken.go.jp/.

  6. 10. international mouse genome conference

    SciTech Connect

    Meisler, M.H.

    1996-12-31

    Ten years after hosting the First International Mammalian Genome Conference in Paris in 1986, Dr. Jean-Louis Guenet presided over the Tenth Conference at the Pasteur Institute, October 7--10, 1996. The 1986 conference was a satellite to the Human Gene Mapping Workshop and had approximately 50 attendees. The 1996 meeting was attended by 300 scientists from around the world. In the interim, the number of mapped loci in the mouse increased from 1,000 to over 20,000. This report contains a listing of the program and its participants, and two articles that review the meeting and the role of the laboratory mouse in the Human Genome project. More than 200 papers were presented at the conference covering the following topics: International mouse chromosome committee meetings; Mutant generation and identification; Physical and genetic maps; New technology and resources; Chromatin structure and gene regulation; Rate and hamster genetic maps; Informatics and databases; and Quantitative trait analysis.

  7. A physical map of the mouse genome.

    PubMed

    Gregory, Simon G; Sekhon, Mandeep; Schein, Jacqueline; Zhao, Shaying; Osoegawa, Kazutoyo; Scott, Carol E; Evans, Richard S; Burridge, Paul W; Cox, Tony V; Fox, Christopher A; Hutton, Richard D; Mullenger, Ian R; Phillips, Kimbly J; Smith, James; Stalker, Jim; Threadgold, Glen J; Birney, Ewan; Wylie, Kristine; Chinwalla, Asif; Wallis, John; Hillier, LaDeana; Carter, Jason; Gaige, Tony; Jaeger, Sara; Kremitzki, Colin; Layman, Dan; Maas, Jason; McGrane, Rebecca; Mead, Kelly; Walker, Rebecca; Jones, Steven; Smith, Michael; Asano, Jennifer; Bosdet, Ian; Chan, Susanna; Chittaranjan, Suganthi; Chiu, Readman; Fjell, Chris; Fuhrmann, Dan; Girn, Noreen; Gray, Catharine; Guin, Ran; Hsiao, Letticia; Krzywinski, Martin; Kutsche, Reta; Lee, Soo Sen; Mathewson, Carrie; McLeavy, Candice; Messervier, Steve; Ness, Steven; Pandoh, Pawan; Prabhu, Anna-Liisa; Saeedi, Parvaneh; Smailus, Duane; Spence, Lorraine; Stott, Jeff; Taylor, Sheryl; Terpstra, Wesley; Tsai, Miranda; Vardy, Jill; Wye, Natasja; Yang, George; Shatsman, Sofiya; Ayodeji, Bola; Geer, Keita; Tsegaye, Getahun; Shvartsbeyn, Alla; Gebregeorgis, Elizabeth; Krol, Margaret; Russell, Daniel; Overton, Larry; Malek, Joel A; Holmes, Mike; Heaney, Michael; Shetty, Jyoti; Feldblyum, Tamara; Nierman, William C; Catanese, Joseph J; Hubbard, Tim; Waterston, Robert H; Rogers, Jane; de Jong, Pieter J; Fraser, Claire M; Marra, Marco; McPherson, John D; Bentley, David R

    2002-08-15

    A physical map of a genome is an essential guide for navigation, allowing the location of any gene or other landmark in the chromosomal DNA. We have constructed a physical map of the mouse genome that contains 296 contigs of overlapping bacterial clones and 16,992 unique markers. The mouse contigs were aligned to the human genome sequence on the basis of 51,486 homology matches, thus enabling use of the conserved synteny (correspondence between chromosome blocks) of the two genomes to accelerate construction of the mouse map. The map provides a framework for assembly of whole-genome shotgun sequence data, and a tile path of clones for generation of the reference sequence. Definition of the human-mouse alignment at this level of resolution enables identification of a mouse clone that corresponds to almost any position in the human genome. The human sequence may be used to facilitate construction of other mammalian genome maps using the same strategy.

  8. Qualitative assessment of gene expression in affymetrix genechip arrays

    NASA Astrophysics Data System (ADS)

    Nagarajan, Radhakrishnan; Upreti, Meenakshi

    2007-01-01

    Affymetrix Genechip microarrays are used widely to determine the simultaneous expression of genes in a given biological paradigm. Probes on the Genechip array are atomic entities which by definition are randomly distributed across the array and in turn govern the gene expression. In the present study, we make several interesting observations. We show that there is considerable correlation between the probe intensities across the array which defy the independence assumption. While the mechanism behind such correlations is unclear, we show that scaling behavior and the profiles of perfect match (PM) as well as mismatch (MM) probes are similar and immune-to-background subtraction. We believe that the observed correlations are possibly an outcome of inherent non-stationarities or patchiness in the array devoid of biological significance. This is demonstrated by inspecting their scaling behavior and profiles of the PM and MM probe intensities obtained from publicly available Genechip arrays from three eukaryotic genomes, namely: Drosophila melanogaster (fruit fly), Homo sapiens (humans) and Mus musculus (house mouse) across distinct biological paradigms and across laboratories, with and without background subtraction. The fluctuation functions were estimated using detrended fluctuation analysis (DFA) with fourth-order polynomial detrending. The results presented in this study provide new insights into correlation signatures of PM and MM probe intensities and suggests the choice of DFA as a tool for qualitative assessment of Affymetrix Genechip microarrays prior to their analysis. A more detailed investigation is necessary in order to understand the source of these correlations.

  9. Insights from Human/Mouse genome comparisons

    SciTech Connect

    Pennacchio, Len A.

    2003-03-30

    Large-scale public genomic sequencing efforts have provided a wealth of vertebrate sequence data poised to provide insights into mammalian biology. These include deep genomic sequence coverage of human, mouse, rat, zebrafish, and two pufferfish (Fugu rubripes and Tetraodon nigroviridis) (Aparicio et al. 2002; Lander et al. 2001; Venter et al. 2001; Waterston et al. 2002). In addition, a high-priority has been placed on determining the genomic sequence of chimpanzee, dog, cow, frog, and chicken (Boguski 2002). While only recently available, whole genome sequence data have provided the unique opportunity to globally compare complete genome contents. Furthermore, the shared evolutionary ancestry of vertebrate species has allowed the development of comparative genomic approaches to identify ancient conserved sequences with functionality. Accordingly, this review focuses on the initial comparison of available mammalian genomes and describes various insights derived from such analysis.

  10. The Riken mouse genome encyclopedia project.

    PubMed

    Hayashizaki, Yoshihide

    2003-01-01

    The Riken mouse genome encyclopedia a comprehensive full-length cDNA collection and sequence database. High-level functional annotation is based on sequence homology search, expression profiling, mapping and protein-protein interactions. More than 1000000 clones prepared from 163 tissues were end-sequenced and classified into 128000 clusters, and 60000 representative clones were fully sequenced representing 24000 clear protein-encoding genes. The application of the mouse genome database for positional cloning and gene network regulation analysis is reported.

  11. Mouse genome engineering using designer nucleases.

    PubMed

    Hermann, Mario; Cermak, Tomas; Voytas, Daniel F; Pelczar, Pawel

    2014-04-02

    Transgenic mice carrying site-specific genome modifications (knockout, knock-in) are of vital importance for dissecting complex biological systems as well as for modeling human diseases and testing therapeutic strategies. Recent advances in the use of designer nucleases such as zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9 system for site-specific genome engineering open the possibility to perform rapid targeted genome modification in virtually any laboratory species without the need to rely on embryonic stem (ES) cell technology. A genome editing experiment typically starts with identification of designer nuclease target sites within a gene of interest followed by construction of custom DNA-binding domains to direct nuclease activity to the investigator-defined genomic locus. Designer nuclease plasmids are in vitro transcribed to generate mRNA for microinjection of fertilized mouse oocytes. Here, we provide a protocol for achieving targeted genome modification by direct injection of TALEN mRNA into fertilized mouse oocytes.

  12. Identification of structural variation in mouse genomes

    PubMed Central

    Keane, Thomas M.; Wong, Kim; Adams, David J.; Flint, Jonathan; Reymond, Alexandre; Yalcin, Binnaz

    2014-01-01

    Structural variation is variation in structure of DNA regions affecting DNA sequence length and/or orientation. It generally includes deletions, insertions, copy-number gains, inversions, and transposable elements. Traditionally, the identification of structural variation in genomes has been challenging. However, with the recent advances in high-throughput DNA sequencing and paired-end mapping (PEM) methods, the ability to identify structural variation and their respective association to human diseases has improved considerably. In this review, we describe our current knowledge of structural variation in the mouse, one of the prime model systems for studying human diseases and mammalian biology. We further present the evolutionary implications of structural variation on transposable elements. We conclude with future directions on the study of structural variation in mouse genomes that will increase our understanding of molecular architecture and functional consequences of structural variation. PMID:25071822

  13. A report from the Sixth International Mouse Genome Conference

    SciTech Connect

    Brown, S.

    1992-12-31

    The Sixth Annual Mouse Genome Conference was held in October, 1992 at Buffalo, USA. The mouse is one of the primary model organisms in the Human Genome Project. Through the use of gene targeting studies the mouse has become a powerful biological model for the study of gene function and, in addition, the comparison of the many homologous mutations identified in human and mouse have widened our understanding of the biology of these two organisms. A primary goal in the mouse genome program has been to create a genetic map of STSs of high resolution (<1cM) that would form the basis for the physical mapping of the whole mouse genome. Buffalo saw substantial new progress towards the goal of a very high density genetic map and the beginnings of substantive efforts towards physical mapping in chromosome regions with a high density of genetic markers.

  14. Initial sequencing and comparative analysis of the mouse genome

    SciTech Connect

    Waterston, Robert H.; Lindblad-Toh, Kerstin; Birney, Ewan; Rogers, Jane; Abril, Josep F.; Agarwal, Pankaj; Agarwala, Richa; Ainscough, Rachel; Alexandersson, Marina; An, Peter; Antonarakis, Stylianos E.; Attwood, John; Baertsch, Robert; Bailey, Jonathon; Barlow, Karen; Beck, Stephan; Berry, Eric; Birren, Bruce; Bloom, Toby; Bork, Peer; Botcherby, Marc; Bray, Nicolas; Brent, Michael R.; Brown, Daniel G.; Brown, Stephen D.; Bult, Carol; Burton, John; Butler, Jonathan; Campbell, Robert D.; Carninci, Piero; Cawley, Simon; Chiaromonte, Francesca; Chinwalla, Asif T.; Church, Deanna M.; Clamp, Michele; Clee, Christopher; Collins, Francis S.; Cook, Lisa L.; Copley, Richard R.; Coulson, Alan; Couronne, Olivier; Cuff, James; Curwen, Val; Cutts, Tim; Daly, Mark; David, Robert; Davies, Joy; Delehaunty, Kimberly D.; Deri, Justin; Dermitzakis, Emmanouil T.; Dewey, Colin; Dickens, Nicholas J.; Diekhans, Mark; Dodge, Sheila; Dubchak, Inna; Dunn, Diane M.; Eddy, Sean R.; Elnitski, Laura; Emes, Richard D.; Eswara, Pallavi; Eyras, Eduardo; Felsenfeld, Adam; Fewell, Ginger A.; Flicek, Paul; Foley, Karen; Frankel, Wayne N.; Fulton, Lucinda A.; Fulton, Robert S.; Furey, Terrence S.; Gage, Diane; Gibbs, Richard A.; Glusman, Gustavo; Gnerre, Sante; Goldman, Nick; Goodstadt, Leo; Grafham, Darren; Graves, Tina A.; Green, Eric D.; Gregory, Simon; Guigo, Roderic; Guyer, Mark; Hardison, Ross C.; Haussler, David; Hayashizaki, Yoshihide; Hillier, LaDeana W.; Hinrichs, Angela; Hlavina, Wratko; Holzer, Timothy; Hsu, Fan; Hua, Axin; Hubbard, Tim; Hunt, Adrienne; Jackson, Ian; Jaffe, David B.; Johnson, L. Steven; Jones, Matthew; Jones, Thomas A.; Joy, Ann; Kamal, Michael; Karlsson, Elinor K.; Karolchik, Donna; Kasprzyk, Arkadiusz; Kawai, Jun; Keibler, Evan; Kells, Cristyn; Kent, W. James; Kirby, Andrew; Kolbe, Diana L.; Korf, Ian; Kucherlapati, Raju S.; Kulbokas III, Edward J.; Kulp, David; Landers, Tom; Leger, J.P.; Leonard, Steven; Letunic, Ivica; Levine, Rosie; et al.

    2002-12-15

    The sequence of the mouse genome is a key informational tool for understanding the contents of the human genome and a key experimental tool for biomedical research. Here, we report the results of an international collaboration to produce a high-quality draft sequence of the mouse genome. We also present an initial comparative analysis of the mouse and human genomes, describing some of the insights that can be gleaned from the two sequences. We discuss topics including the analysis of the evolutionary forces shaping the size, structure and sequence of the genomes; the conservation of large-scale synteny across most of the genomes; the much lower extent of sequence orthology covering less than half of the genomes; the proportions of the genomes under selection; the number of protein-coding genes; the expansion of gene families related to reproduction and immunity; the evolution of proteins; and the identification of intraspecies polymorphism.

  15. VIZARD: analysis of Affymetrix Arabidopsis GeneChip data

    NASA Technical Reports Server (NTRS)

    Moseyko, Nick; Feldman, Lewis J.

    2002-01-01

    SUMMARY: The Affymetrix GeneChip Arabidopsis genome array has proved to be a very powerful tool for the analysis of gene expression in Arabidopsis thaliana, the most commonly studied plant model organism. VIZARD is a Java program created at the University of California, Berkeley, to facilitate analysis of Arabidopsis GeneChip data. It includes several integrated tools for filtering, sorting, clustering and visualization of gene expression data as well as tools for the discovery of regulatory motifs in upstream sequences. VIZARD also includes annotation and upstream sequence databases for the majority of genes represented on the Affymetrix Arabidopsis GeneChip array. AVAILABILITY: VIZARD is available free of charge for educational, research, and not-for-profit purposes, and can be downloaded at http://www.anm.f2s.com/research/vizard/ CONTACT: moseyko@uclink4.berkeley.edu.

  16. Meeting Report: The Twelfth International Mouse Genome Conference

    SciTech Connect

    Manolakou, Katerina; Cross, Sally H.; Simpson, Eleanor H.; Jackson, Ian J.

    1998-10-01

    The annual International Mouse Genome Conference (IMGC) is where, scientifically speaking, classical mouse genetics meets the relative newcomer of genomics. The 12th meeting took place last October in the delightful Bavarian village of Garmisch-Partenkirchen, and we were greeted by the sight on the mountains of the first snowfall of the season. However the discussions left little time for exploration. Minds of participants in Garmisch were focused by a recent document produced by the NIH and by discussions within other funding agencies worldwide. If implemented, the proposals will further enhance the status of the mouse as the principal model for study of the function of the human genome.

  17. The Mouse Genome Database (MGD): facilitating mouse as a model for human biology and disease.

    PubMed

    Eppig, Janan T; Blake, Judith A; Bult, Carol J; Kadin, James A; Richardson, Joel E

    2015-01-01

    The Mouse Genome Database (MGD, http://www.informatics.jax.org) serves the international biomedical research community as the central resource for integrated genomic, genetic and biological data on the laboratory mouse. To facilitate use of mouse as a model in translational studies, MGD maintains a core of high-quality curated data and integrates experimentally and computationally generated data sets. MGD maintains a unified catalog of genes and genome features, including functional RNAs, QTL and phenotypic loci. MGD curates and provides functional and phenotype annotations for mouse genes using the Gene Ontology and Mammalian Phenotype Ontology. MGD integrates phenotype data and associates mouse genotypes to human diseases, providing critical mouse-human relationships and access to repositories holding mouse models. MGD is the authoritative source of nomenclature for genes, genome features, alleles and strains following guidelines of the International Committee on Standardized Genetic Nomenclature for Mice. A new addition to MGD, the Human-Mouse: Disease Connection, allows users to explore gene-phenotype-disease relationships between human and mouse. MGD has also updated search paradigms for phenotypic allele attributes, incorporated incidental mutation data, added a module for display and exploration of genes and microRNA interactions and adopted the JBrowse genome browser. MGD resources are freely available to the scientific community.

  18. The Mouse Genome Database (MGD): facilitating mouse as a model for human biology and disease

    PubMed Central

    Eppig, Janan T.; Blake, Judith A.; Bult, Carol J.; Kadin, James A.; Richardson, Joel E.

    2015-01-01

    The Mouse Genome Database (MGD, http://www.informatics.jax.org) serves the international biomedical research community as the central resource for integrated genomic, genetic and biological data on the laboratory mouse. To facilitate use of mouse as a model in translational studies, MGD maintains a core of high-quality curated data and integrates experimentally and computationally generated data sets. MGD maintains a unified catalog of genes and genome features, including functional RNAs, QTL and phenotypic loci. MGD curates and provides functional and phenotype annotations for mouse genes using the Gene Ontology and Mammalian Phenotype Ontology. MGD integrates phenotype data and associates mouse genotypes to human diseases, providing critical mouse–human relationships and access to repositories holding mouse models. MGD is the authoritative source of nomenclature for genes, genome features, alleles and strains following guidelines of the International Committee on Standardized Genetic Nomenclature for Mice. A new addition to MGD, the Human–Mouse: Disease Connection, allows users to explore gene–phenotype–disease relationships between human and mouse. MGD has also updated search paradigms for phenotypic allele attributes, incorporated incidental mutation data, added a module for display and exploration of genes and microRNA interactions and adopted the JBrowse genome browser. MGD resources are freely available to the scientific community. PMID:25348401

  19. Engineering subtle targeted mutations into the mouse genome.

    PubMed

    Menke, Douglas B

    2013-09-01

    Homologous recombination in embryonic stem (ES) cells offers an exquisitely precise mechanism to introduce targeted modifications to the mouse genome. This ability to produce specific alterations to the mouse genome has become an essential tool for the analysis of gene function and the development of mouse models of human disease. Of the many thousands of mouse alleles that have been generated by gene targeting, the majority are designed to completely ablate gene function, to create conditional alleles that are inactivated in the presence of Cre recombinase, or to produce reporter alleles that label-specific tissues or cell populations (Eppig et al., 2012, Nucleic Acids Res 40:D881-D886). However, there is a variety of powerful motivations for the introduction of subtle targeted mutations (STMs) such as point mutations, small deletions, or small insertions into the mouse genome. The introduction of STMs allows the ablation of specific transcript isoforms, permits the functional investigation of particular domains or amino acids within a protein, provides the ability to study the role of specific sites with in cis-regulatory elements, and can result in better mouse models of human genetic disorders. In this review, I examine the current strategies that are commonly used to introduce STMs into the mouse genome and highlight new gene targeting technologies, including TALENs and CRISPR/Cas, which are likely to influence the future of gene targeting in mice.

  20. Mouse Genome Database: From sequence to phenotypes and disease models

    PubMed Central

    Richardson, Joel E.; Kadin, James A.; Smith, Cynthia L.; Blake, Judith A.; Bult, Carol J.

    2015-01-01

    Summary The Mouse Genome Database (MGD, www.informatics.jax.org) is the international scientific database for genetic, genomic, and biological data on the laboratory mouse to support the research requirements of the biomedical community. To accomplish this goal, MGD provides broad data coverage, serves as the authoritative standard for mouse nomenclature for genes, mutants, and strains, and curates and integrates many types of data from literature and electronic sources. Among the key data sets MGD supports are: the complete catalog of mouse genes and genome features, comparative homology data for mouse and vertebrate genes, the authoritative set of Gene Ontology (GO) annotations for mouse gene functions, a comprehensive catalog of mouse mutations and their phenotypes, and a curated compendium of mouse models of human diseases. Here, we describe the data acquisition process, specifics about MGD's key data areas, methods to access and query MGD data, and outreach and user help facilities. genesis 53:458–473, 2015. © 2015 The Authors. Genesis Published by Wiley Periodicals, Inc. PMID:26150326

  1. Mouse Genome Editing Using the CRISPR/Cas System.

    PubMed

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

    2014-10-01

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

  2. Mouse Genome Editing using CRISPR/Cas System

    PubMed Central

    Harms, Donald W; Quadros, Rolen M; Seruggia, Davide; Ohtsuka, Masato; Takahashi, Gou

    2015-01-01

    The availability of techniques to create desired genetic mutations has enabled the laboratory mouse as an extensively used model organism in biomedical research including human genetics. A new addition to this existing technical repertoire is the CRISPR/Cas system. Specifically, this system allows editing of the mouse genome much faster than the previously used techniques and more importantly multiple mutations can be created in a single experiment. Here we provide protocols for preparation of CRISPR/Cas reagents and microinjection into one cell mouse embryos to create knockout or knock-in mouse models. PMID:25271839

  3. Mouse Genome Informatics (MGI): Resources for Mining Mouse Genetic, Genomic, and Biological Data in Support of Primary and Translational Research.

    PubMed

    Eppig, Janan T; Smith, Cynthia L; Blake, Judith A; Ringwald, Martin; Kadin, James A; Richardson, Joel E; Bult, Carol J

    2017-01-01

    The Mouse Genome Informatics (MGI), resource ( www.informatics.jax.org ) has existed for over 25 years, and over this time its data content, informatics infrastructure, and user interfaces and tools have undergone dramatic changes (Eppig et al., Mamm Genome 26:272-284, 2015). Change has been driven by scientific methodological advances, rapid improvements in computational software, growth in computer hardware capacity, and the ongoing collaborative nature of the mouse genomics community in building resources and sharing data. Here we present an overview of the current data content of MGI, describe its general organization, and provide examples using simple and complex searches, and tools for mining and retrieving sets of data.

  4. A Comparative Encyclopedia of DNA Elements in the Mouse Genome

    PubMed Central

    Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D.; Shen, Yin; Pervouchine, Dmitri D.; Djebali, Sarah; Thurman, Bob; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K.; Williams, Brian A.; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M. A.; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T.; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D.; Bansal, Mukul S.; Keller, Cheryl A.; Morrissey, Christapher S.; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S.; Cayting, Philip; Kawli, Trupti; Boyle, Alan P.; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S.; Cline, Melissa S.; Erickson, Drew T.; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A.; Rosenbloom, Kate R.; de Sousa, Beatriz Lacerda; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W. James; Santos, Miguel Ramalho; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J.; Wilken, Matthew S.; Reh, Thomas A.; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P.; Neph, Shane; Humbert, Richard; Hansen, R. Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E.; Orkin, Stuart H.; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J.; Blobel, Gerd A.; Good, Peter J.; Lowdon, Rebecca F.; Adams, Leslie B.; Zhou, Xiao-Qiao; Pazin, Michael J.; Feingold, Elise A.; Wold, Barbara; Taylor, James; Kellis, Manolis; Mortazavi, Ali; Weissman, Sherman M.; Stamatoyannopoulos, John; Snyder, Michael P.; Guigo, Roderic; Gingeras, Thomas R.; Gilbert, David M.; Hardison, Ross C.; Beer, Michael A.; Ren, Bing

    2014-01-01

    Summary As the premier model organism in biomedical research, the laboratory mouse shares the majority of protein-coding genes with humans, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications, and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of other sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases. PMID:25409824

  5. A comparative encyclopedia of DNA elements in the mouse genome.

    PubMed

    Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D; Shen, Yin; Pervouchine, Dmitri D; Djebali, Sarah; Thurman, Robert E; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K; Williams, Brian A; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M A; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D; Bansal, Mukul S; Kellis, Manolis; Keller, Cheryl A; Morrissey, Christapher S; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S; Cayting, Philip; Kawli, Trupti; Boyle, Alan P; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S; Cline, Melissa S; Erickson, Drew T; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A; Rosenbloom, Kate R; Lacerda de Sousa, Beatriz; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W James; Ramalho Santos, Miguel; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J; Wilken, Matthew S; Reh, Thomas A; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P; Neph, Shane; Humbert, Richard; Hansen, R Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E; Orkin, Stuart H; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J; Blobel, Gerd A; Cao, Xiaoyi; Zhong, Sheng; Wang, Ting; Good, Peter J; Lowdon, Rebecca F; Adams, Leslie B; Zhou, Xiao-Qiao; Pazin, Michael J; Feingold, Elise A; Wold, Barbara; Taylor, James; Mortazavi, Ali; Weissman, Sherman M; Stamatoyannopoulos, John A; Snyder, Michael P; Guigo, Roderic; Gingeras, Thomas R; Gilbert, David M; Hardison, Ross C; Beer, Michael A; Ren, Bing

    2014-11-20

    The laboratory mouse shares the majority of its protein-coding genes with humans, making it the premier model organism in biomedical research, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.

  6. A unified gene catalog for the laboratory mouse reference genome.

    PubMed

    Zhu, Y; Richardson, J E; Hale, P; Baldarelli, R M; Reed, D J; Recla, J M; Sinclair, R; Reddy, T B K; Bult, C J

    2015-08-01

    We report here a semi-automated process by which mouse genome feature predictions and curated annotations (i.e., genes, pseudogenes, functional RNAs, etc.) from Ensembl, NCBI and Vertebrate Genome Annotation database (Vega) are reconciled with the genome features in the Mouse Genome Informatics (MGI) database (http://www.informatics.jax.org) into a comprehensive and non-redundant catalog. Our gene unification method employs an algorithm (fjoin--feature join) for efficient detection of genome coordinate overlaps among features represented in two annotation data sets. Following the analysis with fjoin, genome features are binned into six possible categories (1:1, 1:0, 0:1, 1:n, n:1, n:m) based on coordinate overlaps. These categories are subsequently prioritized for assessment of annotation equivalencies and differences. The version of the unified catalog reported here contains more than 59,000 entries, including 22,599 protein-coding coding genes, 12,455 pseudogenes, and 24,007 other feature types (e.g., microRNAs, lincRNAs, etc.). More than 23,000 of the entries in the MGI gene catalog have equivalent gene models in the annotation files obtained from NCBI, Vega, and Ensembl. 12,719 of the features are unique to NCBI relative to Ensembl/Vega; 11,957 are unique to Ensembl/Vega relative to NCBI, and 3095 are unique to MGI. More than 4000 genome features fall into categories that require manual inspection to resolve structural differences in the gene models from different annotation sources. Using the MGI unified gene catalog, researchers can easily generate a comprehensive report of mouse genome features from a single source and compare the details of gene and transcript structure using MGI's mouse genome browser.

  7. Discovery and mapping of single feature polymorphisms in wheat using affymetrix arrays

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Single feature polymorphisms (SFPs) can be a rich source of markers for gene mapping and function studies. To explore the feasibility of using the Affymetrix GeneChip to discover and map SFPs in the large hexaploid wheat genome, six wheat varieties of diverse origins were analyzed for significant pr...

  8. Rawcopy: Improved copy number analysis with Affymetrix arrays

    PubMed Central

    Mayrhofer, Markus; Viklund, Björn; Isaksson, Anders

    2016-01-01

    Microarray data is subject to noise and systematic variation that negatively affects the resolution of copy number analysis. We describe Rawcopy, an R package for processing of Affymetrix CytoScan HD, CytoScan 750k and SNP 6.0 microarray raw intensities (CEL files). Noise characteristics of a large number of reference samples are used to estimate log ratio and B-allele frequency for total and allele-specific copy number analysis. Rawcopy achieves better signal-to-noise ratio and higher proportion of validated alterations than commonly used free and proprietary alternatives. In addition, Rawcopy visualizes each microarray sample for assessment of technical quality, patient identity and genome-wide absolute copy number states. Software and instructions are available at http://rawcopy.org. PMID:27796336

  9. Endonucleases: new tools to edit the mouse genome.

    PubMed

    Wijshake, Tobias; Baker, Darren J; van de Sluis, Bart

    2014-10-01

    Mouse transgenesis has been instrumental in determining the function of genes in the pathophysiology of human diseases and modification of genes by homologous recombination in mouse embryonic stem cells remains a widely used technology. However, this approach harbors a number of disadvantages, as it is time-consuming and quite laborious. Over the last decade a number of new genome editing technologies have been developed, including zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/CRISPR-associated (CRISPR/Cas). These systems are characterized by a designed DNA binding protein or RNA sequence fused or co-expressed with a non-specific endonuclease, respectively. The engineered DNA binding protein or RNA sequence guides the nuclease to a specific target sequence in the genome to induce a double strand break. The subsequent activation of the DNA repair machinery then enables the introduction of gene modifications at the target site, such as gene disruption, correction or insertion. Nuclease-mediated genome editing has numerous advantages over conventional gene targeting, including increased efficiency in gene editing, reduced generation time of mutant mice, and the ability to mutagenize multiple genes simultaneously. Although nuclease-driven modifications in the genome are a powerful tool to generate mutant mice, there are concerns about off-target cleavage, especially when using the CRISPR/Cas system. Here, we describe the basic principles of these new strategies in mouse genome manipulation, their inherent advantages, and their potential disadvantages compared to current technologies used to study gene function in mouse models. This article is part of a Special Issue entitled: From Genome to Function.

  10. Mouse genome annotation by the RefSeq project.

    PubMed

    McGarvey, Kelly M; Goldfarb, Tamara; Cox, Eric; Farrell, Catherine M; Gupta, Tripti; Joardar, Vinita S; Kodali, Vamsi K; Murphy, Michael R; O'Leary, Nuala A; Pujar, Shashikant; Rajput, Bhanu; Rangwala, Sanjida H; Riddick, Lillian D; Webb, David; Wright, Mathew W; Murphy, Terence D; Pruitt, Kim D

    2015-10-01

    Complete and accurate annotation of the mouse genome is critical to the advancement of research conducted on this important model organism. The National Center for Biotechnology Information (NCBI) develops and maintains many useful resources to assist the mouse research community. In particular, the reference sequence (RefSeq) database provides high-quality annotation of multiple mouse genome assemblies using a combinatorial approach that leverages computation, manual curation, and collaboration. Implementation of this conservative and rigorous approach, which focuses on representation of only full-length and non-redundant data, produces high-quality annotation products. RefSeq records explicitly link sequences to current knowledge in a timely manner, updating public records regularly and rapidly in response to nomenclature updates, addition of new relevant publications, collaborator discussion, and user feedback. Whole genome re-annotation is also conducted at least every 12-18 months, and often more frequently in response to assembly updates or availability of informative data. This article highlights key features and advantages of RefSeq genome annotation products and presents an overview of NCBI processes to generate these data. Further discussion of NCBI's resources highlights useful features and the best methods for accessing our data.

  11. Interspecific introgressive origin of genomic diversity in the house mouse.

    PubMed

    Liu, Kevin J; Steinberg, Ethan; Yozzo, Alexander; Song, Ying; Kohn, Michael H; Nakhleh, Luay

    2015-01-06

    We report on a genome-wide scan for introgression between the house mouse (Mus musculus domesticus) and the Algerian mouse (Mus spretus), using samples from the ranges of sympatry and allopatry in Africa and Europe. Our analysis reveals wide variability in introgression signatures along the genomes, as well as across the samples. We find that fewer than half of the autosomes in each genome harbor all detectable introgression, whereas the X chromosome has none. Further, European mice carry more M. spretus alleles than the sympatric African ones. Using the length distribution and sharing patterns of introgressed genomic tracts across the samples, we infer, first, that at least three distinct hybridization events involving M. spretus have occurred, one of which is ancient, and the other two are recent (one presumably due to warfarin rodenticide selection). Second, several of the inferred introgressed tracts contain genes that are likely to confer adaptive advantage. Third, introgressed tracts might contain driver genes that determine the evolutionary fate of those tracts. Further, functional analysis revealed introgressed genes that are essential to fitness, including the Vkorc1 gene, which is implicated in rodenticide resistance, and olfactory receptor genes. Our findings highlight the extent and role of introgression in nature and call for careful analysis and interpretation of house mouse data in evolutionary and genetic studies.

  12. Mouse Genome Database (MGD)-2017: community knowledge resource for the laboratory mouse

    PubMed Central

    Blake, Judith A.; Eppig, Janan T.; Kadin, James A.; Richardson, Joel E.; Smith, Cynthia L.; Bult, Carol J.

    2017-01-01

    The Mouse Genome Database (MGD: http://www.informatics.jax.org) is the primary community data resource for the laboratory mouse. It provides a highly integrated and highly curated system offering a comprehensive view of current knowledge about mouse genes, genetic markers and genomic features as well as the associations of those features with sequence, phenotypes, functional and comparative information, and their relationships to human diseases. MGD continues to enhance access to these data, to extend the scope of data content and visualizations, and to provide infrastructure and user support that ensures effective and efficient use of MGD in the advancement of scientific knowledge. Here, we report on recent enhancements made to the resource and new features. PMID:27899570

  13. An annotation infrastructure for the analysis and interpretation of Affymetrix exon array data.

    PubMed

    Okoniewski, Michał J; Yates, Tim; Dibben, Siân; Miller, Crispin J

    2007-01-01

    Affymetrix exon arrays contain probesets intended to target every known and predicted exon in the entire genome, posing significant challenges for high-throughput genome-wide data analysis. X:MAP http://xmap.picr.man.ac.uk, an annotation database, and exonmap http://www.bioconductor.org/packages/2.0/bioc/html/exonmap.html, a BioConductor/R package, are designed to support fine-grained analysis of exon array data. The system supports the application of standard statistical techniques, prior to the use of genome scale annotation to provide gene-, transcript- and exon-level summaries and visualization tools.

  14. An annotation infrastructure for the analysis and interpretation of Affymetrix exon array data

    PubMed Central

    Okoniewski, Michał J; Yates, Tim; Dibben, Siân; Miller, Crispin J

    2007-01-01

    Affymetrix exon arrays contain probesets intended to target every known and predicted exon in the entire genome, posing significant challenges for high-throughput genome-wide data analysis. X:MAP , an annotation database, and exonmap , a BioConductor/R package, are designed to support fine-grained analysis of exon array data. The system supports the application of standard statistical techniques, prior to the use of genome scale annotation to provide gene-, transcript- and exon-level summaries and visualization tools. PMID:17498294

  15. Comparison of the genomes of human and mouse lays the foundation of genome zoology.

    PubMed

    Emes, Richard D; Goodstadt, Leo; Winter, Eitan E; Ponting, Chris P

    2003-04-01

    The extensive similarities between the genomes of human and model organisms are the foundation of much of modern biology, with model organism experimentation permitting valuable insights into biological function and the aetiology of human disease. In contrast, differences among genomes have received less attention. Yet these can be expected to govern the physiological and morphological distinctions apparent among species, especially if such differences are the result of evolutionary adaptation. A recent comparison of the draft sequences of mouse and human genomes has shed light on the selective forces that have predominated in their recent evolutionary histories. In particular, mouse-specific clusters of homologues associated with roles in reproduction, immunity and host defence appear to be under diversifying positive selective pressure, as indicated by high ratios of non-synonymous to synonymous substitution rates. These clusters are also frequently punctuated by homologous pseudogenes. They thus have experienced numerous gene death, as well as gene birth, events. These regions appear, therefore, to have borne the brunt of adaptive evolution that underlies physiological and behavioural innovation in mice. We predict that the availability of numerous animal genomes will give rise to a new field of genome zoology in which differences in animal physiology and ethology are illuminated by the study of genomic sequence variations.

  16. Micro-Analyzer: automatic preprocessing of Affymetrix microarray data.

    PubMed

    Guzzi, Pietro Hiram; Cannataro, Mario

    2013-08-01

    A current trend in genomics is the investigation of the cell mechanism using different technologies, in order to explain the relationship among genes, molecular processes and diseases. For instance, the combined use of gene-expression arrays and genomic arrays has been demonstrated as an effective instrument in clinical practice. Consequently, in a single experiment different kind of microarrays may be used, resulting in the production of different types of binary data (images and textual raw data). The analysis of microarray data requires an initial preprocessing phase, that makes raw data suitable for use on existing analysis platforms, such as the TIGR M4 (TM4) Suite. An additional challenge to be faced by emerging data analysis platforms is the ability to treat in a combined way those different microarray formats coupled with clinical data. In fact, resulting integrated data may include both numerical and symbolic data (e.g. gene expression and SNPs regarding molecular data), as well as temporal data (e.g. the response to a drug, time to progression and survival rate), regarding clinical data. Raw data preprocessing is a crucial step in analysis but is often performed in a manual and error prone way using different software tools. Thus novel, platform independent, and possibly open source tools enabling the semi-automatic preprocessing and annotation of different microarray data are needed. The paper presents Micro-Analyzer (Microarray Analyzer), a cross-platform tool for the automatic normalization, summarization and annotation of Affymetrix gene expression and SNP binary data. It represents the evolution of the μ-CS tool, extending the preprocessing to SNP arrays that were not allowed in μ-CS. The Micro-Analyzer is provided as a Java standalone tool and enables users to read, preprocess and analyse binary microarray data (gene expression and SNPs) by invoking TM4 platform. It avoids: (i) the manual invocation of external tools (e.g. the Affymetrix Power

  17. Celsius: a community resource for Affymetrix microarray data.

    PubMed

    Day, Allen; Carlson, Marc R J; Dong, Jun; O'Connor, Brian D; Nelson, Stanley F

    2007-01-01

    Celsius is a data warehousing system to aggregate Affymetrix CEL files and associated metadata. It provides mechanisms for importing, storing, querying, and exporting large volumes of primary and pre-processed microarray data. Celsius contains ten billion assay measurements and affiliated metadata. It is the largest publicly available source of Affymetrix microarray data, and through sheer volume it allows a sophisticated, broad view of transcription that has not previously been possible.

  18. Chromosomal rearrangements associated with LINE elements in the mouse genome.

    PubMed Central

    Shyman, S; Weaver, S

    1985-01-01

    Two segments of DNA that have apparently inserted in the interval between the two adult beta-globin genes in BALB/c (Hbbd haplotype) but not in C57B1/10 (Hbbs haplotype) mouse strains have been described (1). These putative insertions, each about 1000 bp in length, mapped near a repetitive element. To determine the precise position of these alleged insertions, their target sites, and the nature of their boundaries, we cloned and sequenced the appropriate regions of both chromosomes. One of the two segments is not an insertion but rather a region between two independently integrated L1 repetitive elements (LINEs) (2), one in Hbbd and the other in the Hbbs chromosome. The other segment is an insertion of 940 bp which is located within the L1 element in the Hbbd chromosome. This insert is unusual in that it exists in only one copy in the BALB/c genome. PMID:2991852

  19. Efficient analysis of mouse genome sequences reveal many nonsense variants

    PubMed Central

    Steeland, Sophie; Timmermans, Steven; Van Ryckeghem, Sara; Hulpiau, Paco; Saeys, Yvan; Van Montagu, Marc; Vandenbroucke, Roosmarijn E.; Libert, Claude

    2016-01-01

    Genetic polymorphisms in coding genes play an important role when using mouse inbred strains as research models. They have been shown to influence research results, explain phenotypical differences between inbred strains, and increase the amount of interesting gene variants present in the many available inbred lines. SPRET/Ei is an inbred strain derived from Mus spretus that has ∼1% sequence difference with the C57BL/6J reference genome. We obtained a listing of all SNPs and insertions/deletions (indels) present in SPRET/Ei from the Mouse Genomes Project (Wellcome Trust Sanger Institute) and processed these data to obtain an overview of all transcripts having nonsynonymous coding sequence variants. We identified 8,883 unique variants affecting 10,096 different transcripts from 6,328 protein-coding genes, which is about 28% of all coding genes. Because only a subset of these variants results in drastic changes in proteins, we focused on variations that are nonsense mutations that ultimately resulted in a gain of a stop codon. These genes were identified by in silico changing the C57BL/6J coding sequences to the SPRET/Ei sequences, converting them to amino acid (AA) sequences, and comparing the AA sequences. All variants and transcripts affected were also stored in a database, which can be browsed using a SPRET/Ei M. spretus variants web tool (www.spretus.org), including a manual. We validated the tool by demonstrating the loss of function of three proteins predicted to be severely truncated, namely Fas, IRAK2, and IFNγR1. PMID:27147605

  20. Disease model curation improvements at Mouse Genome Informatics

    PubMed Central

    Bello, Susan M.; Richardson, Joel E.; Davis, Allan P.; Wiegers, Thomas C.; Mattingly, Carolyn J.; Dolan, Mary E.; Smith, Cynthia L.; Blake, Judith A.; Eppig, Janan T.

    2012-01-01

    Optimal curation of human diseases requires an ontology or structured vocabulary that contains terms familiar to end users, is robust enough to support multiple levels of annotation granularity, is limited to disease terms and is stable enough to avoid extensive reannotation following updates. At Mouse Genome Informatics (MGI), we currently use disease terms from Online Mendelian Inheritance in Man (OMIM) to curate mouse models of human disease. While OMIM provides highly detailed disease records that are familiar to many in the medical community, it lacks structure to support multilevel annotation. To improve disease annotation at MGI, we evaluated the merged Medical Subject Headings (MeSH) and OMIM disease vocabulary created by the Comparative Toxicogenomics Database (CTD) project. Overlaying MeSH onto OMIM provides hierarchical access to broad disease terms, a feature missing from the OMIM. We created an extended version of the vocabulary to meet the genetic disease-specific curation needs at MGI. Here we describe our evaluation of the CTD application, the extensions made by MGI and discuss the strengths and weaknesses of this approach. Database URL: http://www.informatics.jax.org/ PMID:22434831

  1. Heterogeneity in rates of recombination across the mouse genome

    SciTech Connect

    Nachman, M.W.; Churchill, G.A.

    1996-02-01

    If loci are randomly distributed on a physical map, the density of markers on a genetic map will be inversely proportional to recombination rate. First proposed by Mary Lyon, we have used this idea to estimate recombination rates from the Drosophila melanogaster linkage map. These results were compared with results of two other studies that estimated regional recombination rates in D. melanogaster using both physical and genetic maps. The three methods were largely concordant in identifying large-scale genomic patterns of recombination. The marker density method was then applied to the Mus musculus microsatellite linkage map. The distribution of microsatellites provided evidence for heterogeneity in recombination rates. Centromeric regions for several mouse chromosomes had significantly greater numbers of markers than expected, suggesting that recombination rates were lower in these regions. In contrast, most telomeric regions contained significantly fewer markers than expected. This indicates that recombination rates are elevated at the telomeres of many mouse chromosomes and is consistent with a comparison of the genetic and cytogenetic maps in these regions. The density of markers on a genetic map may provide a generally useful way to estimate regional recombination rates in species for which genetic, but not physical, maps are available. 44 refs., 5 figs., 4 tabs.

  2. Complete Genome Sequences of 12 Species of Stable Defined Moderately Diverse Mouse Microbiota 2

    PubMed Central

    Uchimura, Yasuhiro; Wyss, Madeleine; Brugiroux, Sandrine; Limenitakis, Julien P.; Stecher, Bärbel; McCoy, Kathy D.

    2016-01-01

    We report here the complete genome sequences of 12 bacterial species of stable defined moderately diverse mouse microbiota 2 (sDMDMm2) used to colonize germ-free mice with defined microbes. Whole-genome sequencing of these species was performed using the PacBio sequencing platform yielding circularized genome sequences of all 12 species. PMID:27634994

  3. The mouse genome database: genotypes, phenotypes, and models of human disease.

    PubMed

    Bult, Carol J; Eppig, Janan T; Blake, Judith A; Kadin, James A; Richardson, Joel E

    2013-01-01

    The laboratory mouse is the premier animal model for studying human biology because all life stages can be accessed experimentally, a completely sequenced reference genome is publicly available and there exists a myriad of genomic tools for comparative and experimental research. In the current era of genome scale, data-driven biomedical research, the integration of genetic, genomic and biological data are essential for realizing the full potential of the mouse as an experimental model. The Mouse Genome Database (MGD; http://www.informatics.jax.org), the community model organism database for the laboratory mouse, is designed to facilitate the use of the laboratory mouse as a model system for understanding human biology and disease. To achieve this goal, MGD integrates genetic and genomic data related to the functional and phenotypic characterization of mouse genes and alleles and serves as a comprehensive catalog for mouse models of human disease. Recent enhancements to MGD include the addition of human ortholog details to mouse Gene Detail pages, the inclusion of microRNA knockouts to MGD's catalog of alleles and phenotypes, the addition of video clips to phenotype images, providing access to genotype and phenotype data associated with quantitative trait loci (QTL) and improvements to the layout and display of Gene Ontology annotations.

  4. The Tennessee Mouse Genome Consortium: Identification of ocular mutants

    SciTech Connect

    Jablonski, Monica M.; Wang, Xiaofei; Lu, Lu; Miller, Darla R; Rinchik, Eugene M; Williams, Robert; Goldowitz, Daniel

    2005-06-01

    The Tennessee Mouse Genome Consortium (TMGC) is in its fifth year of a ethylnitrosourea (ENU)-based mutagenesis screen to detect recessive mutations that affect the eye and brain. Each pedigree is tested by various phenotyping domains including the eye, neurohistology, behavior, aging, ethanol, drug, social behavior, auditory, and epilepsy domains. The utilization of a highly efficient breeding protocol and coordination of various universities across Tennessee makes it possible for mice with ENU-induced mutations to be evaluated by nine distinct phenotyping domains within this large-scale project known as the TMGC. Our goal is to create mutant lines that model human diseases and disease syndromes and to make the mutant mice available to the scientific research community. Within the eye domain, mice are screened for anterior and posterior segment abnormalities using slit-lamp biomicroscopy, indirect ophthalmoscopy, fundus photography, eye weight, histology, and immunohistochemistry. As of January 2005, we have screened 958 pedigrees and 4800 mice, excluding those used in mapping studies. We have thus far identified seven pedigrees with primary ocular abnormalities. Six of the mutant pedigrees have retinal or subretinal aberrations, while the remaining pedigree presents with an abnormal eye size. Continued characterization of these mutant mice should in most cases lead to the identification of the mutated gene, as well as provide insight into the function of each gene. Mice from each of these pedigrees of mutant mice are available for distribution to researchers for independent study.

  5. NIG_MoG: a mouse genome navigator for exploring intersubspecific genetic polymorphisms.

    PubMed

    Takada, Toyoyuki; Yoshiki, Atsushi; Obata, Yuichi; Yamazaki, Yukiko; Shiroishi, Toshihiko

    2015-08-01

    The National Institute of Genetics Mouse Genome database (NIG_MoG; http://molossinus.lab.nig.ac.jp/msmdb/) primarily comprises the whole-genome sequence data of two inbred mouse strains, MSM/Ms and JF1/Ms. These strains were established at NIG and originated from the Japanese subspecies Mus musculus molossinus. NIG_MoG provides visualized genome polymorphism information, browsing single-nucleotide polymorphisms and short insertions and deletions in the genomes of MSM/Ms and JF1/Ms with respect to C57BL/6J (whose genome is predominantly derived from the West European subspecies M. m. domesticus). This allows users, especially wet-lab biologists, to intuitively recognize intersubspecific genome divergence in these mouse strains using visual data. The database also supports the in silico screening of bacterial artificial chromosome (BAC) clones that contain genomic DNA from MSM/Ms and the standard classical laboratory strain C57BL/6N. NIG_MoG is thus a valuable navigator for exploring mouse genome polymorphisms and BAC clones that are useful for studies of gene function and regulation based on intersubspecific genome divergence.

  6. The paternal genome in mouse zygotes is less sensitive to ENU mutagenesis than the maternal genome.

    PubMed

    Russell, L B; Bangham, J W

    1991-05-01

    The two parental genomes lie separate within the zygote and may be differentially affected by environmental influences. We have shown earlier (Russell et al., 1988) that the maternal genome within the mouse zygote is exquisitely sensitive to the induction of point mutations by N-ethyl-N-nitrosourea (ENU), and that the initial lesion probably occurs in one strand of the DNA. The present experiment measured specific-locus mutation induction in the paternal genome. Zygotes containing a multiple-recessive maternal genome (a; b; p cch; d se; s) and the corresponding wild-type alleles in the paternal one were exposed to 50 mg ENU/kg in vivo at one of two stages: the presumed times of sperm entry and early pronuclear stage. At weaning age, the resulting mice were examined for mutations at the marked loci as well as for other mutations producing externally visible phenotypes. At the marked loci, one possible mosaic (for b) was observed among 2113 classified offspring that had been treated with ENU as zygotes; this animal failed to transmit a mutation. By contrast, in the reciprocal cross (which tests the maternal genome) we had observed 8 specific-locus mutations (6 of them mosaics) among 1555 offspring that had received the same dose of ENU during sperm entry (and completion of oocyte meiosis II). In the present experiment, we also found one mutation at other loci (two at other loci in the reciprocal cross). The frequency of offspring with small white belly spots was significantly greater in the treated groups (3.5 and 1.9% at the earlier and later stage, respectively) than in the control (1.0%), the excess being almost entirely due to daughters. Genetic tests of a large number of such offspring failed to find a genetic cause. Instead, it appears that this phenotype may be influenced by factors in the intrauterine environment. It is concluded that shortly after sperm entry, the paternal genome of the zygote is less sensitive than the maternal one to the induction of

  7. The mouse QTL map helps interpret human genome-wide association studies for HDL cholesterol.

    PubMed

    Leduc, Magalie S; Lyons, Malcolm; Darvishi, Katayoon; Walsh, Kenneth; Sheehan, Susan; Amend, Sarah; Cox, Allison; Orho-Melander, Marju; Kathiresan, Sekar; Paigen, Beverly; Korstanje, Ron

    2011-06-01

    Genome-wide association (GWA) studies represent a powerful strategy for identifying susceptibility genes for complex diseases in human populations but results must be confirmed and replicated. Because of the close homology between mouse and human genomes, the mouse can be used to add evidence to genes suggested by human studies. We used the mouse quantitative trait loci (QTL) map to interpret results from a GWA study for genes associated with plasma HDL cholesterol levels. We first positioned single nucleotide polymorphisms (SNPs) from a human GWA study on the genomic map for mouse HDL QTL. We then used mouse bioinformatics, sequencing, and expression studies to add evidence for one well-known HDL gene (Abca1) and three newly identified genes (Galnt2, Wwox, and Cdh13), thus supporting the results of the human study. For GWA peaks that occur in human haplotype blocks with multiple genes, we examined the homologous regions in the mouse to prioritize the genes using expression, sequencing, and bioinformatics from the mouse model, showing that some genes were unlikely candidates and adding evidence for candidate genes Mvk and Mmab in one haplotype block and Fads1 and Fads2 in the second haplotype block. Our study highlights the value of mouse genetics for evaluating genes found in human GWA studies.

  8. Lineage-Specific Biology Revealed by a Finished Genome Assembly of the Mouse

    PubMed Central

    Hillier, LaDeana W.; Zody, Michael C.; Goldstein, Steve; She, Xinwe; Bult, Carol J.; Agarwala, Richa; Cherry, Joshua L.; DiCuccio, Michael; Hlavina, Wratko; Kapustin, Yuri; Meric, Peter; Maglott, Donna; Birtle, Zoë; Marques, Ana C.; Graves, Tina; Zhou, Shiguo; Teague, Brian; Potamousis, Konstantinos; Churas, Christopher; Place, Michael; Herschleb, Jill; Runnheim, Ron; Forrest, Daniel; Amos-Landgraf, James; Schwartz, David C.; Cheng, Ze; Lindblad-Toh, Kerstin; Eichler, Evan E.; Ponting, Chris P.

    2009-01-01

    The mouse (Mus musculus) is the premier animal model for understanding human disease and development. Here we show that a comprehensive understanding of mouse biology is only possible with the availability of a finished, high-quality genome assembly. The finished clone-based assembly of the mouse strain C57BL/6J reported here has over 175,000 fewer gaps and over 139 Mb more of novel sequence, compared with the earlier MGSCv3 draft genome assembly. In a comprehensive analysis of this revised genome sequence, we are now able to define 20,210 protein-coding genes, over a thousand more than predicted in the human genome (19,042 genes). In addition, we identified 439 long, non–protein-coding RNAs with evidence for transcribed orthologs in human. We analyzed the complex and repetitive landscape of 267 Mb of sequence that was missing or misassembled in the previously published assembly, and we provide insights into the reasons for its resistance to sequencing and assembly by whole-genome shotgun approaches. Duplicated regions within newly assembled sequence tend to be of more recent ancestry than duplicates in the published draft, correcting our initial understanding of recent evolution on the mouse lineage. These duplicates appear to be largely composed of sequence regions containing transposable elements and duplicated protein-coding genes; of these, some may be fixed in the mouse population, but at least 40% of segmentally duplicated sequences are copy number variable even among laboratory mouse strains. Mouse lineage-specific regions contain 3,767 genes drawn mainly from rapidly-changing gene families associated with reproductive functions. The finished mouse genome assembly, therefore, greatly improves our understanding of rodent-specific biology and allows the delineation of ancestral biological functions that are shared with human from derived functions that are not. PMID:19468303

  9. Lineage-specific biology revealed by a finished genome assembly of the mouse.

    PubMed

    Church, Deanna M; Goodstadt, Leo; Hillier, Ladeana W; Zody, Michael C; Goldstein, Steve; She, Xinwe; Bult, Carol J; Agarwala, Richa; Cherry, Joshua L; DiCuccio, Michael; Hlavina, Wratko; Kapustin, Yuri; Meric, Peter; Maglott, Donna; Birtle, Zoë; Marques, Ana C; Graves, Tina; Zhou, Shiguo; Teague, Brian; Potamousis, Konstantinos; Churas, Christopher; Place, Michael; Herschleb, Jill; Runnheim, Ron; Forrest, Daniel; Amos-Landgraf, James; Schwartz, David C; Cheng, Ze; Lindblad-Toh, Kerstin; Eichler, Evan E; Ponting, Chris P

    2009-05-05

    The mouse (Mus musculus) is the premier animal model for understanding human disease and development. Here we show that a comprehensive understanding of mouse biology is only possible with the availability of a finished, high-quality genome assembly. The finished clone-based assembly of the mouse strain C57BL/6J reported here has over 175,000 fewer gaps and over 139 Mb more of novel sequence, compared with the earlier MGSCv3 draft genome assembly. In a comprehensive analysis of this revised genome sequence, we are now able to define 20,210 protein-coding genes, over a thousand more than predicted in the human genome (19,042 genes). In addition, we identified 439 long, non-protein-coding RNAs with evidence for transcribed orthologs in human. We analyzed the complex and repetitive landscape of 267 Mb of sequence that was missing or misassembled in the previously published assembly, and we provide insights into the reasons for its resistance to sequencing and assembly by whole-genome shotgun approaches. Duplicated regions within newly assembled sequence tend to be of more recent ancestry than duplicates in the published draft, correcting our initial understanding of recent evolution on the mouse lineage. These duplicates appear to be largely composed of sequence regions containing transposable elements and duplicated protein-coding genes; of these, some may be fixed in the mouse population, but at least 40% of segmentally duplicated sequences are copy number variable even among laboratory mouse strains. Mouse lineage-specific regions contain 3,767 genes drawn mainly from rapidly-changing gene families associated with reproductive functions. The finished mouse genome assembly, therefore, greatly improves our understanding of rodent-specific biology and allows the delineation of ancestral biological functions that are shared with human from derived functions that are not.

  10. A whole-genome mouse BAC microarray with 1-Mb resolution for analysis of DNA copy number changes by array comparative genomic hybridization.

    PubMed

    Chung, Yeun-Jun; Jonkers, Jos; Kitson, Hannah; Fiegler, Heike; Humphray, Sean; Scott, Carol; Hunt, Sarah; Yu, Yuejin; Nishijima, Ichiko; Velds, Arno; Holstege, Henne; Carter, Nigel; Bradley, Allan

    2004-01-01

    Microarray-based comparative genomic hybridization (CGH) has become a powerful method for the genome-wide detection of chromosomal imbalances. Although BAC microarrays have been used for mouse CGH studies, the resolving power of these analyses was limited because high-density whole-genome mouse BAC microarrays were not available. We therefore developed a mouse BAC microarray containing 2803 unique BAC clones from mouse genomic libraries at 1-Mb intervals. For the general amplification of BAC clone DNA prior to spotting, we designed a set of three novel degenerate oligonucleotide-primed (DOP) PCR primers that preferentially amplify mouse genomic sequences while minimizing unwanted amplification of contaminating Escherichia coli DNA. The resulting 3K mouse BAC microarrays reproducibly identified DNA copy number alterations in cell lines and primary tumors, such as single-copy deletions, regional amplifications, and aneuploidy.

  11. Fine-scale maps of recombination rates and hotspots in the mouse genome.

    PubMed

    Brunschwig, Hadassa; Levi, Liat; Ben-David, Eyal; Williams, Robert W; Yakir, Benjamin; Shifman, Sagiv

    2012-07-01

    Recombination events are not uniformly distributed and often cluster in narrow regions known as recombination hotspots. Several studies using different approaches have dramatically advanced our understanding of recombination hotspot regulation. Population genetic data have been used to map and quantify hotspots in the human genome. Genetic variation in recombination rates and hotspots usage have been explored in human pedigrees, mouse intercrosses, and by sperm typing. These studies pointed to the central role of the PRDM9 gene in hotspot modulation. In this study, we used single nucleotide polymorphisms (SNPs) from whole-genome resequencing and genotyping studies of mouse inbred strains to estimate recombination rates across the mouse genome and identified 47,068 historical hotspots--an average of over 2477 per chromosome. We show by simulation that inbred mouse strains can be used to identify positions of historical hotspots. Recombination hotspots were found to be enriched for the predicted binding sequences for different alleles of the PRDM9 protein. Recombination rates were on average lower near transcription start sites (TSS). Comparing the inferred historical recombination hotspots with the recent genome-wide mapping of double-strand breaks (DSBs) in mouse sperm revealed a significant overlap, especially toward the telomeres. Our results suggest that inbred strains can be used to characterize and study the dynamics of historical recombination hotspots. They also strengthen previous findings on mouse recombination hotspots, and specifically the impact of sequence variants in Prdm9.

  12. De novo DNA methylation of the paternal genome in 2-cell mouse embryos.

    PubMed

    Ma, X S; Wang, X G; Qin, L; Song, C L; Lin, F; Song, J M; Zhu, C C; Liu, H L

    2014-10-27

    The developmental dynamics of DNA methylation events have been well studied. Active demethylation of the paternal genome occurs in the zygote, passive demethylation occurs during cleavage stages, and de novo methylation occurs by the blastocyst stage. It is believed that the paternal genome has lower levels of methylation during early development than the maternal genome. However, in this study, we provide direct and indirect evidence of genome-wide de novo DNA methylation of the paternal genome after the first cell cycle in mouse embryos. Although very little methylation was detected within the male pronucleus in zygotes, an intense methylation signal was clearly visible within the androgenetic 2-cell embryos. Moreover, the DNA methylation level of the paternal genome in the post-zygotic metaphase embryos was similar to that of the maternal genome. Using indirect immunofluorescence with an antibody to methylated lysine 9 in histone H3, we provided new evidence to support the concept of spatial compartmentalization of parental genomes in 2-cell mouse embryos. Nevertheless, the transient segregation of parental genomes was not observed by determining the DNA methylation distribution in the 2-cell embryos even though DNA methylation asymmetry between the maternal and paternal pronucleus existed in the 1-cell stage. The disappearance of separate immunofluorescence signals of 5-methyl cytosine in the 2-cell embryos might be attributed to the de novo methylation of the paternal genome during the first mitotic cycle.

  13. Characteristics of the mouse genomic histamine H1 receptor gene

    SciTech Connect

    Inoue, Isao; Taniuchi, Ichiro; Kitamura, Daisuke

    1996-08-15

    We report here the molecular cloning of a mouse histamine H1 receptor gene. The protein deduced from the nucleotide sequence is composed of 488 amino acid residues with characteristic properties of GTP binding protein-coupled receptors. Our results suggest that the mouse histamine H1 receptor gene is a single locus, and no related sequences were detected. Interspecific backcross analysis indicated that the mouse histamine H1 receptor gene (Hrh1) is located in the central region of mouse Chromosome 6 linked to microphthalmia (Mitfmi), ras-related fibrosarcoma oncogene 1 (Raf1), and ret proto-oncogene (Ret) in a region of homology with human chromosome 3p. 12 refs., 3 figs.

  14. Cross Species Genomic Analysis Identifies a Mouse Model as Undifferentiated Pleomorphic Sarcoma/Malignant Fibrous Histiocytoma

    PubMed Central

    Mito, Jeffrey K.; Riedel, Richard F.; Dodd, Leslie; Lahat, Guy; Lazar, Alexander J.; Dodd, Rebecca D.; Stangenberg, Lars; Eward, William C.; Hornicek, Francis J.; Yoon, Sam S.; Brigman, Brian E.; Jacks, Tyler; Lev, Dina; Mukherjee, Sayan; Kirsch, David G.

    2009-01-01

    Undifferentiated pleomorphic sarcoma/Malignant Fibrous Histiocytoma (MFH) is one of the most common subtypes of human soft tissue sarcoma. Using cross species genomic analysis, we define a geneset from the LSL-KrasG12D; Trp53Flox/Flox mouse model of soft tissue sarcoma that is highly enriched in human MFH. With this mouse geneset as a filter, we identify expression of the RAS target FOXM1 in human MFH. Expression of Foxm1 is elevated in mouse sarcomas that metastasize to the lung and tissue microarray analysis of human MFH correlates overexpression of FOXM1 with metastasis. These results suggest that genomic alterations present in human MFH are conserved in the LSL-KrasG12D; p53Flox/Flox mouse model of soft tissue sarcoma and demonstrate the utility of this pre-clinical model. PMID:19956606

  15. Comparative analysis of genome maintenance genes in naked mole rat, mouse, and human.

    PubMed

    MacRae, Sheila L; Zhang, Quanwei; Lemetre, Christophe; Seim, Inge; Calder, Robert B; Hoeijmakers, Jan; Suh, Yousin; Gladyshev, Vadim N; Seluanov, Andrei; Gorbunova, Vera; Vijg, Jan; Zhang, Zhengdong D

    2015-04-01

    Genome maintenance (GM) is an essential defense system against aging and cancer, as both are characterized by increased genome instability. Here, we compared the copy number variation and mutation rate of 518 GM-associated genes in the naked mole rat (NMR), mouse, and human genomes. GM genes appeared to be strongly conserved, with copy number variation in only four genes. Interestingly, we found NMR to have a higher copy number of CEBPG, a regulator of DNA repair, and TINF2, a protector of telomere integrity. NMR, as well as human, was also found to have a lower rate of germline nucleotide substitution than the mouse. Together, the data suggest that the long-lived NMR, as well as human, has more robust GM than mouse and identifies new targets for the analysis of the exceptional longevity of the NMR.

  16. Automated whole-genome multiple alignment of rat, mouse, and human

    SciTech Connect

    Brudno, Michael; Poliakov, Alexander; Salamov, Asaf; Cooper, Gregory M.; Sidow, Arend; Rubin, Edward M.; Solovyev, Victor; Batzoglou, Serafim; Dubchak, Inna

    2004-07-04

    We have built a whole genome multiple alignment of the three currently available mammalian genomes using a fully automated pipeline which combines the local/global approach of the Berkeley Genome Pipeline and the LAGAN program. The strategy is based on progressive alignment, and consists of two main steps: (1) alignment of the mouse and rat genomes; and (2) alignment of human to either the mouse-rat alignments from step 1, or the remaining unaligned mouse and rat sequences. The resulting alignments demonstrate high sensitivity, with 87% of all human gene-coding areas aligned in both mouse and rat. The specificity is also high: <7% of the rat contigs are aligned to multiple places in human and 97% of all alignments with human sequence > 100kb agree with a three-way synteny map built independently using predicted exons in the three genomes. At the nucleotide level <1% of the rat nucleotides are mapped to multiple places in the human sequence in the alignment; and 96.5% of human nucleotides within all alignments agree with the synteny map. The alignments are publicly available online, with visualization through the novel Multi-VISTA browser that we also present.

  17. Exon array data analysis using Affymetrix power tools and R statistical software

    PubMed Central

    2011-01-01

    The use of microarray technology to measure gene expression on a genome-wide scale has been well established for more than a decade. Methods to process and analyse the vast quantity of expression data generated by a typical microarray experiment are similarly well-established. The Affymetrix Exon 1.0 ST array is a relatively new type of array, which has the capability to assess expression at the individual exon level. This allows a more comprehensive analysis of the transcriptome, and in particular enables the study of alternative splicing, a gene regulation mechanism important in both normal conditions and in diseases. Some aspects of exon array data analysis are shared with those for standard gene expression data but others present new challenges that have required development of novel tools. Here, I will introduce the exon array and present a detailed example tutorial for analysis of data generated using this platform. PMID:21498550

  18. Exon array data analysis using Affymetrix power tools and R statistical software.

    PubMed

    Lockstone, Helen E

    2011-11-01

    The use of microarray technology to measure gene expression on a genome-wide scale has been well established for more than a decade. Methods to process and analyse the vast quantity of expression data generated by a typical microarray experiment are similarly well-established. The Affymetrix Exon 1.0 ST array is a relatively new type of array, which has the capability to assess expression at the individual exon level. This allows a more comprehensive analysis of the transcriptome, and in particular enables the study of alternative splicing, a gene regulation mechanism important in both normal conditions and in diseases. Some aspects of exon array data analysis are shared with those for standard gene expression data but others present new challenges that have required development of novel tools. Here, I will introduce the exon array and present a detailed example tutorial for analysis of data generated using this platform.

  19. A radiation hybrid transcript map of the mouse genome.

    PubMed

    Avner, P; Bruls, T; Poras, I; Eley, L; Gas, S; Ruiz, P; Wiles, M V; Sousa-Nunes, R; Kettleborough, R; Rana, A; Morissette, J; Bentley, L; Goldsworthy, M; Haynes, A; Herbert, E; Southam, L; Lehrach, H; Weissenbach, J; Manenti, G; Rodriguez-Tome, P; Beddington, R; Dunwoodie, S; Cox, R D

    2001-10-01

    Expressed-sequence tag (EST) maps are an adjunct to sequence-based analytical methods of gene detection and localization for those species for which such data are available, and provide anchors for high-density homology and orthology mapping in species for which large-scale sequencing has yet to be done. Species for which radiation hybrid-based transcript maps have been established include human, rat, mouse, dog, cat and zebrafish. We have established a comprehensive first-generation-placement radiation hybrid map of the mouse consisting of 5,904 mapped markers (3,993 ESTs and 1,911 sequence-tagged sites (STSs)). The mapped ESTs, which often originate from small-EST clusters, are enriched for genes expressed during early mouse embryogenesis and are probably different from those localized in humans. We have confirmed by in situ hybridization that even singleton ESTs, which are usually not retained for mapping studies, may represent bona fide transcribed sequences. Our studies on mouse chromosomes 12 and 14 orthologous to human chromosome 14 show the power of our radiation hybrid map as a predictive tool for orthology mapping in humans.

  20. Complete Genome Sequence of Acinetobacter sp. Strain NCu2D-2 Isolated from a Mouse

    PubMed Central

    Blaschke, Ulrike

    2017-01-01

    ABSTRACT Whole-genome sequencing of Acinetobacter sp. strain NCu2D-2, isolated from the trachea of a mouse, revealed the presence of a plasmid of 309,964 bp with little overall similarity to known plasmids and enriched in insertion sequences (ISs) closely related to IS elements known from the nosocomial pathogen Acinetobacter baumannii. PMID:28126932

  1. Detailed comparative map of human chromosome 19q and related regions of the mouse genome.

    PubMed

    Stubbs, L; Carver, E A; Shannon, M E; Kim, J; Geisler, J; Generoso, E E; Stanford, B G; Dunn, W C; Mohrenweiser, H; Zimmermann, W; Watt, S M; Ashworth, L K

    1996-08-01

    One of the larger contiguous blocks of mouse-human genomic homology includes the proximal portion of mouse chromosome 7 and the long arm of human chromosome 19. Previous studies have demonstrated the close relationship between the two regions, but have also indicated significant rearrangements in the relative orders of homologous mouse and human genes. Here we present the genetic locations of the homologs of 42 human chromosome 19q markers in the mouse, with an emphasis on genes also included in the human chromosome 19 physical map. Our results demonstrate that despite an overall inversion of sequences relative to the centromere, apparent "transpositions" of three gene-rich segments, and a local inversion of markers mapping near the 19q telomere, gene content, order, and spacing are remarkably well conserved throughout the lengths of these related mouse and human regions. Although most human 19q markers have remained genetically linked in mouse, one small human segment forms a separate region of homology between human chromosome 19q and mouse chromosome 17. Three of the four rearrangements of mouse versus human 19q sequences involve segments that are located directly adjacent to each other in 19q13.3-q13.4, suggesting either the coincident occurrence of these events or their common association with unstable DNA sequences. These data permit an unusually in-depth examination of this large region of mouse-human genomic homology and provide an important new tool to aid in the mapping of genes and associated phenotypes in both species.

  2. Creating reference gene annotation for the mouse C57BL6/J genome assembly.

    PubMed

    Mudge, Jonathan M; Harrow, Jennifer

    2015-10-01

    Annotation on the reference genome of the C57BL6/J mouse has been an ongoing project ever since the draft genome was first published. Initially, the principle focus was on the identification of all protein-coding genes, although today the importance of describing long non-coding RNAs, small RNAs, and pseudogenes is recognized. Here, we describe the progress of the GENCODE mouse annotation project, which combines manual annotation from the HAVANA group with Ensembl computational annotation, alongside experimental and in silico validation pipelines from other members of the consortium. We discuss the more recent incorporation of next-generation sequencing datasets into this workflow, including the usage of mass-spectrometry data to potentially identify novel protein-coding genes. Finally, we will outline how the C57BL6/J genebuild can be used to gain insights into the variant sites that distinguish different mouse strains and species.

  3. Functional organization of the genome may shape the species boundary in the house mouse.

    PubMed

    Janoušek, Václav; Munclinger, Pavel; Wang, Liuyang; Teeter, Katherine C; Tucker, Priscilla K

    2015-05-01

    Genomic features such as rate of recombination and differentiation have been suggested to play a role in species divergence. However, the relationship of these phenomena to functional organization of the genome in the context of reproductive isolation remains unexplored. Here, we examine genomic characteristics of the species boundaries between two house mouse subspecies (Mus musculus musculus/M. m. domesticus). These taxa form a narrow semipermeable zone of secondary contact across Central Europe. Due to the incomplete nature of reproductive isolation, gene flow in the zone varies across the genome. We present an analysis of genomic differentiation, rate of recombination, and functional composition of genes relative to varying amounts of introgression. We assessed introgression using 1,316 autosomal single nucleotide polymorphism markers, previously genotyped in hybrid populations from three transects. We found a significant relationship between amounts of introgression and both genomic differentiation and rate of recombination with genomic regions of reduced introgression associated with higher genomic differentiation and lower rates of recombination, and the opposite for genomic regions of extensive introgression. We also found a striking functional polarization of genes based on where they are expressed in the cell. Regions of elevated introgression exhibit a disproportionate number of genes involved in signal transduction functioning at the cell periphery, among which olfactory receptor genes were found to be the most prominent group. Conversely, genes expressed intracellularly and involved in DNA binding were the most prevalent in regions of reduced introgression. We hypothesize that functional organization of the genome is an important driver of species divergence.

  4. Rat Genome Database: a unique resource for rat, human, and mouse quantitative trait locus data.

    PubMed

    Nigam, Rajni; Laulederkind, Stanley J F; Hayman, G Thomas; Smith, Jennifer R; Wang, Shur-Jen; Lowry, Timothy F; Petri, Victoria; De Pons, Jeff; Tutaj, Marek; Liu, Weisong; Jayaraman, Pushkala; Munzenmaier, Diane H; Worthey, Elizabeth A; Dwinell, Melinda R; Shimoyama, Mary; Jacob, Howard J

    2013-09-16

    The rat has been widely used as a disease model in a laboratory setting, resulting in an abundance of genetic and phenotype data from a wide variety of studies. These data can be found at the Rat Genome Database (RGD, http://rgd.mcw.edu/), which provides a platform for researchers interested in linking genomic variations to phenotypes. Quantitative trait loci (QTLs) form one of the earliest and core datasets, allowing researchers to identify loci harboring genes associated with disease. These QTLs are not only important for those using the rat to identify genes and regions associated with disease, but also for cross-organism analyses of syntenic regions on the mouse and the human genomes to identify potential regions for study in these organisms. Currently, RGD has data on >1,900 rat QTLs that include details about the methods and animals used to determine the respective QTL along with the genomic positions and markers that define the region. RGD also curates human QTLs (>1,900) and houses>4,000 mouse QTLs (imported from Mouse Genome Informatics). Multiple ontologies are used to standardize traits, phenotypes, diseases, and experimental methods to facilitate queries, analyses, and cross-organism comparisons. QTLs are visualized in tools such as GBrowse and GViewer, with additional tools for analysis of gene sets within QTL regions. The QTL data at RGD provide valuable information for the study of mapped phenotypes and identification of candidate genes for disease associations.

  5. Construction of a mouse whole-genome radiation hybrid panel and application to MMU11

    SciTech Connect

    Schmitt, K.; Foster, J.W.; Feakes, R.W.

    1996-06-01

    Whole-genome radiation hybrids have been used to construct human genome maps that integrate different types of markers. To investigate this methodology in mammalian species other than humans, panel of 164 mouse x hamster whole-genome radiation hybrids was constructed. This set of hybrids was used to produce a high-resolution map of a region on MMU11 that included microsatellite markers and cDNA sequences. The mouse homologue of the human SRY-related gene SOX9 was mapped to an interval of approximately 1.1 cM flanked by the microsatellite markers D11Mit11 and D11Mit291. This interval includes the region containing the mouse Tail-short mutation, a possible homologue of the human syndrome campomelic dysplasia, which is caused by mutations in SOX9. Our results suggest that whole-genome radiation hybrid technology will be a useful adjunct to mapping the genomes of nonhuman mammalian species. 31 refs., 2 figs.

  6. Highly Efficient Mouse Genome Editing by CRISPR Ribonucleoprotein Electroporation of Zygotes.

    PubMed

    Chen, Sean; Lee, Benjamin; Lee, Angus Yiu-Fai; Modzelewski, Andrew J; He, Lin

    2016-07-08

    The CRISPR/Cas9 system has been employed to efficiently edit the genomes of diverse model organisms. CRISPR-mediated mouse genome editing is typically accomplished by microinjection of Cas9 DNA/RNA and single guide RNA (sgRNA) into zygotes to generate modified animals in one step. However, microinjection is a technically demanding, labor-intensive, and costly procedure with poor embryo viability. Here, we describe a simple and economic electroporation-based strategy to deliver Cas9/sgRNA ribonucleoproteins into mouse zygotes with 100% efficiency for in vivo genome editing. Our methodology, designated as CRISPR RNP Electroporation of Zygotes (CRISPR-EZ), enables highly efficient and high-throughput genome editing in vivo, with a significant improvement in embryo viability compared with microinjection. Using CRISPR-EZ, we generated a variety of editing schemes in mouse embryos, including indel (insertion/deletion) mutations, point mutations, large deletions, and small insertions. In a proof-of-principle experiment, we used CRISPR-EZ to target the tyrosinase (Tyr) gene, achieving 88% bi-allelic editing and 42% homology-directed repair-mediated precise sequence modification in live mice. Taken together, CRISPR-EZ is simple, economic, high throughput, and highly efficient with the potential to replace microinjection for in vivo genome editing in mice and possibly in other mammals.

  7. In vivo genome editing restores haemostasis in a mouse model of haemophilia.

    PubMed

    Li, Hojun; Haurigot, Virginia; Doyon, Yannick; Li, Tianjian; Wong, Sunnie Y; Bhagwat, Anand S; Malani, Nirav; Anguela, Xavier M; Sharma, Rajiv; Ivanciu, Lacramiora; Murphy, Samuel L; Finn, Jonathan D; Khazi, Fayaz R; Zhou, Shangzhen; Paschon, David E; Rebar, Edward J; Bushman, Frederic D; Gregory, Philip D; Holmes, Michael C; High, Katherine A

    2011-06-26

    Editing of the human genome to correct disease-causing mutations is a promising approach for the treatment of genetic disorders. Genome editing improves on simple gene-replacement strategies by effecting in situ correction of a mutant gene, thus restoring normal gene function under the control of endogenous regulatory elements and reducing risks associated with random insertion into the genome. Gene-specific targeting has historically been limited to mouse embryonic stem cells. The development of zinc finger nucleases (ZFNs) has permitted efficient genome editing in transformed and primary cells that were previously thought to be intractable to such genetic manipulation. In vitro, ZFNs have been shown to promote efficient genome editing via homology-directed repair by inducing a site-specific double-strand break (DSB) at a target locus, but it is unclear whether ZFNs can induce DSBs and stimulate genome editing at a clinically meaningful level in vivo. Here we show that ZFNs are able to induce DSBs efficiently when delivered directly to mouse liver and that, when co-delivered with an appropriately designed gene-targeting vector, they can stimulate gene replacement through both homology-directed and homology-independent targeted gene insertion at the ZFN-specified locus. The level of gene targeting achieved was sufficient to correct the prolonged clotting times in a mouse model of haemophilia B, and remained persistent after induced liver regeneration. Thus, ZFN-driven gene correction can be achieved in vivo, raising the possibility of genome editing as a viable strategy for the treatment of genetic disease.

  8. A detailed genome-wide reconstruction of mouse metabolism based on human Recon 1

    PubMed Central

    2010-01-01

    Background Well-curated and validated network reconstructions are extremely valuable tools in systems biology. Detailed metabolic reconstructions of mammals have recently emerged, including human reconstructions. They raise the question if the various successful applications of microbial reconstructions can be replicated in complex organisms. Results We mapped the published, detailed reconstruction of human metabolism (Recon 1) to other mammals. By searching for genes homologous to Recon 1 genes within mammalian genomes, we were able to create draft metabolic reconstructions of five mammals, including the mouse. Each draft reconstruction was created in compartmentalized and non-compartmentalized version via two different approaches. Using gap-filling algorithms, we were able to produce all cellular components with three out of four versions of the mouse metabolic reconstruction. We finalized a functional model by iterative testing until it passed a predefined set of 260 validation tests. The reconstruction is the largest, most comprehensive mouse reconstruction to-date, accounting for 1,415 genes coding for 2,212 gene-associated reactions and 1,514 non-gene-associated reactions. We tested the mouse model for phenotype prediction capabilities. The majority of predicted essential genes were also essential in vivo. However, our non-tissue specific model was unable to predict gene essentiality for many of the metabolic genes shown to be essential in vivo. Our knockout simulation of the lipoprotein lipase gene correlated well with experimental results, suggesting that softer phenotypes can also be simulated. Conclusions We have created a high-quality mouse genome-scale metabolic reconstruction, iMM1415 (Mus Musculus, 1415 genes). We demonstrate that the mouse model can be used to perform phenotype simulations, similar to models of microbe metabolism. Since the mouse is an important experimental organism, this model should become an essential tool for studying metabolic

  9. Malignant glioma: lessons from genomics, mouse models, and stem cells.

    PubMed

    Chen, Jian; McKay, Renée M; Parada, Luis F

    2012-03-30

    Eighty percent of malignant tumors that develop in the central nervous system are malignant gliomas, which are essentially incurable. Here, we discuss how recent sequencing studies are identifying unexpected drivers of gliomagenesis, including mutations in isocitrate dehydrogenase 1 and the NF-κB pathway, and how genome-wide analyses are reshaping the classification schemes for tumors and enhancing prognostic value of molecular markers. We discuss the controversies surrounding glioma stem cells and explore how the integration of new molecular data allows for the generation of more informative animal models to advance our knowledge of glioma's origin, progression, and treatment.

  10. Mobilization of giant piggyBac transposons in the mouse genome.

    PubMed

    Li, Meng Amy; Turner, Daniel J; Ning, Zemin; Yusa, Kosuke; Liang, Qi; Eckert, Sabine; Rad, Lena; Fitzgerald, Tomas W; Craig, Nancy L; Bradley, Allan

    2011-12-01

    The development of technologies that allow the stable delivery of large genomic DNA fragments in mammalian systems is important for genetic studies as well as for applications in gene therapy. DNA transposons have emerged as flexible and efficient molecular vehicles to mediate stable cargo transfer. However, the ability to carry DNA fragments >10 kb is limited in most DNA transposons. Here, we show that the DNA transposon piggyBac can mobilize 100-kb DNA fragments in mouse embryonic stem (ES) cells, making it the only known transposon with such a large cargo capacity. The integrity of the cargo is maintained during transposition, the copy number can be controlled and the inserted giant transposons express the genomic cargo. Furthermore, these 100-kb transposons can also be excised from the genome without leaving a footprint. The development of piggyBac as a large cargo vector will facilitate a wider range of genetic and genomic applications.

  11. Manual Gene Ontology annotation workflow at the Mouse Genome Informatics Database.

    PubMed

    Drabkin, Harold J; Blake, Judith A

    2012-01-01

    The Mouse Genome Database, the Gene Expression Database and the Mouse Tumor Biology database are integrated components of the Mouse Genome Informatics (MGI) resource (http://www.informatics.jax.org). The MGI system presents both a consensus view and an experimental view of the knowledge concerning the genetics and genomics of the laboratory mouse. From genotype to phenotype, this information resource integrates information about genes, sequences, maps, expression analyses, alleles, strains and mutant phenotypes. Comparative mammalian data are also presented particularly in regards to the use of the mouse as a model for the investigation of molecular and genetic components of human diseases. These data are collected from literature curation as well as downloads of large datasets (SwissProt, LocusLink, etc.). MGI is one of the founding members of the Gene Ontology (GO) and uses the GO for functional annotation of genes. Here, we discuss the workflow associated with manual GO annotation at MGI, from literature collection to display of the annotations. Peer-reviewed literature is collected mostly from a set of journals available electronically. Selected articles are entered into a master bibliography and indexed to one of eight areas of interest such as 'GO' or 'homology' or 'phenotype'. Each article is then either indexed to a gene already contained in the database or funneled through a separate nomenclature database to add genes. The master bibliography and associated indexing provide information for various curator-reports such as 'papers selected for GO that refer to genes with NO GO annotation'. Once indexed, curators who have expertise in appropriate disciplines enter pertinent information. MGI makes use of several controlled vocabularies that ensure uniform data encoding, enable robust analysis and support the construction of complex queries. These vocabularies range from pick-lists to structured vocabularies such as the GO. All data associations are supported

  12. Transcript copy number estimation using a mouse whole-genome oligonucleotide microarray

    PubMed Central

    Carter, Mark G; Sharov, Alexei A; VanBuren, Vincent; Dudekula, Dawood B; Carmack, Condie E; Nelson, Charlie; Ko, Minoru SH

    2005-01-01

    The ability to quantitatively measure the expression of all genes in a given tissue or cell with a single assay is an exciting promise of gene-expression profiling technology. An in situ-synthesized 60-mer oligonucleotide microarray designed to detect transcripts from all mouse genes was validated, as well as a set of exogenous RNA controls derived from the yeast genome (made freely available without restriction), which allow quantitative estimation of absolute endogenous transcript abundance. PMID:15998450

  13. Efficient genome editing in the mouse brain by local delivery of engineered Cas9 ribonucleoprotein complexes.

    PubMed

    Staahl, Brett T; Benekareddy, Madhurima; Coulon-Bainier, Claire; Banfal, Ashwin A; Floor, Stephen N; Sabo, Jennifer K; Urnes, Cole; Munares, Gabriela Acevedo; Ghosh, Anirvan; Doudna, Jennifer A

    2017-02-13

    We demonstrate editing of post-mitotic neurons in the adult mouse brain following injection of Cas9 ribonucleoprotein (RNP) complexes in the hippocampus, striatum and cortex. Engineered variants of Cas9 with multiple SV40 nuclear localization sequences enabled a tenfold increase in the efficiency of neuronal editing in vivo. These advances indicate the potential of genome editing in the brain to correct or inactivate the underlying genetic causes of neurological diseases.

  14. CEL_INTERROGATOR: A FREE AND OPEN SOURCE PACKAGE FOR AFFYMETRIX CEL FILE PARSING

    Technology Transfer Automated Retrieval System (TEKTRAN)

    CEL_Interrogator Package is a suite of programs designed to extract the average probe intensity and other information for each probe sequence from an Affymetrix GeneChip CEL file and unite them with their human-readable Affymetrix consensus sequence names. The resulting text file is suitable for di...

  15. Exploring the utility of human DNA methylation arrays for profiling mouse genomic DNA.

    PubMed

    Wong, Nicholas C; Ng, Jane; Hall, Nathan E; Lunke, Sebastian; Salmanidis, Marika; Brumatti, Gabriela; Ekert, Paul G; Craig, Jeffrey M; Saffery, Richard

    2013-07-01

    Illumina Infinium Human Methylation (HM) BeadChips are widely used for measuring genome-scale DNA methylation, particularly in relation to epigenome-wide association studies (EWAS) studies. The methylation profile of human samples can be assessed accurately and reproducibly using the HM27 BeadChip (27,578 CpG sites) or its successor, the HM450 BeadChip (482,421 CpG sites). To date no mouse equivalent has been developed, greatly hindering the application of this methodology to the wide range of valuable murine models of disease and development currently in existence. We found 1308 and 13,715 probes from HM27 and HM450 BeadChip respectively, uniquely matched the bisulfite converted reference mouse genome (mm9). We demonstrate reproducible measurements of DNA methylation at these probes in a range of mouse tissue samples and in a murine cell line model of acute myeloid leukaemia. In the absence of a mouse counterpart, the Infinium Human Methylation BeadChip arrays have utility for methylation profiling in non-human species.

  16. Improving mRNA 5' coding sequence determination in the mouse genome.

    PubMed

    Piovesan, Allison; Caracausi, Maria; Pelleri, Maria Chiara; Vitale, Lorenza; Martini, Silvia; Bassani, Chiara; Gurioli, Annalisa; Casadei, Raffaella; Soldà, Giulia; Strippoli, Pierluigi

    2014-04-01

    The incomplete determination of the mRNA 5' end sequence may lead to the incorrect assignment of the first AUG codon and to errors in the prediction of the encoded protein product. Due to the significance of the mouse as a model organism in biomedical research, we performed a systematic identification of coding regions at the 5' end of all known mouse mRNAs, using an automated expressed sequence tag (EST)-based approach which we have previously described. By parsing almost 4 million BLAT alignments we found 351 mouse loci, out of 20,221 analyzed, in which an extension of the mRNA 5' coding region was identified. Proof-of-concept confirmation was obtained by in vitro cloning and sequencing for Apc2 and Mknk2 cDNAs. We also generated a list of 16,330 mouse mRNAs where the presence of an in-frame stop codon upstream of the known start codon indicates completeness of the coding sequence at 5' end in the current form. Systematic searches in the main mouse genome databases and genome browsers showed that 82% of our results are original and have not been identified by their annotation pipelines. Moreover, the same information is not easily derivable from RNA-Seq data, due to short sequence length and laboriousness in building full-length transcript structures. In conclusion, our results improve the determination of full-length 5' coding sequences and might be useful in order to reduce errors when studying mouse gene structure and function in biomedical research.

  17. Computational analysis of full-length mouse cDNAs compared with human genome sequences.

    PubMed

    Kondo, S; Shinagawa, A; Saito, T; Kiyosawa, H; Yamanaka, I; Aizawa, K; Fukuda, S; Hara, A; Itoh, M; Kawai, J; Shibata, K; Hayashizaki, Y

    2001-09-01

    Although the sequencing of the human genome is complete, identification of encoded genes and determination of their structures remain a major challenge. In this report, we introduce a method that effectively uses full-length mouse cDNAs to complement efforts in carrying out these difficult tasks. A total of 61,227 RIKEN mouse cDNAs (21,076 full-length and 40,151 EST sequences containing certain redundancies) were aligned with the draft human sequences. We found 35,141 non-redundant genomic regions that showed a significant alignment with the mouse cDNAs. We analyzed the structures and compositional properties of the regions detected by the full-length cDNAs, including cross-species comparisons, and noted a systematic bias of GENSCAN against exons of small size and/or low GC-content. Of the cDNAs locating the 35,141 genomic regions, 3,217 did not match any sequences of the known human genes or ESTs. Among those 3,217 cDNAs, 1,141 did not show any significant similarity to any protein sequence in the GenBank non-redundant protein database and thus are candidates for novel genes.

  18. High correspondence between Affymetrix exon and standard expression arrays.

    PubMed

    Okoniewski, Michał J; Hey, Yvonne; Pepper, Stuart D; Miller, Crispin J

    2007-02-01

    Exon arrays aim to provide comprehensive gene expression data at the level of individual exons, similar to that provided on a per-gene basis by existing expression arrays. This report describes the performance of Affymetrix GeneChip Human Exon 1.0 ST array by using replicated RNA samples from two human cell lines, MCF7 and MCF10A, hybridized both to Exon 1.0 ST and to HG-U133 Plus2 arrays. Cross-comparison between array types requires an appropriate mapping to be found between individual probe sets. Three possible mappings were considered, reflecting different strategies for dealing with probe sets that target different parts of the same transcript. Irrespective of the mapping used, Exon 1.0 ST and HG-U133 Plus2 arrays show a high degree of correspondence. More than 80% of HG-U133 Plus2 probe sets may be mapped to the Exon chip, and fold changes are found well preserved for over 96% of those probe sets detected present. Since HG-U133 Plus2 arrays have already been extensively validated, these results lend a significant degree of confidence to exon arrays.

  19. The genomic landscape shaped by selection on transposable elements across 18 mouse strains

    PubMed Central

    2012-01-01

    Background Transposable element (TE)-derived sequence dominates the landscape of mammalian genomes and can modulate gene function by dysregulating transcription and translation. Our current knowledge of TEs in laboratory mouse strains is limited primarily to those present in the C57BL/6J reference genome, with most mouse TEs being drawn from three distinct classes, namely short interspersed nuclear elements (SINEs), long interspersed nuclear elements (LINEs) and the endogenous retrovirus (ERV) superfamily. Despite their high prevalence, the different genomic and gene properties controlling whether TEs are preferentially purged from, or are retained by, genetic drift or positive selection in mammalian genomes remain poorly defined. Results Using whole genome sequencing data from 13 classical laboratory and 4 wild-derived mouse inbred strains, we developed a comprehensive catalogue of 103,798 polymorphic TE variants. We employ this extensive data set to characterize TE variants across the Mus lineage, and to infer neutral and selective processes that have acted over 2 million years. Our results indicate that the majority of TE variants are introduced though the male germline and that only a minority of TE variants exert detectable changes in gene expression. However, among genes with differential expression across the strains there are twice as many TE variants identified as being putative causal variants as expected. Conclusions Most TE variants that cause gene expression changes appear to be purged rapidly by purifying selection. Our findings demonstrate that past TE insertions have often been highly deleterious, and help to prioritize TE variants according to their likely contribution to gene expression or phenotype variation. PMID:22703977

  20. Regulatory Features for Odorant Receptor Genes in the Mouse Genome

    PubMed Central

    Degl’Innocenti, Andrea; D’Errico, Anna

    2017-01-01

    The odorant receptor genes, seven transmembrane receptor genes constituting the vastest mammalian gene multifamily, are expressed monogenically and monoallelicaly in each sensory neuron in the olfactory epithelium. This characteristic, often referred to as the one neuron–one receptor rule, is driven by mostly uncharacterized molecular dynamics, generally named odorant receptor gene choice. Much attention has been paid by the scientific community to the identification of sequences regulating the expression of odorant receptor genes within their loci, where related genes are usually arranged in genomic clusters. A number of studies identified transcription factor binding sites on odorant receptor promoter sequences. Similar binding sites were also found on a number of enhancers that regulate in cis their transcription, but have been proposed to form interchromosomal networks. Odorant receptor gene choice seems to occur via the local removal of strongly repressive epigenetic markings, put in place during the maturation of the sensory neuron on each odorant receptor locus. Here we review the fast-changing state of art for the study of regulatory features for odorant receptor genes. PMID:28270833

  1. Regulatory Features for Odorant Receptor Genes in the Mouse Genome.

    PubMed

    Degl'Innocenti, Andrea; D'Errico, Anna

    2017-01-01

    The odorant receptor genes, seven transmembrane receptor genes constituting the vastest mammalian gene multifamily, are expressed monogenically and monoallelicaly in each sensory neuron in the olfactory epithelium. This characteristic, often referred to as the one neuron-one receptor rule, is driven by mostly uncharacterized molecular dynamics, generally named odorant receptor gene choice. Much attention has been paid by the scientific community to the identification of sequences regulating the expression of odorant receptor genes within their loci, where related genes are usually arranged in genomic clusters. A number of studies identified transcription factor binding sites on odorant receptor promoter sequences. Similar binding sites were also found on a number of enhancers that regulate in cis their transcription, but have been proposed to form interchromosomal networks. Odorant receptor gene choice seems to occur via the local removal of strongly repressive epigenetic markings, put in place during the maturation of the sensory neuron on each odorant receptor locus. Here we review the fast-changing state of art for the study of regulatory features for odorant receptor genes.

  2. Significant gene content variation characterizes the genomes of inbred mouse strains

    PubMed Central

    Cutler, Gene; Marshall, Lisa A.; Chin, Ni; Baribault, Helene; Kassner, Paul D.

    2007-01-01

    The contribution to genetic diversity of genomic segmental copy number variations (CNVs) is less well understood than that of single-nucleotide polymorphisms (SNPs). While less frequent than SNPs, CNVs have greater potential to affect phenotype. In this study, we have performed the most comprehensive survey to date of CNVs in mice, analyzing the genomes of 42 Mouse Phenome Consortium priority strains. This microarray comparative genomic hybridization (CGH)-based analysis has identified 2094 putative CNVs, with an average of 10 Mb of DNA in 51 CNVs when individual mouse strains were compared to the reference strain C57BL/6J. This amount of variation results in gene content that can differ by hundreds of genes between strains. These genes include members of large families such as the major histocompatibility and pheromone receptor genes, but there are also many singleton genes including genes with expected phenotypic consequences from their deletion or amplification. Using a whole-genome association analysis, we demonstrate that complex multigenic phenotypes, such as food intake, can be associated with specific copy number changes. PMID:17989247

  3. Translating human genetics into mouse: the impact of ultra-rapid in vivo genome editing.

    PubMed

    Aida, Tomomi; Imahashi, Risa; Tanaka, Kohichi

    2014-01-01

    Gene-targeted mutant animals, such as knockout or knockin mice, have dramatically improved our understanding of the functions of genes in vivo and the genetic diversity that characterizes health and disease. However, the generation of targeted mice relies on gene targeting in embryonic stem (ES) cells, which is a time-consuming, laborious, and expensive process. The recent groundbreaking development of several genome editing technologies has enabled the targeted alteration of almost any sequence in any cell or organism. These technologies have now been applied to mouse zygotes (in vivo genome editing), thereby providing new avenues for simple, convenient, and ultra-rapid production of knockout or knockin mice without the need for ES cells. Here, we review recent achievements in the production of gene-targeted mice by in vivo genome editing.

  4. CRISPR/Cas9-Mediated Genome Editing of Mouse Small Intestinal Organoids.

    PubMed

    Schwank, Gerald; Clevers, Hans

    2016-01-01

    The CRISPR/Cas9 system is an RNA-guided genome-editing tool that has been recently developed based on the bacterial CRISPR-Cas immune defense system. Due to its versatility and simplicity, it rapidly became the method of choice for genome editing in various biological systems, including mammalian cells. Here we describe a protocol for CRISPR/Cas9-mediated genome editing in murine small intestinal organoids, a culture system in which somatic stem cells are maintained by self-renewal, while giving rise to all major cell types of the intestinal epithelium. This protocol allows the study of gene function in intestinal epithelial homeostasis and pathophysiology and can be extended to epithelial organoids derived from other internal mouse and human organs.

  5. A Genetic Linkage Map of the Mouse Using Restriction Landmark Genomic Scanning (Rlgs)

    PubMed Central

    Hayashizaki, Y.; Hirotsune, S.; Okazaki, Y.; Shibata, H.; Akasako, A.; Muramatsu, M.; Kawai, J.; Hirasawa, T.; Watanabe, S.; Shiroishi, T.; Moriwaki, K.; Taylor, B. A.; Matsuda, Y.; Elliott, R. W.; Manly, K. F.; Chapman, V. M.

    1994-01-01

    We have developed a multiplex method of genome analysis, restriction landmark genomic scanning (RLGS) that has been used to construct genetic maps in mice. Restriction landmarks are end-labeled restriction fragments of genomic DNA that are separated by using high resolution, two-dimensional gel electrophoresis identifying as many as two thousand landmark loci in a single gel. Variation for several hundred of these loci has been identified between laboratory strains and between these strains and Mus spretus. The segregation of more than 1100 RLGS loci has been analyxed in recombinant inbred (RI) strains and in two separate interspecific genetic crosses. Genetic maps have been derived that link 1045 RLGS loci to reference loci on all of the autosomes and the X chromosome of the mouse genome. The RLGS method can be applied to genome analysis in many different organisms to identify genomic loci because it used end-labeling of restriction landmarks rather than probe hybridization. Different combinations of restriction enzymes yield different sets of RLGS loci providing expanded power for genetic mapping. PMID:7896102

  6. Comparative genome mapping of the deer mouse (Peromyscus maniculatus) reveals greater similarity to rat (Rattus norvegicus) than to the lab mouse (Mus musculus)

    PubMed Central

    2008-01-01

    Background Deer mice (Peromyscus maniculatus) and congeneric species are the most common North American mammals. They represent an emerging system for the genetic analyses of the physiological and behavioral bases of habitat adaptation. Phylogenetic evidence suggests a much more ancient divergence of Peromyscus from laboratory mice (Mus) and rats (Rattus) than that separating latter two. Nevertheless, early karyotypic analyses of the three groups suggest Peromyscus to be exhibit greater similarities with Rattus than with Mus. Results Comparative linkage mapping of an estimated 35% of the deer mouse genome was done with respect to the Rattus and Mus genomes. We particularly focused on regions that span synteny breakpoint regions between the rat and mouse genomes. The linkage analysis revealed the Peromyscus genome to have a higher degree of synteny and gene order conservation with the Rattus genome. Conclusion These data suggest that: 1. the Rattus and Peromyscus genomes more closely represent ancestral Muroid and rodent genomes than that of Mus. 2. the high level of genome rearrangement observed in Muroid rodents is especially pronounced in Mus. 3. evolution of genome organization can operate independently of more commonly assayed measures of genetic change (e.g. SNP frequency). PMID:18302785

  7. Comparison of mouse and human genomes followed by experimental verification yields an estimated 1,019 additional genes.

    PubMed

    Guigo, Roderic; Dermitzakis, Emmanouil T; Agarwal, Pankaj; Ponting, Chris P; Parra, Genis; Reymond, Alexandre; Abril, Josep F; Keibler, Evan; Lyle, Robert; Ucla, Catherine; Antonarakis, Stylianos E; Brent, Michael R

    2003-02-04

    A primary motivation for sequencing the mouse genome was to accelerate the discovery of mammalian genes by using sequence conservation between mouse and human to identify coding exons. Achieving this goal proved challenging because of the large proportion of the mouse and human genomes that is apparently conserved but apparently does not code for protein. We developed a two-stage procedure that exploits the mouse and human genome sequences to produce a set of genes with a much higher rate of experimental verification than previously reported prediction methods. RT-PCR amplification and direct sequencing applied to an initial sample of mouse predictions that do not overlap previously known genes verified the regions flanking one intron in 139 predictions, with verification rates reaching 76%. On average, the confirmed predictions show more restricted expression patterns than the mouse orthologs of known human genes, and two-thirds lack homologs in fish genomes, demonstrating the sensitivity of this dual-genome approach to hard-to-find genes. We verified 112 previously unknown homologs of known proteins, including two homeobox proteins relevant to developmental biology, an aquaporin, and a homolog of dystrophin. We estimate that transcription and splicing can be verified for >1,000 gene predictions identified by this method that do not overlap known genes. This is likely to constitute a significant fraction of the previously unknown, multiexon mammalian genes.

  8. Effects of whole genome duplication on cell size and gene expression in mouse embryonic stem cells

    PubMed Central

    IMAI, Hiroyuki; FUJII, Wataru; KUSAKABE, Ken Takeshi; KISO, Yasuo; KANO, Kiyoshi

    2016-01-01

    Alterations in ploidy tend to influence cell physiology, which in the long-term, contribute to species adaptation and evolution. Polyploid cells are observed under physiological conditions in the nerve and liver tissues, and in tumorigenic processes. Although tetraploid cells have been studied in mammalian cells, the basic characteristics and alterations caused by whole genome duplication are still poorly understood. The purpose of this study was to acquire basic knowledge about the effect of whole genome duplication on the cell cycle, cell size, and gene expression. Using flow cytometry, we demonstrate that cell cycle subpopulations in mouse tetraploid embryonic stem cells (TESCs) were similar to those in embryonic stem cells (ESCs). We performed smear preparations and flow cytometric analysis to identify cell size alterations. These indicated that the relative cell volume of TESCs was approximately 2.2–2.5 fold that of ESCs. We also investigated the effect of whole genome duplication on the expression of housekeeping and pluripotency marker genes using quantitative real-time PCR with external RNA. We found that the target transcripts were 2.2 times more abundant in TESCs than those in ESCs. This indicated that gene expression and cell volume increased in parallel. Our findings suggest the existence of a homeostatic mechanism controlling the cytoplasmic transcript levels in accordance with genome volume changes caused by whole genome duplication. PMID:27569766

  9. Passage Number is a Major Contributor to Genomic Structural Variations in Mouse iPS Cells

    PubMed Central

    Liu, Pengfei; Kaplan, Anna; Yuan, Bo; Hanna, Jacob H.; Lupski, James R.; Reiner, Orly

    2014-01-01

    Emergence of genomic instability is a practical issue in preparing neural stem cells (NSCs) and induced pluripotent stem cells (iPSCs). However, it is still not fully understood what the origins and mechanisms for formation are for the genomic alternations observed. Here, we studied the extent of genomic variation on the scale of individual cells originating from the same animal. We used mouse NSCs grown from embryonic cells and iPSCs generated from embryonic brain cells, B cells or fibroblasts, and performed comparative analysis with cultures of fibroblasts from the same mouse. In the first passage of these cell lines, aneuploidies were only observed for chromosomes 6, 11, 12, 19 and Y, which is overall at a rate lower than previously reported; de novo copy number variations (CNVs) were observed in 4.3% of neural iPSCs, 29% of B cell iPSCs, 10% of fibroblast iPSCs, and 1.3% of neurospheres. In contrast, propagation of these first passage cells to a later passage induced additional aneuploidies and CNVs. Breakpoint sequencing analysis suggested that the majority of the detected CNVs arose by replicative mechanisms. Interestingly, we detected identical de novo CNVs in different single cell colonies that appeared to have arisen independently from each other, which suggests a novel CNV formation mechanism in these cells. Our findings provide insights into mechanisms of CNV formation during reprogramming, and suggest that replicative mechanisms for CNV formation accompany mitotic divisions. PMID:24965231

  10. Comparative analysis of dioxin response elements in human, mouse and rat genomic sequences.

    PubMed

    Sun, Y V; Boverhof, D R; Burgoon, L D; Fielden, M R; Zacharewski, T R

    2004-01-01

    Comparative approaches were used to identify human, mouse and rat dioxin response elements (DREs) in genomic sequences unambiguously assigned to a nucleotide RefSeq accession number. A total of 13 bona fide DREs, all including the substitution intolerant core sequence (GCGTG) and adjacent variable sequences, were used to establish a position weight matrix and a matrix similarity (MS) score threshold to rank identified DREs. DREs with MS scores above the threshold were disproportionately distributed in close proximity to the transcription start site in all three species. Gene expression assays in hepatic mouse tissue confirmed the responsiveness of 192 genes possessing a putative DRE. Previously identified functional DREs in well-characterized AhR-regulated genes including Cyp1a1 and Cyp1b1 were corroborated. Putative DREs were identified in 48 out of 2437 human-mouse-rat orthologous genes between -1500 and the transcriptional start site, of which 19 of these genes possessed positionally conserved DREs as determined by multiple sequence alignment. Seven of these nineteen genes exhibited 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated regulation, although there were significant discrepancies between in vivo and in vitro results. Interestingly, of the mouse-rat orthologous genes with a DRE between -1500 and +1500, only 37% had an equivalent human ortholog. These results suggest that AhR-mediated gene expression may not be well conserved across species, which could have significant implications in human risk assessment.

  11. Novel skin phenotypes revealed by a genome-wide mouse reverse genetic screen

    PubMed Central

    Liakath-Ali, Kifayathullah; Vancollie, Valerie E.; Heath, Emma; Smedley, Damian P.; Estabel, Jeanne; Sunter, David; DiTommaso, Tia; White, Jacqueline K.; Ramirez-Solis, Ramiro; Smyth, Ian; Steel, Karen P.; Watt, Fiona M.

    2014-01-01

    Permanent stop-and-shop large-scale mouse mutant resources provide an excellent platform to decipher tissue phenogenomics. Here we analyse skin from 538 knockout mouse mutants generated by the Sanger Institute Mouse Genetics Project. We optimize immunolabelling of tail epidermal wholemounts to allow systematic annotation of hair follicle, sebaceous gland and interfollicular epidermal abnormalities using ontology terms from the Mammalian Phenotype Ontology. Of the 50 mutants with an epidermal phenotype, 9 map to human genetic conditions with skin abnormalities. Some mutant genes are expressed in the skin, whereas others are not, indicating systemic effects. One phenotype is affected by diet and several are incompletely penetrant. In-depth analysis of three mutants, Krt76, Myo5a (a model of human Griscelli syndrome) and Mysm1, provides validation of the screen. Our study is the first large-scale genome-wide tissue phenotype screen from the International Knockout Mouse Consortium and provides an open access resource for the scientific community. PMID:24721909

  12. Analysis of segmental duplications reveals a distinct pattern of continuation-of-synteny between human and mouse genomes.

    PubMed

    Mehan, Michael R; Almonte, Maricel; Slaten, Erin; Freimer, Nelson B; Rao, P Nagesh; Ophoff, Roel A

    2007-03-01

    About 5% of the human genome consists of large-scale duplicated segments of almost identical sequences. Segmental duplications (SDs) have been proposed to be involved in non-allelic homologous recombination leading to recurrent genomic variation and disease. It has also been suggested that these SDs are associated with syntenic rearrangements that have shaped the human genome. We have analyzed 14 members of a single family of closely related SDs in the human genome, some of which are associated with common inversion polymorphisms at chromosomes 8p23 and 4p16. Comparative analysis with the mouse genome revealed syntenic inversions for these two human polymorphic loci. In addition, 12 of the 14 SDs, while absent in the mouse genome, occur at the breaks of synteny; suggesting a non-random involvement of these sequences in genome evolution. Furthermore, we observed a syntenic familial relationship between 8 and 12 breakpoint-loci, where broken synteny that ends at one family member resumes at another, even across different chromosomes. Subsequent genome-wide assessment revealed that this relationship, which we named continuation-of-synteny, is not limited to the 8p23 family and occurs 46 times in the human genome with high frequency at specific chromosomes. Our analysis supports a non-random breakage model of genomic evolution with an active involvement of segmental duplications for specific regions of the human genome.

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

    PubMed Central

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

    2016-01-01

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

  14. Deconstructing Mus gemischus: advances in understanding ancestry, structure, and variation in the genome of the laboratory mouse

    PubMed Central

    2013-01-01

    The laboratory mouse is an artificial construct with a complex relationship to its natural ancestors. In 2002, the mouse became the first mammalian model organism with a reference genome. Importantly, the mouse genome sequence was assembled from data on a single inbred laboratory strain, C57BL/6. Several large-scale genetic variant discovery efforts have been conducted, resulting in a catalog of tens of millions of SNPs and structural variants. High-density genotyping arrays covering a subset of those variants have been used to produce hundreds of millions of genotypes in laboratory stocks and a small number of wild mice. These landmark resources now enable us to determine relationships among laboratory mice, assign local ancestry at fine scale, resolve important controversies, and identify a new set of challenges—most importantly, the troubling scarcity of genetic data on the very natural populations from which the laboratory mouse was derived. Our aim with this review is to provide the reader with an historical context for the mouse as a model organism and to explain how practical decisions made in the past have influenced both the architecture of the laboratory mouse genome and the design and execution of current large-scale resources. We also provide examples on how the accomplishments of the past decade can be used by researchers to streamline the use of mice in their experiments and correctly interpret results. Finally, we propose future steps that will enable the mouse community to extend its successes in the decade to come. PMID:23223940

  15. In vivo genome editing improves muscle function in a mouse model of Duchenne muscular dystrophy.

    PubMed

    Nelson, Christopher E; Hakim, Chady H; Ousterout, David G; Thakore, Pratiksha I; Moreb, Eirik A; Castellanos Rivera, Ruth M; Madhavan, Sarina; Pan, Xiufang; Ran, F Ann; Yan, Winston X; Asokan, Aravind; Zhang, Feng; Duan, Dongsheng; Gersbach, Charles A

    2016-01-22

    Duchenne muscular dystrophy (DMD) is a devastating disease affecting about 1 out of 5000 male births and caused by mutations in the dystrophin gene. Genome editing has the potential to restore expression of a modified dystrophin gene from the native locus to modulate disease progression. In this study, adeno-associated virus was used to deliver the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system to the mdx mouse model of DMD to remove the mutated exon 23 from the dystrophin gene. This includes local and systemic delivery to adult mice and systemic delivery to neonatal mice. Exon 23 deletion by CRISPR-Cas9 resulted in expression of the modified dystrophin gene, partial recovery of functional dystrophin protein in skeletal myofibers and cardiac muscle, improvement of muscle biochemistry, and significant enhancement of muscle force. This work establishes CRISPR-Cas9-based genome editing as a potential therapy to treat DMD.

  16. The mouse formin (Fmn) gene: Genomic structure, novel exons, and genetic mapping

    SciTech Connect

    Wang, C.C.; Chan, D.C.; Leder, P.

    1997-02-01

    Mutations in the mouse formin (Fmn) gene, formerly known as the limb deformity (ld) gene, give rise to recessively inherited limb deformities and renal malformations or aplasia. The Fmn gene encodes many differentially processed transcripts that are expressed in both adult and embryonic tissues. To study the genomic organization of the Fmn locus, we have used Fmn probes to isolate and characterize genomic clones spanning 500 kb. Our analysis of these clones shows that the Fmn gene is composed of at least 24 exons and spans 400 kb. We have identified two novel exons that are expressed in the developing embryonic limb bud as well as adult tissues such as brain and kidney. We have also used a microsatellite polymorphism from within the Fmn gene to map it genetically to a 2.2-cM interval between D2Mit58 and D2Mit103. 36 refs., 6 figs., 1 tab.

  17. Cre Fused with RVG Peptide Mediates Targeted Genome Editing in Mouse Brain Cells In Vivo

    PubMed Central

    Zou, Zhiyuan; Sun, Zhaolin; Li, Pan; Feng, Tao; Wu, Sen

    2016-01-01

    Cell penetrating peptides (CPPs) are short peptides that can pass through cell membranes. CPPs can facilitate the cellular entry of proteins, macromolecules, nanoparticles and drugs. RVG peptide (RVG hereinafter) is a 29-amino-acid CPP derived from a rabies virus glycoprotein that can cross the blood-brain barrier (BBB) and enter brain cells. However, whether RVG can be used for genome editing in the brain has not been reported. In this work, we combined RVG with Cre recombinase for bacterial expression. The purified RVG-Cre protein cut plasmids in vitro and traversed cell membranes in cultured Neuro2a cells. By tail vein-injecting RVG-Cre into Cre reporter mouse lines mTmG and Rosa26lacZ, we demonstrated that RVG-Cre could target brain cells and achieve targeted somatic genome editing in adult mice. This direct delivery of the gene-editing enzyme protein into mouse brains with RVG is much safer than plasmid- or viral-based methods, holding promise for further applications in the treatment of various brain diseases. PMID:27983648

  18. Comparative genomics of the human, macaque and mouse major histocompatibility complex.

    PubMed

    Shiina, Takashi; Blancher, Antoine; Inoko, Hidetoshi; Kulski, Jerzy K

    2017-02-01

    The MHC is a highly polymorphic genomic region that encodes the transplantation and immune regulatory molecules. It receives special attention for genetic investigation because of its important role in the regulation of innate and adaptive immune responses and its strong association with numerous infectious and/or autoimmune diseases. The MHC locus was first discovered in the mouse and for the past 50 years it has been studied most intensively in both mice and humans. However, in recent years the macaque species have emerged as some of the more important and advanced experimental animal models for biomedical research into MHC with important human immunodeficiency virus/simian immunodeficiency virus and transplantation studies undertaken in association with precise MHC genotyping and haplotyping methods using Sanger sequencing and next-generation sequencing. Here, in this special issue on 'Macaque Immunology' we provide a short review of the genomic similarities and differences among the human, macaque and mouse MHC class I and class II regions, with an emphasis on the association of the macaque class I region with MHC polymorphism, haplotype structure and function.

  19. Molecular cloning, genomic organization and cell‐binding characteristics of mouse Spα

    PubMed Central

    Gebe, J A; Llewellyn, M‐B C; Hoggatt, H; Aruffo, A

    2000-01-01

    Several group B scavenger receptor cysteine‐rich (SRCR) proteins have been shown to function as modulators in the immune response. Recently, we reported the cloning of a new member of this family, human Spα (hSpα). Herein we report the cloning and characterization of the mouse homologue of hSpα. Like its human counterpart, mouse Spα (mSpα), is a secreted protein containing three SRCR domains. Most lymphoid tissues express RNA transcripts encoding mSpα. Characterization of a genomic clone encoding the mature mSpα protein showed that each of the SRCR domains of mSpα is encoded by a single exon. Comparison of the sequence of mSPα with those of other published proteins indicates that it is the same as the recently reported protein named AIM (apoptosis inhibitor expressed by macrophages). Cell‐binding studies with a mSpα immunoglobulin (mSpα‐Rγ) fusion protein indicated that mSpα is capable of binding to spleen‐derived CD19+ B cells and minimally to peritoneal cavity‐derived CD19+ B cells but not to peripheral blood‐derived B cells. Spleen‐derived CD3+ T cells also bound mSpα‐Rγ; however, no binding was observed to either peripheral blood mononuclear cells or peritoneal cavity‐derived CD3+ T cells. The mSpα‐Rγ fusion protein was also shown to bind to the mouse cell lines WEHI3 (monocytic) and EL‐4 (thymoma, T cell). The cloning of cDNA and genomic clones encoding mSpα and the identification of cells expressing a putative mSpα receptor(s) should facilitate in vivo studies designed to investigate the function of Spα in the immune compartment. PMID:10651944

  20. Molecular cloning, genomic organization and cell-binding characteristics of mouse Spalpha.

    PubMed

    Gebe, J A; Llewellyn, M; Hoggatt, H; Aruffo, A

    2000-01-01

    Several group B scavenger receptor cysteine-rich (SRCR) proteins have been shown to function as modulators in the immune response. Recently, we reported the cloning of a new member of this family, human Spalpha (hSpalpha). Herein we report the cloning and characterization of the mouse homologue of hSpalpha. Like its human counterpart, mouse Spalpha (mSpalpha), is a secreted protein containing three SRCR domains. Most lymphoid tissues express RNA transcripts encoding mSpalpha. Characterization of a genomic clone encoding the mature mSpalpha protein showed that each of the SRCR domains of mSpalpha is encoded by a single exon. Comparison of the sequence of mSPalpha with those of other published proteins indicates that it is the same as the recently reported protein named AIM (apoptosis inhibitor expressed by macrophages). Cell-binding studies with a mSpalpha immunoglobulin (mSpalpha-Rgamma) fusion protein indicated that mSpalpha is capable of binding to spleen-derived CD19+ B cells and minimally to peritoneal cavity-derived CD19+ B cells but not to peripheral blood-derived B cells. Spleen-derived CD3+ T cells also bound mSpalpha-Rgamma; however, no binding was observed to either peripheral blood mononuclear cells or peritoneal cavity-derived CD3+ T cells. The mSpalpha-Rgamma fusion protein was also shown to bind to the mouse cell lines WEHI3 (monocytic) and EL-4 (thymoma, T cell). The cloning of cDNA and genomic clones encoding mSpalpha and the identification of cells expressing a putative mSpalpha receptor(s) should facilitate in vivo studies designed to investigate the function of Spalpha in the immune compartment.

  1. Sequences homologous to retrovirus-like genes of the mouse are present in multiple copies in the Syrian hamster genome.

    PubMed Central

    Lueders, K K; Kuff, E L

    1981-01-01

    The genome of M. musculus contains many copies of DNA sequences homologous to the 35S RNA of intracisternal type-A particles (IAP) (1,2). A major class of IAP genes has been identified and isolated from a mouse library in Charon 4A (3). Cloned mouse IAP genes were used as probes to study homologous sequences in the DNA of other species. Sequences related to mouse IAP genes were detected in the DNAs from a variety of animal cells. DNAs from rat, gerbil, and hamster cells all gave strong reactions which could be localized to discrete restriction fragments on genomic blots. The reaction of Syrian hamster DNA was particularly strong. Fragments derived from different parts of the IAP gene all reacted with Syrian hamster DNA, and the reactive restriction fragments in the Syrian hamster DNA could be ordered with reference to the known restriction map of the IAP genes. The data suggest that sequences related to mouse IAP genes make up a 7 Kb unit in the Syrian hamster genome. Since the majority of the hamster sequences are quite divergent from those in the mouse, the ease with which they are detected suggests that they must be reiterated in the hamster genome. Images PMID:7198224

  2. Protein-coding potential of mouse mammary tumor virus genome RNA as examined by in vitro translation.

    PubMed Central

    Dickson, C; Peters, G

    1981-01-01

    The protein-coding capacity of the mouse mammary tumor virus genome has been examined by in vitro translation of genome length and polyadenylated subgenomic fragments of viral RNA. Intact genome RNA of about 35S programmed synthesis of the Pr77gag, Pr110gag and Pr160gag/pol precursors seen in infected cells in vivo. Polyadenylated RNA fragments of 18 to 28S encoded products whose tryptic peptide maps resembled those of the nonglycosylated precursor to the envelope glycoproteins, confirming the gene order 5'-gag-pol-env-3'. Translation of polyadenylated RNA fragments smaller than 18S yielded a series of related proteins whose peptide maps bore no resemblance to any of the virion structural proteins. Thus, a region of the mouse mammary tumor virus genome distal to the env gene appears to have an open reading frame sufficient to encode at least 36,000 daltons of protein as of yet unknown function. Images PMID:6260988

  3. Co-incident insertion enables high efficiency genome engineering in mouse embryonic stem cells

    PubMed Central

    Shy, Brian R.; MacDougall, Matthew S.; Clarke, Ryan; Merrill, Bradley J.

    2016-01-01

    CRISPR/Cas9 nucleases have enabled powerful, new genome editing capabilities; however, the preponderance of non-homologous end joining (NHEJ) mediated repair events over homology directed repair (HDR) in most cell types limits the ability to engineer precise changes in mammalian genomes. Here, we increase the efficiency of isolating precise HDR-mediated events in mouse embryonic stem (ES) cells by more than 20-fold through the use of co-incidental insertion (COIN) of independent donor DNA sequences. Analysis of on:off-target frequencies at the Lef1 gene revealed that bi-allelic insertion of a PGK-Neo cassette occurred more frequently than expected. Using various selection cassettes targeting multiple loci, we show that the insertion of a selectable marker at one control site frequently coincided with an insertion at an unlinked, independently targeted site, suggesting enrichment of a sub-population of HDR-proficient cells. When individual cell events were tracked using flow cytometry and fluorescent protein markers, individual cells frequently performed either a homology-dependent insertion event or a homology-independent event, but rarely both types of insertions in a single cell. Thus, when HDR-dependent selection donors are used, COIN enriches for HDR-proficient cells among heterogeneous cell populations. When combined with a self-excising selection cassette, COIN provides highly efficient and scarless genome editing. PMID:27484482

  4. Asymmetric parental genome engineering by Cas9 during mouse meiotic exit

    PubMed Central

    Suzuki, Toru; Asami, Maki; Perry, Anthony C. F.

    2014-01-01

    Mammalian genomes can be edited by injecting pronuclear embryos with Cas9 cRNA and guide RNA (gRNA) but it is unknown whether editing can also occur during the onset of embryonic development, prior to pronuclear embryogenesis. We here report Cas9-mediated editing during sperm-induced meiotic exit and the initiation of development. Injection of unfertilized, mouse metaphase II (mII) oocytes with Cas9 cRNA, gRNA and sperm enabled efficient editing of transgenic and native alleles. Pre-loading oocytes with Cas9 increased sensitivity to gRNA ~100-fold. Paternal allelic editing occurred as an early event: single embryo genome analysis revealed editing within 3 h of sperm injection, coinciding with sperm chromatin decondensation during the gamete-to-embryo transition but prior to pronucleus formation. Maternal alleles underwent editing after the first round of DNA replication, resulting in mosaicism. Asymmetric editing of maternal and paternal alleles suggests a novel strategy for discriminatory targeting of parental genomes. PMID:25532495

  5. Genome Sequence of “Candidatus Arthromitus” sp. Strain SFB-Mouse-NL, a Commensal Bacterium with a Key Role in Postnatal Maturation of Gut Immune Functions

    PubMed Central

    Bolotin, Alexander; de Wouters, Tomas; Schnupf, Pamela; Bouchier, Christiane; Loux, Valentin; Rhimi, Moez; Jamet, Alexandre; Dervyn, Rozenn; Boudebbouze, Samira; Blottière, Hervé M.; Sorokin, Alexei; Snel, Johannes; Cerf-Bensussan, Nadine; Gaboriau-Routhiau, Valérie; van de Guchte, Maarten

    2014-01-01

    “Candidatus Arthromitus” sp. strain SFB-mouse-NL (SFB, segmented filamentous bacteria) is a commensal bacterium necessary for inducing the postnatal maturation of homeostatic innate and adaptive immune responses in the mouse gut. Here, we report the genome sequence of this bacterium, which sets it apart from earlier sequenced mouse SFB isolates. PMID:25035333

  6. Epigenetic Basis of Regeneration: Analysis of Genomic DNA Methylation Profiles in the MRL/MpJ Mouse

    PubMed Central

    Górnikiewicz, Bartosz; Ronowicz, Anna; Podolak, Justyna; Madanecki, Piotr; Stanisławska-Sachadyn, Anna; Sachadyn, PaweŁ

    2013-01-01

    Epigenetic regulation plays essential role in cell differentiation and dedifferentiation, which are the intrinsic processes involved in regeneration. To investigate the epigenetic basis of regeneration capacity, we choose DNA methylation as one of the most important epigenetic mechanisms and the MRL/MpJ mouse as a model of mammalian regeneration known to exhibit enhanced regeneration response in different organs. We report the comparative analysis of genomic DNA methylation profiles of the MRL/MpJ and the control C57BL/6J mouse. Methylated DNA immunoprecipitation followed by microarray analysis using the Nimblegen ‘3 × 720 K CpG Island Plus RefSeq Promoter’ platform was applied in order to carry out genome-wide DNA methylation profiling covering 20 404 promoter regions. We identified hundreds of hypo- and hypermethylated genes and CpG islands in the heart, liver, and spleen, and 37 of them in the three tissues. Decreased inter-tissue diversification and the shift of DNA methylation balance upstream the genes distinguish the genomic methylation patterns of the MRL/MpJ mouse from the C57BL/6J. Homeobox genes and a number of other genes involved in embryonic morphogenesis are significantly overrepresented among the genes hypomethylated in the MRL/MpJ mouse. These findings indicate that epigenetic patterning might be a likely molecular basis of regeneration capability in the MRL/MpJ mouse. PMID:23929942

  7. Complete Genome Sequence of Turicibacter sp. Strain H121, Isolated from the Feces of a Contaminated Germ-Free Mouse

    PubMed Central

    Auchtung, T. A.; Holder, M. E.; Gesell, J. R.; Ajami, N. J.; Duarte, R. T. D.; Itoh, K.; Caspi, R. R.; Petrosino, J. F.; Horai, R.

    2016-01-01

    Turicibacter bacteria are commonly detected in the gastrointestinal tracts and feces of humans and animals, but their phylogeny, ecological role, and pathogenic potential remain unclear. We present here the first complete genome sequence of Turicibacter sp. strain H121, which was isolated from the feces of a mouse line contaminated following germ-free derivation. PMID:27013036

  8. Population genomics of the Anthropocene: urbanization is negatively associated with genome-wide variation in white-footed mouse populations.

    PubMed

    Munshi-South, Jason; Zolnik, Christine P; Harris, Stephen E

    2016-04-01

    Urbanization results in pervasive habitat fragmentation and reduces standing genetic variation through bottlenecks and drift. Loss of genomewide variation may ultimately reduce the evolutionary potential of animal populations experiencing rapidly changing conditions. In this study, we examined genomewide variation among 23 white-footed mouse (Peromyscus leucopus) populations sampled along an urbanization gradient in the New York City metropolitan area. Genomewide variation was estimated as a proxy for evolutionary potential using more than 10 000 single nucleotide polymorphism (SNP) markers generated by ddRAD-Seq. We found that genomewide variation is inversely related to urbanization as measured by percent impervious surface cover, and to a lesser extent, human population density. We also report that urbanization results in enhanced genomewide differentiation between populations in cities. There was no pattern of isolation by distance among these populations, but an isolation by resistance model based on impervious surface significantly explained patterns of genetic differentiation. Isolation by environment modeling also indicated that urban populations deviate much more strongly from global allele frequencies than suburban or rural populations. This study is the first to examine loss of genomewide SNP variation along an urban-to-rural gradient and quantify urbanization as a driver of population genomic patterns.

  9. Integration of the Rat Recombination and EST Maps in the Rat Genomic Sequence and Comparative Mapping Analysis With the Mouse Genome

    PubMed Central

    Wilder, Steven P.; Bihoreau, Marie-Thérèse; Argoud, Karène; Watanabe, Takeshi K.; Lathrop, Mark; Gauguier, Dominique

    2004-01-01

    Inbred strains of the laboratory rat are widely used for identifying genetic regions involved in the control of complex quantitative phenotypes of biomedical importance. The draft genomic sequence of the rat now provides essential information for annotating rat quantitative trait locus (QTL) maps. Following the survey of unique rat microsatellite (11,585 including 1648 new markers) and EST (10,067) markers currently available, we have incorporated a selection of 7952 rat EST sequences in an improved version of the integrated linkage-radiation hybrid map of the rat containing 2058 microsatellite markers which provided over 10,000 potential anchor points between rat QTL and the genomic sequence of the rat. A total of 996 genetic positions were resolved (avg. spacing 1.77 cM) in a single large intercross and anchored in the rat genomic sequence (avg. spacing 1.62 Mb). Comparative genome maps between rat and mouse were constructed by successful computational alignment of 6108 mapped rat ESTs in the mouse genome. The integration of rat linkage maps in the draft genomic sequence of the rat and that of other species represents an essential step for translating rat QTL intervals into human chromosomal targets. PMID:15060020

  10. A genome-scale map of expression for a mouse brain section obtained using voxelation

    PubMed Central

    Chin, Mark H.; Geng, Alex B.; Khan, Arshad H.; Qian, Wei-Jun; Petyuk, Vladislav A.; Boline, Jyl; Levy, Shawn; Toga, Arthur W.; Smith, Richard D.; Leahy, Richard M.; Smith, Desmond J.

    2011-01-01

    Gene expression signatures in the mammalian brain hold the key to understanding neural development and neurological diseases. We have reconstructed two-dimensional images of gene expression for 20,000 genes in a coronal slice of the mouse brain at the level of the striatum by using microarrays in combination with voxelation at a resolution of 1 mm3. Good reliability of the microarray results were confirmed using multiple replicates, subsequent quantitative RT-PCR voxelation, mass spectrometry voxelation, and publicly available in situ hybridization data. Known and novel genes were identified with expression patterns localized to defined substructures within the brain. In addition, genes with unexpected patterns were identified, and cluster analysis identified a set of genes with a gradient of dorsal/ventral expression not restricted to known anatomical boundaries. The genome-scale maps of gene expression obtained using voxelation will be a valuable tool for the neuroscience community. PMID:17504947

  11. A genome-scale map of expression for a mouse brain section obtained using voxelation

    SciTech Connect

    Chin, Mark H.; Geng, Alex B.; Khan, Arshad H.; Qian, Weijun; Petyuk, Vladislav A.; Boline, Jyl; Levy, Shawn; Toga, Arthur W.; Smith, Richard D.; Leahy, Richard M.; Smith, Desmond J.

    2007-08-20

    Gene expression signatures in the mammalian brain hold the key to understanding neural development and neurological diseases. We have reconstructed 2- dimensional images of gene expression for 20,000 genes in a coronal slice of the mouse brain at the level of the striatum by using microarrays in combination with voxelation at a resolution of 1 mm3. Good reliability of the microarray results were confirmed using multiple replicates, subsequent quantitative RT-PCR voxelation, mass spectrometry voxelation and publicly available in situ hybridization data. Known and novel genes were identified with expression patterns localized to defined substructures within the brain. In addition, genes with unexpected patterns were identified and cluster analysis identified a set of genes with a gradient of dorsal/ventral expression not restricted to known anatomical boundaries. The genome-scale maps of gene expression obtained using voxelation will be a valuable tool for the neuroscience community.

  12. Human diseases versus mouse models: insights into the regulation of genomic imprinting at the human 11p15/mouse distal chromosome 7 region.

    PubMed

    Shmela, Mansur Ennuri; Gicquel, Christine F

    2013-01-01

    The 11p15 region is organised into two independent imprinted domains controlled by imprinting control regions, which carry opposite germline imprints. Dysregulation of 11p15 genomic imprinting results in two human fetal growth disorders (Silver-Russell syndrome (SRS, MIM 180860) and Beckwith-Wiedemann syndrome (BWS, MIM 130650)) with opposite growth phenotypes. The mouse orthologous region on distal chromosome 7 (dist7) is well conserved in its organisation and its regulation. Targeted mutagenesis in mice has provided highly valuable clues in terms of the mechanisms involved in the regulation of genomic imprinting of the 11p15/dist7 imprinted region. On the other hand, the recent identification of unexpected genetic defects in BWS and SRS patients also brought new insights into the mechanisms of 11p15 imprinting regulation. However, some mouse models and human genetic defects show contradictions in term of growth phenotypes and parental transmission. In this review, we extensively analyse those various mouse and human models and more particularly models with mutations affecting the two imprinting centres, in order to improve our understanding of regulation of 11p15/dist7 genomic imprinting.

  13. Comparative analysis of dioxin response elements in human, mouse and rat genomic sequences

    PubMed Central

    Sun, Y. V.; Boverhof, D. R.; Burgoon, L. D.; Fielden, M. R.; Zacharewski, T. R.

    2004-01-01

    Comparative approaches were used to identify human, mouse and rat dioxin response elements (DREs) in genomic sequences unambiguously assigned to a nucleotide RefSeq accession number. A total of 13 bona fide DREs, all including the substitution intolerant core sequence (GCGTG) and adjacent variable sequences, were used to establish a position weight matrix and a matrix similarity (MS) score threshold to rank identified DREs. DREs with MS scores above the threshold were disproportionately distributed in close proximity to the transcription start site in all three species. Gene expression assays in hepatic mouse tissue confirmed the responsiveness of 192 genes possessing a putative DRE. Previously identified functional DREs in well-characterized AhR-regulated genes including Cyp1a1 and Cyp1b1 were corroborated. Putative DREs were identified in 48 out of 2437 human–mouse–rat orthologous genes between −1500 and the transcriptional start site, of which 19 of these genes possessed positionally conserved DREs as determined by multiple sequence alignment. Seven of these nineteen genes exhibited 2,3,7,8-tetrachlorodibenzo-p-dioxin-mediated regulation, although there were significant discrepancies between in vivo and in vitro results. Interestingly, of the mouse–rat orthologous genes with a DRE between −1500 and +1500, only 37% had an equivalent human ortholog. These results suggest that AhR-mediated gene expression may not be well conserved across species, which could have significant implications in human risk assessment. PMID:15328365

  14. New mouse models for metabolic bone diseases generated by genome-wide ENU mutagenesis.

    PubMed

    Sabrautzki, Sibylle; Rubio-Aliaga, Isabel; Hans, Wolfgang; Fuchs, Helmut; Rathkolb, Birgit; Calzada-Wack, Julia; Cohrs, Christian M; Klaften, Matthias; Seedorf, Hartwig; Eck, Sebastian; Benet-Pagès, Ana; Favor, Jack; Esposito, Irene; Strom, Tim M; Wolf, Eckhard; Lorenz-Depiereux, Bettina; Hrabě de Angelis, Martin

    2012-08-01

    Metabolic bone disorders arise as primary diseases or may be secondary due to a multitude of organ malfunctions. Animal models are required to understand the molecular mechanisms responsible for the imbalances of bone metabolism in disturbed bone mineralization diseases. Here we present the isolation of mutant mouse models for metabolic bone diseases by phenotyping blood parameters that target bone turnover within the large-scale genome-wide Munich ENU Mutagenesis Project. A screening panel of three clinical parameters, also commonly used as biochemical markers in patients with metabolic bone diseases, was chosen. Total alkaline phosphatase activity and total calcium and inorganic phosphate levels in plasma samples of F1 offspring produced from ENU-mutagenized C3HeB/FeJ male mice were measured. Screening of 9,540 mice led to the identification of 257 phenodeviants of which 190 were tested by genetic confirmation crosses. Seventy-one new dominant mutant lines showing alterations of at least one of the biochemical parameters of interest were confirmed. Fifteen mutations among three genes (Phex, Casr, and Alpl) have been identified by positional-candidate gene approaches and one mutation of the Asgr1 gene, which was identified by next-generation sequencing. All new mutant mouse lines are offered as a resource for the scientific community.

  15. Data for identification of porcine X-chromosome inactivation center, XIC, by genomic comparison with human and mouse XIC

    PubMed Central

    Hwang, Jae Yeon; Choi, Kwang-Hwan; Lee, Chang-Kyu

    2015-01-01

    The data included in this article shows homologies of genes in porcine X-chromosome inactivation center, XIC, to each orthologue in human and mouse XIC. Open sequences of XIC-linked genes in human and mouse were compared to porcine genome and sequence homology of each orthologue to porcine genome was calculated. Sequence information of porcine genes encoded in the genomic regions having sequence homology with the human XIC-linked genes and their 2 Kb upstream regions were downloaded. Obtained information was used to design primer pairs for expression and methylation pattern analyses of XIC-linked genes in pigs. The data represented in here is related and applied to the research article entitled “Dosage compensation of X-chromosome inactivation center, XIC,-linked genes in porcine preimplantation embryos: Non-chromosome wide initiation of X-chromosome inactivation in blastocysts”, published in Mechanisms of Development Hwang et al., 2015 [1]. PMID:26793753

  16. Data for identification of porcine X-chromosome inactivation center, XIC, by genomic comparison with human and mouse XIC.

    PubMed

    Hwang, Jae Yeon; Choi, Kwang-Hwan; Lee, Chang-Kyu

    2015-12-01

    The data included in this article shows homologies of genes in porcine X-chromosome inactivation center, XIC, to each orthologue in human and mouse XIC. Open sequences of XIC-linked genes in human and mouse were compared to porcine genome and sequence homology of each orthologue to porcine genome was calculated. Sequence information of porcine genes encoded in the genomic regions having sequence homology with the human XIC-linked genes and their 2 Kb upstream regions were downloaded. Obtained information was used to design primer pairs for expression and methylation pattern analyses of XIC-linked genes in pigs. The data represented in here is related and applied to the research article entitled "Dosage compensation of X-chromosome inactivation center, XIC,-linked genes in porcine preimplantation embryos: Non-chromosome wide initiation of X-chromosome inactivation in blastocysts", published in Mechanisms of Development Hwang et al., 2015 [1].

  17. Generation of mouse models of myeloid malignancy with combinatorial genetic lesions using CRISPR-Cas9 genome editing.

    PubMed

    Heckl, Dirk; Kowalczyk, Monika S; Yudovich, David; Belizaire, Roger; Puram, Rishi V; McConkey, Marie E; Thielke, Anne; Aster, Jon C; Regev, Aviv; Ebert, Benjamin L

    2014-09-01

    Genome sequencing studies have shown that human malignancies often bear mutations in four or more driver genes, but it is difficult to recapitulate this degree of genetic complexity in mouse models using conventional breeding. Here we use the CRISPR-Cas9 system of genome editing to overcome this limitation. By delivering combinations of small guide RNAs (sgRNAs) and Cas9 with a lentiviral vector, we modified up to five genes in a single mouse hematopoietic stem cell (HSC), leading to clonal outgrowth and myeloid malignancy. We thereby generated models of acute myeloid leukemia (AML) with cooperating mutations in genes encoding epigenetic modifiers, transcription factors and mediators of cytokine signaling, recapitulating the combinations of mutations observed in patients. Our results suggest that lentivirus-delivered sgRNA:Cas9 genome editing should be useful to engineer a broad array of in vivo cancer models that better reflect the complexity of human disease.

  18. Generation of mouse models of myeloid malignancy with combinatorial genetic lesions using CRISPR-Cas9 genome editing

    PubMed Central

    Heckl, Dirk; Kowalczyk, Monika S.; Yudovich, David; Belizaire, Roger; Puram, Rishi V.; McConkey, Marie E.; Thielke, Anne; Aster, Jon C.; Regev, Aviv; Ebert, Benjamin L.

    2014-01-01

    Genome sequencing studies have shown that human malignancies often bear mutations in four or more driver genes1, but it is difficult to recapitulate this degree of genetic complexity in mouse models using conventional breeding. Here we use the CRISPR-Cas9 system of genome editing2–4 to overcome this limitation. By delivering combinations of small guide RNAs (sgRNAs) and Cas9 with a lentiviral vector, we modified up to five genes in a single mouse hematopoietic stem cell (HSC), leading to clonal outgrowth and myeloid malignancy. We thereby generated models of acute myeloid leukemia (AML) with cooperating mutations in genes encoding epigenetic modifiers, transcription factors, and mediators of cytokine signaling, recapitulating the combinations of mutations observed in the human disease. Our results suggest that lentivirus-delivered sgRNA:Cas9 genome editing should be useful to engineer a broad array of in vivo cancer models that better reflect the complexity of human disease. PMID:24952903

  19. The mouse Fau gene: genomic structure, chromosomal localization, and characterization of two retropseudogenes.

    PubMed

    Casteels, D; Poirier, C; Guénet, J L; Merregaert, J

    1995-01-01

    The Fau gene is the cellular homolog of the fox sequence of the Finkel-Biskis-Reilly murine sarcoma virus (FBR-MuSV). FBR-MuSV acquired the Fau gene by transduction in a transcriptional orientation opposite to that of the genomic Fau gene. The genomic structure of the mouse Fau gene (MMFAU) and its upstream elements have been determined and are similar to those of the human FAU gene. The gene consists of five exons and is located on chromosome 19. The first exon is not translated. The promoter region has no well-defined TATA box but contains the polypyrimidine initiator flanked by regions of high GC content (65%) and shows all of the characteristics of a housekeeping gene. The 5' end of the mRNA transcript was determined by 5' RACE analysis and is located, as expected, in the polypyrimidine initiator site. Furthermore, the sequences of two retropseudogenes (Fau-ps1 and Fau-ps2) are reported. Both pseudogenes are approximately 75% identical to the Fau cDNA, but both are shorter due to a deletion at the 5' end and do not encode a functional protein. Fau-prs is interrupted by an AG-rich region of about 350 bp within the S30 region of the Fau cDNA. Fau-ps1 was localized on chromosome 1 and Fau-ps2 on chromosome 7.

  20. Genome-wide assembly and analysis of alternative transcripts in mouse

    PubMed Central

    Sharov, Alexei A.; Dudekula, Dawood B.; Ko, Minoru S.H.

    2005-01-01

    To build a mouse gene index with the most comprehensive coverage of alternative transcription/splicing (ATS), we developed an algorithm and a fully automated computational pipeline for transcript assembly from expressed sequences aligned to the genome. We identified 191,946 genomic loci, which included 27,497 protein-coding genes and 11,906 additional gene candidates (e.g., nonprotein-coding, but multiexon). Comparison of the resulting gene index with TIGR, UniGene, DoTS, and ESTGenes databases revealed that it had a greater number of transcripts, a greater average number of exons and introns with proper splicing sites per gene, and longer ORFs. The 27,497 protein-coding genes had 77,138 transcripts, i.e., 2.8 transcripts per gene on average. Close examination of transcripts led to a combinatorial table of 23 types of ATS units, only nine of which were previously described, i.e., 14 types of alternative splicing, seven types of alternative starts, and two types of alternative termination. The 47%, 18%, and 14% of 20,323 multiexon protein-coding genes with proper splice sites had alternative splicings, alternative starts, and alternative terminations, respectively. The gene index with the comprehensive ATS will provide a useful platform for analyzing the nature and mechanism of ATS, as well as for designing the accurate exon-based DNA microarrays. PMID:15867436

  1. Genome Editing in Mouse Spermatogonial Stem/Progenitor Cells Using Engineered Nucleases

    PubMed Central

    Fanslow, Danielle A.; Wirt, Stacey E.; Barker, Jenny C.; Connelly, Jon P.; Porteus, Matthew H.; Dann, Christina Tenenhaus

    2014-01-01

    Editing the genome to create specific sequence modifications is a powerful way to study gene function and promises future applicability to gene therapy. Creation of precise modifications requires homologous recombination, a very rare event in most cell types that can be stimulated by introducing a double strand break near the target sequence. One method to create a double strand break in a particular sequence is with a custom designed nuclease. We used engineered nucleases to stimulate homologous recombination to correct a mutant gene in mouse “GS” (germline stem) cells, testicular derived cell cultures containing spermatogonial stem cells and progenitor cells. We demonstrated that gene-corrected cells maintained several properties of spermatogonial stem/progenitor cells including the ability to colonize following testicular transplantation. This proof of concept for genome editing in GS cells impacts both cell therapy and basic research given the potential for GS cells to be propagated in vitro, contribute to the germline in vivo following testicular transplantation or become reprogrammed to pluripotency in vitro. PMID:25409432

  2. Genome-wide disruption of 5-hydroxymethylcytosine in a mouse model of autism

    PubMed Central

    Papale, Ligia A.; Zhang, Qi; Li, Sisi; Chen, Kailei; Keleş, Sündüz; Alisch, Reid S.

    2015-01-01

    The autism spectrum disorders (ASD) comprise a broad group of behaviorally related neurodevelopmental disorders affecting as many as 1 in 68 children. The hallmarks of ASD consist of impaired social and communication interactions, pronounced repetitive behaviors and restricted patterns of interests. Family, twin and epidemiological studies suggest a polygenetic and epistatic susceptibility model involving the interaction of many genes; however, the etiology of ASD is likely to be complex and include both epigenetic and environmental factors. 5-hydroxymethylcytosine (5hmC) is a novel environmentally sensitive DNA modification that is highly enriched in post-mitotic neurons and is associated with active transcription of neuronal genes. Here, we used an established chemical labeling and affinity purification method coupled with high-throughput sequencing technology to generate a genome-wide profile of striatal 5hmC in an autism mouse model (Cntnap2−/− mice) and found that at 9 weeks of age the Cntnap2−/− mice have a genome-wide disruption in 5hmC, primarily in genic regions and repetitive elements. Annotation of differentially hydroxymethylated regions (DhMRs) to genes revealed a significant overlap with known ASD genes (e.g. Nrxn1 and Reln) that carried an enrichment of neuronal ontological functions, including axonogenesis and neuron projection morphogenesis. Finally, sequence motif predictions identified associations with transcription factors that have a high correlation with important genes in neuronal developmental and functional pathways. Together, our data implicate a role for 5hmC-mediated epigenetic modulation in the pathogenesis of autism and represent a critical step toward understanding the genome-wide molecular consequence of the Cntnap2 mutation, which results in an autism-like phenotype. PMID:26423458

  3. TEX11 is mutated in infertile men with azoospermia and regulates genome-wide recombination rates in mouse

    PubMed Central

    Yang, Fang; Silber, Sherman; Leu, N Adrian; Oates, Robert D; Marszalek, Janet D; Skaletsky, Helen; Brown, Laura G; Rozen, Steve; Page, David C; Wang, P Jeremy

    2015-01-01

    Genome-wide recombination is essential for genome stability, evolution, and speciation. Mouse Tex11, an X-linked meiosis-specific gene, promotes meiotic recombination and chromosomal synapsis. Here, we report that TEX11 is mutated in infertile men with non-obstructive azoospermia and that an analogous mutation in the mouse impairs meiosis. Genetic screening of a large cohort of idiopathic infertile men reveals that TEX11 mutations, including frameshift and splicing acceptor site mutations, cause infertility in 1% of azoospermic men. Functional evaluation of three analogous human TEX11 missense mutations in transgenic mouse models identified one mutation (V748A) as a potential infertility allele and found two mutations non-causative. In the mouse model, an intronless autosomal Tex11 transgene functionally substitutes for the X-linked Tex11 gene, providing genetic evidence for the X-to-autosomal retrotransposition evolution phenomenon. Furthermore, we find that TEX11 protein levels modulate genome-wide recombination rates in both sexes. These studies indicate that TEX11 alleles affecting expression level or substituting single amino acids may contribute to variations in recombination rates between sexes and among individuals in humans. PMID:26136358

  4. A genome-wide survey on basic helix-loop-helix transcription factors in rat and mouse.

    PubMed

    Zheng, Xiaodong; Zheng, X; Wang, Yong; Wang, Y; Yao, Qin; Yao, Q; Yang, Zhe; Yang, Z; Chen, Keping; Chen, K

    2009-04-01

    The basic helix-loop-helix (bHLH) proteins play essential roles in a wide range of developmental processes in higher organisms. bHLH family members have been identified in over 20 organisms, including nematode, fruit fly, and human. Our study identified 114 rat and 14 additional mouse bHLH members in rat and mouse genomes, respectively. Phylogenetic analyses revealed that both rat and mouse had 49, 26, 15, 4, 12, and 4 bHLH members in groups A, B, C, D, E, and F, respectively. Only the rat Mxi1 gene has two copies in the genome. All other rat bHLH genes and all mouse bHLH genes are single-copy genes. The chromosomal distribution pattern of mouse, rat, and human bHLH genes suggests the emergence of some bHLH genes through gene duplication, which probably happened at least before the divergence of vertebrates from invertebrates. The present study provides useful information for future studies using rat as a model animal for mammalian development.

  5. A comparative phenotypic and genomic analysis of C57BL/6J and C57BL/6N mouse strains

    PubMed Central

    2013-01-01

    Background The mouse inbred line C57BL/6J is widely used in mouse genetics and its genome has been incorporated into many genetic reference populations. More recently large initiatives such as the International Knockout Mouse Consortium (IKMC) are using the C57BL/6N mouse strain to generate null alleles for all mouse genes. Hence both strains are now widely used in mouse genetics studies. Here we perform a comprehensive genomic and phenotypic analysis of the two strains to identify differences that may influence their underlying genetic mechanisms. Results We undertake genome sequence comparisons of C57BL/6J and C57BL/6N to identify SNPs, indels and structural variants, with a focus on identifying all coding variants. We annotate 34 SNPs and 2 indels that distinguish C57BL/6J and C57BL/6N coding sequences, as well as 15 structural variants that overlap a gene. In parallel we assess the comparative phenotypes of the two inbred lines utilizing the EMPReSSslim phenotyping pipeline, a broad based assessment encompassing diverse biological systems. We perform additional secondary phenotyping assessments to explore other phenotype domains and to elaborate phenotype differences identified in the primary assessment. We uncover significant phenotypic differences between the two lines, replicated across multiple centers, in a number of physiological, biochemical and behavioral systems. Conclusions Comparison of C57BL/6J and C57BL/6N demonstrates a range of phenotypic differences that have the potential to impact upon penetrance and expressivity of mutational effects in these strains. Moreover, the sequence variants we identify provide a set of candidate genes for the phenotypic differences observed between the two strains. PMID:23902802

  6. Zooming in on the human-mouse comparative map: genome conservation re-examined on a high-resolution scale.

    PubMed

    Carver, E A; Stubbs, L

    1997-12-01

    Over the past decade, conservation of genetic linkage groups has been shown in mammals and used to great advantage, fueling significant exchanges of gene mapping and functional information especially between the genomes of humans and mice. As human physical maps increase in resolution from chromosome bands to nucleotide sequence, comparative alignments of mouse and human regions have revealed striking similarities and surprising differences between the genomes of these two best-mapped mammalian species. Whereas, at present, very few mouse and human regions have been compared on the physical level, existing studies provide intriguing insights to genome evolution, including the observation of recent duplications and deletions of genes that may play significant roles in defining some of the biological differences between the two species. Although high-resolution conserved marker-based maps are currently available only for human and mouse, a variety of new methods and resources are speeding the development of comparative maps of additional organisms. These advances mark the first step toward establishment of the human genome as a reference map for vertebrate species, providing evolutionary and functional annotation to human sequence and vast new resources for genetic analysis of a variety of commercially, medically, and ecologically important animal models.

  7. SFP Genotyping from Affymetrix Arrays is Robust but Largely Detects Cis-acting Expression Regulators

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The recent development of Affymetrix chips designed from assembled EST sequences has spawned considerable interest in identifying single-feature polymorphisms (SFPs) from transcriptome data. SFPs are valuable genetic markers that potentially offer a physical link to the structural genes themselves....

  8. Large Genomic Fragment Deletions and Insertions in Mouse Using CRISPR/Cas9

    PubMed Central

    Satheka, Achim Cchitvsanzwhoh; Togo, Jacques; An, Yao; Humphrey, Mabwi; Ban, Luying; Ji, Yan; Jin, Honghong; Feng, Xuechao; Zheng, Yaowu

    2015-01-01

    ZFN, TALENs and CRISPR/Cas9 system have been used to generate point mutations and large fragment deletions and insertions in genomic modifications. CRISPR/Cas9 system is the most flexible and fast developing technology that has been extensively used to make mutations in all kinds of organisms. However, the most mutations reported up to date are small insertions and deletions. In this report, CRISPR/Cas9 system was used to make large DNA fragment deletions and insertions, including entire Dip2a gene deletion, about 65kb in size, and β-galactosidase (lacZ) reporter gene insertion of larger than 5kb in mouse. About 11.8% (11/93) are positive for 65kb deletion from transfected and diluted ES clones. High targeting efficiencies in ES cells were also achieved with G418 selection, 46.2% (12/26) and 73.1% (19/26) for left and right arms respectively. Targeted large fragment deletion efficiency is about 21.4% of live pups or 6.0% of injected embryos. Targeted insertion of lacZ reporter with NEO cassette showed 27.1% (13/48) of targeting rate by ES cell transfection and 11.1% (2/18) by direct zygote injection. The procedures have bypassed in vitro transcription by directly co-injection of zygotes or co-transfection of embryonic stem cells with circular plasmid DNA. The methods are technically easy, time saving, and cost effective in generating mouse models and will certainly facilitate gene function studies. PMID:25803037

  9. Functional and structural analyses of mouse genomic regions screened by the morphological specific-locus test.

    PubMed

    Russell, L B

    1989-05-01

    Genetic analyses of certain classes of mutations recovered in the mouse specific-locus test (SLT) have characterized arrays of deletions, overlapping at the marked loci. Complementation maps, generated for several of the regions, have identified a number of functional units surrounding each marked locus and have ordered the mutations into complementation groups. Molecular entry to all but one of the marked regions has been achieved by (1) identifying proviral integrations in, or close to, the specific loci (d, se, a, c); (2) mapping random anonymous clones from appropriately enriched libraries to the longest deleted segments, then submapping to more limited segments on the basis of complementation and deletion-breakpoint maps (c, p); (3) similarly mapping known clones thought to be located in pertinent chromosomal regions (p, c, d); and (4) cloning specific genes that reside in regions corresponding to the deletions (b, c, p). The molecular analyses have confirmed that genetically-inferred deletions are structural deletions of DNA. The emerging physical maps are concordant with the complementation maps, and in several cases have discriminated among members of a complementation group with respect to breakpoint positions. Deletion-breakpoint-fusion fragments have prove to be highly useful for making large chromosomal jumps to facilitate physical mapping. The recent advances toward correlating physical and functional maps of specific regions of the mouse genome owe much to the existence of arrays of mutations involving loci marked in the SLT. In turn, the characterizations of these regions have made it possible to demonstrate qualitative differences among mutations resulting from different treatments. This new capability for qualitative analysis, which will increase as the molecular studies proceed, further enhances the value of the SLT, which has been extensively used for quantitative studies in germ-cell mutagenesis.

  10. Accelerating discovery for complex neurological and behavioral disorders through systems genetics and integrative genomics in the laboratory mouse.

    PubMed

    Bubier, Jason A; Chesler, Elissa J

    2012-04-01

    Recent advances in systems genetics and integrative functional genomics have greatly improved the study of complex neurological and behavioral traits. The methods developed for the integrated characterization of new, high-resolution mouse genetic reference populations and systems genetics enable behavioral geneticists an unprecedented opportunity to address questions of the molecular basis of neurological and psychiatric disorders and their comorbidities. Integrative genomics augment these strategies by enabling rapid informatics-assisted candidate gene prioritization, cross-species translation, and mechanistic comparison across related disorders from a wealth of existing data in mouse and other model organisms. Ultimately, through these complementary approaches, finding the mechanisms and sources of genetic variation underlying complex neurobehavioral disease related traits is becoming tractable. Furthermore, these methods enable categorization of neurobehavioral disorders through their underlying biological basis. Together, these model organism-based approaches can lead to a refinement of diagnostic categories and targeted treatment of neurological and psychiatric disease.

  11. Human and mouse introns are linked to the same processes and functions through each genome's most frequent non-conserved motifs.

    PubMed

    Tsirigos, Aristotelis; Rigoutsos, Isidore

    2008-06-01

    We identified the most frequent, variable-length DNA sequence motifs in the human and mouse genomes and sub-selected those with multiple recurrences in the intergenic and intronic regions and at least one additional exonic instance in the corresponding genome. We discovered that these motifs have virtually no overlap with intronic sequences that are conserved between human and mouse, and thus are genome-specific. Moreover, we found that these motifs span a substantial fraction of previously uncharacterized human and mouse intronic space. Surprisingly, we found that these genome-specific motifs are over-represented in the introns of genes belonging to the same biological processes and molecular functions in both the human and mouse genomes even though the underlying sequences are not conserved between the two genomes. In fact, the processes and functions that are linked to these genome-specific sequence-motifs are distinct from the processes and functions which are associated with intronic regions that are conserved between human and mouse. The findings show that intronic regions from different genomes are linked to the same processes and functions in the absence of underlying sequence conservation. We highlight the ramifications of this observation with a concrete example that involves the microsatellite instability gene MLH1.

  12. Genome wide analysis of inbred mouse lines identifies a locus containing Ppar-gamma as contributing to enhanced malaria survival.

    PubMed

    Bopp, Selina E R; Ramachandran, Vandana; Henson, Kerstin; Luzader, Angelina; Lindstrom, Merle; Spooner, Muriel; Steffy, Brian M; Suzuki, Oscar; Janse, Chris; Waters, Andrew P; Zhou, Yingyao; Wiltshire, Tim; Winzeler, Elizabeth A

    2010-05-28

    The genetic background of a patient determines in part if a person develops a mild form of malaria and recovers, or develops a severe form and dies. We have used a mouse model to detect genes involved in the resistance or susceptibility to Plasmodium berghei malaria infection. To this end we first characterized 32 different mouse strains infected with P. berghei and identified survival as the best trait to discriminate between the strains. We found a locus on chromosome 6 by linking the survival phenotypes of the mouse strains to their genetic variations using genome wide analyses such as haplotype associated mapping and the efficient mixed-model for association. This new locus involved in malaria resistance contains only two genes and confirms the importance of Ppar-gamma in malaria infection.

  13. Genomic organization of the mouse fibroblast growth factor receptor 3 (Fgfr3) gene

    SciTech Connect

    Perez-Castro, A.V.; Wilson, J.; Altherr, M.R.

    1995-11-20

    The fibroblast growth factor receptor 3 (Fgfr3) protein is a tyrosine kinase receptor involved in the signal transduction of various fibroblast growth factors. Recent studies suggest its important role in normal development. In humans, mutation in Fgfr3 is responsible for growth disorders such as achondroplasia, hypoachondroplasia, and thanatophoric dysplasia. Here, we report the complete genomic organization of the mouse Fgfr3 gene. The murine gene spans approximately 15 kb and consists of 19 exons and 18 introns. One major and one minor transcription initiation site were identified. Position +1 is located 614 nucleotides upstream from the ATG initiation codon. The translation initiation and termination sites are located in exons 2 and 19, respectively. Five Sp1 sites, two AP2 sites, one Zeste site, and one Krox 24 site were observed in the 5{prime}-flanking region. The Fgfr3 promoter appears to be contained within a CpG island and, as is common in genes having multiple Sp1-binding sites, lacks a TATA box. 35 refs., 3 figs., 1 tab.

  14. Longitudinal assessment of neuronal 3D genomes in mouse prefrontal cortex

    PubMed Central

    Mitchell, Amanda C.; Javidfar, Behnam; Bicks, Lucy K.; Neve, Rachael; Garbett, Krassimira; Lander, Sharon S.; Mirnics, Karoly; Morishita, Hirofumi; Wood, Marcelo A.; Jiang, Yan; Gaisler-Salomon, Inna; Akbarian, Schahram

    2016-01-01

    Neuronal epigenomes, including chromosomal loopings moving distal cis-regulatory elements into proximity of target genes, could serve as molecular proxy linking present-day-behaviour to past exposures. However, longitudinal assessment of chromatin state is challenging, because conventional chromosome conformation capture assays essentially provide single snapshots at a given time point, thus reflecting genome organization at the time of brain harvest and therefore are non-informative about the past. Here we introduce ‘NeuroDam' to assess epigenome status retrospectively. Short-term expression of the bacterial DNA adenine methyltransferase Dam, tethered to the Gad1 gene promoter in mouse prefrontal cortex neurons, results in stable GmethylATC tags at Gad1-bound chromosomal contacts. We show by NeuroDam that mice with defective cognition 4 months after pharmacological NMDA receptor blockade already were affected by disrupted chromosomal conformations shortly after drug exposure. Retrospective profiling of neuronal epigenomes is likely to illuminate epigenetic determinants of normal and diseased brain development in longitudinal context. PMID:27597321

  15. Genomic Analyses as a Guide to Target Identification and Preclinical Testing of Mouse Models of Breast Cancer

    PubMed Central

    Bennett, Christina N; Green, Jeffrey E.

    2012-01-01

    Cross-species genomic analyses have proven useful for identifying common genomic alterations that occur in human cancers and mouse models designed to recapitulate human tumor development. High-throughput molecular analyses provide a valuable tool for identifying particular animal models that may represent aspects of specific subtypes of human cancers. Corresponding alterations in gene copy number and expression in tumors from mouse and human suggest that these conserved changes may be mechanistically essential for cancer development and progression, and therefore, they may be critical targets for therapeutic intervention. Using a cross-species analysis approach, mouse models in which the functions of p53, Rb, and BRCA1 have been disrupted demonstrate molecular features of human, triple-negative (ER-, PR-, and ERBB2-), basal-type breast cancer. Using mouse tumor models based on the targeted abrogation of p53 and Rb function, we identified a large, integrated genetic network that correlates to poor outcome in several human epithelial cancers. This gene signature is highly enriched for genes involved in DNA replication and repair, chromosome maintenance, cell cycle regulation, and apoptosis. Current studies are determining whether inactivation of specific members within this signature, using drugs or siRNA, will identify potentially important new targets to inhibit triple-negative, basal-type breast cancer for which no targeted therapies currently exist. PMID:20080934

  16. Analysis of segmental duplications, mouse genome synteny and recurrent cancer-associated amplicons in human chromosome 6p21-p12.

    PubMed

    Martin, J W; Yoshimoto, M; Ludkovski, O; Thorner, P S; Zielenska, M; Squire, J A; Nuin, P A S

    2010-06-01

    It has been proposed that regions of microhomology in the human genome could facilitate genomic rearrangements, copy number transitions, and rapid genomic change during tumor progression. To investigate this idea, this study examines the role of repetitive sequence elements, and corresponding syntenic mouse genomic features, in targeting cancer-associated genomic instability of specific regions of the human genome. Automated database-mining algorithms designed to search for frequent copy number transitions and genomic breakpoints were applied to 2 publicly-available online databases and revealed that 6p21-p12 is one of the regions of the human genome most frequently involved in tumor-specific alterations. In these analyses, 6p21-p12 exhibited the highest frequency of genomic amplification in osteosarcomas. Analysis of repetitive elements in regions of homology between human chromosome 6p and the syntenic regions of the mouse genome revealed a strong association between the location of segmental duplications greater than 5 kilobase-pairs and the position of discontinuities at the end of the syntenic region. The presence of clusters of segmental duplications flanking these syntenic regions also correlated with a high frequency of amplification and genomic alteration. Collectively, the experimental findings, in silico analyses, and comparative genomic studies presented here suggest that segmental duplications may facilitate cancer-associated copy number transitions and rearrangements at chromosome 6p21-p12. This process may involve homology-dependent DNA recombination and/or repair, which may also contribute towards the overall plasticity of the human genome.

  17. Promoter-Specific Expression and Genomic Structure of IgLON Family Genes in Mouse

    PubMed Central

    Vanaveski, Taavi; Singh, Katyayani; Narvik, Jane; Eskla, Kattri-Liis; Visnapuu, Tanel; Heinla, Indrek; Jayaram, Mohan; Innos, Jürgen; Lilleväli, Kersti; Philips, Mari-Anne; Vasar, Eero

    2017-01-01

    IgLON family is composed of five genes: Lsamp, Ntm, Opcml, Negr1, and Iglon5; encoding for five highly homologous neural adhesion proteins that regulate neurite outgrowth and synapse formation. In the current study we performed in silico analysis revealing that Ntm and Opcml display similar genomic structure as previously reported for Lsamp, characterized by two alternative promotors 1a and 1b. Negr1 and Iglon5 transcripts have uniform 5′ region, suggesting single promoter. Iglon5, the recently characterized family member, shares high level of conservation and structural qualities characteristic to IgLON family such as N-terminal signal peptide, three Ig domains, and GPI anchor binding site. By using custom 5′-isoform-specific TaqMan gene-expression assay, we demonstrated heterogeneous expression of IgLON transcripts in different areas of mouse brain and several-fold lower expression in selected tissues outside central nervous system. As an example, the expression of IgLON transcripts in urogenital and reproductive system is in line with repeated reports of urogenital tumors accompanied by mutations in IgLON genes. Considering the high levels of intra-family homology shared by IgLONs, we investigated potential compensatory effects at the level of IgLON isoforms in the brains of mice deficient of one or two family members. We found that the lack of IgLONs is not compensated by a systematic quantitative increase of the other family members. On the contrary, the expression of Ntm 1a transcript and NEGR1 protein was significantly reduced in the frontal cortex of Lsamp-deficient mice suggesting that the expression patterns within IgLON family are balanced coherently. The actions of individual IgLONs, however, can be antagonistic as demonstrated by differential expression of Syp in deletion mutants of IgLONs. In conclusion, we show that the genomic twin-promoter structure has impact on both anatomical distribution and intra-family interactions of IgLON family members

  18. Maternal-foetal genomic conflict and speciation: no evidence for hybrid placental dysplasia in crosses between two house mouse subspecies.

    PubMed

    Kropáčková, L; Piálek, J; Gergelits, V; Forejt, J; Reifová, R

    2015-03-01

    Interspecific hybridization between closely related mammalian species, including various species of the genus Mus, is commonly associated with abnormal growth of the placenta and hybrid foetuses, a phenomenon known as hybrid placental dysplasia (HPD). The role of HPD in speciation is anticipated but still poorly understood. Here, we studied placental and foetal growth in F1 crosses between four inbred mouse strains derived from two house mouse subspecies, Mus musculus musculus and Mus musculus domesticus. These subspecies are in the early stage of speciation and still hybridize in nature. In accordance with the maternal-foetal genomic conflict hypothesis, we found different parental influences on placental and foetal development, with placental weight most affected by the father's body weight and foetal weight by the mother's body weight. After removing the effects of parents' body weight, we did not find any significant differences in foetal or placental weights between intra-subspecific and inter-subspecific F1 crosses. Nevertheless, we found that the variability in placental weight in inter-subspecific crosses is linked to the X chromosome, similarly as for HPD in interspecific mouse crosses. Our results suggest that maternal-foetal genomic conflict occurs in the house mouse system, but has not yet diverged sufficiently to cause abnormalities in placental and foetal growth in inter-subspecific crosses. HPD is thus unlikely to contribute to speciation in the house mouse system. However, we cannot rule out that it might have contributed to other speciation events in the genus Mus, where differences in the levels of polyandry exist between the species.

  19. Genome wide chromatin occupancy of mrhl RNA and its role in gene regulation in mouse spermatogonial cells

    PubMed Central

    Akhade, Vijay Suresh; Arun, Gayatri; Donakonda, Sainitin; Satyanarayana Rao, Manchanahalli R

    2014-01-01

    Mrhl RNA is a nuclear lncRNA encoded in the mouse genome and negatively regulates Wnt signaling in spermatogonial cells through p68/Ddx5 RNA helicase. Mrhl RNA is present in the chromatin fraction of mouse spermatogonial Gc1-Spg cells and genome wide chromatin occupancy of mrhl RNA by ChOP (Chromatin oligo affinity precipitation) technique identified 1370 statistically significant genomic loci. Among these, genes at 37 genomic loci also showed altered expression pattern upon mrhl RNA down regulation which are referred to as GRPAM (Genes Regulated by Physical Association of Mrhl RNA). p68 interacted with mrhl RNA in chromatin at these GRPAM loci. p68 silencing drastically reduced mrhl RNA occupancy at 27 GRPAM loci and also perturbed the expression of GRPAM suggesting a role for p68 mediated mrhl RNA occupancy in regulating GRPAM expression. Wnt3a ligand treatment of Gc1-Spg cells down regulated mrhl RNA expression and also perturbed expression of these 27 GRPAM genes that included genes regulating Wnt signaling pathway and spermatogenesis, one of them being Sox8, a developmentally important transcription factor. We also identified interacting proteins of mrhl RNA associated chromatin fraction which included Pc4, a chromatin organizer protein and hnRNP A/B and hnRNP A2/B1 which have been shown to be associated with lincRNA-Cox2 function in gene regulation. Our findings in the Gc1-Spg cell line also correlate with the results from analysis of mouse testicular tissue which further highlights the in vivo physiological significance of mrhl RNA in the context of gene regulation during mammalian spermatogenesis. PMID:25584904

  20. Genome-Scale Assessment of Age-Related DNA Methylation Changes in Mouse Spermatozoa

    PubMed Central

    Kobayashi, Norio; Okae, Hiroaki; Hiura, Hitoshi; Chiba, Hatsune; Shirakata, Yoshiki; Hara, Kenshiro; Tanemura, Kentaro; Arima, Takahiro

    2016-01-01

    DNA methylation plays important roles in the production and functioning of spermatozoa. Recent studies have suggested that DNA methylation patterns in spermatozoa can change with age, but the regions susceptible to age-related methylation changes remain to be fully elucidated. In this study, we conducted genome-scale DNA methylation profiling of spermatozoa obtained from C57BL/6N mice at 8 weeks (8w), 18 weeks (18w) and 17 months of age (17m). There was no substantial difference in the global DNA methylation patterns between 18w and 17m samples except for a slight increase of methylation levels in long interspersed nuclear elements in the 17m samples. We found that maternally methylated imprinting control regions (mICRs) and spermatogenesis-related gene promoters had 5–10% higher methylation levels in 8w samples than in 18w or 17m samples. Analysis of individual sequence reads suggested that these regions were fully methylated (80–100%) in a subset of 8w spermatozoa. These regions are also known to be highly methylated in a subset of postnatal spermatogonia, which might be the source of the increased DNA methylation in 8w spermatozoa. Another possible source was contamination by somatic cells. Although we carefully purified the spermatozoa, it was difficult to completely exclude the possibility of somatic cell contamination. Further studies are needed to clarify the source of the small increase in DNA methylation in the 8w samples. Overall, our findings suggest that DNA methylation patterns in mouse spermatozoa are relatively stable throughout reproductive life. PMID:27880848

  1. An encyclopedia of mouse DNA elements (Mouse ENCODE).

    PubMed

    Stamatoyannopoulos, John A; Snyder, Michael; Hardison, Ross; Ren, Bing; Gingeras, Thomas; Gilbert, David M; Groudine, Mark; Bender, Michael; Kaul, Rajinder; Canfield, Theresa; Giste, Erica; Johnson, Audra; Zhang, Mia; Balasundaram, Gayathri; Byron, Rachel; Roach, Vaughan; Sabo, Peter J; Sandstrom, Richard; Stehling, A Sandra; Thurman, Robert E; Weissman, Sherman M; Cayting, Philip; Hariharan, Manoj; Lian, Jin; Cheng, Yong; Landt, Stephen G; Ma, Zhihai; Wold, Barbara J; Dekker, Job; Crawford, Gregory E; Keller, Cheryl A; Wu, Weisheng; Morrissey, Christopher; Kumar, Swathi A; Mishra, Tejaswini; Jain, Deepti; Byrska-Bishop, Marta; Blankenberg, Daniel; Lajoie, Bryan R; Jain, Gaurav; Sanyal, Amartya; Chen, Kaun-Bei; Denas, Olgert; Taylor, James; Blobel, Gerd A; Weiss, Mitchell J; Pimkin, Max; Deng, Wulan; Marinov, Georgi K; Williams, Brian A; Fisher-Aylor, Katherine I; Desalvo, Gilberto; Kiralusha, Anthony; Trout, Diane; Amrhein, Henry; Mortazavi, Ali; Edsall, Lee; McCleary, David; Kuan, Samantha; Shen, Yin; Yue, Feng; Ye, Zhen; Davis, Carrie A; Zaleski, Chris; Jha, Sonali; Xue, Chenghai; Dobin, Alex; Lin, Wei; Fastuca, Meagan; Wang, Huaien; Guigo, Roderic; Djebali, Sarah; Lagarde, Julien; Ryba, Tyrone; Sasaki, Takayo; Malladi, Venkat S; Cline, Melissa S; Kirkup, Vanessa M; Learned, Katrina; Rosenbloom, Kate R; Kent, W James; Feingold, Elise A; Good, Peter J; Pazin, Michael; Lowdon, Rebecca F; Adams, Leslie B

    2012-08-13

    To complement the human Encyclopedia of DNA Elements (ENCODE) project and to enable a broad range of mouse genomics efforts, the Mouse ENCODE Consortium is applying the same experimental pipelines developed for human ENCODE to annotate the mouse genome.

  2. The first-generation whole-genome radiation hybrid map in the horse identifies conserved segments in human and mouse genomes.

    PubMed

    Chowdhary, Bhanu P; Raudsepp, Terje; Kata, Srinivas R; Goh, Glenda; Millon, Lee V; Allan, Veronica; Piumi, François; Guérin, Gérard; Swinburne, June; Binns, Matthew; Lear, Teri L; Mickelson, Jim; Murray, James; Antczak, Douglas F; Womack, James E; Skow, Loren C

    2003-04-01

    A first-generation radiation hybrid (RH) map of the equine (Equus caballus) genome was assembled using 92 horse x hamster hybrid cell lines and 730 equine markers. The map is the first comprehensive framework map of the horse that (1) incorporates type I as well as type II markers, (2) integrates synteny, cytogenetic, and meiotic maps into a consensus map, and (3) provides the most detailed genome-wide information to date on the organization and comparative status of the equine genome. The 730 loci (258 type I and 472 type II) included in the final map are clustered in 101 RH groups distributed over all equine autosomes and the X chromosome. The overall marker retention frequency in the panel is approximately 21%, and the possibility of adding any new marker to the map is approximately 90%. On average, the mapped markers are distributed every 19 cR (4 Mb) of the equine genome--a significant improvement in resolution over previous maps. With 69 new FISH assignments, a total of 253 cytogenetically mapped loci physically anchor the RH map to various chromosomal segments. Synteny assignments of 39 gene loci complemented the RH mapping of 27 genes. The results added 12 new loci to the horse gene map. Lastly, comparison of the assembly of 447 equine genes (256 linearly ordered RH-mapped and additional 191 FISH-mapped) with the location of draft sequences of their human and mouse orthologs provides the most extensive horse-human and horse-mouse comparative map to date. We expect that the foundation established through this map will significantly facilitate rapid targeted expansion of the horse gene map and consequently, mapping and positional cloning of genes governing traits significant to the equine industry.

  3. Specific sequence deletions in two classes of murine leukemia virus-related proviruses in the mouse genome.

    PubMed

    Ch'ang, L Y; Yang, W K; Myer, F E; Koh, C K; Boone, L R

    1989-02-01

    Characteristic long terminal repeats (LTR) of approximately 700 and 750 bp were found, respectively, in the two classes (polytropic and modified polytropic) of murine leukemia virus (MuLV)-related nonecotropic nonxenotropic proviral sequences in eight individual molecular clones of RFM/Un mouse chromosomal DNA fragments. Three proviral clones, two polytropic and one modified polytropic, contained sequence deletions in the viral structural genes. Nucleotide sequence analysis revealed that 7-bp direct repeats occur at both ends of deleted sequences in intact structures and one of the repeats remains in genomes with the deletion. Specifically, the deleted sequences were a 1487-bp gag-pol sequence with ACTGCCC repeat, a 113-bp mid-pol sequence with CAGGCAA repeat, and a 1811-bp env sequence with GGTCCAG repeat. The same specific sequence deletions were found in both classes of MuLV-related proviral structures. Examination of chromosomal DNA from eight inbred laboratory mouse strains and six wild mouse species showed that a minor population of proviruses with these specific deletions were present in Mus musculus and Mus spretus, all of which contain prominent 700-bp LTR polytropic proviral structures. The 750-bp LTR modified polytropic proviral structures were phylogenetically more restricted, being equally predominant in Mus musculus domesticus mice, but minor to undetectable in Mus spretus subspecies, and absent in other wild mouse populations.

  4. Promising genomic transfectant into Xeroderma pigmentosum group A with highly amplified mouse DNA and intermediate UV resistance turns revertant

    SciTech Connect

    Blum, M.; Baumann, I.; Lohrer, H.; Rahmsdorf, H.J.; Herrlich, P.

    1989-04-28

    Following transfection of genomic mouse DNA into an SV40 transformed fibroblast cell line from a patient with Xeroderma pigmentosum (complementation group A, XPA), a single UV resistant cell clone was isolated out of a total of 10(4) independent transfectants. The recipient XPA cell line has as yet not produced spontaneous revertants among 2.2 x 10(8) cells. The isolated cell clone contains 50-70 kb of mouse sequences which are heavily amplified (500-fold), and has acquired both intermediate resistance to UV killing and intermediate unscheduled DNA synthesis (UDS) capacity. By continued passage without selective pressure, cells were generated, which had lost both the dominant marker gene and repetitive mouse sequences. Single colonies of these cells were still intermediately resistant to UV suggesting that either undetected unique mouse DNA had segregated from the bulk of repetitive DNA, or, more likely, that the initially isolated transfectant was a spontaneous revertant. This documents that a persuasive clone isolated can still be a false positive (spontaneous revertant) and that an extremely laborious approach may lead into a dead end.

  5. Pharmacokinetic and Genomic Effects of Arsenite in Drinking Water on Mouse Lung in a 30-Day Exposure.

    PubMed

    Chilakapati, Jaya; Wallace, Kathleen; Hernandez-Zavala, Araceli; Moore, Tanya; Ren, Hongzu; Kitchin, Kirk T

    2015-01-01

    The 2 objectives of this subchronic study were to determine the arsenite drinking water exposure dependent increases in female C3H mouse liver and lung tissue arsenicals and to characterize the dose response (to 0, 0.05, 0.25, 1, 10, and 85 ppm arsenite in drinking water for 30 days and a purified AIN-93M diet) for genomic mouse lung expression patterns. Mouse lungs were analyzed for inorganic arsenic, monomethylated, and dimethylated arsenicals by hydride generation atomic absorption spectroscopy. The total lung mean arsenical levels were 1.4, 22.5, 30.1, 50.9, 105.3, and 316.4 ng/g lung tissue after 0, 0.05, 0.25, 1, 10, and 85 ppm, respectively. At 85 ppm, the total mean lung arsenical levels increased 14-fold and 131-fold when compared to either the lowest noncontrol dose (0.05 ppm) or the control dose, respectively. We found that arsenic exposure elicited minimal numbers of differentially expressed genes (DEGs; 77, 38, 90, 87, and 87 DEGs) after 0.05, 0.25, 1, 10, and 85 ppm, respectively, which were associated with cardiovascular disease, development, differentiation, apoptosis, proliferation, and stress response. After 30 days of arsenite exposure, this study showed monotonic increases in mouse lung arsenical (total arsenic and dimethylarsinic acid) concentrations but no clear dose-related increases in DEG numbers.

  6. Genome-guided design of a defined mouse microbiota that confers colonization resistance against Salmonella enterica serovar Typhimurium.

    PubMed

    Brugiroux, Sandrine; Beutler, Markus; Pfann, Carina; Garzetti, Debora; Ruscheweyh, Hans-Joachim; Ring, Diana; Diehl, Manuel; Herp, Simone; Lötscher, Yvonne; Hussain, Saib; Bunk, Boyke; Pukall, Rüdiger; Huson, Daniel H; Münch, Philipp C; McHardy, Alice C; McCoy, Kathy D; Macpherson, Andrew J; Loy, Alexander; Clavel, Thomas; Berry, David; Stecher, Bärbel

    2016-11-21

    Protection against enteric infections, also termed colonization resistance, results from mutualistic interactions of the host and its indigenous microbes. The gut microbiota of humans and mice is highly diverse and it is therefore challenging to assign specific properties to its individual members. Here, we have used a collection of murine bacterial strains and a modular design approach to create a minimal bacterial community that, once established in germ-free mice, provided colonization resistance against the human enteric pathogen Salmonella enterica serovar Typhimurium (S. Tm). Initially, a community of 12 strains, termed Oligo-Mouse-Microbiota (Oligo-MM(12)), representing members of the major bacterial phyla in the murine gut, was selected. This community was stable over consecutive mouse generations and provided colonization resistance against S. Tm infection, albeit not to the degree of a conventional complex microbiota. Comparative (meta)genome analyses identified functions represented in a conventional microbiome but absent from the Oligo-MM(12). By genome-informed design, we created an improved version of the Oligo-MM community harbouring three facultative anaerobic bacteria from the mouse intestinal bacterial collection (miBC) that provided conventional-like colonization resistance. In conclusion, we have established a highly versatile experimental system that showed efficacy in an enteric infection model. Thus, in combination with exhaustive bacterial strain collections and systems-based approaches, genome-guided design can be used to generate insights into microbe-microbe and microbe-host interactions for the investigation of ecological and disease-relevant mechanisms in the intestine.

  7. Evolution of the vertebrate genome as reflected in paralogous chromosomal regions in man and the house mouse

    SciTech Connect

    Lundin, L.G. )

    1993-04-01

    Gene constellations on several human chromosomes are interpreted as indications of large regional duplications that took place during evolution of the vertebrate genome. Four groups of paralogous chromosomal regions in man and the house mouse are suggested and are believed to be conserved remnants of the two or three rounds of tetraploidization that are likely to have occurred during evolution of the vertebrates. The phenomenon of differential silencing of genes is described. The importance of conservation of linkage of particular genes is discussed in relation to genetic regulation and cell differentiation. 120 refs., 5 tabs.

  8. Genomic cloning of mouse MIF (macrophage inhibitory factor) and genetic mapping of the human and mouse expressed gene and nine mouse pseudogenes

    SciTech Connect

    Kozak, C.A.; Adamson, M.C.; Buckler, C.E.

    1995-06-10

    The single functional mouse gene for MIF (macrophage migration inhibitory factor) has been cloned from a P1 library, and its exon/intron structure determined and shown to resemble that of the human gene. The gene was mapped to chromosome 10 using two multilocus crosses between laboratory strains and either Mus musculus or Mus spretus. Nine additional loci containing related sequences, apparently all processed pseudogenes, were also mapped to chromosomes 1, 2, 3, 7, 8, 9, 12, 17, and 19. While most of these pseudogenes were also found in inbred mice and M. spretus, some are species specific. This suggests that there have been active phases of pseudogene formation in Mus both before and after the separation of musculus and spretus. The human gene contains no pseudogene; we assigned the human gene to chromosome 19, consistent with the location of mouse and human functional genes for MIF in a region of conserved linkage. 43 refs., 4 figs., 1 tab.

  9. Beyond genome-wide association studies: the usefulness of mouse genetics in understanding the complex etiology of atherosclerosis.

    PubMed

    Welch, Carrie L

    2012-02-01

    The development of population-based genome-wide association studies has led to the rapid identification of large numbers of genetic variants associated with coronary artery disease (CAD) and related traits. Together with large-scale gene-centric studies, at least 35 loci associated with CAD per se have been identified with replication. The majority of these associations are with common single-nucleotide polymorphisms exhibiting modest effects on relative risk. The modest nature of the effects, coupled with ethical/practical constraints associated with human sampling, makes it difficult to answer important questions beyond gene/locus localization and allele frequency via human genetic studies. Questions related to gene function, disease-causing mechanism(s), and effective interventions will likely require studies in model organisms. The use of the mouse model for further detailed studies of CAD-associated loci identified by genome-wide association studies is highlighted herein.

  10. Novel insights into the relationships between dendritic cell subsets in human and mouse revealed by genome-wide expression profiling

    PubMed Central

    Robbins, Scott H; Walzer, Thierry; Dembélé, Doulaye; Thibault, Christelle; Defays, Axel; Bessou, Gilles; Xu, Huichun; Vivier, Eric; Sellars, MacLean; Pierre, Philippe; Sharp, Franck R; Chan, Susan; Kastner, Philippe; Dalod, Marc

    2008-01-01

    Background Dendritic cells (DCs) are a complex group of cells that play a critical role in vertebrate immunity. Lymph-node resident DCs (LN-DCs) are subdivided into conventional DC (cDC) subsets (CD11b and CD8α in mouse; BDCA1 and BDCA3 in human) and plasmacytoid DCs (pDCs). It is currently unclear if these various DC populations belong to a unique hematopoietic lineage and if the subsets identified in the mouse and human systems are evolutionary homologs. To gain novel insights into these questions, we sought conserved genetic signatures for LN-DCs and in vitro derived granulocyte-macrophage colony stimulating factor (GM-CSF) DCs through the analysis of a compendium of genome-wide expression profiles of mouse or human leukocytes. Results We show through clustering analysis that all LN-DC subsets form a distinct branch within the leukocyte family tree, and reveal a transcriptomal signature evolutionarily conserved in all LN-DC subsets. Moreover, we identify a large gene expression program shared between mouse and human pDCs, and smaller conserved profiles shared between mouse and human LN-cDC subsets. Importantly, most of these genes have not been previously associated with DC function and many have unknown functions. Finally, we use compendium analysis to re-evaluate the classification of interferon-producing killer DCs, lin-CD16+HLA-DR+ cells and in vitro derived GM-CSF DCs, and show that these cells are more closely linked to natural killer and myeloid cells, respectively. Conclusion Our study provides a unique database resource for future investigation of the evolutionarily conserved molecular pathways governing the ontogeny and functions of leukocyte subsets, especially DCs. PMID:18218067

  11. High-throughput single nucleotide variant discovery in E14 mouse embryonic stem cells provides a new reference genome assembly.

    PubMed

    Incarnato, Danny; Krepelova, Anna; Neri, Francesco

    2014-08-01

    Mouse E14 embryonic stem cells (ESCs) are a well-characterized and widespread used ESC line, often employed for genome-wide studies involving next generation sequencing analysis. More than 2×10(9) sequences made on Illumina platform derived from the genome of E14 ESCs were used to build a database of about 2.7×10(6) single nucleotide variants (SNVs). The identified variants are enriched in intergenic regions, but several thousands reside in gene exons and regulatory regions, such as promoters, enhancers, splicing sites and untranslated regions of RNA, thus indicating high probability of an important functional impact on the molecular biology of these cells. We created a new E14 genome assembly reference that increases the number of mapped reads of about 5%. We performed a Reduced Representation Bisulfite Sequencing on E14 ESCs and we obtained an increase of about 120,000 called CpGs and avoided about 20,000 wrong CpG calls with respect to the mm9 genome reference.

  12. Genomic resources for wild populations of the house mouse, Mus musculus and its close relative Mus spretus.

    PubMed

    Harr, Bettina; Karakoc, Emre; Neme, Rafik; Teschke, Meike; Pfeifle, Christine; Pezer, Željka; Babiker, Hiba; Linnenbrink, Miriam; Montero, Inka; Scavetta, Rick; Abai, Mohammad Reza; Molins, Marta Puente; Schlegel, Mathias; Ulrich, Rainer G; Altmüller, Janine; Franitza, Marek; Büntge, Anna; Künzel, Sven; Tautz, Diethard

    2016-09-13

    Wild populations of the house mouse (Mus musculus) represent the raw genetic material for the classical inbred strains in biomedical research and are a major model system for evolutionary biology. We provide whole genome sequencing data of individuals representing natural populations of M. m. domesticus (24 individuals from 3 populations), M. m. helgolandicus (3 individuals), M. m. musculus (22 individuals from 3 populations) and M. spretus (8 individuals from one population). We use a single pipeline to map and call variants for these individuals and also include 10 additional individuals of M. m. castaneus for which genomic data are publically available. In addition, RNAseq data were obtained from 10 tissues of up to eight adult individuals from each of the three M. m. domesticus populations for which genomic data were collected. Data and analyses are presented via tracks viewable in the UCSC or IGV genome browsers. We also provide information on available outbred stocks and instructions on how to keep them in the laboratory.

  13. Genomic resources for wild populations of the house mouse, Mus musculus and its close relative Mus spretus

    PubMed Central

    Harr, Bettina; Karakoc, Emre; Neme, Rafik; Teschke, Meike; Pfeifle, Christine; Pezer, Željka; Babiker, Hiba; Linnenbrink, Miriam; Montero, Inka; Scavetta, Rick; Abai, Mohammad Reza; Molins, Marta Puente; Schlegel, Mathias; Ulrich, Rainer G.; Altmüller, Janine; Franitza, Marek; Büntge, Anna; Künzel, Sven; Tautz, Diethard

    2016-01-01

    Wild populations of the house mouse (Mus musculus) represent the raw genetic material for the classical inbred strains in biomedical research and are a major model system for evolutionary biology. We provide whole genome sequencing data of individuals representing natural populations of M. m. domesticus (24 individuals from 3 populations), M. m. helgolandicus (3 individuals), M. m. musculus (22 individuals from 3 populations) and M. spretus (8 individuals from one population). We use a single pipeline to map and call variants for these individuals and also include 10 additional individuals of M. m. castaneus for which genomic data are publically available. In addition, RNAseq data were obtained from 10 tissues of up to eight adult individuals from each of the three M. m. domesticus populations for which genomic data were collected. Data and analyses are presented via tracks viewable in the UCSC or IGV genome browsers. We also provide information on available outbred stocks and instructions on how to keep them in the laboratory. PMID:27622383

  14. Efficient genome engineering by targeted homologous recombination in mouse embryos using transcription activator-like effector nucleases.

    PubMed

    Sommer, Daniel; Peters, Annika; Wirtz, Tristan; Mai, Maren; Ackermann, Justus; Thabet, Yasser; Schmidt, Jürgen; Weighardt, Heike; Wunderlich, F Thomas; Degen, Joachim; Schultze, Joachim L; Beyer, Marc

    2014-01-01

    Generation of mouse models by introducing transgenes using homologous recombination is critical for understanding fundamental biology and pathology of human diseases. Here we investigate whether artificial transcription activator-like effector nucleases (TALENs)-powerful tools that induce DNA double-strand breaks at specific genomic locations-can be combined with a targeting vector to induce homologous recombination for the introduction of a transgene in embryonic stem cells and fertilized murine oocytes. We describe the generation of a conditional mouse model using TALENs, which introduce double-strand breaks at the genomic locus of the special AT-rich sequence-binding protein-1 in combination with a large 14.4 kb targeting template vector. We report successful germline transmission of this allele and demonstrate its recombination in primary cells in the presence of Cre-recombinase. These results suggest that TALEN-assisted induction of DNA double-strand breaks can facilitate homologous recombination of complex targeting constructs directly in oocytes.

  15. Transgenerational developmental effects and genomic instability after X-irradiation of preimplantation embryos: studies on two mouse strains.

    PubMed

    Jacquet, P; Buset, J; Neefs, M; Vankerkom, J; Benotmane, M A; Derradji, H; Hildebrandt, G; Baatout, S

    2010-05-01

    Recent results have shown that irradiation of a single cell, the zygote or 1-cell embryo of various mouse strains, could lead to congenital anomalies in the fetuses. In the Heiligenberger strain, a link between the radiation-induced congenital anomalies and the development of a genomic instability was also suggested. Moreover, further studies showed that in that strain, both congenital anomalies and genomic instability could be transmitted to the next generation. The aim of the experiments described in this paper was to investigate whether such non-targeted transgenerational effects could also be observed in two other radiosensitive mouse strains (CF1 and ICR), using lower radiation doses. Irradiation of the CF1 and ICR female zygotes with 0.2 or 0.4Gy did not result in a decrease of their fertility after birth, when they had reached sexual maturity. Moreover, females of both strains that had been X-irradiated with 0.2Gy exhibited higher rates of pregnancy, less resorptions and more living fetuses. Additionally, the mean weight of living fetuses in these groups had significantly increased. Exencephaly and dwarfism were observed in CF1 fetuses issued from control and X-irradiated females. In the control group of that strain, polydactyly and limb deformity were also found. The yields of abnormal fetuses did not differ significantly between the control and X-irradiated groups. Polydactyly, exencephaly and dwarfism were observed in fetuses issued from ICR control females. In addition to these anomalies, gastroschisis, curly tail and open eye were observed at low frequencies in ICR fetuses issued from X-irradiated females. Again, the frequencies of abnormal fetuses found in the different groups did not differ significantly. In both CF1 and ICR mouse strains, irradiation of female zygotes did not result in the development of a genomic instability in the next generation embryos. Overall, our results suggest that, at the moderate doses used, developmental defects observed

  16. Cognition and Hippocampal Plasticity in the Mouse Is Altered by Monosomy of a Genomic Region Implicated in Down Syndrome

    PubMed Central

    Sahún, Ignasi; Marechal, Damien; Pereira, Patricia Lopes; Nalesso, Valérie; Gruart, Agnes; Garcia, José Maria Delgado; Antonarakis, Stylianos E.; Dierssen, Mara; Herault, Yann

    2014-01-01

    Down syndrome (DS) is due to increased copy number of human chromosome 21. The contribution of different genetic regions has been tested using mouse models. As shown previously, the Abcg1-U2af1 genetic region contributes to cognitive defects in working and short-term recognition memory in Down syndrome mouse models. Here we analyzed the impact of monosomy of the same genetic interval, using a new mouse model, named Ms2Yah. We used several cognitive paradigms and did not detect defects in the object recognition or the Morris water maze tests. However, surprisingly, Ms2Yah mice displayed increased associative memory in a pure contextual fear-conditioning test and decreased social novelty interaction along with a larger long-term potentiation recorded in the CA1 area following stimulation of Schaffer collaterals. Whole-genome expression studies carried out on hippocampus showed that the transcription of only a small number of genes is affected, mainly from the genetic interval (Cbs, Rsph1, Wdr4), with a few additional ones, including the postsynaptic Gabrr2, Gabbr1, Grid2p, Park2, and Dlg1 and the components of the Ubiquitin-mediated proteolysis (Anapc1, Rnf7, Huwe1, Park2). The Abcg1–U2af1 region is undeniably encompassing dosage-sensitive genes or elements whose change in copy number directly affects learning and memory, synaptic function, and autistic related behavior. PMID:24752061

  17. Integration of mouse and human genome-wide association data identifies KCNIP4 as an asthma gene.

    PubMed

    Himes, Blanca E; Sheppard, Keith; Berndt, Annerose; Leme, Adriana S; Myers, Rachel A; Gignoux, Christopher R; Levin, Albert M; Gauderman, W James; Yang, James J; Mathias, Rasika A; Romieu, Isabelle; Torgerson, Dara G; Roth, Lindsey A; Huntsman, Scott; Eng, Celeste; Klanderman, Barbara; Ziniti, John; Senter-Sylvia, Jody; Szefler, Stanley J; Lemanske, Robert F; Zeiger, Robert S; Strunk, Robert C; Martinez, Fernando D; Boushey, Homer; Chinchilli, Vernon M; Israel, Elliot; Mauger, David; Koppelman, Gerard H; Postma, Dirkje S; Nieuwenhuis, Maartje A E; Vonk, Judith M; Lima, John J; Irvin, Charles G; Peters, Stephen P; Kubo, Michiaki; Tamari, Mayumi; Nakamura, Yusuke; Litonjua, Augusto A; Tantisira, Kelan G; Raby, Benjamin A; Bleecker, Eugene R; Meyers, Deborah A; London, Stephanie J; Barnes, Kathleen C; Gilliland, Frank D; Williams, L Keoki; Burchard, Esteban G; Nicolae, Dan L; Ober, Carole; DeMeo, Dawn L; Silverman, Edwin K; Paigen, Beverly; Churchill, Gary; Shapiro, Steve D; Weiss, Scott T

    2013-01-01

    Asthma is a common chronic respiratory disease characterized by airway hyperresponsiveness (AHR). The genetics of asthma have been widely studied in mouse and human, and homologous genomic regions have been associated with mouse AHR and human asthma-related phenotypes. Our goal was to identify asthma-related genes by integrating AHR associations in mouse with human genome-wide association study (GWAS) data. We used Efficient Mixed Model Association (EMMA) analysis to conduct a GWAS of baseline AHR measures from males and females of 31 mouse strains. Genes near or containing SNPs with EMMA p-values <0.001 were selected for further study in human GWAS. The results of the previously reported EVE consortium asthma GWAS meta-analysis consisting of 12,958 diverse North American subjects from 9 study centers were used to select a subset of homologous genes with evidence of association with asthma in humans. Following validation attempts in three human asthma GWAS (i.e., Sepracor/LOCCS/LODO/Illumina, GABRIEL, DAG) and two human AHR GWAS (i.e., SHARP, DAG), the Kv channel interacting protein 4 (KCNIP4) gene was identified as nominally associated with both asthma and AHR at a gene- and SNP-level. In EVE, the smallest KCNIP4 association was at rs6833065 (P-value 2.9e-04), while the strongest associations for Sepracor/LOCCS/LODO/Illumina, GABRIEL, DAG were 1.5e-03, 1.0e-03, 3.1e-03 at rs7664617, rs4697177, rs4696975, respectively. At a SNP level, the strongest association across all asthma GWAS was at rs4697177 (P-value 1.1e-04). The smallest P-values for association with AHR were 2.3e-03 at rs11947661 in SHARP and 2.1e-03 at rs402802 in DAG. Functional studies are required to validate the potential involvement of KCNIP4 in modulating asthma susceptibility and/or AHR. Our results suggest that a useful approach to identify genes associated with human asthma is to leverage mouse AHR association data.

  18. Improvements to previous algorithms to predict gene structure and isoform concentrations using Affymetrix Exon arrays

    PubMed Central

    2010-01-01

    Background Exon arrays provide a way to measure the expression of different isoforms of genes in an organism. Most of the procedures to deal with these arrays are focused on gene expression or on exon expression. Although the only biological analytes that can be properly assigned a concentration are transcripts, there are very few algorithms that focus on them. The reason is that previously developed summarization methods do not work well if applied to transcripts. In addition, gene structure prediction, i.e., the correspondence between probes and novel isoforms, is a field which is still unexplored. Results We have modified and adapted a previous algorithm to take advantage of the special characteristics of the Affymetrix exon arrays. The structure and concentration of transcripts -some of them possibly unknown- in microarray experiments were predicted using this algorithm. Simulations showed that the suggested modifications improved both specificity (SP) and sensitivity (ST) of the predictions. The algorithm was also applied to different real datasets showing its effectiveness and the concordance with PCR validated results. Conclusions The proposed algorithm shows a substantial improvement in the performance over the previous version. This improvement is mainly due to the exploitation of the redundancy of the Affymetrix exon arrays. An R-Package of SPACE with the updated algorithms have been developed and is freely available. PMID:21110835

  19. Genome-wide coexpression of steroid receptors in the mouse brain: Identifying signaling pathways and functionally coordinated regions

    PubMed Central

    Lelieveldt, Boudewijn P. F.; Grefhorst, Aldo; van Weert, Lisa T. C. M.; Mol, Isabel M.; Sips, Hetty C. M.; van den Heuvel, José K.; Datson, Nicole A.; Visser, Jenny A.; Meijer, Onno C.

    2016-01-01

    Steroid receptors are pleiotropic transcription factors that coordinate adaptation to different physiological states. An important target organ is the brain, but even though their effects are well studied in specific regions, brain-wide steroid receptor targets and mediators remain largely unknown due to the complexity of the brain. Here, we tested the idea that novel aspects of steroid action can be identified through spatial correlation of steroid receptors with genome-wide mRNA expression across different regions in the mouse brain. First, we observed significant coexpression of six nuclear receptors (NRs) [androgen receptor (Ar), estrogen receptor alpha (Esr1), estrogen receptor beta (Esr2), glucocorticoid receptor (Gr), mineralocorticoid receptor (Mr), and progesterone receptor (Pgr)] with sets of steroid target genes that were identified in single brain regions. These coexpression relationships were also present in distinct other brain regions, suggestive of as yet unidentified coordinate regulation of brain regions by, for example, glucocorticoids and estrogens. Second, coexpression of a set of 62 known NR coregulators and the six steroid receptors in 12 nonoverlapping mouse brain regions revealed selective downstream pathways, such as Pak6 as a mediator for the effects of Ar and Gr on dopaminergic transmission. Third, Magel2 and Irs4 were identified and validated as strongly responsive targets to the estrogen diethylstilbestrol in the mouse hypothalamus. The brain- and genome-wide correlations of mRNA expression levels of six steroid receptors that we provide constitute a rich resource for further predictions and understanding of brain modulation by steroid hormones. PMID:26811448

  20. A genome survey sequencing of the Java mouse deer (Tragulus javanicus) adds new aspects to the evolution of lineage specific retrotransposons in Ruminantia (Cetartiodactyla).

    PubMed

    Gallus, S; Kumar, V; Bertelsen, M F; Janke, A; Nilsson, M A

    2015-10-25

    Ruminantia, the ruminating, hoofed mammals (cow, deer, giraffe and allies) are an unranked artiodactylan clade. Around 50-60 million years ago the BovB retrotransposon entered the ancestral ruminantian genome through horizontal gene transfer. A survey genome screen using 454-pyrosequencing of the Java mouse deer (Tragulus javanicus) and the lesser kudu (Tragelaphus imberbis) was done to investigate and to compare the landscape of transposable elements within Ruminantia. The family Tragulidae (mouse deer) is the only representative of Tragulina and phylogenetically important, because it represents the earliest divergence in Ruminantia. The data analyses show that, relative to other ruminantian species, the lesser kudu genome has seen an expansion of BovB Long INterspersed Elements (LINEs) and BovB related Short INterspersed Elements (SINEs) like BOVA2. In comparison the genome of Java mouse deer has fewer BovB elements than other ruminants, especially Bovinae, and has in addition a novel CHR-3 SINE most likely propagated by LINE-1. By contrast the other ruminants have low amounts of CHR SINEs but high numbers of actively propagating BovB-derived and BovB-propagated SINEs. The survey sequencing data suggest that the transposable element landscape in mouse deer (Tragulina) is unique among Ruminantia, suggesting a lineage specific evolutionary trajectory that does not involve BovB mediated retrotransposition. This shows that the genomic landscape of mobile genetic elements can rapidly change in any lineage.

  1. 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

  2. Genome-wide alteration of 5-hydroxymethylcytosine in a mouse model of fragile X-associated tremor/ataxia syndrome

    PubMed Central

    Yao, Bing; Lin, Li; Street, R. Craig; Zalewski, Zachary A.; Galloway, Jocelyn N.; Wu, Hao; Nelson, David L.; Jin, Peng

    2014-01-01

    Fragile X-associated tremor/ataxia syndrome (FXTAS) is a late-onset neurodegenerative disorder in which patients carry premutation alleles of 55–200 CGG repeats in the FMR1 gene. To date, whether alterations in epigenetic regulation modulate FXTAS has gone unexplored. 5-Hydroxymethylcytosine (5hmC) converted from 5-methylcytosine (5mC) by the ten-eleven translocation (TET) family of proteins has been found recently to play key roles in neuronal functions. Here, we undertook genome-wide profiling of cerebellar 5hmC in a FXTAS mouse model (rCGG mice) and found that rCGG mice at 16 weeks showed overall reduced 5hmC levels genome-wide compared with age-matched wild-type littermates. However, we also observed gain-of-5hmC regions in repetitive elements, as well as in cerebellum-specific enhancers, but not in general enhancers. Genomic annotation and motif prediction of wild-type- and rCGG-specific differential 5-hydroxymethylated regions (DhMRs) revealed their high correlation with genes and transcription factors that are important in neuronal developmental and functional pathways. DhMR-associated genes partially overlapped with genes that were differentially associated with ribosomes in CGG mice identified by bacTRAP ribosomal profiling. Taken together, our data strongly indicate a functional role for 5hmC-mediated epigenetic modulation in the etiology of FXTAS, possibly through the regulation of transcription. PMID:24108107

  3. Characterization of AKR murine leukemia virus sequences in AKR mouse substrains and structure of integrated recombinant genomes in tumor tissues.

    PubMed Central

    Quint, W; Quax, W; van der Putten, H; Berns, A

    1981-01-01

    A specific cDNA probe of AKR murine leukemia virus (AKR-MLV) was prepared to detect AKR-MLV sequences in normal and tumor tissues in a variety of AKR mouse substrains. AKR strains contained up to six endogenous AKR-MLV genomes. All substrains tested had one AKR-MLV locus in common, and closely related substrains had several proviruses integrated in an identical site. Virus-induced tumors in the AKR/FuRdA and AKR/JS strains showed a reintegration pattern of AKR-MLV sequences unique for the individual animal, suggesting a monoclonal origin for the outgrown tumors. An analysis of tumor DNAs from the AKR/FuRdA and AKR/JS substrains with restriction enzymes cleaving within the proviral genome revealed a new EcoRI restriction site and BamHI restriction site not present in normal tissues. The positions of these sites corresponded both with cleavage sites of EcoRI and BamHI in integrated Moloney recombinants and with the structure of isolated AKR mink cell focus-forming viruses. All tumors analyzed to data contain nearly identical integrated recombinant genomes, suggesting a causal relationship between the formation of recombinants and the leukemogenic process. Images PMID:6268802

  4. Legal Agreements and the Governance of Research Commons: Lessons from Materials Sharing in Mouse Genomics

    PubMed Central

    Mishra, Amrita

    2014-01-01

    Abstract Omics research infrastructure such as databases and bio-repositories requires effective governance to support pre-competitive research. Governance includes the use of legal agreements, such as Material Transfer Agreements (MTAs). We analyze the use of such agreements in the mouse research commons, including by two large-scale resource development projects: the International Knockout Mouse Consortium (IKMC) and International Mouse Phenotyping Consortium (IMPC). We combine an analysis of legal agreements and semi-structured interviews with 87 members of the mouse model research community to examine legal agreements in four contexts: (1) between researchers; (2) deposit into repositories; (3) distribution by repositories; and (4) exchanges between repositories, especially those that are consortium members of the IKMC and IMPC. We conclude that legal agreements for the deposit and distribution of research reagents should be kept as simple and standard as possible, especially when minimal enforcement capacity and resources exist. Simple and standardized legal agreements reduce transactional bottlenecks and facilitate the creation of a vibrant and sustainable research commons, supported by repositories and databases. PMID:24552652

  5. Proteomic Analysis of Propiconazole Responses in Mouse Liver-Comparison of Genomic and Proteomic Profiles

    EPA Science Inventory

    We have performed for the first time a comprehensive profiling of changes in protein expression of soluble proteins in livers from mice treated with the mouse liver tumorigen, propiconazole, to uncover the pathways and networks altered by this commonly used fungicide. Utilizing t...

  6. Proteomic analysis of propiconazole responses in mouse liver: comparison of genomic and proteomic profiles

    EPA Science Inventory

    We have performed for the first time a comprehensive profiling of changes in protein expression of soluble proteins in livers from mice treated with the mouse liver tumorigen, propiconazole, to uncover the pathways and networks altered by this fungicide. Utilizing twodimensional...

  7. Functionally Charged Polystyrene Particles Activate Immortalized Mouse Microglia (BV2): Cellular and Genomic Response

    EPA Science Inventory

    The effect of particle surface charge on the biological activation of immortalized mouse microglia (BV2) was examined. Same size (~850-950 nm) spherical polystyrene microparticles (SPM) with net negative (carboxyl, COOH-) or positive (dimethyl amino, CH3)2

  8. Understanding the physics of oligonucleotide microarrays: the Affymetrix spike-in data reanalysed

    NASA Astrophysics Data System (ADS)

    Burden, Conrad J.

    2008-03-01

    The Affymetrix U95 and U133 Latin-Square spike-in datasets are reanalysed, together with a dataset from a version of the U95 spike-in experiment without a complex non-specific background. The approach uses a physico-chemical model which includes the effects of the specific and non-specific hybridization and probe folding at the microarray surface, target folding and hybridization in the bulk RNA target solution and duplex dissociation during the post-hybridization washing phase. The model predicts a three-parameter hyperbolic response function that fits well with fluorescence intensity data from all the three datasets. The importance of the various hybridization and washing effects in determining each of the three parameters is examined, and some guidance is given as to how a practical algorithm for determining specific target concentrations might be developed.

  9. MAAMD: a workflow to standardize meta-analyses and comparison of affymetrix microarray data

    PubMed Central

    2014-01-01

    Background Mandatory deposit of raw microarray data files for public access, prior to study publication, provides significant opportunities to conduct new bioinformatics analyses within and across multiple datasets. Analysis of raw microarray data files (e.g. Affymetrix CEL files) can be time consuming, complex, and requires fundamental computational and bioinformatics skills. The development of analytical workflows to automate these tasks simplifies the processing of, improves the efficiency of, and serves to standardize multiple and sequential analyses. Once installed, workflows facilitate the tedious steps required to run rapid intra- and inter-dataset comparisons. Results We developed a workflow to facilitate and standardize Meta-Analysis of Affymetrix Microarray Data analysis (MAAMD) in Kepler. Two freely available stand-alone software tools, R and AltAnalyze were embedded in MAAMD. The inputs of MAAMD are user-editable csv files, which contain sample information and parameters describing the locations of input files and required tools. MAAMD was tested by analyzing 4 different GEO datasets from mice and drosophila. MAAMD automates data downloading, data organization, data quality control assesment, differential gene expression analysis, clustering analysis, pathway visualization, gene-set enrichment analysis, and cross-species orthologous-gene comparisons. MAAMD was utilized to identify gene orthologues responding to hypoxia or hyperoxia in both mice and drosophila. The entire set of analyses for 4 datasets (34 total microarrays) finished in ~ one hour. Conclusions MAAMD saves time, minimizes the required computer skills, and offers a standardized procedure for users to analyze microarray datasets and make new intra- and inter-dataset comparisons. PMID:24621103

  10. Creation of targeted genomic deletions using TALEN or CRISPR/Cas nuclease pairs in one-cell mouse embryos

    PubMed Central

    Brandl, Christina; Ortiz, Oskar; Röttig, Bernhard; Wefers, Benedikt; Wurst, Wolfgang; Kühn, Ralf

    2014-01-01

    The use of TALEN and CRISPR/CAS nucleases is becoming increasingly popular as a means to edit single target sites in one-cell mouse embryos. Nevertheless, an area that has received less attention concerns the engineering of structural genome variants and the necessary religation of two distant double-strand breaks. Herein, we applied pairs of TALEN or sgRNAs and Cas9 to create deletions in the Rab38 gene. We found that the deletion of 3.2 or 9.3 kb, but not of 30 kb, occurs at a frequency of 6–37%. This is sufficient for the direct production of mutants by embryo microinjection. Therefore, deletions up to ∼10 kb can be readily achieved for modeling human disease alleles. This work represents an important step towards the establishment of new protocols that support the ligation of remote DSB ends to achieve even larger rearrangements. PMID:25685662

  11. Genome-wide mapping in a house mouse hybrid zone reveals hybrid sterility loci and Dobzhansky-Muller interactions.

    PubMed

    Turner, Leslie M; Harr, Bettina

    2014-12-09

    Mapping hybrid defects in contact zones between incipient species can identify genomic regions contributing to reproductive isolation and reveal genetic mechanisms of speciation. The house mouse features a rare combination of sophisticated genetic tools and natural hybrid zones between subspecies. Male hybrids often show reduced fertility, a common reproductive barrier between incipient species. Laboratory crosses have identified sterility loci, but each encompasses hundreds of genes. We map genetic determinants of testis weight and testis gene expression using offspring of mice captured in a hybrid zone between M. musculus musculus and M. m. domesticus. Many generations of admixture enables high-resolution mapping of loci contributing to these sterility-related phenotypes. We identify complex interactions among sterility loci, suggesting multiple, non-independent genetic incompatibilities contribute to barriers to gene flow in the hybrid zone.

  12. Genome-wide mapping in a house mouse hybrid zone reveals hybrid sterility loci and Dobzhansky-Muller interactions

    PubMed Central

    Turner, Leslie M; Harr, Bettina

    2014-01-01

    Mapping hybrid defects in contact zones between incipient species can identify genomic regions contributing to reproductive isolation and reveal genetic mechanisms of speciation. The house mouse features a rare combination of sophisticated genetic tools and natural hybrid zones between subspecies. Male hybrids often show reduced fertility, a common reproductive barrier between incipient species. Laboratory crosses have identified sterility loci, but each encompasses hundreds of genes. We map genetic determinants of testis weight and testis gene expression using offspring of mice captured in a hybrid zone between M. musculus musculus and M. m. domesticus. Many generations of admixture enables high-resolution mapping of loci contributing to these sterility-related phenotypes. We identify complex interactions among sterility loci, suggesting multiple, non-independent genetic incompatibilities contribute to barriers to gene flow in the hybrid zone. DOI: http://dx.doi.org/10.7554/eLife.02504.001 PMID:25487987

  13. Mouse genome-wide association study identifies polymorphisms on chromosomes 4, 11 and 15 for age-related cardiac fibrosis

    PubMed Central

    Li, Qiaoli; Berndt, Annerose; Sundberg, Beth A.; Silva, Kathleen A.; Kennedy, Victoria E.; Cario, Clinton L; Richardson, Matthew A.; Chase, Thomas H.; Schofield, Paul N.; Uitto, Jouni; Sundberg, John P.

    2017-01-01

    Dystrophic cardiac calcinosis (DCC), also called epicardial and myocardial fibrosis and mineralization, has been detected in mice of a number of laboratory inbred strains, most commonly C3H/HeJ and DBA/2J. In previous mouse breeding studies between these DCC susceptible and the DCC resistant strain C57BL/6J, 4 genetic loci harboring genes involved in DCC inheritance were identified and subsequently termed Dyscal loci 1 through 4. Here we report susceptibility to cardiac fibrosis, a sub-phenotype of DCC, at 12 and 20 months of age and close to natural death in a survey of 28 inbred mouse strains. Eight strains showed cardiac fibrosis with highest frequency and severity in the moribund mice. Using genotype and phenotype information of the 28 investigated strains we performed genome-wide association studies (GWAS) and identified the most significant associations on chromosome (Chr) 15 at 72 million base pairs (Mb) (P < 10−13) and Chr 4 at 122 Mb (P < 10−11) and 134 Mb (P < 10−7). At the Chr 15 locus Col22a1 and Kcnk9 were identified. Both have been reported to be morphologically and functionally important in the heart muscle. The strongest Chr 4 associations were located approximate 6 Mb away from the Dyscal 2 quantitative trait locus peak within the boundaries of the Extl1 gene and in close proximity to the Trim63 and Cap1 genes. In addition, a single nucleotide polymorphism association was found on chromosome 11. This study provides evidence for more than the previously reported 4 genetic loci determining cardiac fibrosis and DCC. The study also highlights the power of GWAS in the mouse for dissecting complex genetic traits. PMID:27126641

  14. Characterization of the genomic structure of the mouse APLP1 gene

    SciTech Connect

    Zhong, Sue; Wu, Kuo; Black, I.B.; Schaar, D.G.

    1996-02-15

    This article reports on the organization of the mouse APLP1 gene, an evolutionarily conserved amyloid precursor-like protein. The amyloid beta protein, important in Alzheimer diseases, is derived from these precursor proteins. By investigating the expression and structure of this murine gene, it is hoped that more will be learned about the function and regulation of the human homologue. 27 refs., 2 figs.

  15. Application of oocyte cryopreservation technology in TALEN-mediated mouse genome editing.

    PubMed

    Nakagawa, Yoshiko; Sakuma, Tetsushi; Nakagata, Naomi; Yamasaki, Sho; Takeda, Naoki; Ohmuraya, Masaki; Yamamoto, Takashi

    2014-01-01

    Reproductive engineering techniques, such as in vitro fertilization (IVF) and cryopreservation of embryos or spermatozoa, are essential for preservation, reproduction, and transportation of genetically engineered mice. However, it has not yet been elucidated whether these techniques can be applied for the generation of genome-edited mice using engineered nucleases such as transcription activator-like effector nucleases (TALENs). Here, we demonstrate the usefulness of frozen oocytes fertilized in vitro using frozen sperm for TALEN-mediated genome editing in mice. We examined side-by-side comparisons concerning sperm (fresh vs. frozen), fertilization method (mating vs. IVF), and fertilized oocytes (fresh vs. frozen) for the source of oocytes used for TALEN injection; we found that fertilized oocytes created under all tested conditions were applicable for TALEN-mediated mutagenesis. In addition, we investigated whether the ages in weeks of parental female mice can affect the efficiency of gene modification, by comparing 5-week-old and 8-12-week-old mice as the source of oocytes used for TALEN injection. The genome editing efficiency of an endogenous gene was consistently 95-100% when either 5-week-old or 8-12-week-old mice were used with or without freezing the oocytes. Thus, our report describes the availability of freeze-thawed oocytes and oocytes from female mice at various weeks of age for TALEN-mediated genome editing, thus boosting the convenience of such innovative gene targeting strategies.

  16. Chromosomal evolution of Arvicolinae (Cricetidae, Rodentia). I. The genome homology of tundra vole, field vole, mouse and golden hamster revealed by comparative chromosome painting.

    PubMed

    Sitnikova, Natalia A; Romanenko, Svetlana A; O'Brien, Patricia C M; Perelman, Polina L; Fu, Beiyuan; Rubtsova, Nadezhda V; Serdukova, Natalya A; Golenishchev, Feodor N; Trifonov, Vladimir A; Ferguson-Smith, Malcolm A; Yang, Fengtang; Graphodatsky, Alexander S

    2007-01-01

    Cross-species chromosome painting has become the mainstay of comparative cytogenetic and chromosome evolution studies. Here we have made a set of chromosomal painting probes for the field vole (Microtus agrestis) by DOP-PCR amplification of flow-sorted chromosomes. Together with painting probes of golden hamster (Mesocricetus auratus) and mouse (Mus musculus), the field vole probes have been hybridized onto the metaphases of the tundra vole (Microtus oeconomus). A comparative chromosome map between these two voles, golden hamster and mouse has been established based on the results of cross-species chromosome painting and G-banding comparisons. The sets of paints from the field vole, golden hamster and mouse identified a total of 27, 40 and 47 homologous autosomal regions, respectively, in the genome of tundra vole; 16, 41 and 51 fusion/fission rearrangements differentiate the karyotype of the tundra vole from the karyotypes of the field vole, golden hamster and mouse, respectively.

  17. Genome-Wide and Species-Wide In Silico Screening for Intragenic MicroRNAs in Human, Mouse and Chicken

    PubMed Central

    Godnic, Irena; Zorc, Minja; Jevsinek Skok, Dasa; Calin, George Adrian; Horvat, Simon; Dovc, Peter; Kovac, Milena; Kunej, Tanja

    2013-01-01

    MicroRNAs (miRNAs) are non-coding RNAs (ncRNAs) involved in regulation of gene expression. Intragenic miRNAs, especially those exhibiting a high degree of evolutionary conservation, have been shown to be coordinately regulated and/or expressed with their host genes, either with synergistic or antagonistic correlation patterns. However, the degree of cross-species conservation of miRNA/host gene co-location is not known and co-expression information is incomplete and fragmented among several studies. Using the genomic resources (miRBase and Ensembl) we performed a genome-wide in silico screening (GWISS) for miRNA/host gene pairs in three well-annotated vertebrate species: human, mouse, and chicken. Approximately half of currently annotated miRNA genes resided within host genes: 53.0% (849/1,600) in human, 48.8% (418/855) in mouse, and 42.0% (210/499) in chicken, which we present in a central publicly available Catalog of intragenic miRNAs (http://www.integratomics-time.com/miR-host/catalog). The miRNA genes resided within either protein-coding or ncRNA genes, which include long intergenic ncRNAs (lincRNAs) and small nucleolar RNAs (snoRNAs). Twenty-seven miRNA genes were found to be located within the same host genes in all three species and the data integration from literature and databases showed that most (26/27) have been found to be co-expressed. Particularly interesting are miRNA genes located within genes encoding for miRNA silencing machinery (DGCR8, DICER1, and SND1 in human and Cnot3, Gdcr8, Eif4e, Tnrc6b, and Xpo5 in mouse). We furthermore discuss a potential for phenotype misattribution of miRNA host gene polymorphism or gene modification studies due to possible collateral effects on miRNAs hosted within them. In conclusion, the catalog of intragenic miRNAs and identified 27 miRNA/host gene pairs with cross-species conserved co-location, co-expression, and potential co-regulation, provide excellent candidates for further functional annotation of

  18. Experimental validation of the regulated expression of large numbers of non-coding RNAs from the mouse genome

    PubMed Central

    Ravasi, Timothy; Suzuki, Harukazu; Pang, Ken C.; Katayama, Shintaro; Furuno, Masaaki; Okunishi, Rie; Fukuda, Shiro; Ru, Kelin; Frith, Martin C.; Gongora, M. Milena; Grimmond, Sean M.; Hume, David A.; Hayashizaki, Yoshihide; Mattick, John S.

    2006-01-01

    Recent large-scale analyses of mainly full-length cDNA libraries generated from a variety of mouse tissues indicated that almost half of all representative cloned sequences did not contain an apparent protein-coding sequence, and were putatively derived from non-protein-coding RNA (ncRNA) genes. However, many of these clones were singletons and the majority were unspliced, raising the possibility that they may be derived from genomic DNA or unprocessed pre-mRNA contamination during library construction, or alternatively represent nonspecific “transcriptional noise.” Here we show, using reverse transcriptase-dependent PCR, microarray, and Northern blot analyses, that many of these clones were derived from genuine transcripts of unknown function whose expression appears to be regulated. The ncRNA transcripts have larger exons and fewer introns than protein-coding transcripts. Analysis of the genomic landscape around these sequences indicates that some cDNA clones were produced not from terminal poly(A) tracts but internal priming sites within longer transcripts, only a minority of which is encompassed by known genes. A significant proportion of these transcripts exhibit tissue-specific expression patterns, as well as dynamic changes in their expression in macrophages following lipopolysaccharide stimulation. Taken together, the data provide strong support for the conclusion that ncRNAs are an important, regulated component of the mammalian transcriptome. PMID:16344565

  19. Whole Genome Expression Analysis in a Mouse Model of Tauopathy Identifies MECP2 as a Possible Regulator of Tau Pathology

    PubMed Central

    Maphis, Nicole M.; Jiang, Shanya; Binder, Jessica; Wright, Carrie; Gopalan, Banu; Lamb, Bruce T.; Bhaskar, Kiran

    2017-01-01

    Increasing evidence suggests that hyperphosphorylation and aggregation of microtubule-associated protein tau (MAPT or tau) correlates with the development of cognitive impairment in Alzheimer’s disease (AD) and related tauopathies. While numerous attempts have been made to model AD-relevant tau pathology in various animal models, there has been very limited success for these models to fully recapitulate the progression of disease as seen in human tauopathies. Here, we performed whole genome gene expression in a genomic mouse model of tauopathy that expressed human MAPT gene under the control of endogenous human MAPT promoter and also were complete knockout for endogenous mouse tau [referred to as ‘hTauMaptKO(Duke)′ mice]. First, whole genome expression analysis revealed 64 genes, which were differentially expressed (32 up-regulated and 32 down-regulated) in the hippocampus of 6-month-old hTauMaptKO(Duke) mice compared to age-matched non-transgenic controls. Genes relevant to neuronal function or neurological disease include up-regulated genes: PKC-alpha (Prkca), MECP2 (Mecp2), STRN4 (Strn4), SLC40a1 (Slc40a1), POLD2 (Pold2), PCSK2 (Pcsk2), and down-regulated genes: KRT12 (Krt12), LASS1 (Cers1), PLAT (Plat), and NRXN1 (Nrxn1). Second, network analysis suggested anatomical structure development, cellular metabolic process, cell death, signal transduction, and stress response were significantly altered biological processes in the hTauMaptKO(Duke) mice as compared to age-matched non-transgenic controls. Further characterization of a sub-group of significantly altered genes revealed elevated phosphorylation of MECP2 (methyl-CpG-binding protein-2), which binds to methylated CpGs and associates with chromatin, in hTauMaptKO(Duke) mice compared to age-matched controls. Third, phoshpho-MECP2 was elevated in autopsy brain samples from human AD compared to healthy controls. Finally, siRNA-mediated knockdown of MECP2 in human tau expressing N2a cells resulted in a

  20. Reselection of a genomic upstream open reading frame in mouse hepatitis coronavirus 5'-untranslated-region mutants.

    PubMed

    Wu, Hung-Yi; Guan, Bo-Jhih; Su, Yu-Pin; Fan, Yi-Hsin; Brian, David A

    2014-01-01

    An AUG-initiated upstream open reading frame (uORF) encoding a potential polypeptide of 3 to 13 amino acids (aa) is found within the 5' untranslated region (UTR) of >75% of coronavirus genomes based on 38 reference strains. Potential CUG-initiated uORFs are also found in many strains. The AUG-initiated uORF is presumably translated following genomic 5'-end cap-dependent ribosomal scanning, but its function is unknown. Here, in a reverse-genetics study with mouse hepatitis coronavirus, the following were observed. (i) When the uORF AUG-initiating codon was replaced with a UAG stop codon along with a U112A mutation to maintain a uORF-harboring stem-loop 4 structure, an unimpaired virus with wild-type (WT) growth kinetics was recovered. However, reversion was found at all mutated sites within five virus passages. (ii) When the uORF was fused with genomic (main) ORF1 by converting three in-frame stop codons to nonstop codons, a uORF-ORF1 fusion protein was made, and virus replicated at WT levels. However, a frameshifting G insertion at virus passage 7 established a slightly 5'-extended original uORF. (iii) When uAUG-eliminating deletions of 20, 30, or 51 nucleotides (nt) were made within stem-loop 4, viable but debilitated virus was recovered. However, a C80U mutation in the first mutant and an A77G mutation in the second appeared by passage 10, which generated alternate uORFs that correlated with restored WT growth kinetics. In vitro, the uORF-disrupting nondeletion mutants showed enhanced translation of the downstream ORF1 compared with the WT. These results together suggest that the uORF represses ORF1 translation yet plays a beneficial but nonessential role in coronavirus replication in cell culture.

  1. Mouse scrapie responsive gene 1 (Scrg1): genomic organization, physical linkage to sap30, genetic mapping on chromosome 8, and expression in neuronal primary cell cultures.

    PubMed

    Dron, M; Tartare, X; Guillo, F; Haik, S; Barbin, G; Maury, C; Tovey, M; Dandoy-Dron, F

    2000-11-15

    We have previously reported a transcript of a novel mouse gene (Scrg1) with increased expression in transmissible spongiform encephalopathies and the cloning of the human mRNA analogue. In this paper, we present the genomic organization of the mouse and human SCRG1 loci, which exhibit a high degree of conservation. The genes are composed of three exons; the two downstream exons contain the protein coding region. The mouse gene is expressed in brain tissue essentially as a 0.7-kb message but also as a minor 2.6-kb mRNA. We have sequenced 20 kb of DNA at the mouse Scrg1 locus and found that the longer transcript is the prolongation of the 0.7-kb mRNA to a polyadenylation site located about 2 kb further downstream. Sequencing revealed that the mouse Scrg1 gene is physically linked to Sap30, a gene that encodes a protein of the histone deacetylase complex, and genetic linkage mapping assigned the localization of Scrg1 to chromosome 8 between Ant1 and Hmg2. Northern blot analysis showed that Scrg1 is under strict developmental control in mouse embryo and is expressed by cells of neuronal origin in vitro. Comparison of the rat, mouse, and human SCRG1 proteins identified a box of 35 identical contiguous amino acids and a characteristic cysteine distribution pattern defining a new protein signature.

  2. A gene expression resource generated by genome-wide lacZ profiling in the mouse

    PubMed Central

    Tuck, Elizabeth; Estabel, Jeanne; Oellrich, Anika; Maguire, Anna Karin; Adissu, Hibret A.; Souter, Luke; Siragher, Emma; Lillistone, Charlotte; Green, Angela L.; Wardle-Jones, Hannah; Carragher, Damian M.; Karp, Natasha A.; Smedley, Damian; Adams, Niels C.; Bussell, James N.; Adams, David J.; Ramírez-Solis, Ramiro; Steel, Karen P.; Galli, Antonella; White, Jacqueline K.

    2015-01-01

    ABSTRACT Knowledge of the expression profile of a gene is a critical piece of information required to build an understanding of the normal and essential functions of that gene and any role it may play in the development or progression of disease. High-throughput, large-scale efforts are on-going internationally to characterise reporter-tagged knockout mouse lines. As part of that effort, we report an open access adult mouse expression resource, in which the expression profile of 424 genes has been assessed in up to 47 different organs, tissues and sub-structures using a lacZ reporter gene. Many specific and informative expression patterns were noted. Expression was most commonly observed in the testis and brain and was most restricted in white adipose tissue and mammary gland. Over half of the assessed genes presented with an absent or localised expression pattern (categorised as 0-10 positive structures). A link between complexity of expression profile and viability of homozygous null animals was observed; inactivation of genes expressed in ≥21 structures was more likely to result in reduced viability by postnatal day 14 compared with more restricted expression profiles. For validation purposes, this mouse expression resource was compared with Bgee, a federated composite of RNA-based expression data sets. Strong agreement was observed, indicating a high degree of specificity in our data. Furthermore, there were 1207 observations of expression of a particular gene in an anatomical structure where Bgee had no data, indicating a large amount of novelty in our data set. Examples of expression data corroborating and extending genotype-phenotype associations and supporting disease gene candidacy are presented to demonstrate the potential of this powerful resource. PMID:26398943

  3. Genetic‐Genomic Replication to Identify Candidate Mouse Atherosclerosis Modifier Genes

    PubMed Central

    Hsu, Jeffrey; Smith, Jonathan D.

    2013-01-01

    Objective Genetics plays a large role in atherosclerosis susceptibility in humans and mice. We attempted to confirm previously determined mouse atherosclerosis‐associated loci and use bioinformatics and transcriptomics to create a catalog of candidate atherosclerosis modifier genes at these loci. Methods and Results A strain intercross was performed between AKR and DBA/2 mice on the apoE−/− background generating 166 F2 progeny. Using the phenotype log10 of the aortic root lesion area, we identified 3 suggestive atherosclerosis quantitative trait loci (Ath QTLs). When combined with our prior strain intercross, we confirmed 3 significant Ath QTLs on chromosomes 2, 15, and 17, with combined logarithm of odds scores of 5.9, 5.3, and 5.6, respectively, which each met the genome‐wide 5% false discovery rate threshold. We identified all of the protein coding differences between these 2 mouse strains within the Ath QTL intervals. Microarray gene expression profiling was performed on macrophages and endothelial cells from this intercross to identify expression QTLs (eQTLs), the loci that are associated with variation in the expression levels of specific transcripts. Cross tissue eQTLs and macrophage eQTLs that replicated from a prior strain intercross were identified. These bioinformatic and eQTL analyses produced a comprehensive list of candidate genes that may be responsible for the Ath QTLs. Conclusions Replication studies for clinical traits as well as gene expression traits are worthwhile in identifying true versus false genetic associations. We have replicated 3 loci on mouse chromosomes 2, 15, and 17 that are associated with atherosclerosis. We have also identified protein coding differences and multiple replicated eQTLs, which may be useful in the identification of atherosclerosis modifier genes. PMID:23525445

  4. Estimation of mutation induction rates in AT-rich sequences using a genome scanning approach after X irradiation of mouse spermatogonia.

    PubMed

    Asakawa, Jun-ichi; Nakamura, Nori; Katayama, Hiroaki; Cullings, Harry M

    2007-08-01

    We have previously used NotI as the marker enzyme (recognizing GCGGCCGC) in a genome scanning approach for detection of mutations induced in mouse spermatogonia and estimated the mutation induction rate as about 0.7 x 10(-5) per locus per Gy. To see whether different parts of the genome have different sensitivities for mutation induction, we used AflII (recognizing CTTAAG) as the marker enzyme in the present study. After the screening of 1,120 spots in each mouse offspring, we found five mutations among 92,655 spots from the unirradiated paternal genome, five mutations among 218,411 spots from the unirradiated maternal genome, and 13 mutations among 92,789 spots from 5 Gy-exposed paternal genome. Among the 23 mutations, 11 involved mouse satellite DNA sequences (AT-rich), and the remaining 12 mutations also involved AT-rich but non-satellite sequences. Both types of sequences were found as multiple, similar-sequence blocks in the genome. Counting each member of cluster mutations separately and excluding results on one hypermutable spot, the spontaneous mutation rates were estimated as 3.2 (+/- 1.9) x 10(-5) and 2.3 (+/- 1.0) x 10(-5) per locus per generation in the male and female genomes, respectively, and the mutation induction rate as 1.1 (+/- 1.2) x 10(-5) per locus per Gy. The induction rate would be reduced to 0.9 x 10(-5) per locus per Gy if satellite sequence mutations were excluded from this analysis. The results indicate that mutation induction rates do not largely differ between GC-rich and AT-rich regions: 1 x 10(-5) per locus per Gy or less, which is close to 1.08 x 10(-5) per locus per Gy, the current estimate for the mean mutation induction rate in mice.

  5. Concerted copy number variation balances ribosomal DNA dosage in human and mouse genomes

    PubMed Central

    Gibbons, John G.; Branco, Alan T.; Godinho, Susana A.; Yu, Shoukai; Lemos, Bernardo

    2015-01-01

    Tandemly repeated ribosomal DNA (rDNA) arrays are among the most evolutionary dynamic loci of eukaryotic genomes. The loci code for essential cellular components, yet exhibit extensive copy number (CN) variation within and between species. CN might be partly determined by the requirement of dosage balance between the 5S and 45S rDNA arrays. The arrays are nonhomologous, physically unlinked in mammals, and encode functionally interdependent RNA components of the ribosome. Here we show that the 5S and 45S rDNA arrays exhibit concerted CN variation (cCNV). Despite 5S and 45S rDNA elements residing on different chromosomes and lacking sequence similarity, cCNV between these loci is strong, evolutionarily conserved in humans and mice, and manifested across individual genotypes in natural populations and pedigrees. Finally, we observe that bisphenol A induces rapid and parallel modulation of 5S and 45S rDNA CN. Our observations reveal a novel mode of genome variation, indicate that natural selection contributed to the evolution and conservation of cCNV, and support the hypothesis that 5S CN is partly determined by the requirement of dosage balance with the 45S rDNA array. We suggest that human disease variation might be traced to disrupted rDNA dosage balance in the genome. PMID:25583482

  6. Genomic analysis of sexual dimorphism of gene expression in the mouse adrenal gland.

    PubMed

    El Wakil, A; Mari, B; Barhanin, J; Lalli, E

    2013-11-01

    A relevant gender difference exists in adrenal physiology and propensity to disease. In mice, a remarkable sexual dimorphism is present in several components of the hypothalamic-pituitary-adrenal axis, with females displaying higher adrenal weight, plasma ACTH, corticosterone, and aldosterone levels than males. The molecular bases of this sexual dimorphism are little known. We have compared global gene expression profiles in males vs. female mouse adrenal glands and also studied the effect that testosterone treatment and castration have on adrenal gene expression in female vs. male mice, respectively. Our study evidenced a set of 71 genes that are coordinately modulated according to sex and hormonal treatments and represent the core sexually dimorphic expression program in the mouse adrenal gland. Moreover, we show that some genes involved in steroid metabolism have a remarkable sexual dimorphic expression and identify new potential markers for the adrenal X-zone, a transitory cellular layer in the inner adrenal cortex, which spontaneously regresses at puberty in males and during the first pregnancy in females and has an uncertain physiological role. Finally, sexually dimorphic expression of the transcriptional regulators Nr5a1 and Nr0b1 may explain at least in part the differences in adrenal steroidogenesis between sexes.

  7. Comparison of gene coverage of mouse oligonucleotide microarray platforms

    PubMed Central

    Verdugo, Ricardo A; Medrano, Juan F

    2006-01-01

    Background The increasing use of DNA microarrays for genetical genomics studies generates a need for platforms with complete coverage of the genome. We have compared the effective gene coverage in the mouse genome of different commercial and noncommercial oligonucleotide microarray platforms by performing an in-house gene annotation of probes. We only used information about probes that is available from vendors and followed a process that any researcher may take to find the gene targeted by a given probe. In order to make consistent comparisons between platforms, probes in each microarray were annotated with an Entrez Gene id and the chromosomal position for each gene was obtained from the UCSC Genome Browser Database. Gene coverage was estimated as the percentage of Entrez Genes with a unique position in the UCSC Genome database that is tested by a given microarray platform. Results A MySQL relational database was created to store the mapping information for 25,416 mouse genes and for the probes in five microarray platforms (gene coverage level in parenthesis): Affymetrix430 2.0 (75.6%), ABI Genome Survey (81.24%), Agilent (79.33%), Codelink (78.09%), Sentrix (90.47%); and four array-ready oligosets: Sigma (47.95%), Operon v.3 (69.89%), Operon v.4 (84.03%), and MEEBO (84.03%). The differences in coverage between platforms were highly conserved across chromosomes. Differences in the number of redundant and unspecific probes were also found among arrays. The database can be queried to compare specific genomic regions using a web interface. The software used to create, update and query the database is freely available as a toolbox named ArrayGene. Conclusion The software developed here allows researchers to create updated custom databases by using public or proprietary information on genes for any organisms. ArrayGene allows easy comparisons of gene coverage between microarray platforms for any region of the genome. The comparison presented here reveals that the

  8. A Single-Array-Based Method for Detecting Copy Number Variants Using Affymetrix High Density SNP Arrays and its Application to Breast Cancer

    PubMed Central

    Li, Ming; Wen, Yalu; Fu, Wenjiang

    2014-01-01

    Cumulative evidence has shown that structural variations, due to insertions, deletions, and inversions of DNA, may contribute considerably to the development of complex human diseases, such as breast cancer. High-throughput genotyping technologies, such as Affymetrix high density single-nucleotide polymorphism (SNP) arrays, have produced large amounts of genetic data for genome-wide SNP genotype calling and copy number estimation. Meanwhile, there is a great need for accurate and efficient statistical methods to detect copy number variants. In this article, we introduce a hidden-Markov-model (HMM)-based method, referred to as the PICR-CNV, for copy number inference. The proposed method first estimates copy number abundance for each single SNP on a single array based on the raw fluorescence values, and then standardizes the estimated copy number abundance to achieve equal footing among multiple arrays. This method requires no between-array normalization, and thus, maintains data integrity and independence of samples among individual subjects. In addition to our efforts to apply new statistical technology to raw fluorescence values, the HMM has been applied to the standardized copy number abundance in order to reduce experimental noise. Through simulations, we show our refined method is able to infer copy number variants accurately. Application of the proposed method to a breast cancer dataset helps to identify genomic regions significantly associated with the disease. PMID:26279618

  9. Establishment of paternal genomic imprinting in mouse prospermatogonia analyzed by nuclear transfer.

    PubMed

    Kamimura, Satoshi; Hatanaka, Yuki; Hirasawa, Ryutaro; Matsumoto, Kazuya; Oikawa, Mami; Lee, Jiyoung; Matoba, Shogo; Mizutani, Eiji; Ogonuki, Narumi; Inoue, Kimiko; Kohda, Takashi; Ishino, Fumitoshi; Ogura, Atsuo

    2014-11-01

    In mice, the establishment of paternal genomic imprinting in male germ cells starts at midgestation, as suggested by DNA methylation analyses of differentially methylated regions (DMRs). However, this information is based on averages from mixed populations of germ cells, and the DNA methylation pattern might not always provide a full representation of imprinting status. To obtain more detailed information on the establishment of paternal imprinting, single prospermatogonia at Embryonic Days 15.5 (E15.5), E16.5, and E17.5 and at Day 0.5 after birth were cloned using nuclear transfer; previous reports suggested that cloned embryos reflected the donor's genomic imprinting status. Then, the resultant fetuses (E9.5) were analyzed for the DNA methylation pattern of three paternal DMRs (IG-DMR, H19 DMR, and Rasgrf1 DMR) and the expression pattern of imprinted genes therein. The overall data indicated that establishment of genomic imprinting in all paternally imprinted regions was completed by E17.5, following a short intermediate period at E16.5. Furthermore, comparison between the methylation status of DMRs and the expression profiles of imprinted genes suggested that methylation of the IG-DMR, but not the H19 DMR, solely governed the control of its imprinted gene cluster. The Rasgrf1 DMR seemed to be imprinted later than the other two genes. We also found that the methylation status of the Gtl2 DMR, the secondary DMR that acquires DNA methylation after fertilization, was likely to follow the methylation status of the upstream IG-DMR. Thus, the systematic analyses of prospermatogonium-derived embryos provided additional important information on the establishment of paternal imprinting.

  10. Identifying the impact of G-quadruplexes on Affymetrix 3' arrays using cloud computing.

    PubMed

    Memon, Farhat N; Owen, Anne M; Sanchez-Graillet, Olivia; Upton, Graham J G; Harrison, Andrew P

    2010-01-15

    A tetramer quadruplex structure is formed by four parallel strands of DNA/ RNA containing runs of guanine. These quadruplexes are able to form because guanine can Hoogsteen hydrogen bond to other guanines, and a tetrad of guanines can form a stable arrangement. Recently we have discovered that probes on Affymetrix GeneChips that contain runs of guanine do not measure gene expression reliably. We associate this finding with the likelihood that quadruplexes are forming on the surface of GeneChips. In order to cope with the rapidly expanding size of GeneChip array datasets in the public domain, we are exploring the use of cloud computing to replicate our experiments on 3' arrays to look at the effect of the location of G-spots (runs of guanines). Cloud computing is a recently introduced high-performance solution that takes advantage of the computational infrastructure of large organisations such as Amazon and Google. We expect that cloud computing will become widely adopted because it enables bioinformaticians to avoid capital expenditure on expensive computing resources and to only pay a cloud computing provider for what is used. Moreover, as well as financial efficiency, cloud computing is an ecologically-friendly technology, it enables efficient data-sharing and we expect it to be faster for development purposes. Here we propose the advantageous use of cloud computing to perform a large data-mining analysis of public domain 3' arrays.

  11. The Affymetrix DMET Plus Platform Reveals Unique Distribution of ADME-Related Variants in Ethnic Arabs

    PubMed Central

    Wakil, Salma M.; Nguyen, Cao; Muiya, Nzioka P.; Andres, Editha; Lykowska-Tarnowska, Agnieszka; Baz, Batoul; Meyer, Brian F.; Morahan, Grant

    2015-01-01

    Background. The Affymetrix Drug Metabolizing Enzymes and Transporters (DMET) Plus Premier Pack has been designed to genotype 1936 gene variants thought to be essential for screening patients in personalized drug therapy. These variants include the cytochrome P450s (CYP450s), the key metabolizing enzymes, many other enzymes involved in phase I and phase II pharmacokinetic reactions, and signaling mediators associated with variability in clinical response to numerous drugs not only among individuals, but also between ethnic populations. Materials and Methods. We genotyped 600 Saudi individuals for 1936 variants on the DMET platform to evaluate their clinical potential in personalized medicine in ethnic Arabs. Results. Approximately 49% each of the 437 CYP450 variants, 56% of the 581 transporters, 56% of 419 transferases, 48% of the 104 dehydrogenases, and 58% of the remaining 390 variants were detected. Several variants, such as rs3740071, rs6193, rs258751, rs6199, rs11568421, and rs8187797, exhibited significantly either higher or lower minor allele frequencies (MAFs) than those in other ethnic groups. Discussion. The present study revealed some unique distribution trends for several variants in Arabs, which displayed partly inverse allelic prevalence compared to other ethnic populations. The results point therefore to the need to verify and ascertain the prevalence of a variant as a prerequisite for engaging it in clinical routine screening in personalized medicine in any given population. PMID:25802476

  12. Polycomb repressive complex PRC1 spatially constrains the mouse embryonic stem cell genome

    PubMed Central

    Mifsud, Borbala; Dimitrova, Emilia; Matheson, Louise; Tavares-Cadete, Filipe; Furlan-Magaril, Mayra; Segonds-Pichon, Anne; Jurkowski, Wiktor; Wingett, Steven W.; Tabbada, Kristina; Andrews, Simon; Herman, Bram; LeProust, Emily; Osborne, Cameron S.; Koseki, Haruhiko; Fraser, Peter; Luscombe, Nicholas M.; Elderkin, Sarah

    2016-01-01

    The Polycomb Repressive Complexes PRC1 and PRC2 maintain embryonic stem cell (ESC) pluripotency by silencing lineage-specifying developmental regulator genes1. Emerging evidence suggests that Polycomb complexes act through controlling spatial genome organisation2–9. We show that PRC1 functions as a master regulator of ESC genome architecture by organizing genes in three-dimensional interaction networks. The strongest spatial network is composed of the four Hox clusters and early developmental transcription factor genes, the majority of which contact poised enhancers. Removal of Polycomb repression leads to disruption of promoter-promoter contacts in the Hox network. In contrast, promoter-enhancer contacts are maintained, accompanied by widespread acquisition of active chromatin signatures at network enhancers and pronounced transcriptional up-regulation of network genes. Thus, PRC1 physically constrains developmental transcription factor genes and their enhancers in a silenced but poised spatial network. We propose that selective release of genes from this spatial network underlies cell fate specification during early embryonic development. PMID:26323060

  13. Grape seed polyphenols and curcumin reduce genomic instability events in a transgenic mouse model for Alzheimer's disease.

    PubMed

    Thomas, Philip; Wang, Yan-Jiang; Zhong, Jin-Hua; Kosaraju, Shantha; O'Callaghan, Nathan J; Zhou, Xin-Fu; Fenech, Michael

    2009-02-10

    The study set out to determine (a) whether DNA damage is elevated in mice that carry mutations in the amyloid precursor protein (APP695swe) and presenilin 1 (PSEN1-dE9) that predispose to Alzheimer's disease (AD) relative to non-transgenic control mice, and (b) whether increasing the intake of dietary polyphenols from curcumin or grape seed extract could reduce genomic instability events in a transgenic mouse model for AD. DNA damage was measured using the micronucleus (MN) assay in both buccal mucosa and erythrocytes and an absolute telomere length assay for both buccal mucosa and olfactory bulb tissue. MN frequency tended to be higher in AD mice in both buccal mucosa (1.7-fold) and polychromatic erythrocytes (1.3-fold) relative to controls. Telomere length was significantly reduced by 91% (p=0.04) and non-significantly reduced by 50% in buccal mucosa and olfactory bulbs respectively in AD mice relative to controls. A significant 10-fold decrease in buccal MN frequency (p=0.01) was found for AD mice fed diets containing curcumin (CUR) or micro-encapsulated grape seed extract (MGSE) and a 7-fold decrease (p=0.02) for AD mice fed unencapsulated grape seed extract (GSE) compared to the AD group on control diet. Similarly, in polychromatic erythrocytes a significant reduction in MN frequency was found for the MGSE cohort (65.3%) (p<0.05), whereas the AD CUR and AD GSE groups were non-significantly reduced by 39.2 and 34.8% respectively compared to the AD Control. A non-significant 2-fold increase in buccal cell telomere length was evident for the CUR, GSE and MGSE groups compared to the AD control group. Olfactory bulb telomere length was found to be non-significantly 2-fold longer in mice fed on the CUR diet compared to controls. These results suggest potential protective effects of polyphenols against genomic instability events in different somatic tissues of a transgenic mouse model for AD.

  14. Systems analysis of primary Sjögren's syndrome pathogenesis in salivary glands identifies shared pathways in human and a mouse model

    PubMed Central

    2012-01-01

    Introduction Primary Sjögren's syndrome (pSS) is a chronic autoimmune disease with complex etiopathogenesis. Despite extensive studies to understand the disease process utilizing human and mouse models, the intersection between these species remains elusive. To address this gap, we utilized a novel systems biology approach to identify disease-related gene modules and signaling pathways that overlap between humans and mice. Methods Parotid gland tissues were harvested from 24 pSS and 16 non-pSS sicca patients and 25 controls. For mouse studies, salivary glands were harvested from C57BL/6.NOD-Aec1Aec2 mice at various times during development of pSS-like disease. RNA was analyzed with Affymetrix HG U133+2.0 arrays for human samples and with MOE430+2.0 arrays for mouse samples. The images were processed with Affymetrix software. Weighted-gene co-expression network analysis was used to identify disease-related and functional pathways. Results Nineteen co-expression modules were identified in human parotid tissue, of which four were significantly upregulated and three were downregulated in pSS patients compared with non-pSS sicca patients and controls. Notably, one of the human disease-related modules was highly preserved in the mouse model, and was enriched with genes involved in immune and inflammatory responses. Further comparison between these two species led to the identification of genes associated with leukocyte recruitment and germinal center formation. Conclusion Our systems biology analysis of genome-wide expression data from salivary gland tissue of pSS patients and from a pSS mouse model identified common dysregulated biological pathways and molecular targets underlying critical molecular alterations in pSS pathogenesis. PMID:23116360

  15. Utilization of a whole genome SNP panel for efficient genetic mapping in the mouse

    PubMed Central

    Moran, Jennifer L.; Bolton, Andrew D.; Tran, Pamela V.; Brown, Alison; Dwyer, Noelle D.; Manning, Danielle K.; Bjork, Bryan C.; Li, Cheng; Montgomery, Kate; Siepka, Sandra M.; Vitaterna, Martha Hotz; Takahashi, Joseph S.; Wiltshire, Tim; Kwiatkowski, David J.; Kucherlapati, Raju; Beier, David R.

    2006-01-01

    Phenotype-driven genetics can be used to create mouse models of human disease and birth defects. However, the utility of these mutant models is limited without identification of the causal gene. To facilitate genetic mapping, we developed a fixed single nucleotide polymorphism (SNP) panel of 394 SNPs as an alternative to analyses using simple sequence length polymorphism (SSLP) marker mapping. With the SNP panel, chromosomal locations for 22 monogenic mutants were identified. The average number of affected progeny genotyped for mapped monogenic mutations is nine. Map locations for several mutants have been obtained with as few as four affected progeny. The average size of genetic intervals obtained for these mutants is 43 Mb, with a range of 17–83 Mb. Thus, our SNP panel allows for identification of moderate resolution map position with small numbers of mice in a high-throughput manner. Importantly, the panel is suitable for mapping crosses from many inbred and wild-derived inbred strain combinations. The chromosomal localizations obtained with the SNP panel allow one to quickly distinguish between potentially novel loci or remutations in known genes, and facilitates fine mapping and positional cloning. By using this approach, we identified DNA sequence changes in two ethylnitrosourea-induced mutants. PMID:16461637

  16. Genetic and molecular characterization of genomic regions surrounding specific loci of the mouse

    SciTech Connect

    Russell, L.B.; Rinchik, E.M.

    1987-01-01

    Mutations detected by the mouse specific-locus test (SLT) include multilocus deletions as well as intragenic lesions. Genetic analyses have characterized sets of presumed overlapping deletions and have mapped previously unrecognized genes to the regions surrounding each of several specific loci. Molecular entry to one of these regions, d se, was achieved by utilizing a viral integration at, or near, a marker locus. Presumed deletions were shown to be, in fact deleted for DNA sequences, and the physical map was oriented relative to the earlier functional map. Presently, a random-clone approach is being used for initiating molecular characterization of regions, which, in aggregate, span a minimum of 9 cM. Mapping to subregions already identified by functional units will facilitate the generation of comprehensive molecular maps and the identification of numerous structure-function correlations for the regions. Results of the genetic and molecular analyses of multilocus deletions have enhanced the value of the SLT by adding qualitative to quantitative capabilities. Studies of the heterozygous effects of deletions (which are the predominant lesions induced by many mutagens) provide information important to assessment of genetic risk. Long deletions are, further, providing tools for targeted mutagenesis studies that will generate information on the number of loci within segments of defined length that are capable of mutating to detectable alleles, as well as providing new mutations important for strategies of refining molecular and functional maps. 28 refs., 2 tabs.

  17. Lentivirus-meditated frataxin gene delivery reverses genome instability in Friedreich ataxia patient and mouse model fibroblasts

    PubMed Central

    Khonsari, H; Schneider, M; Al-Mahdawi, S; Chianea, Y G; Themis, M; Parris, C; Pook, M A; Themis, M

    2016-01-01

    Friedreich ataxia (FRDA) is a progressive neurodegenerative disease caused by deficiency of frataxin protein, with the primary sites of pathology being the large sensory neurons of the dorsal root ganglia and the cerebellum. FRDA is also often accompanied by severe cardiomyopathy and diabetes mellitus. Frataxin is important in mitochondrial iron–sulfur cluster (ISC) biogenesis and low-frataxin expression is due to a GAA repeat expansion in intron 1 of the FXN gene. FRDA cells are genomically unstable, with increased levels of reactive oxygen species and sensitivity to oxidative stress. Here we report the identification of elevated levels of DNA double strand breaks (DSBs) in FRDA patient and YG8sR FRDA mouse model fibroblasts compared to normal fibroblasts. Using lentivirus FXN gene delivery to FRDA patient and YG8sR cells, we obtained long-term overexpression of FXN mRNA and frataxin protein levels with reduced DSB levels towards normal. Furthermore, γ-irradiation of FRDA patient and YG8sR cells revealed impaired DSB repair that was recovered on FXN gene transfer. This suggests that frataxin may be involved in DSB repair, either directly by an unknown mechanism, or indirectly via ISC biogenesis for DNA repair enzymes, which may be essential for the prevention of neurodegeneration. PMID:27518705

  18. Identification of Cis-regulatory elements in the mouse Pax9/Nkx2-9 genomic region: implication for evolutionary conserved synteny.

    PubMed Central

    Santagati, Fabio; Abe, Kuniya; Schmidt, Volker; Schmitt-John, Thomas; Suzuki, Misao; Yamamura, Ken-Ichi; Imai, Kenji

    2003-01-01

    We previously reported close physical linkage between Pax9 and Nkx2-9 in the human, mouse, and pufferfish (Fugu rubripes) genomes. In this study, we analyzed cis-regulatory elements of the two genes by comparative sequencing in the three species and by transgenesis in the mouse. We identified two regions including conserved noncoding sequences that possessed specific enhancer activities for expression of Pax9 in the medial nasal process and of Nkx2-9 in the ventral neural tube. Remarkably, the latter contained the consensus Gli-binding motif. Interestingly, the identified Pax9 cis-regulatory sequences were located in an intron of the neighboring gene Slc25a21. Close examination of an extended genomic interval around Pax9 revealed the presence of strong synteny conservation in the human, mouse, and Fugu genomes. We propose such an intersecting organization of cis-regulatory sequences in multigenic regions as a possible mechanism that maintains evolutionary conserved synteny. PMID:14504231

  19. Adaptation of the targeted capture Methyl-Seq platform for the mouse genome identifies novel tissue-specific DNA methylation patterns of genes involved in neurodevelopment

    PubMed Central

    Hing, Benjamin; Ramos, Enrique; Braun, Patricia; McKane, Melissa; Jancic, Dubravka; Tamashiro, Kellie L K; Lee, Richard S; Michaelson, Jacob J; Druley, Todd E; Potash, James B

    2015-01-01

    Methyl-Seq was recently developed as a targeted approach to assess DNA methylation (DNAm) at a genome-wide level in human. We adapted it for mouse and sought to examine DNAm differences across liver and 2 brain regions: cortex and hippocampus. A custom hybridization array was designed to isolate 99 Mb of CpG islands, shores, shelves, and regulatory elements in the mouse genome. This was followed by bisulfite conversion and sequencing on the Illumina HiSeq2000. The majority of differentially methylated cytosines (DMCs) were present at greater than expected frequency in introns, intergenic regions, near CpG islands, and transcriptional enhancers. Liver-specific enhancers were observed to be methylated in cortex, while cortex specific enhancers were methylated in the liver. Interestingly, commonly shared enhancers were differentially methylated between the liver and cortex. Gene ontology and pathway analysis showed that genes that were hypomethylated in the cortex and hippocampus were enriched for neuronal components and neuronal function. In contrast, genes that were hypomethylated in the liver were enriched for cellular components important for liver function. Bisulfite-pyrosequencing validation of 75 DMCs from 19 different loci showed a correlation of r = 0.87 with Methyl-Seq data. We also identified genes involved in neurodevelopment that were not previously reported to be differentially methylated across brain regions. This platform constitutes a valuable tool for future genome-wide studies involving mouse models of disease. PMID:25985232

  20. Schizophrenia: a neurodevelopmental disorder--integrative genomic hypothesis and therapeutic implications from a transgenic mouse model.

    PubMed

    Stachowiak, M K; Kucinski, A; Curl, R; Syposs, C; Yang, Y; Narla, S; Terranova, C; Prokop, D; Klejbor, I; Bencherif, M; Birkaya, B; Corso, T; Parikh, A; Tzanakakis, E S; Wersinger, S; Stachowiak, E K

    2013-02-01

    Schizophrenia is a neurodevelopmental disorder featuring complex aberrations in the structure, wiring, and chemistry of multiple neuronal systems. The abnormal developmental trajectory of the brain appears to be established during gestation, long before clinical symptoms of the disease appear in early adult life. Many genes are associated with schizophrenia, however, altered expression of no one gene has been shown to be present in a majority of schizophrenia patients. How does altered expression of such a variety of genes lead to the complex set of abnormalities observed in the schizophrenic brain? We hypothesize that the protein products of these genes converge on common neurodevelopmental pathways that affect the development of multiple neural circuits and neurotransmitter systems. One such neurodevelopmental pathway is Integrative Nuclear FGFR1 Signaling (INFS). INFS integrates diverse neurogenic signals that direct the postmitotic development of embryonic stem cells, neural progenitors and immature neurons, by direct gene reprogramming. Additionally, FGFR1 and its partner proteins link multiple upstream pathways in which schizophrenia-linked genes are known to function and interact directly with those genes. A th-fgfr1(tk-) transgenic mouse with impaired FGF receptor signaling establishes a number of important characteristics that mimic human schizophrenia - a neurodevelopmental origin, anatomical abnormalities at birth, a delayed onset of behavioral symptoms, deficits across multiple domains of the disorder and symptom improvement with typical and atypical antipsychotics, 5-HT antagonists, and nicotinic receptor agonists. Our research suggests that altered FGF receptor signaling plays a central role in the developmental abnormalities underlying schizophrenia and that nicotinic agonists are an effective class of compounds for the treatment of schizophrenia.

  1. Genome-wide association for fear conditioning in an advanced intercross mouse line.

    PubMed

    Parker, Clarissa C; Sokoloff, Greta; Cheng, Riyan; Palmer, Abraham A

    2012-05-01

    Fear conditioning (FC) may provide a useful model for some components of post-traumatic stress disorder (PTSD). We used a C57BL/6J × DBA/2J F(2) intercross (n = 620) and a C57BL/6J × DBA/2J F(8) advanced intercross line (n = 567) to fine-map quantitative trait loci (QTL) associated with FC. We conducted an integrated genome-wide association analysis in QTLRel and identified five highly significant QTL affecting freezing to context as well as four highly significant QTL associated with freezing to cue. The average percent decrease in QTL width between the F(2) and the integrated analysis was 59.2%. Next, we exploited bioinformatic sequence and expression data to identify candidate genes based on the existence of non-synonymous coding polymorphisms and/or expression QTLs. We identified numerous candidate genes that have been previously implicated in either fear learning in animal models (Bcl2, Btg2, Dbi, Gabr1b, Lypd1, Pam and Rgs14) or PTSD in humans (Gabra2, Oprm1 and Trkb); other identified genes may represent novel findings. The integration of F(2) and AIL data maintains the advantages of studying FC in model organisms while significantly improving resolution over previous approaches.

  2. Differential genomic imprinting regulates paracrine and autocrine roles of IGF2 in mouse adult neurogenesis

    PubMed Central

    Ferrón, S. R.; Radford, E. J.; Domingo-Muelas, A.; Kleine, I.; Ramme, A.; Gray, D.; Sandovici, I.; Constancia, M.; Ward, A.; Menheniott, T. R.; Ferguson-Smith, A. C.

    2015-01-01

    Genomic imprinting is implicated in the control of gene dosage in neurogenic niches. Here we address the importance of Igf2 imprinting for murine adult neurogenesis in the subventricular zone (SVZ) and in the subgranular zone (SGZ) of the hippocampus in vivo. In the SVZ, paracrine IGF2 is a cerebrospinal fluid and endothelial-derived neurogenic factor requiring biallelic expression, with mutants having reduced activation of the stem cell pool and impaired olfactory bulb neurogenesis. In contrast, Igf2 is imprinted in the hippocampus acting as an autocrine factor expressed in neural stem cells (NSCs) solely from the paternal allele. Conditional mutagenesis of Igf2 in blood vessels confirms that endothelial-derived IGF2 contributes to NSC maintenance in SVZ but not in the SGZ, and that this is regulated by the biallelic expression of IGF2 in the vascular compartment. Our findings indicate that a regulatory decision to imprint or not is a functionally important mechanism of transcriptional dosage control in adult neurogenesis. PMID:26369386

  3. Comparative genomics between fly, mouse, and cattle identifies genes associated with sire conception rate.

    PubMed

    Li, G; Peñagaricano, F; Weigel, K A; Zhang, Y; Rosa, G; Khatib, H

    2012-10-01

    The decline in reproductive performance in cattle is of major concern to farmers and the dairy industry worldwide. Most fertility studies in cattle have focused on fertility of the cow, whereas the genetics of male fertility have not been thoroughly investigated. The present study hypothesizes that the high conservation of spermatogenesis genes from fly to human implies important roles of these genes in male fertility in cattle. To test this hypothesis, we performed an association analysis between highly conserved spermatogenesis genes and sire conception rate (SCR) in US Holsteins as a measure of bull fertility. Sequencing analysis revealed 24 single nucleotide polymorphisms (SNP) in 9 genes in the bull population using the pooled DNA sequencing approach. Five SNP previously identified in 5 genes from the POU1F1 pathway were also included in this study because they have shown significant associations with female and male fertility traits. Overall, 29 SNP located in 14 candidate genes were tested for association with sire conception rate in a population of 1,988 bulls. Three SNP located in MAP1B and 1 SNP in PPP1R11 showed significant associations with SCR. For the POU1F1 pathway, single gene analysis revealed significant associations of POU1F1 and STAT5A with SCR. Analysis of genotypic interactions between adjacent genes in the pathway revealed significant associations of STAT5A and UTMP genotypic combinations with SCR. The most significant spermatogenesis gene, MAP1B, was found to be associated with fertilization and blastocyst rates. Thus, the association of these genes with bull fertility testifies to the usefulness of the comparative genomics approach in selecting candidate male fertility genes.

  4. Genetic analysis of albuminuria in the aging mouse and concordance with loci for diabetic nephropathy found in a genome-wide association scan

    PubMed Central

    Tsaih, Shirng-Wern; Pezzolesi, Marcus G.; Yuan, Rong; Warram, James H.; Krolewski, Andrzej S.; Korstanje, Ron

    2009-01-01

    Aging is a physiological process involving both genetic factors and environmental agents that can lead to function loss in organs. In the kidney, aging can cause leakage of proteins in urine, starting with albumin. Discovering molecular mechanisms responsible for albuminuria during aging could offer new perspectives on the etiology of this abnormality. Haplotype association mapping in the mouse is a novel approach which uses the haplotypes of the relatively closely related mouse inbred strains and the variation of the phenotypes among these strains to find associations between haplotypes and phenotype. Albumin-to-creatinine ratios, measures of urinary albumin excretion, were determined in 30 inbred mouse strains at 12, 18, and 24 months of age. To determine genetic loci that are involved in albuminuria, haplotype association mapping was performed for males and females separately at all 3 time points using a set of 63,222 SNPs. One significant and 8 suggestive loci were identified, some of which map to previously identified loci for traits associated with kidney damage in the mouse, but with a much higher resolution, which narrowed the mapped loci. These 9 loci were then investigated in the data of the genome-wide association scan for diabetic nephropathy in human type 1 diabetes. Two of the 9 mouse loci were found to be significantly associated with diabetic nephropathy, suggesting common underlying genes predisposing to kidney disease in mice and humans. PMID:19924099

  5. A sequence-based identification of the genes detected by probesets on the Affymetrix U133 plus 2.0 array.

    PubMed

    Harbig, Jeremy; Sprinkle, Robert; Enkemann, Steven A

    2005-02-18

    One of the biggest problems facing microarray experiments is the difficulty of translating results into other microarray formats or comparing microarray results to other biochemical methods. We believe that this is largely the result of poor gene identification. We re-identified the probesets on the Affymetrix U133 plus 2.0 GeneChip array. This identification was based on the sequence of the probes and the sequence of the human genome. Using the BLAST program, we matched probes with documented and postulated human transcripts. This resulted in the redefinition of approximately 37% of the probes on the U133 plus 2.0 array. This updated identification specifically points out where the identification is complicated by cross-hybridization from splice variants or closely related genes. More than 5000 probesets detect multiple transcripts and therefore the exact protein affected cannot be readily concluded from the performance of one probeset alone. This makes naming difficult and impacts any downstream analysis such as associating gene ontologies, mapping affected pathways or simply validating expression changes. We have now automated the sequence-based identification and can more appropriately annotate any array where the sequence on each spot is known.

  6. Genome-Wide Scleral Micro- and Messenger-RNA Regulation During Myopia Development in the Mouse

    PubMed Central

    Metlapally, Ravikanth; Park, Han Na; Chakraborty, Ranjay; Wang, Kevin K.; Tan, Christopher C.; Light, Jacob G.; Pardue, Machelle T.; Wildsoet, Christine F.

    2016-01-01

    Purpose MicroRNA (miRNAs) have been previously implicated in scleral remodeling in normal eye growth. They have the potential to be therapeutic targets for prevention/retardation of exaggerated eye growth in myopia by modulating scleral matrix remodeling. To explore this potential, genome-wide miRNA and messenger RNA (mRNA) scleral profiles in myopic and control eyes from mice were studied. Methods C57BL/6J mice (n = 7; P28) reared under a 12L:12D cycle were form-deprived (FD) unilaterally for 2 weeks. Refractive error and axial length changes were measured using photorefraction and 1310-nm spectral-domain optical coherence tomography, respectively. Scleral RNA samples from FD and fellow control eyes were processed for microarray assay. Statistical analyses were performed using National Institute of Aging array analysis tool; group comparisons were made using ANOVA, and gene ontologies were identified using software available on the Web. Findings were confirmed using quantitative PCR in a separate group of mice (n = 7). Results Form-deprived eyes showed myopic shifts in refractive error (−2.02 ± 0.47 D; P < 0.01). Comparison of the scleral RNA profiles of test eyes with those of control eyes revealed 54 differentially expressed miRNAs and 261 mRNAs fold-change >1.25 (maximum fold change = 1.63 and 2.7 for miRNAs and mRNAs, respectively) (P < 0.05; minimum, P = 0.0001). Significant ontologies showing gene over-representation (P < 0.05) included intermediate filament organization, scaffold protein binding, detection of stimuli, calcium ion, G protein, and phototransduction. Significant differential expression of Let-7a and miR-16-2, and Smok4a, Prph2, and Gnat1 were confirmed. Conclusions Scleral mi- and mRNAs showed differential expression linked to myopia, supporting the involvement of miRNAs in eye growth regulation. The observed general trend of relatively small fold-changes suggests a tightly controlled, regulatory mechanism for scleral gene expression. PMID

  7. Intracisternal A particle genes: Distribution in the mouse genome, active subtypes, and potential roles as species-specific mediators of susceptibility to cancer.

    PubMed

    Qin, Chunhua; Wang, Zhibin; Shang, Jin; Bekkari, Kavitha; Liu, Rong; Pacchione, Stephen; McNulty, Kathleen A; Ng, Alan; Barnum, John E; Storer, Richard D

    2010-01-01

    Rodents, mice and rats in particular, are the species of choice for evaluating chemical carcinogenesis. However, different species and strains often respond very differently, undermining the logic of extrapolation of animal results to humans and complicating risk assessment. Intracisternal A particles (IAPs), endogenous retroviral sequences, are an important class of transposable elements that induce genomic mutations and cell transformation by disrupting gene expression. Several lines of evidence support a role of IAPs as mouse-specific genetic factors in responses to toxicity and expression of disease phenotypes. Since multiple subtypes and copies of IAPs are present in the mouse genome, their activity and locations relative to functional genes are of critical importance. This study identified the major "active" subtypes of IAPs (subtype 1/1a) that are responsible for newly transposed IAP insertions described in the literature, and confirmed that (1) polymorphisms for IAP insertions exist among different mouse strains and (2) promoter activity of the LTRs can be modulated by chemicals. This study further identified all the genes in the C57BL/6 mouse genome with IAP subtype 1 and 1a sequences inserted in their proximity, and the major biofunctional categories and cellular signaling networks of those genes. Since many "IAP-associated genes" play important roles in the regulation of cell proliferation, cell cycle, and cell death, the associated IAPs, upon activation, can affect cellular responses to xenobiotics and disease processes, especially carcinogenesis. This systemic analysis provides a solid foundation for further investigations of the role of IAPs as species- and strain-specific disease susceptibility factors.

  8. Transcriptome networks in the mouse retina: An exon level BXD RI database

    PubMed Central

    King, Rebecca; Lu, Lu; Williams, Robert W.

    2015-01-01

    Purpose Differences in gene expression provide diverse retina phenotypes and may also contribute to susceptibility to injury and disease. The present study defines the transcriptome of the retina in the BXD RI strain set, using the Affymetrix Mouse Gene 2.0 ST array to investigate all exons of traditional protein coding genes, non-coding RNAs, and microRNAs. These data are presented in a highly interactive database on the GeneNetwork website. Methods In the Normal Retina Database, the mRNA levels of the transcriptome from retinas was quantified using the Affymetrix Mouse Gene 2.0 ST array. This database consists of data from male and female mice. The data set includes a total of 52 BXD RI strains, the parental strains (C57BL/6J and DBA/2J), and a reciprocal cross. Results In combination with GeneNetwork, the Department of Defense (DoD) Congressionally Directed Medical Research Programs (CDMRP) Normal Retina Database provides a large resource for mapping, graphing, analyzing, and testing complex genetic networks. Protein-coding and non-coding RNAs can be used to map quantitative trait loci (QTLs) that contribute to expression differences among the BXD strains and to establish links between classical ocular phenotypes associated with differences in the genomic sequence. Using this resource, we extracted transcriptome signatures for retinal cells and defined genetic networks associated with the maintenance of the normal retina. Furthermore, we examined differentially expressed exons within a single gene. Conclusions The high level of variation in mRNA levels found among the BXD RI strains makes it possible to identify expression networks that underline differences in retina structure and function. Ultimately, we will use this database to define changes that occur following blast injury to the retina. PMID:26604663

  9. A partial genomic DNA clone for the alpha subunit of the mouse complement receptor type 3 and cellular adhesion molecule Mac-1.

    PubMed Central

    Sastre, L; Roman, J M; Teplow, D B; Dreyer, W J; Gee, C E; Larson, R S; Roberts, T M; Springer, T A

    1986-01-01

    A genomic clone coding for the alpha subunit of the mouse complement receptor type 3 and the cellular adhesion molecule Mac-1 has been isolated directly from a genomic library using synthetic oligonucleotide probes based on the amino-terminal amino acid sequence of the protein. The identity of the clone has been established by DNA sequencing and in vitro translation of hybrid-selected mRNA. The gene is present in a single copy in the murine genome. The region containing the amino-terminal exon has been sequenced. RNA gel blotting shows that the Mac-1 alpha-subunit mRNA is 6 kilobases in length. Mac-1 alpha-subunit mRNA is present in macrophages but not T lymphoma or L cells. During gamma interferon-stimulated maturation of the mouse premyelocytic cell line M1, Mac-1 alpha-subunit mRNA is induced. This corresponds with the tissue distribution of the Mac-1 alpha subunit, showing expression is regulated at least partially at the message level. Images PMID:2942940

  10. Systematic Expression Profiling of the Mouse Transcriptome Using RIKEN cDNA Microarrays

    PubMed Central

    Bono, Hidemasa; Yagi, Ken; Kasukawa, Takeya; Nikaido, Itoshi; Tominaga, Naoko; Miki, Rika; Mizuno, Yosuke; Tomaru, Yasuhiro; Goto, Hitoshi; Nitanda, Hiroyuki; Shimizu, Daisuke; Makino, Hirochika; Morita, Tomoyuki; Fujiyama, Junshin; Sakai, Takehito; Shimoji, Takashi; Hume, David A.; Hayashizaki, Yoshihide; Okazaki, Yasushi

    2003-01-01

    The number of known mRNA transcripts in the mouse has been greatly expanded by the RIKEN Mouse Gene Encyclopedia project. Validation of their reproducible expression in a tissue is an important contribution to the study of functional genomics. In this report, we determine the expression profile of 57,931 clones on 20 mouse tissues using cDNA microarrays. Of these 57,931 clones, 22,928 clones correspond to the FANTOM2 clone set. The set represents 20,234 transcriptional units (TUs) out of 33,409 TUs in the FANTOM2 set. We identified 7206 separate clones that satisfied stringent criteria for tissue-specific expression. Gene Ontology terms were assigned for these 7206 clones, and the proportion of `molecular function' ontology for each tissue-specific clone was examined. These data will provide insights into the function of each tissue. Tissue-specific gene expression profiles obtained using our cDNA microarrays were also compared with the data extracted from the GNF Expression Atlas based on Affymetrix microarrays. One major outcome of the RIKEN transcriptome analysis is the identification of numerous nonprotein-coding mRNAs. The expression profile was also used to obtain evidence of expression for putative noncoding RNAs. In addition, 1926 clones (70%) of 2768 clones that were categorized as “unknown EST,” and 1969 (58%) clones of 3388 clones that were categorized as “unclassifiable” were also shown to be reproducibly expressed. PMID:12819129

  11. Genome-wide siRNA screen of genes regulating the LPS-induced NF-κB and TNF-α responses in mouse macrophages

    PubMed Central

    Li, Ning; Katz, Samuel; Dutta, Bhaskar; Benet, Zachary L.; Sun, Jing; Fraser, Iain D.C.

    2017-01-01

    The mammalian innate immune system senses many bacterial stimuli through the toll-like receptor (TLR) family. Activation of the TLR4 receptor by bacterial lipopolysaccharide (LPS) is the most widely studied TLR pathway due to its central role in host responses to gram-negative bacterial infection and its contribution to endotoxemia and sepsis. Here we describe a genome-wide siRNA screen to identify genes regulating the mouse macrophage TNF-α and NF-κB responses to LPS. We include a secondary validation screen conducted with six independent siRNAs per gene to facilitate removal of off-target screen hits. We also provide microarray data from the same LPS-treated macrophage cells to facilitate downstream data analysis. These data provide a resource for analyzing gene function in the predominant pathway driving inflammatory signaling and cytokine expression in mouse macrophages. PMID:28248925

  12. Evaluating the performance of Affymetrix SNP Array 6.0 platform with 400 Japanese individuals

    PubMed Central

    Nishida, Nao; Koike, Asako; Tajima, Atsushi; Ogasawara, Yuko; Ishibashi, Yoshimi; Uehara, Yasuka; Inoue, Ituro; Tokunaga, Katsushi

    2008-01-01

    Background With improvements in genotyping technologies, genome-wide association studies with hundreds of thousands of SNPs allow the identification of candidate genetic loci for multifactorial diseases in different populations. However, genotyping errors caused by genotyping platforms or genotype calling algorithms may lead to inflation of false associations between markers and phenotypes. In addition, the number of SNPs available for genome-wide association studies in the Japanese population has been investigated using only 45 samples in the HapMap project, which could lead to an inaccurate estimation of the number of SNPs with low minor allele frequencies. We genotyped 400 Japanese samples in order to estimate the number of SNPs available for genome-wide association studies in the Japanese population and to examine the performance of the current SNP Array 6.0 platform and the genotype calling algorithm "Birdseed". Results About 20% of the 909,622 SNP markers on the array were revealed to be monomorphic in the Japanese population. Consequently, 661,599 SNPs were available for genome-wide association studies in the Japanese population, after excluding the poorly behaving SNPs. The Birdseed algorithm accurately determined the genotype calls of each sample with a high overall call rate of over 99.5% and a high concordance rate of over 99.8% using more than 48 samples after removing low-quality samples by adjusting QC criteria. Conclusion Our results confirmed that the SNP Array 6.0 platform reached the level reported by the manufacturer, and thus genome-wide association studies using the SNP Array 6.0 platform have considerable potential to identify candidate susceptibility or resistance genetic factors for multifactorial diseases in the Japanese population, as well as in other populations. PMID:18803882

  13. Genome-Wide Deletion Screening with the Array CGH Method in Mouse Offspring Derived from Irradiated Spermatogonia Indicates that Mutagenic Responses are Highly Variable among Genes.

    PubMed

    Asakawa, Jun-Ichi; Kodaira, Mieko; Miura, Akiko; Tsuji, Takahiro; Nakamoto, Yoshiko; Imanaka, Masaaki; Kitamura, Jun; Cullings, Harry; Nishimura, Mayumi; Shimada, Yoshiya; Nakamura, Nori

    2016-12-01

    Until the end of the 20th century, mouse germ cell data on induced mutation rates, which were collected using classical genetic methods at preselected specific loci, provided the principal basis for estimates of genetic risks from radiation in humans. The work reported on here is an extension of earlier efforts in this area using molecular methods. It focuses on validating the use of array comparative genomic hybridization (array CGH) methods for identifying radiation-induced copy number variants (CNVs) and specifically for DNA deletions. The emphasis on deletions stems from the view that it constitutes the predominant type of radiation-induced genetic damage, which is relevant for estimating genetic risks in humans. In the current study, deletion mutations were screened in the genomes of F1 mice born to unirradiated or 4 Gy irradiated sires at the spermatogonia stage (100 offspring each). The array CGH analysis was performed using a "2M array" with over 2 million probes with a mean interprobe distance of approximately 1 kb. The results provide evidence of five molecularly-confirmed paternally-derived deletions in the irradiated group (5/100) and one in the controls (1/100). These data support a calculation, which estimates that the mutation rate is 1 × 10(-2)/Gy per genome for induced deletions; this is much lower than would be expected if one assumes that the specific locus rate of 1 × 10(-5)/locus per Gy (at 34 loci) is applicable to other genes in the genome. The low observed rate of induced deletions suggests that the effective number of genes/genomic regions at which recoverable deletions could be induced would be only approximately 1,000. This estimate is far lower than expected from the size of the mouse genome (>20,000 genes). Such a discrepancy between observation and expectation can occur if the genome contains numerous genes that are far less sensitive to radiation-induced deletions, if many deletion-bearing offspring are not viable or if the current

  14. Inhibition of phosphorylated Ser473-Akt from translocating into the nucleus contributes to 2-cell arrest and defective zygotic genome activation in mouse preimplantation embryogenesis.

    PubMed

    Chen, Junming; Lian, Xiuli; Du, Juan; Xu, Songhua; Wei, Jianen; Pang, Lili; Song, Chanchan; He, Lin; Wang, Shie

    2016-04-01

    Phosphorylated Ser473-Akt (p-Ser473-Akt) is extensively studied as a correlate for the activity of Akt, which plays an important role in mouse oogenesis and preimplantation embryogenesis. However, little progress has been made about its effect on the mouse zygotic genome activation (ZGA) of 2-cell stage in mouse preimplantation embryos. In this study, we confirmed its localization in the pronuclei of 1-cell embryos and found that p-Ser473-Akt acquired prominent nucleus localization in 2-cell embryos physiologically. Akt specific inhibitors API-2 and MK2206 could inhibit the development of mouse preimplantation embryos in vitro, and induce 2-cell arrest at certain concentrations. 2-cell embryos exposed to 2.0 μmol/L API-2 or 30 μmol/L MK2206 displayed attenuated immunofluorescence intensity of p-Ser473-Akt in the nucleus. Simultaneously, qRT-PCR results revealed that 2.0 μmol/L API-2 treatment significantly downregulated the mRNA pattern of MuERV-L and eIF-1A, two marker genes of ZGA, suggesting a defect in ZGA compared with that of control group. Collectively, our work demonstrated the nuclear localization of p-Ser473-Akt during major ZGA, and Akt specific inhibitors API-2 and MK2206 which led to 2-cell arrest inhibited p-Ser473-Akt from translocating into the nucleus of 2-cell embryos with defective ZGA as well, implying p-Ser473-Akt may be a potential player in the major ZGA of 2-cell mouse embryos.

  15. Affymetrix Whole-Transcript Human Gene 1.0 ST array is highly concordant with standard 3' expression arrays.

    PubMed

    Pradervand, Sylvain; Paillusson, Alexandra; Thomas, Jérôme; Weber, Johann; Wirapati, Pratyaksha; Hagenbüchle, Otto; Harshman, Keith

    2008-05-01

    The recently released Affymetrix Human Gene 1.0 ST array has two major differences compared with standard 3' based arrays: (i) it interrogates the entire mRNA transcript, and (ii) it uses DNA targets. To assess the impact of these differences on array performance, we performed a series of comparative hybridizations between the Human Gene 1.0 ST and the Affymetrix HG-U133 Plus 2.0 and the Illumina HumanRef-8 BeadChip arrays. Additionally, both RNA and DNA targets were hybridized on HG-U133 Plus 2.0 arrays. The results show that the overall reproducibility of the Gene 1.0 ST array is best. When looking only at the high intensity probes, the reproducibility of the Gene 1.0 ST array and the Illumina BeadChip array is equally good. Concordance of array results was assessed using different inter-platform mappings. Agreements are best between the two labeling protocols using HG-U133 Plus 2.0 array. The Gene 1.0 ST array is most concordant with the HG-U133 array hybridized with cDNA targets. This may reflect the impact of the target type. Overall, the high degree of correspondence provides strong evidence for the reliability of the Gene 1.0 ST array.

  16. Mining Affymetrix microarray data for long non-coding RNAs: altered expression in the nucleus accumbens of heroin abusers.

    PubMed

    Michelhaugh, Sharon K; Lipovich, Leonard; Blythe, Jason; Jia, Hui; Kapatos, Gregory; Bannon, Michael J

    2011-02-01

    Although recent data suggest that some long non-coding RNAs (lncRNAs) exert widespread effects on gene expression and organelle formation, lncRNAs as a group constitute a sizable but poorly characterized fraction of the human transcriptome. We investigated whether some human lncRNA sequences were fortuitously represented on commonly used microarrays, then used this annotation to assess lncRNA expression in human brain. A computational and annotation pipeline was developed to identify lncRNA transcripts represented on Affymetrix U133 arrays. A previously published dataset derived from human nucleus accumbens was then examined for potential lncRNA expression. Twenty-three lncRNAs were determined to be represented on U133 arrays. Of these, dataset analysis revealed that five lncRNAs were consistently detected in samples of human nucleus accumbens. Strikingly, the abundance of these lncRNAs was up-regulated in human heroin abusers compared to matched drug-free control subjects, a finding confirmed by quantitative PCR. This study presents a paradigm for examining existing Affymetrix datasets for the detection and potential regulation of lncRNA expression, including changes associated with human disease. The finding that all detected lncRNAs were up-regulated in heroin abusers is consonant with the proposed role of lncRNAs as mediators of widespread changes in gene expression as occur in drug abuse.

  17. The mouse p97 (CDC48) gene. Genomic structure, definition of transcriptional regulatory sequences, gene expression, and characterization of a pseudogene.

    PubMed

    Müller, J M; Meyer, H H; Ruhrberg, C; Stamp, G W; Warren, G; Shima, D T

    1999-04-09

    Here we present the first description of the genomic organization, transcriptional regulatory sequences, and adult and embryonic gene expression for the mouse p97(CDC48) AAA ATPase. Clones representing two distinct p97 genes were isolated in a genomic library screen, one of them likely representing a non-functional processed pseudogene. The coding region of the gene encoding the functional mRNA is interrupted by 16 introns and encompasses 20.4 kilobase pairs. Definition of the transcriptional initiation site and sequence analysis showed that the gene contains a TATA-less, GC-rich promoter region with an initiator element spanning the transcription start site. Cis-acting elements necessary for basal transcription activity reside within 410 base pairs of the flanking region as determined by transient transfection assays. In immunohistological analyses, p97 was widely expressed in embryos and adults, but protein levels were tightly controlled in a cell type- and cell differentiation-dependent manner. A remarkable heterogeneity in p97 immunostaining was found on a cellular level within a given tissue, and protein amounts in the cytoplasm and nucleus varied widely, suggesting a highly regulated and intermittent function for p97. This study provides the basis for a detailed analysis of the complex regulation of p97 and the reagents required for assessing its functional significance using targeted gene manipulation in the mouse.

  18. AAV genome loss from dystrophic mouse muscles during AAV-U7 snRNA-mediated exon-skipping therapy.

    PubMed

    Le Hir, Maëva; Goyenvalle, Aurélie; Peccate, Cécile; Précigout, Guillaume; Davies, Kay E; Voit, Thomas; Garcia, Luis; Lorain, Stéphanie

    2013-08-01

    In the context of future adeno-associated viral (AAV)-based clinical trials for Duchenne myopathy, AAV genome fate in dystrophic muscles is of importance considering the viral capsid immunogenicity that prohibits recurring treatments. We showed that AAV genomes encoding non-therapeutic U7 were lost from mdx dystrophic muscles within 3 weeks after intramuscular injection. In contrast, AAV genomes encoding U7ex23 restoring expression of a slightly shortened dystrophin were maintained endorsing that the arrest of the dystrophic process is crucial for maintaining viral genomes in transduced fibers. Indeed, muscles treated with low doses of AAV-U7ex23, resulting in sub-optimal exon skipping, displayed much lower titers of viral genomes, showing that sub-optimal dystrophin restoration does not prevent AAV genome loss. We also followed therapeutic viral genomes in severe dystrophic dKO mice over time after systemic treatment with scAAV9-U7ex23. Dystrophin restoration decreased significantly between 3 and 12 months in various skeletal muscles, which was correlated with important viral genome loss, except in the heart. Altogether, these data show that the success of future AAV-U7 therapy for Duchenne patients would require optimal doses of AAV-U7 to induce substantial levels of dystrophin to stabilize the treated fibers and maintain the long lasting effect of the treatment.

  19. Genomic analyses of the Formosan harvest mouse (Micromys minutus) and comparisons to the brown Norway rat (Rattus norvegicus) and the house mouse (Mus musculus).

    PubMed

    Lin, Liang-Kong; Ma, Gwo-Chin; Chen, Tze-Ho; Lin, Wen-Hsiang; Lee, Dong-Jay; Wen, Pao-Ying; Wu, Sheng-Hai; Chen, Ming

    2013-10-01

    The harvest mouse, Micromys minutus (MMIN), has a very wide range of distribution (from the British Isles across the Euroasian continent to Japan and Taiwan). We studied an isolated population of MMIN in Taiwan, which is at the southeastern margin of the species' geographic distribution, and compared its genetic complement with those of the same species previously reported from other geographic locations and with two model rodent species, the house mouse (Mus musculus) and the brown Norway rat (Rattus norvegicus). The diploid number (2N) of MMIN was 68, consistent with that reported for other populations. However, variations were noted in the fundamental number (FN) and the shape and banding patterns of the individual chromosomes among populations. The FN of MMIN was estimated to be 72, including 2 bi-armed autosomes, 31 one-armed autosomes, and one pair of one-armed sex chromosomes. Here, we propose the first ideogram for MMIN. C-banding, Ag-NOR, and the locations of 18S rRNA gene sequences (MMIN chromosomes no. 10, 14, 19, 29, 31, 33, and X) mapped by fluorescence in situ hybridization (FISH) are also reported. Additionally, we compared the 18S rDNA sequences and performed cross-species X chromosome painting (FISH) for M. minutus, M. musculus, and R. norvegicus. The results indicate that both genetic elements are rather conserved across species. Thus, implications for the phylogenetic position of Micromys were limited.

  20. Genome-Wide Association Mapping for Intelligence in Military Working Dogs: Canine Cohort, Canine Intelligence Assessment Regimen, Genome-Wide Single Nucleotide Polymorphism (SNP) Typing, and Unsupervised Classification Algorithm for Genome-Wide Association Data Analysis

    DTIC Science & Technology

    2011-09-01

    were down-selected and successfully genotyped for whole genome (WG) single nucleotide polymorphism (SNP) markers by means of the Affymetrix Canine...SUBJECT TERMS Military working dog genome-wide association study genetic marker intelligence... marker , intelligence, Canine Intelligence Testing Protocol, classification technique, clustering analysis Technical Report: September 2011 2

  1. Analysis of Genome-Wide Monoallelic Expression Patterns in Three Major Cell Types of Mouse Visual Cortex Using Laser Capture Microdissection.

    PubMed

    Lin, Chia-Yi; Huang, Shih-Chuan; Tung, Chun-Che; Chou, Chih-Hsuan; Gau, Susan Shur-Fen; Huang, Hsien-Sung

    Genomic imprinting is an epigenetic mechanism causing monoallelic expression in a parent-of-origin-specific manner. Disruption of imprinted genes causes various neurological and psychiatric disorders. However, the role of imprinted genes in the brain is largely unknown. Different cell types within distinct brain regions can influence the genomic imprinting status, but imprinted genes in single cell types within distinct brain regions have not been characterized on a genome-wide scale. To address this critical question, we used a multi-stage approach, which combined genetically engineered mice with fluorescence-based laser capture microdissection (LCM) to capture excitatory neurons, inhibitory neurons and astrocytes as single cells in layer 2/3 of mouse visual cortex. RNA sequencing determined parental expression patterns on a genome-wide scale in the captured cells within specific brain regions. The expression level of cell-type-specific genes for excitatory neurons (13 genes), inhibitory neurons (16 genes) and astrocytes (20 genes) confirmed the LCM-captured cells maintained their cellular identities. The parent-of-origin-specific expression pattern of imprinted genes, including maternally expressed Meg3 and paternally expressed Peg3, provided evidence that the status of known imprinted genes was also maintained. Although our platform remains to be improved, our findings demonstrate the parental expression pattern can be analysed not only at the level of a single cell type but also at the level of specific cortical layers. Our approach has the potential to reveal novel regulatory modules associated with plasticity through genomic imprinting mechanisms in different cell types, not only in the visual cortex but also in other brain regions.

  2. Analysis of Genome-Wide Monoallelic Expression Patterns in Three Major Cell Types of Mouse Visual Cortex Using Laser Capture Microdissection

    PubMed Central

    Gau, Susan Shur-Fen

    2016-01-01

    Genomic imprinting is an epigenetic mechanism causing monoallelic expression in a parent-of-origin-specific manner. Disruption of imprinted genes causes various neurological and psychiatric disorders. However, the role of imprinted genes in the brain is largely unknown. Different cell types within distinct brain regions can influence the genomic imprinting status, but imprinted genes in single cell types within distinct brain regions have not been characterized on a genome-wide scale. To address this critical question, we used a multi-stage approach, which combined genetically engineered mice with fluorescence-based laser capture microdissection (LCM) to capture excitatory neurons, inhibitory neurons and astrocytes as single cells in layer 2/3 of mouse visual cortex. RNA sequencing determined parental expression patterns on a genome-wide scale in the captured cells within specific brain regions. The expression level of cell-type-specific genes for excitatory neurons (13 genes), inhibitory neurons (16 genes) and astrocytes (20 genes) confirmed the LCM-captured cells maintained their cellular identities. The parent-of-origin-specific expression pattern of imprinted genes, including maternally expressed Meg3 and paternally expressed Peg3, provided evidence that the status of known imprinted genes was also maintained. Although our platform remains to be improved, our findings demonstrate the parental expression pattern can be analysed not only at the level of a single cell type but also at the level of specific cortical layers. Our approach has the potential to reveal novel regulatory modules associated with plasticity through genomic imprinting mechanisms in different cell types, not only in the visual cortex but also in other brain regions. PMID:27662371

  3. Erasure of DNA methylation, genomic imprints, and epimutations in a primordial germ-cell model derived from mouse pluripotent stem cells

    PubMed Central

    Miyoshi, Norikatsu; Stel, Jente M.; Shioda, Keiko; Qu, Na; Odajima, Junko; Mitsunaga, Shino; Zhang, Xiangfan; Nagano, Makoto; Hochedlinger, Konrad; Isselbacher, Kurt J.; Shioda, Toshi

    2016-01-01

    The genome-wide depletion of 5-methylcytosines (5meCs) caused by passive dilution through DNA synthesis without daughter strand methylation and active enzymatic processes resulting in replacement of 5meCs with unmethylated cytosines is a hallmark of primordial germ cells (PGCs). Although recent studies have shown that in vitro differentiation of pluripotent stem cells (PSCs) to PGC-like cells (PGCLCs) mimics the in vivo differentiation of epiblast cells to PGCs, how DNA methylation status of PGCLCs resembles the dynamics of 5meC erasure in embryonic PGCs remains controversial. Here, by differential detection of genome-wide 5meC and 5-hydroxymethylcytosine (5hmeC) distributions by deep sequencing, we show that PGCLCs derived from mouse PSCs recapitulated the process of genome-wide DNA demethylation in embryonic PGCs, including significant demethylation of imprint control regions (ICRs) associated with increased mRNA expression of the corresponding imprinted genes. Although 5hmeCs were also significantly diminished in PGCLCs, they retained greater amounts of 5hmeCs than intragonadal PGCs. The genomes of both PGCLCs and PGCs selectively retained both 5meCs and 5hmeCs at a small number of repeat sequences such as GSAT_MM, of which the significant retention of bisulfite-resistant cytosines was corroborated by reanalysis of previously published whole-genome bisulfite sequencing data for intragonadal PGCs. PSCs harboring abnormal hypermethylation at ICRs of the Dlk1-Gtl2-Dio3 imprinting cluster diminished these 5meCs upon differentiation to PGCLCs, resulting in transcriptional reactivation of the Gtl2 gene. These observations support the usefulness of PGCLCs in studying the germline epigenetic erasure including imprinted genes, epimutations, and erasure-resistant loci, which may be involved in transgenerational epigenetic inheritance. PMID:27486249

  4. Genomic DNA of MCF-7 breast cancer cells not an ideal choice as positive control for PCR amplification based detection of Mouse Mammary Tumor Virus-Like Sequences.

    PubMed

    Kulkarni, Bhushan B; Hiremath, Shivaprakash V; Kulkarni, Suyamindra S; Hallikeri, Umesh R; Patil, Basavaraj R; Gai, Pramod B

    2013-11-01

    The identification of the etiology of breast cancer is a crucial research issue for the development of an effective preventive and treatment strategies. Researchers are exploring the possible involvement of Mouse Mammary Tumor Virus (MMTV) in causing human breast cancer. Hence, it becomes very important to use a consistent positive control agent in PCR amplification based detection of MMTV-Like Sequence (MMTV-LS) in human breast cancer for accurate and reproducible results. This study was done to investigate the feasibility of using genomic DNA of MCF-7 breast cancer cells to detect MMTV-LS using PCR amplification based detection. MMTV env and SAG gene located at the 3' long terminal repeat (LTR) sequences were targeted for the PCR based detection. No amplification was observed in case of the genomic DNA of MCF-7 breast cancer cells. However, the 2.7 kb DNA fragment comprising MMTV env and SAG LTR sequences yielded the products of desired size. From these results it can be concluded that Genomic DNA of MCF-7 cell is not a suitable choice as positive control for PCR or RT-PCR based detection of MMTV-LS. It is also suggested that plasmids containing the cloned genes or sequences of MMTV be used as positive control for detection of MMTV-LS.

  5. EzArray: A web-based highly automated Affymetrix expression array data management and analysis system

    PubMed Central

    Zhu, Yuerong; Zhu, Yuelin; Xu, Wei

    2008-01-01

    Background Though microarray experiments are very popular in life science research, managing and analyzing microarray data are still challenging tasks for many biologists. Most microarray programs require users to have sophisticated knowledge of mathematics, statistics and computer skills for usage. With accumulating microarray data deposited in public databases, easy-to-use programs to re-analyze previously published microarray data are in high demand. Results EzArray is a web-based Affymetrix expression array data management and analysis system for researchers who need to organize microarray data efficiently and get data analyzed instantly. EzArray organizes microarray data into projects that can be analyzed online with predefined or custom procedures. EzArray performs data preprocessing and detection of differentially expressed genes with statistical methods. All analysis procedures are optimized and highly automated so that even novice users with limited pre-knowledge of microarray data analysis can complete initial analysis quickly. Since all input files, analysis parameters, and executed scripts can be downloaded, EzArray provides maximum reproducibility for each analysis. In addition, EzArray integrates with Gene Expression Omnibus (GEO) and allows instantaneous re-analysis of published array data. Conclusion EzArray is a novel Affymetrix expression array data analysis and sharing system. EzArray provides easy-to-use tools for re-analyzing published microarray data and will help both novice and experienced users perform initial analysis of their microarray data from the location of data storage. We believe EzArray will be a useful system for facilities with microarray services and laboratories with multiple members involved in microarray data analysis. EzArray is freely available from . PMID:18218103

  6. Concordance in Genomic Changes Between Mouse Lungs and Human Airway Epithelial Cells Exposed to Diesel Exhaust Particles

    EPA Science Inventory

    Human and animal toxicity studies have shown that exposure to diesel exhaust particles (DEP) or their constituents affect multiple biological processes including immune and inflammatory pathways, mutagenesis and in some cases carcinogenesis. This study compared genomic changes by...

  7. Genome-wide random regression analysis for parent-of-origin effects of body composition allometries in mouse

    PubMed Central

    Zhao, Jingli; Li, Shuling; Wang, Lijuan; Jiang, Li; Yang, Runqing; Cui, Yuehua

    2017-01-01

    Genomic imprinting underlying growth and development traits has been recognized, with a focus on the form of absolute or pure growth. However, little is known about the effect of genomic imprinting on relative growth. In this study, we proposed a random regression model to estimate genome-wide imprinting effects on the relative growth of multiple tissues and organs to body weight in mice. Joint static allometry scaling equation as sub-model is nested within the genetic effects of markers and polygenic effects caused by a pedigree. Both chromosome-wide and genome-wide statistical tests were conducted to identify imprinted quantitative trait nucleotides (QTNs) associated with relative growth of individual tissues and organs to body weight. Real data analysis showed that three of six analysed tissues and organs are significantly associated with body weight in terms of phenotypic relative growth. At the chromosome-wide level, a total 122 QTNs were associated with allometries of kidney, spleen and liver weights to body weight, 36 of which were imprinted with different imprinting fashions. Further, only two imprinted QTNs responsible for relative growth of spleen and liver were verified by genome-wide test. Our approach provides a general framework for statistical inference of genomic imprinting underlying allometry scaling in animals. PMID:28338098

  8. Tet3 and DNA replication mediate demethylation of both the maternal and paternal genomes in mouse zygotes.

    PubMed

    Shen, Li; Inoue, Azusa; He, Jin; Liu, Yuting; Lu, Falong; Zhang, Yi

    2014-10-02

    With the exception of imprinted genes and certain repeats, DNA methylation is globally erased during preimplantation development. Recent studies have suggested that Tet3-mediated oxidation of 5-methylcytosine (5mC) and DNA replication-dependent dilution both contribute to global paternal DNA demethylation, but demethylation of the maternal genome occurs via replication. Here we present genome-scale DNA methylation maps for both the paternal and maternal genomes of Tet3-depleted and/or DNA replication-inhibited zygotes. In both genomes, we found that inhibition of DNA replication blocks DNA demethylation independently from Tet3 function and that Tet3 facilitates DNA demethylation largely by coupling with DNA replication. For both genomes, our data indicate that replication-dependent dilution is the major contributor to demethylation, but Tet3 plays an important role, particularly at certain loci. Our study thus defines the respective functions of Tet3 and DNA replication in paternal DNA demethylation and reveals an unexpected contribution of Tet3 to demethylation of the maternal genome.

  9. Mouse p53-Deficient Cancer Models as Platforms for Obtaining Genomic Predictors of Human Cancer Clinical Outcomes

    PubMed Central

    Dueñas, Marta; Santos, Mirentxu; Aranda, Juan F.; Bielza, Concha; Martínez-Cruz, Ana B.; Lorz, Corina; Taron, Miquel; Ciruelos, Eva M.; Rodríguez-Peralto, José L.; Martín, Miguel; Larrañaga, Pedro; Dahabreh, Jubrail; Stathopoulos, George P.; Rosell, Rafael; Paramio, Jesús M.; García-Escudero, Ramón

    2012-01-01

    Mutations in the TP53 gene are very common in human cancers, and are associated with poor clinical outcome. Transgenic mouse models lacking the Trp53 gene or that express mutant Trp53 transgenes produce tumours with malignant features in many organs. We previously showed the transcriptome of a p53-deficient mouse skin carcinoma model to be similar to those of human cancers with TP53 mutations and associated with poor clinical outcomes. This report shows that much of the 682-gene signature of this murine skin carcinoma transcriptome is also present in breast and lung cancer mouse models in which p53 is inhibited. Further, we report validated gene-expression-based tests for predicting the clinical outcome of human breast and lung adenocarcinoma. It was found that human patients with cancer could be stratified based on the similarity of their transcriptome with the mouse skin carcinoma 682-gene signature. The results also provide new targets for the treatment of p53-defective tumours. PMID:22880004

  10. Gene expression profiles in liver of mouse after chronic exposure to drinking water.

    PubMed

    Wu, Bing; Zhang, Yan; Zhao, Dayong; Zhang, Xuxiang; Kong, Zhiming; Cheng, Shupei

    2009-10-01

    cDNA micorarray approach was applied to hepatic transcriptional profile analysis in male mouse (Mus musculus, ICR) to assess the potential health effects of drinking water in Nanjing, China. Mice were treated with continuous exposure to drinking water for 90 days. Hepatic gene expression was analyzed with Affymetrix Mouse Genome 430A 2.0 arrays, and pathway analysis was carried out by Molecule Annotation System 2.0 and KEGG pathway database. A total of 836 genes were found to be significantly altered (1.5-fold, P < or = 0.05), including 294 up-regulated genes and 542 down-regulated genes. According to biological pathway analysis, drinking water exposure resulted in aberration of gene expression and biological pathways linked to xenobiotic metabolism, signal transduction, cell cycle and oxidative stress response. Further, deregulation of several genes associated with carcinogenesis or tumor progression including Ccnd1, Egfr, Map2k3, Mcm2, Orc2l and Smad2 was observed. Although transcription changes in identified genes are unlikely to be used as a sole indicator of adverse health effects, the results of this study could enhance our understanding of early toxic effects of drinking water exposure and support future studies on drinking water safety.

  11. DNA repair efficiency in germ cells and early mouse embryos and consequences for radiation-induced transgenerational genomic damage

    SciTech Connect

    Marchetti, Francesco; Wyrobek, Andrew J.

    2009-01-18

    Exposure to ionizing radiation and other environmental agents can affect the genomic integrity of germ cells and induce adverse health effects in the progeny. Efficient DNA repair during gametogenesis and the early embryonic cycles after fertilization is critical for preventing transmission of DNA damage to the progeny and relies on maternal factors stored in the egg before fertilization. The ability of the maternal repair machinery to repair DNA damage in both parental genomes in the fertilizing egg is especially crucial for the fertilizing male genome that has not experienced a DNA repair-competent cellular environment for several weeks prior to fertilization. During the DNA repair-deficient period of spermatogenesis, DNA lesions may accumulate in sperm and be carried into the egg where, if not properly repaired, could result in the formation of heritable chromosomal aberrations or mutations and associated birth defects. Studies with female mice deficient in specific DNA repair genes have shown that: (i) cell cycle checkpoints are activated in the fertilized egg by DNA damage carried by the sperm; and (ii) the maternal genotype plays a major role in determining the efficiency of repairing genomic lesions in the fertilizing sperm and directly affect the risk for abnormal reproductive outcomes. There is also growing evidence that implicates DNA damage carried by the fertilizing gamete as a mediator of postfertilization processes that contribute to genomic instability in subsequent generations. Transgenerational genomic instability most likely involves epigenetic mechanisms or error-prone DNA repair processes in the early embryo. Maternal and embryonic DNA repair processes during the early phases of mammalian embryonic development can have far reaching consequences for the genomic integrity and health of subsequent generations.

  12. Generation of a Knockout Mouse Embryonic Stem Cell Line Using a Paired CRISPR/Cas9 Genome Engineering Tool.

    PubMed

    Wettstein, Rahel; Bodak, Maxime; Ciaudo, Constance

    2016-01-01

    CRISPR/Cas9, originally discovered as a bacterial immune system, has recently been engineered into the latest tool to successfully introduce site-specific mutations in a variety of different organisms. Composed only of the Cas9 protein as well as one engineered guide RNA for its functionality, this system is much less complex in its setup and easier to handle than other guided nucleases such as Zinc-finger nucleases or TALENs.Here, we describe the simultaneous transfection of two paired CRISPR sgRNAs-Cas9 plasmids, in mouse embryonic stem cells (mESCs), resulting in the knockout of the selected target gene. Together with a four primer-evaluation system, it poses an efficient way to generate new independent knockout mouse embryonic stem cell lines.

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

    PubMed

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

    2015-07-01

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

  14. Glial cell line-derived neurotrophic factor alters the growth characteristics and genomic imprinting of mouse multipotent adult germline stem cells

    SciTech Connect

    Jung, Yoon Hee

    2010-03-10

    This study evaluated the essentiality of glial cell line-derived neurotrophic factor (GDNF) for in vitro culture of established mouse multipotent adult germline stem (maGS) cell lines by culturing them in the presence of GDNF, leukemia inhibitory factor (LIF) or both. We show that, in the absence of LIF, GDNF slows the proliferation of maGS cells and result in smaller sized colonies without any change in distribution of cells to different cell-cycle stages, expression of pluripotency genes and in vitro differentiation potential. Furthermore, in the absence of LIF, GDNF increased the expression of male germ-line genes and repopulated the empty seminiferous tubule of W/W{sup v} mutant mouse without the formation of teratoma. GDNF also altered the genomic imprinting of Igf2, Peg1, and H19 genes but had no effect on DNA methylation of Oct4, Nanog and Stra8 genes. However, these effects of GDNF were masked in the presence of LIF. GDNF also did not interfere with the multipotency of maGS cells if they are cultured in the presence of LIF. In conclusion, our results suggest that, in the absence of LIF, GDNF alters the growth characteristics of maGS cells and partially impart them some of the germline stem (GS) cell-like characteristics.

  15. Genomic organisation of the mouse gene encoding endothelin-converting enzyme-1 (ECE-1) and mRNA expression of ECE-1 isoforms in murine tissues.

    PubMed

    Lindenau, Steffi; von Langsdorff, Christian; Saxena, Amit; Paul, Martin; Orzechowski, Hans-Dieter

    2006-05-24

    Mouse knockout-models have previously revealed important biological functions of endothelin-converting enzyme-1 (ECE-1) in normal cardiac and craniofacial development. Since human ECE-1 is expressed in various isoforms, termed a, b, c, and d, expression of which is controlled by alternative promoters, we postulated that corresponding isoforms may also be transcribed from the murine Ece1 gene. By comparative sequence analysis using exon-specific sequences of human and rat ECE-1 we have resolved the complete exon-intron structure of the murine Ece1 locus on chromosome 4. The murine Ece1 gene comprises 23 exons distributed over 100 kb of genomic DNA and was found to be structurally highly conserved when compared to the human ECE1 gene. As with the human gene, the exons containing isoform-specific sequences were localised in the 5' terminal region of the murine Ece1 gene. Using specific sense primers, isoform-specific expression of murine ECE-1 mRNA in various mouse tissues was confirmed by RT-PCR. Using real-time PCR we demonstrated that ECE-1c was the most abundantly expressed isoform in most tissues, except for heart and aorta displaying a more even isoform distribution. We detected an additional isoform-specific exon, designated c2, which was apparently constitutively spliced and expressed only as minor fraction of ECE-1c transcripts. Our results provide evidence of structural conservation of mammalian genes encoding ECE-1 and will facilitate a more refined analysis of ECE-1 mRNA expression in the mouse model organism.

  16. Genomic organization and chromosomal localization of the human and mouse genes encoding the alpha receptor component for ciliary neurotrophic factor.

    PubMed

    Valenzuela, D M; Rojas, E; Le Beau, M M; Espinosa, R; Brannan, C I; McClain, J; Masiakowski, P; Ip, N Y; Copeland, N G; Jenkins, N A

    1995-01-01

    Ciliary neurotrophic factor (CNTF) has recently been found to share receptor components with, and to be structurally related to, a family of broadly acting cytokines, including interleukin-6, leukemia inhibitory factor, and oncostatin M. However, the CNTF receptor complex also includes a CNTF-specific component known as CNTF receptor alpha (CNTFR alpha). Here we describe the molecular cloning of the human and mouse genes encoding CNTFR. We report that the human and mouse genes have an identical intron-exon structure that correlates well with the domain structure of CNTFR alpha. That is, the signal peptide and the immunoglobulin-like domain are each encoded by single exons, the cytokine receptor-like domain is distributed among 4 exons, and the C-terminal glycosyl phosphatidylinositol recognition domain is encoded by the final coding exon. The position of the introns within the cytokine receptor-like domain corresponds to those found in other members of the cytokine receptor superfamily. Confirming a recent study using radiation hybrids, we have also mapped the human CNTFR gene to chromosome band 9p13 and the mouse gene to a syntenic region of chromosome 4.

  17. Genome wide analysis of DNA methylation and gene expression changes in the mouse lung following subchronic arsenate exposure

    EPA Science Inventory

    Alterations in DNA methylation have been proposed as a mechanism for the complex toxicological effects of arsenic. In this study, whole genome DNA methylation and gene expression changes were evaluated in lungs from female mice exposed for 90 days to 50 ppm arsenate (As) in drink...

  18. Microarray Analysis of LTR Retrotransposon Silencing Identifies Hdac1 as a Regulator of Retrotransposon Expression in Mouse Embryonic Stem Cells

    PubMed Central

    Madej, Monika J.; Taggart, Mary; Gautier, Philippe; Garcia-Perez, Jose Luis; Meehan, Richard R.; Adams, Ian R.

    2012-01-01

    Retrotransposons are highly prevalent in mammalian genomes due to their ability to amplify in pluripotent cells or developing germ cells. Host mechanisms that silence retrotransposons in germ cells and pluripotent cells are important for limiting the accumulation of the repetitive elements in the genome during evolution. However, although silencing of selected individual retrotransposons can be relatively well-studied, many mammalian retrotransposons are seldom analysed and their silencing in germ cells, pluripotent cells or somatic cells remains poorly understood. Here we show, and experimentally verify, that cryptic repetitive element probes present in Illumina and Affymetrix gene expression microarray platforms can accurately and sensitively monitor repetitive element expression data. This computational approach to genome-wide retrotransposon expression has allowed us to identify the histone deacetylase Hdac1 as a component of the retrotransposon silencing machinery in mouse embryonic stem cells, and to determine the retrotransposon targets of Hdac1 in these cells. We also identify retrotransposons that are targets of other retrotransposon silencing mechanisms such as DNA methylation, Eset-mediated histone modification, and Ring1B/Eed-containing polycomb repressive complexes in mouse embryonic stem cells. Furthermore, our computational analysis of retrotransposon silencing suggests that multiple silencing mechanisms are independently targeted to retrotransposons in embryonic stem cells, that different genomic copies of the same retrotransposon can be differentially sensitive to these silencing mechanisms, and helps define retrotransposon sequence elements that are targeted by silencing machineries. Thus repeat annotation of gene expression microarray data suggests that a complex interplay between silencing mechanisms represses retrotransposon loci in germ cells and embryonic stem cells. PMID:22570599

  19. Effects of DMSA-coated Fe3O4 magnetic nanoparticles on global gene expression of mouse macrophage RAW264.7 cells.

    PubMed

    Liu, Yingxun; Chen, Zhongping; Gu, Ning; Wang, Jinke

    2011-08-28

    Fe(3)O(4) magnetic nanoparticles (MNPs) coated with 2,3-dimercaptosuccinnic acid (DMSA) are considered to be a promising nanomaterial with biocompatibility. In the present study, the effects of DMSA-coated Fe(3)O(4) MNPs on the expression of all identified mouse genes, which regulate various cellular biological processes, were determined to establish whether this nanoparticle is cytotoxic to mammalian cells. Mouse macrophage RAW264.7 cells were treated with 100μg/ml of DMSA-coated Fe(3)O(4) MNPs for 4, 24 and 48h, and the global gene expression was detected via Affymetrix Mouse Genome 430 2.0 GeneChips(®) microarrays. It was found that gene expression of 711, 545 and 434 transcripts was significantly altered by 4-, 24- and 48-h treatments, respectively. Of these genes, 27 were consistently upregulated and 6 were consistently downregulated at the three treatment durations. Bioinformatic analysis of all differentially expressed genes revealed that this nanoparticle can strongly activate inflammatory and immune responses and can inhibit the biosynthesis and metabolism of RAW264.7 cells at a dose of 100μg/ml. These results demonstrated that DMSA-coated Fe(3)O(4) MNPs display cytotoxicity in this type of macrophage at high doses.

  20. Acquisition of biologically relevant gene expression data by Affymetrix microarray analysis of archival formalin-fixed paraffin-embedded tumours

    PubMed Central

    Linton, K M; Hey, Y; Saunders, E; Jeziorska, M; Denton, J; Wilson, C L; Swindell, R; Dibben, S; Miller, C J; Pepper, S D; Radford, J A; Freemont, A J

    2008-01-01

    Robust protocols for microarray gene expression profiling of archival formalin-fixed paraffin-embedded tissue (FFPET) are needed to facilitate research when availability of fresh-frozen tissue is limited. Recent reports attest to the feasibility of this approach, but the clinical value of these data is poorly understood. We employed state-of-the-art RNA extraction and Affymetrix microarray technology to examine 34 archival FFPET primary extremity soft tissue sarcomas. Nineteen arrays met stringent QC criteria and were used to model prognostic signatures for metastatic recurrence. Arrays from two paired frozen and FFPET samples were compared: although FFPET sensitivity was low (∼50%), high specificity (95%) and positive predictive value (92%) suggest that transcript detection is reliable. Good agreement between arrays and real time (RT)–PCR was confirmed, especially for abundant transcripts, and RT–PCR validated the regulation pattern for 19 of 24 candidate genes (overall R2=0.4662). RT–PCR and immunohistochemistry on independent cases validated prognostic significance for several genes including RECQL4, FRRS1, CFH and MET – whose combined expression carried greater prognostic value than tumour grade – and cmet and TRKB proteins. These molecules warrant further evaluation in larger series. Reliable clinically relevant data can be obtained from archival FFPET, but protocol amendments are needed to improve the sensitivity and broad application of this approach. PMID:18382428

  1. Acquisition of biologically relevant gene expression data by Affymetrix microarray analysis of archival formalin-fixed paraffin-embedded tumours.

    PubMed

    Linton, K M; Hey, Y; Saunders, E; Jeziorska, M; Denton, J; Wilson, C L; Swindell, R; Dibben, S; Miller, C J; Pepper, S D; Radford, J A; Freemont, A J

    2008-04-22

    Robust protocols for microarray gene expression profiling of archival formalin-fixed paraffin-embedded tissue (FFPET) are needed to facilitate research when availability of fresh-frozen tissue is limited. Recent reports attest to the feasibility of this approach, but the clinical value of these data is poorly understood. We employed state-of-the-art RNA extraction and Affymetrix microarray technology to examine 34 archival FFPET primary extremity soft tissue sarcomas. Nineteen arrays met stringent QC criteria and were used to model prognostic signatures for metastatic recurrence. Arrays from two paired frozen and FFPET samples were compared: although FFPET sensitivity was low ( approximately 50%), high specificity (95%) and positive predictive value (92%) suggest that transcript detection is reliable. Good agreement between arrays and real time (RT)-PCR was confirmed, especially for abundant transcripts, and RT-PCR validated the regulation pattern for 19 of 24 candidate genes (overall R(2)=0.4662). RT-PCR and immunohistochemistry on independent cases validated prognostic significance for several genes including RECQL4, FRRS1, CFH and MET - whose combined expression carried greater prognostic value than tumour grade - and cmet and TRKB proteins. These molecules warrant further evaluation in larger series. Reliable clinically relevant data can be obtained from archival FFPET, but protocol amendments are needed to improve the sensitivity and broad application of this approach.

  2. The mouse genome displays highly dynamic populations of KRAB-zinc finger protein genes and related genetic units

    PubMed Central

    Kauzlaric, Annamaria; Ecco, Gabriela; Cassano, Marco; Duc, Julien; Imbeault, Michael; Trono, Didier

    2017-01-01

    KRAB-containing poly-zinc finger proteins (KZFPs) constitute the largest family of transcription factors encoded by mammalian genomes, and growing evidence indicates that they fulfill functions critical to both embryonic development and maintenance of adult homeostasis. KZFP genes underwent broad and independent waves of expansion in many higher vertebrates lineages, yet comprehensive studies of members harbored by a given species are scarce. Here we present a thorough analysis of KZFP genes and related units in the murine genome. We first identified about twice as many elements than previously annotated as either KZFP genes or pseudogenes, notably by assigning to this family an entity formerly considered as a large group of Satellite repeats. We then could delineate an organization in clusters distributed throughout the genome, with signs of recombination, translocation, duplication and seeding of new sites by retrotransposition of KZFP genes and related genetic units (KZFP/rGUs). Moreover, we harvested evidence indicating that closely related paralogs had evolved through both drifting and shifting of sequences encoding for zinc finger arrays. Finally, we could demonstrate that the KAP1-SETDB1 repressor complex tames the expression of KZFP/rGUs within clusters, yet that the primary targets of this regulation are not the KZFP/rGUs themselves but enhancers contained in neighboring endogenous retroelements and that, underneath, KZFPs conserve highly individualized patterns of expression. PMID:28334004

  3. Genomic organization and characterization of mouse SAP, the gene that is altered in X-linked lymphoproliferative disease.

    PubMed

    Wu, C; Sayos, J; Wang, N; Howie, D; Coyle, A; Terhorst, C

    2000-08-01

    X-linked lymphoproliferative (XLP) disease is a fatal immunological disorder that renders the immune system unable to respond effectively to Epstein-Barr virus (EBV) infection. The gene that encodes a protein termed SAP or SH2D1A is either deleted or mutated in XLP patients, resulting in uncontrolled B- and T-cell proliferation upon EBV infection. Here, we report the cloning and characterization of the mouse SAP gene. It is localized on the mouse X chromosome and comprises four exons spanning approximately 25 kb. Its expression appears to be restricted to T lymphocytes. Whereas a high level of SAP expression is observed in Thl cells, only small amounts are detectable in Th2 cells. Moreover, SAP expression is down-regulated upon in vitro activation of T cells, including CD4+, CD8+ single-positive T cells, and Thl and Th2 cells. This study provides valuable information for in-depth genetic and biochemical analysis of the function of SAP in the immune system.

  4. Genomic characterisation of Eμ-Myc mouse lymphomas identifies Bcor as a Myc co-operative tumour-suppressor gene.

    PubMed

    Lefebure, Marcus; Tothill, Richard W; Kruse, Elizabeth; Hawkins, Edwin D; Shortt, Jake; Matthews, Geoffrey M; Gregory, Gareth P; Martin, Benjamin P; Kelly, Madison J; Todorovski, Izabela; Doyle, Maria A; Lupat, Richard; Li, Jason; Schroeder, Jan; Wall, Meaghan; Craig, Stuart; Poortinga, Gretchen; Cameron, Don; Bywater, Megan; Kats, Lev; Gearhart, Micah D; Bardwell, Vivian J; Dickins, Ross A; Hannan, Ross D; Papenfuss, Anthony T; Johnstone, Ricky W

    2017-03-06

    The Eμ-Myc mouse is an extensively used model of MYC driven malignancy; however to date there has only been partial characterization of MYC co-operative mutations leading to spontaneous lymphomagenesis. Here we sequence spontaneously arising Eμ-Myc lymphomas to define transgene architecture, somatic mutations, and structural alterations. We identify frequent disruptive mutations in the PRC1-like component and BCL6-corepressor gene Bcor. Moreover, we find unexpected concomitant multigenic lesions involving Cdkn2a loss and other cancer genes including Nras, Kras and Bcor. These findings challenge the assumed two-hit model of Eμ-Myc lymphoma and demonstrate a functional in vivo role for Bcor in suppressing tumorigenesis.

  5. Advantages of using the CRISPR/Cas9 system of genome editing to investigate male reproductive mechanisms using mouse models

    PubMed Central

    Young, Samantha AM; Aitken, R John; Ikawa, Masahito

    2015-01-01

    Gene disruption technology has long been beneficial for the study of male reproductive biology. However, because of the time and cost involved, this technology was not a viable method except in specialist laboratories. The advent of the CRISPR/Cas9 system of gene disruption has ushered in a new era of genetic investigation. Now, it is possible to generate gene-disrupted mouse models in very little time and at very little cost. This Highlight article discusses the application of this technology to study the genetics of male fertility and looks at some of the future uses of this system that could be used to reveal the essential and nonessential genetic components of male reproductive mechanisms. PMID:25994645

  6. Genomic characterisation of Eμ-Myc mouse lymphomas identifies Bcor as a Myc co-operative tumour-suppressor gene

    PubMed Central

    Lefebure, Marcus; Tothill, Richard W.; Kruse, Elizabeth; Hawkins, Edwin D.; Shortt, Jake; Matthews, Geoffrey M.; Gregory, Gareth P.; Martin, Benjamin P.; Kelly, Madison J.; Todorovski, Izabela; Doyle, Maria A.; Lupat, Richard; Li, Jason; Schroeder, Jan; Wall, Meaghan; Craig, Stuart; Poortinga, Gretchen; Cameron, Don; Bywater, Megan; Kats, Lev; Gearhart, Micah D.; Bardwell, Vivian J.; Dickins, Ross A.; Hannan, Ross D.; Papenfuss, Anthony T.; Johnstone, Ricky W.

    2017-01-01

    The Eμ-Myc mouse is an extensively used model of MYC driven malignancy; however to date there has only been partial characterization of MYC co-operative mutations leading to spontaneous lymphomagenesis. Here we sequence spontaneously arising Eμ-Myc lymphomas to define transgene architecture, somatic mutations, and structural alterations. We identify frequent disruptive mutations in the PRC1-like component and BCL6-corepressor gene Bcor. Moreover, we find unexpected concomitant multigenic lesions involving Cdkn2a loss and other cancer genes including Nras, Kras and Bcor. These findings challenge the assumed two-hit model of Eμ-Myc lymphoma and demonstrate a functional in vivo role for Bcor in suppressing tumorigenesis. PMID:28262675

  7. Advantages of using the CRISPR/Cas9 system of genome editing to investigate male reproductive mechanisms using mouse models.

    PubMed

    Young, Samantha A M; Aitken, R John; Ikawa, Masahito

    2015-01-01

    Gene disruption technology has long been beneficial for the study of male reproductive biology. However, because of the time and cost involved, this technology was not a viable method except in specialist laboratories. The advent of the CRISPR/Cas9 system of gene disruption has ushered in a new era of genetic investigation. Now, it is possible to generate gene-disrupted mouse models in very little time and at very little cost. This Highlight article discusses the application of this technology to study the genetics of male fertility and looks at some of the future uses of this system that could be used to reveal the essential and nonessential genetic components of male reproductive mechanisms.

  8. Introduction of a point mutation into the mouse genome by homologous recombination in embryonic stem cells using a replacement type vector with a selectable marker.

    PubMed

    Rubinstein, M; Japón, M A; Low, M J

    1993-06-11

    The introduction of small mutations instead of null alleles into the mouse genome has broad applications to the study of protein structure-function relationships and the creation of animal models of human genetic diseases. To test a simple mutational strategy we designed a targeting vector for the mouse proopiomelanocortin (POMC) gene containing a single nucleotide insertion that converts the initial tyrosine codon of beta-endorphin 1-31 to a premature translational termination codon and introduces a unique Hpal endonuclease restriction site. The targeting vector also contains a neo cassette immediately 3' to the last POMC exon and a herpes simplex virus thymidine kinase cassette to allow positive and negative selection. Homologous recombination occurred at a frequency of 1/30 clones of electroporated embryonic stem cells selected in G418 and gancyclovir. 10/11 clones identified initially by a polymerase chain reaction (PCR) strategy had the predicted structure without evidence of concatemer formation by Southern blot analysis. We used a combination of Hpa I digestion of PCR amplified fragments and direct nucleotide sequencing to further confirm that the point mutation was retained in 9/10 clones. The POMC gene was transcriptionally silent in embryonic stem cells and the targeted allele was not activated by the downstream phosphoglycerate kinase-1 promoter that transcribed the neo gene. Under the electroporation conditions used, we have demonstrated that a point mutation can be introduced with high efficiency and precision into the POMC gene using a replacement type vector containing a retained selectable marker without affecting expression of the allele in the embryonic stem cells. A similar strategy may be useful for a wide range of genes.

  9. Penetrance of craniofacial anomalies in mouse models of Smith-Magenis syndrome is modified by genomic sequence surrounding Rai1: not all null alleles are alike.

    PubMed

    Yan, Jiong; Bi, Weimin; Lupski, James R

    2007-03-01

    Craniofacial abnormality is one of the major clinical manifestations of Smith-Magenis syndrome (SMS). Previous analyses in a mixed genetic background of several SMS mouse models--including Df(11)17/+ and Df(11)17-1/+, which have 2-Mb and 590-kb deletions, respectively, and Rai1(-/+)--revealed that the penetrance of the craniofacial phenotype appears to be influenced by deletion size and genetic background. We generated an additional strain with a 1-Mb deletion intermediate in size between the two described above. Remarkably, the penetrance of its craniofacial anomalies in the mixed background was between those of Df(11)17 and Df(11)17-1. We further analyzed the deletion mutations and the Rai1(-/+) allele in a pure C57BL/6 background, to control for nonlinked modifier loci. The penetrance of the craniofacial anomalies was markedly increased for all the strains in comparison with the mixed background. Mice with Df(11)17 and Df(11)17-1 deletions had a similar penetrance, suggesting that penetrance may be less influenced by deletion size, whereas that of Rai1(-/+) mice was significantly lower than that of the deletion strains. We hypothesize that potential trans-regulatory sequence(s) or gene(s) that reside within the 590-kb genomic interval surrounding Rai1 are the major modifying genetic element(s) affecting the craniofacial penetrance. Moreover, we confirmed the influence of genetic background and different deletion sizes on the phenotype. The complicated control of the penetrance for one phenotype in SMS mouse models provides tools to elucidate molecular mechanisms for penetrance and clearly shows that a null allele caused by chromosomal deletion can have different phenotypic consequences than one caused by gene inactivation.

  10. Genomic and immunologic factors associated with viral pathogenesis in a lethal EV71 infected neonatal mouse model

    PubMed Central

    YUE, YINGYING; LI, PENG; SONG, NANNAN; LI, BINGQING; LI, ZHIHUI; GUO, YUQI; ZHANG, WEIDONG; WEI, MING Q.; GAI, ZHONGTAO; MENG, HONG; WANG, JIWEN; QIN, LIZENG

    2016-01-01

    Hand, foot and mouth disease (HFMD) caused by enterovirus 71 (EV71) has emerged as a major health problem in China and worldwide. The present study aimed to understand the virological features of EV71 and host responses resulting from EV71 infection. Six different EV71 strains were isolated from HFMD patients with severe or mild clinical symptoms, and were analyzed for pathogenicity in vitro and in vivo. The results demonstrated that the six virus strains exhibited similar cytopathogenic effects on susceptible MA104 cells. However, marked differences in histological and immunopathological changes were observed when mice were inoculated with the different virus strains. Thus, the viruses studied were divided into two groups, highly or weakly pathogenic. Two representative virus strains, JN200804 and JN200803 (highly and weakly pathogenic, respectively) were studied further to investigate pathogenicity-associated factors, including genetic mutations and immunopathogenesis. The present study has demonstrated that highly pathogenic strains have stable genome and amino acid sequences. Notably, the present study demonstrated that a highly pathogenic strain induced a significant increase of the bulk CD4 T cell levels at 3 days post-inoculation. In conclusion, the current study demonstrates that genomic and immunologic factors may be responsible for the multiple tissue damage caused by highly pathogenic EV71 infection. PMID:27035332

  11. Genomic and immunologic factors associated with viral pathogenesis in a lethal EV71 infected neonatal mouse model.

    PubMed

    Yue, Yingying; Li, Peng; Song, Nannan; Li, Bingqing; Li, Zhihui; Guo, Yuqi; Zhang, Weidong; Wei, Ming Q; Gai, Zhongtao; Meng, Hong; Wang, Jiwen; Qin, Lizeng

    2016-05-01

    Hand, foot and mouth disease (HFMD) caused by enterovirus 71 (EV71) has emerged as a major health problem in China and worldwide. The present study aimed to understand the virological features of EV71 and host responses resulting from EV71 infection. Six different EV71 strains were isolated from HFMD patients with severe or mild clinical symptoms, and were analyzed for pathogenicity in vitro and in vivo. The results demonstrated that the six virus strains exhibited similar cytopathogenic effects on susceptible MA104 cells. However, marked differences in histological and immunopathological changes were observed when mice were inoculated with the different virus strains. Thus, the viruses studied were divided into two groups, highly or weakly pathogenic. Two representative virus strains, JN200804 and JN200803 (highly and weakly pathogenic, respectively) were studied further to investigate pathogenicity-associated factors, including genetic mutations and immunopathogenesis. The present study has demonstrated that highly pathogenic strains have stable genome and amino acid sequences. Notably, the present study demonstrated that a highly pathogenic strain induced a significant increase of the bulk CD4 T cell levels at 3 days post‑inoculation. In conclusion, the current study demonstrates that genomic and immunologic factors may be responsible for the multiple tissue damage caused by highly pathogenic EV71 infection.

  12. Mutation at the Polymerase Active Site of Mouse DNA Polymerase δ Increases Genomic Instability and Accelerates Tumorigenesis▿

    PubMed Central

    Venkatesan, Ranga N.; Treuting, Piper M.; Fuller, Evan D.; Goldsby, Robert E.; Norwood, Thomas H.; Gooley, Ted A.; Ladiges, Warren C.; Preston, Bradley D.; Loeb, Lawrence A.

    2007-01-01

    Mammalian DNA polymerase δ (Pol δ) is believed to replicate a large portion of the genome and to synthesize DNA in DNA repair and genetic recombination pathways. The effects of mutation in the polymerase domain of this essential enzyme are unknown. Here, we generated mice harboring an L604G or L604K substitution in highly conserved motif A in the polymerase active site of Pol δ. Homozygous Pold1L604G/L604G and Pold1L604K/L604K mice died in utero. However, heterozygous animals were viable and displayed no overall increase in disease incidence, indicative of efficient compensation for the defective mutant polymerase. The life spans of wild-type and heterozygous Pold1+/L604G mice did not differ, while that of Pold1+/L604K mice was reduced by 18%. Cultured embryonic fibroblasts from the heterozygous strains exhibited comparable increases in both spontaneous mutation rate and chromosome aberrations. We observed no significant increase in cancer incidence; however, Pold1+/L604K mice bearing histologically diagnosed tumors died at a younger median age than wild-type mice. Our results indicate that heterozygous mutation at L604 in the polymerase active site of DNA polymerase δ reduces life span, increases genomic instability, and accelerates tumorigenesis in an allele-specific manner, novel findings that have implications for human cancer. PMID:17785453

  13. A genome-wide in situ hybridization map of RNA-binding proteins reveals anatomically restricted expression in the developing mouse brain

    PubMed Central

    McKee, Adrienne E; Minet, Emmanuel; Stern, Charlene; Riahi, Shervin; Stiles, Charles D; Silver, Pamela A

    2005-01-01

    Background In eukaryotic cells, RNA-binding proteins (RBPs) contribute to gene expression by regulating the form, abundance, and stability of both coding and non-coding RNA. In the vertebrate brain, RBPs account for many distinctive features of RNA processing such as activity-dependent transcript localization and localized protein synthesis. Several RBPs with activities that are important for the proper function of adult brain have been identified, but how many RBPs exist and where these genes are expressed in the developing brain is uncharacterized. Results Here we describe a comprehensive catalogue of the unique RBPs encoded in the mouse genome and provide an online database of RBP expression in developing brain. We identified 380 putative RBPs in the mouse genome. Using in situ hybridization, we visualized the expression of 323 of these RBP genes in the brains of developing mice at embryonic day 13.5, when critical fate choice decisions are made and at P0, when major structural components of the adult brain are apparent. We demonstrate i) that 16 of the 323 RBPs examined show neural-specific expression at the stages we examined, and ii) that a far larger subset (221) shows regionally restricted expression in the brain. Of the regionally restricted RBPs, we describe one group that is preferentially expressed in the E13.5 ventricular areas and a second group that shows spatially restricted expression in post-mitotic regions of the embryonic brain. Additionally, we find a subset of RBPs that share the same complex pattern of expression, in proliferating regions of the embryonic and postnatal NS and peripheral tissues. Conclusion Our data show that, in contrast to their proposed ubiquitous involvement in gene regulation, most RBPs are not uniformly expressed. Here we demonstrate the region-specific expression of RBPs in proliferating vs. post-mitotic brain regions as well as cell-type-specific RBP expression. We identify uncharacterized RBPs that exhibit neural

  14. Generating a transgenic mouse line stably expressing human MHC surface antigen from a HAC carrying multiple genomic BACs.

    PubMed

    Hasegawa, Yoshinori; Ishikura, Tomoyuki; Hasegawa, Takanori; Watanabe, Takashi; Suzuki, Junpei; Nakayama, Manabu; Okamura, Yoshiaki; Okazaki, Tuneko; Koseki, Haruhiko; Ohara, Osamu; Ikeno, Masashi; Masumoto, Hiroshi

    2015-03-01

    The human artificial chromosome (HAC) vector is a promising tool to improve the problematic suppression and position effects of transgene expression frequently seen in transgenic cells and animals produced by conventional plasmid or viral vectors. We generated transgenic mice maintaining a single HAC vector carrying two genomic bacterial artificial chromosomes (BACs) from human HLA-DR loci (DRA and DRB1). Both transgenes on the HAC in transgenic mice exhibited tissue-specific expression in kidney, liver, lung, spleen, lymph node, bone marrow, and thymus cells in RT-PCR analysis. Stable functional expression of a cell surface HLA-DR marker from both transgenes, DRA and DRB1 on the HAC, was detected by flow cytometric analysis of splenocytes and maintained through at least eight filial generations. These results indicate that the de novo HAC system can allow us to manipulate multiple BAC transgenes with coordinated expression as a surface antigen through the generation of transgenic animals.

  15. Statistical evaluation of transcriptomic data generated using the Affymetrix one-cycle, two-cycle and IVT-Express RNA labelling protocols with the Arabidopsis ATH1 microarray

    PubMed Central

    2010-01-01

    Background Microarrays are a powerful tool used for the determination of global RNA expression. There is an increasing requirement to focus on profiling gene expression in tissues where it is difficult to obtain large quantities of material, for example individual tissues within organs such as the root, or individual isolated cells. From such samples, it is difficult to produce the amount of RNA required for labelling and hybridisation in microarray experiments, thus a process of amplification is usually adopted. Despite the increasing use of two-cycle amplification for transcriptomic analyses on the Affymetrix ATH1 array, there has been no report investigating any potential bias in gene representation that may occur as a result. Results Here we compare transcriptomic data generated using Affymetrix one-cycle (standard labelling protocol), two-cycle (small-sample protocol) and IVT-Express protocols with the Affymetrix ATH1 array using Arabidopsis root samples. Results obtained with each protocol are broadly similar. However, we show that there are 35 probe sets (of a total of 22810) that are misrepresented in the two-cycle data sets. Of these, 33 probe sets were classed as mis-amplified when comparisons of two independent publicly available data sets were undertaken. Conclusions Given the unreliable nature of the highlighted probes, we caution against using data associated with the corresponding genes in analyses involving transcriptomic data generated with two-cycle amplification protocols. We have shown that the Affymetrix IVT-E labelling protocol produces data with less associated bias than the two-cycle protocol, and as such, would recommend this kit for new experiments that involve small samples. PMID:20230623

  16. Site-specific modification of genome with cell-permeable Cre fusion protein in preimplantation mouse embryo

    SciTech Connect

    Kim, Kyoungmi; Kim, Hwain; Lee, Daekee

    2009-10-09

    Site-specific recombination (SSR) by Cre recombinase and its target sequence, loxP, is a valuable tool in genetic analysis of gene function. Recently, several studies reported successful application of Cre fusion protein containing protein transduction peptide for inducing gene modification in various mammalian cells including ES cell as well as in the whole animal. In this study, we show that a short incubation of preimplantation mouse embryos with purified cell-permeable Cre fusion protein results in efficient SSR. X-Gal staining of preimplantation embryos, heterozygous for Gtrosa26{sup tm1Sor}, revealed that treatment of 1-cell or 2-cell embryos with 3 {mu}M of Cre fusion protein for 2 h leads to Cre-mediated excision in 70-85% of embryos. We have examined the effect of the concentration of the Cre fusion protein and the duration of the treatment on embryonic development, established a condition for full term development and survival to adulthood, and demonstrated the germ line transmission of excised Gtrosa26 allele. Potential applications and advantages of the highly efficient technique described here are discussed.

  17. Non-genomic transmission of paternal behaviour between fathers and sons in the monogamous and biparental California mouse.

    PubMed

    Gleason, Erin D; Marler, Catherine A

    2013-07-22

    Maternal behaviour has profound, long-lasting implications for the health and well-being of developing offspring. In the monogamous California mouse (Peromyscus californicus), care by both parents is critical for offspring survival. We tested the hypothesis that similar to maternal care in rodents, paternal huddling and grooming (HG) behaviour can be transmitted to future generations via behavioural mechanisms. In California mice, testosterone maintains paternal HG behaviour. In the present study, we randomly assigned a group of male California mice to castration or sham-operated conditions and allowed them to raise their offspring normally. Adult sons of these males were paired with a female, and they were observed interacting with their own offspring. We found that like their fathers, the sons of castrated males huddled and groomed their young at lower levels than the sons of sham-operated fathers. The sons of castrates also retrieved pups more frequently. When both parents were present, the sons of castrates also showed a trend towards engaging in less exploratory behaviour. These data support the hypothesis that paternal behaviour, like maternal behaviour, can be transferred to future generations via epigenetic mechanisms and suggest that in a biparental species both parents contribute to offspring behavioural development.

  18. The PRC2-binding long non-coding RNAs in human and mouse genomes are associated with predictive sequence features

    PubMed Central

    Tu, Shiqi; Yuan, Guo-Cheng; Shao, Zhen

    2017-01-01

    Recently, long non-coding RNAs (lncRNAs) have emerged as an important class of molecules involved in many cellular processes. One of their primary functions is to shape epigenetic landscape through interactions with chromatin modifying proteins. However, mechanisms contributing to the specificity of such interactions remain poorly understood. Here we took the human and mouse lncRNAs that were experimentally determined to have physical interactions with Polycomb repressive complex 2 (PRC2), and systematically investigated the sequence features of these lncRNAs by developing a new computational pipeline for sequences composition analysis, in which each sequence is considered as a series of transitions between adjacent nucleotides. Through that, PRC2-binding lncRNAs were found to be associated with a set of distinctive and evolutionarily conserved sequence features, which can be utilized to distinguish them from the others with considerable accuracy. We further identified fragments of PRC2-binding lncRNAs that are enriched with these sequence features, and found they show strong PRC2-binding signals and are more highly conserved across species than the other parts, implying their functional importance. PMID:28139710

  19. The PRC2-binding long non-coding RNAs in human and mouse genomes are associated with predictive sequence features

    NASA Astrophysics Data System (ADS)

    Tu, Shiqi; Yuan, Guo-Cheng; Shao, Zhen

    2017-01-01

    Recently, long non-coding RNAs (lncRNAs) have emerged as an important class of molecules involved in many cellular processes. One of their primary functions is to shape epigenetic landscape through interactions with chromatin modifying proteins. However, mechanisms contributing to the specificity of such interactions remain poorly understood. Here we took the human and mouse lncRNAs that were experimentally determined to have physical interactions with Polycomb repressive complex 2 (PRC2), and systematically investigated the sequence features of these lncRNAs by developing a new computational pipeline for sequences composition analysis, in which each sequence is considered as a series of transitions between adjacent nucleotides. Through that, PRC2-binding lncRNAs were found to be associated with a set of distinctive and evolutionarily conserved sequence features, which can be utilized to distinguish them from the others with considerable accuracy. We further identified fragments of PRC2-binding lncRNAs that are enriched with these sequence features, and found they show strong PRC2-binding signals and are more highly conserved across species than the other parts, implying their functional importance.

  20. Genome-wide analysis of DNA methylation during antagonism of DMOG to MnCl2-induced cytotoxicity in the mouse substantia nigra

    PubMed Central

    Yang, Nannan; Wei, Yang; Wang, Tan; Guo, Jifeng; Sun, Qiying; Hu, Yacen; Yan, Xinxiang; Zhu, Xiongwei; Tang, Beisha; Xu, Qian

    2016-01-01

    Exposure to excessive manganese (Mn) causes manganism, a progressive neurodegenerative disorder similar to idiopathic Parkinson’s disease (IPD). The detailed mechanisms of Mn neurotoxicity in nerve cells, especially in dopaminergic neurons are not yet fully understood. Meanwhile, it is unknown whether there exists a potential antagonist or effective drug for treating neuron damage in manganism. In the present study, we report the discovery of an HIF prolyl-hydroxylase inhibitor, DMOG [N-(2-Methoxy-2-oxoacetyl) glycine methyl ester], that can partially inhibit manganese toxicity not only in the neuroblastoma cell line SH-SY5Y in vitro but also in a mouse model in vivo. A genome-wide methylation DNA analysis was performed using microarray hybridization. Intriguingly, DNA methylation in the promoter region of 226 genes was found to be regulated by MnCl2, while the methylation effects of MnCl2 could be restored with combinatorial DMOG treatment. Furthermore, we found that genes with converted promoter methylation during DMOG antagonism were associated across several categories of molecular function, including mitochondria integrity maintain, cell cycle and DNA damage response, and ion transportation. Collectively, our results serve as the basis of a mechanism analysis of neuron damage in manganism and may supply possible gene targets for clinical therapy. PMID:27380887

  1. Zinc-finger transcription factors are associated with guanine quadruplex motifs in human, chimpanzee, mouse and rat promoters genome-wide.

    PubMed

    Kumar, Pankaj; Yadav, Vinod Kumar; Baral, Aradhita; Kumar, Parveen; Saha, Dhurjhoti; Chowdhury, Shantanu

    2011-10-01

    Function of non-B DNA structures are poorly understood though several bioinformatics studies predict role of the G-quadruplex DNA structure in transcription. Earlier, using transcriptome profiling we found evidence of widespread G-quadruplex-mediated gene regulation. Herein, we asked whether potential G-quadruplex (PG4) motifs associate with transcription factors (TF). This was analyzed using 220 position weight matrices [designated as transcription factor binding sites (TFBS)], representing 187 unique TF, in >75,000 genes in human, chimpanzee, mouse and rat. Results show binding sites of nine TFs, including that of AP-2, SP1, MAZ and VDR, occurred significantly within 100 bases of the PG4 motif (P < 1.24E-10). PG4-TFBS combinations were conserved in 'orthologously' related promoters across all four organisms and were associated with >850 genes in each genome. Remarkably, seven of the nine TFs were zinc-finger binding proteins indicating a novel characteristic of PG4 motifs. To test these findings, transcriptome profiles from human cell lines treated with G-quadruplex-specific molecules were used; 66 genes were significantly differentially expressed across both cell-types, which also harbored conserved PG4 motifs along with one/more of the nine TFBS. In addition, genes regulated by PG4-TFBS combinations were found to be co-regulated in human tissues, further emphasizing the regulatory significance of the associations.

  2. Genome-wide analysis of expression modes and DNA methylation status at sense-antisense transcript loci in mouse.

    PubMed

    Watanabe, Yutaka; Numata, Koji; Murata, Shinya; Osada, Yuko; Saito, Rintaro; Nakaoka, Hajime; Yamamoto, Naoyuki; Watanabe, Kazufumi; Kato, Hidemasa; Abe, Kuniya; Kiyosawa, Hidenori

    2010-12-01

    The functionality of sense-antisense transcripts (SATs), although widespread throughout the mammalian genome, is largely unknown. Here, we analyzed the SATs expression and its associated promoter DNA methylation status by surveying 12 tissues of mice to gain insights into the relationship between expression and DNA methylation of SATs. We have found that sense and antisense expression positively correlate in most tissues. However, in some SATs with tissue-specific expression, the expression level of a transcript from a CpG island-bearing promoter is low when the promoter DNA methylation is present. In these circumstances, the expression level of its opposite-strand transcript, especially when it is poly(A)-negative was coincidentally higher. These observations suggest that, albeit the general tendency of sense-antisense simultaneous expression, some antisense transcripts have coordinated expression with its counterpart sense gene promoter methylation. This cross-strand relationship is not a privilege of imprinted genes but seems to occur widely in SATs.

  3. Genome-Wide Alteration of Histone H3K9 Acetylation Pattern in Mouse Offspring Prenatally Exposed to Arsenic

    PubMed Central

    Cronican, Andrea A.; Fitz, Nicholas F.; Carter, Alexis; Saleem, Muzamil; Shiva, Sruti; Barchowsky, Aaron; Koldamova, Radosveta; Schug, Jonathan; Lefterov, Iliya

    2013-01-01

    Chronic exposure to arsenic in drinking water, especially in utero or perinatal exposure, can initiate neurological and cognitive dysfunction, as well as memory impairment. Several epidemiological studies have demonstrated cognitive and learning deficits in children with early exposure to low to moderate levels of arsenic, but pathogenic mechanisms or etiology for these deficits are poorly understood. Since in vivo studies show a role for histone acetylation in cognitive performance and memory formation, we examined if prenatal exposure to arsenic causes changes in the epigenomic landscape. We exposed C57Bl6/J mice to 100 μg/L arsenic in the drinking water starting 1 week before conception till birth and applied chromatin immunoprecipitation followed by high-throughput massive parallel sequencing (ChIP-seq) to evaluate H3K9 acetylation pattern in the offspring of exposed and control mice. Arsenic exposure during embryonic life caused global hypo-acetylation at H3K9 and changes in functional annotation with highly significant representation of Krüppel associated box (KRAB) transcription factors in brain samples from exposed pups. We also found that arsenic exposure of adult mice impaired spatial and episodic memory, as well as fear conditioning performance. This is the first study to demonstrate: a) genome wide changes in H3K9 acetylation pattern in an offspring prenatally exposed to arsenic, and b) a connection between moderate arsenic exposure and cognitive impairment in adult mice. The results also emphasize the applicability of Next Generation Sequencing methodology in studies aiming to reveal the role of environmental factors, other than dietary restriction, in developmental reprogramming through histone modifications during embryonic development. PMID:23405071

  4. Differential gene expression in mouse liver associated with the hepatoprotective effect of clofibrate

    SciTech Connect

    Moffit, Jeffrey S.; Koza-Taylor, Petra H.; Holland, Ricky D.; Thibodeau, Michael S.; Beger, Richard D.; Lawton, Michael P.; Manautou, Jose E. . E-mail: jose.manautou@uconn.edu

    2007-07-15

    Pretreatment of mice with the peroxisome proliferator clofibrate (CFB) protects against acetaminophen (APAP)-induced hepatotoxicity. Previous studies have shown that activation of the nuclear peroxisome proliferator activated receptor-alpha (PPAR{alpha}) is required for this effect. The present study utilizes gene expression profile analysis to identify potential pathways contributing to PPAR{alpha}-mediated hepatoprotection. Gene expression profiles were compared between wild type and PPAR{alpha}-null mice pretreated with vehicle or CFB (500 mg/kg, i.p., daily for 10 days) and then challenged with APAP (400 mg/kg, p.o.). Total hepatic RNA was isolated 4 h after APAP treatment and hybridized to Affymetrix Mouse Genome MGU74 v2.0 GeneChips. Gene expression analysis was performed utilizing GeneSpring (registered) software. Our analysis identified 53 genes of interest including vanin-1, cell cycle regulators, lipid-metabolizing enzymes, and aldehyde dehydrogenase 2, an acetaminophen binding protein. Vanin-1 could be important for CFB-mediated hepatoprotection because this protein is involved in the synthesis of cysteamine and cystamine. These are potent antioxidants capable of ameliorating APAP toxicity in rodents and humans. HPLC-ESI/MS/MS analysis of liver extracts indicates that enhanced vanin-1 gene expression results in elevated cystamine levels, which could be mechanistically associated with CFB-mediated hepatoprotection.

  5. Genome-Wide Mouse Mutagenesis Reveals CD45-Mediated T Cell Function as Critical in Protective Immunity to HSV-1

    PubMed Central

    Caignard, Grégory; Leiva-Torres, Gabriel A.; Leney-Greene, Michael; Charbonneau, Benoit; Dumaine, Anne; Fodil-Cornu, Nassima; Pyzik, Michal; Cingolani, Pablo; Schwartzentruber, Jeremy; Dupaul-Chicoine, Jeremy; Guo, Huaijian; Saleh, Maya; Veillette, André; Lathrop, Marc; Blanchette, Mathieu; Majewski, Jacek; Pearson, Angela; Vidal, Silvia M.

    2013-01-01

    Herpes simplex encephalitis (HSE) is a lethal neurological disease resulting from infection with Herpes Simplex Virus 1 (HSV-1). Loss-of-function mutations in the UNC93B1, TLR3, TRIF, TRAF3, and TBK1 genes have been associated with a human genetic predisposition to HSE, demonstrating the UNC93B-TLR3-type I IFN pathway as critical in protective immunity to HSV-1. However, the TLR3, UNC93B1, and TRIF mutations exhibit incomplete penetrance and represent only a minority of HSE cases, perhaps reflecting the effects of additional host genetic factors. In order to identify new host genes, proteins and signaling pathways involved in HSV-1 and HSE susceptibility, we have implemented the first genome-wide mutagenesis screen in an in vivo HSV-1 infectious model. One pedigree (named P43) segregated a susceptible trait with a fully penetrant phenotype. Genetic mapping and whole exome sequencing led to the identification of the causative nonsense mutation L3X in the Receptor-type tyrosine-protein phosphatase C gene (PtprcL3X), which encodes for the tyrosine phosphatase CD45. Expression of MCP1, IL-6, MMP3, MMP8, and the ICP4 viral gene were significantly increased in the brain stems of infected PtprcL3X mice accounting for hyper-inflammation and pathological damages caused by viral replication. PtprcL3X mutation drastically affects the early stages of thymocytes development but also the final stage of B cell maturation. Transfer of total splenocytes from heterozygous littermates into PtprcL3X mice resulted in a complete HSV-1 protective effect. Furthermore, T cells were the only cell population to fully restore resistance to HSV-1 in the mutants, an effect that required both the CD4+ and CD8+ T cells and could be attributed to function of CD4+ T helper 1 (Th1) cells in CD8+ T cell recruitment to the site of infection. Altogether, these results revealed the CD45-mediated T cell function as potentially critical for infection and viral spread to the brain, and also for subsequent

  6. Integration of Genome-Wide Computation DRE Search, AhR ChIP-chip and Gene Expression Analyses of TCDD-Elicited Responses in the Mouse Liver

    PubMed Central

    2011-01-01

    Background The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor (TF) that mediates responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Integration of TCDD-induced genome-wide AhR enrichment, differential gene expression and computational dioxin response element (DRE) analyses further elucidate the hepatic AhR regulatory network. Results Global ChIP-chip and gene expression analyses were performed on hepatic tissue from immature ovariectomized mice orally gavaged with 30 μg/kg TCDD. ChIP-chip analysis identified 14,446 and 974 AhR enriched regions (1% false discovery rate) at 2 and 24 hrs, respectively. Enrichment density was greatest in the proximal promoter, and more specifically, within ± 1.5 kb of a transcriptional start site (TSS). AhR enrichment also occurred distal to a TSS (e.g. intergenic DNA and 3' UTR), extending the potential gene expression regulatory roles of the AhR. Although TF binding site analyses identified over-represented DRE sequences within enriched regions, approximately 50% of all AhR enriched regions lacked a DRE core (5'-GCGTG-3'). Microarray analysis identified 1,896 number of TCDD-responsive genes (|fold change| ≥ 1.5, P1(t) > 0.999). Integrating this gene expression data with our ChIP-chip and DRE analyses only identified 625 differentially expressed genes that involved an AhR interaction at a DRE. Functional annotation analysis of differentially regulated genes associated with AhR enrichment identified overrepresented processes related to fatty acid and lipid metabolism and transport, and xenobiotic metabolism, which are consistent with TCDD-elicited steatosis in the mouse liver. Conclusions Details of the AhR regulatory network have been expanded to include AhR-DNA interactions within intragenic and intergenic genomic regions. Moreover, the AhR can interact with DNA independent of a DRE core suggesting there are alternative mechanisms of AhR-mediated gene regulation. PMID:21762485

  7. A Microarray Analysis for Differential Gene Expression in the Soybean Genome Using Bioconductor and R

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper describes specific procedures for conducting quality assessment of Affymetrix GeneChip® soybean genome data and performing analyses to determine differential gene expression using the open-source R language and environment in conjunction with the open-source Bioconductor package. Procedu...

  8. Disruption of O-GlcNAc cycling by deletion of O-GlcNAcase (Oga/Mgea5) changed gene expression pattern in mouse embryonic fibroblast (MEF) cells.

    PubMed

    Keembiyehetty, Chithra

    2015-09-01

    Adding a single O-GlcNAc moiety to a Ser/Thr molecule of a protein by O-GlcNAc transferase and transiently removing it by O-GlcNAcase is referred to as O-GlcNAc cycling (or O-GlcNAcylation). This O-GlcNAc modification is sensitive to nutrient availability and also shows cross talk with phosphorylation signaling, affecting downstream targets. A mouse model system was developed and evaluated to show genome wide transcriptional changes associated with disruption of O-GlcNAc cycling. Mouse embryonic fibroblast cells derived from O-GlcNAcase (Oga) knock out (KO), heterozygous (Het) and wild type (WT) embryos were used for an Affymetrix based microarray. Results are deposited in GEO dataset GSE52721. Data reveals that Oga KO MEFs had 2534 transcripts differentially expressed at 1.5 fold level while Oga heterozygous MEFs had 959 transcripts changed compared to WT MEFs. There were 1835 transcripts differentially expressed at 1.5 fold Het versus WT comparison group. Gene ontology analysis indicated differentially expressed genes enriched in metabolic, growth, and cell proliferation categories.

  9. Did Androgen-Binding Protein Paralogs Undergo Neo- and/or Subfunctionalization as the Abp Gene Region Expanded in the Mouse Genome?

    PubMed Central

    Karn, Robert C.; Chung, Amanda G.; Laukaitis, Christina M.

    2014-01-01

    The Androgen-binding protein (Abp) region of the mouse genome contains 30 Abpa genes encoding alpha subunits and 34 Abpbg genes encoding betagamma subunits, their products forming dimers composed of an alpha and a betagamma subunit. We endeavored to determine how many Abp genes are expressed as proteins in tears and saliva, and as transcripts in the exocrine glands producing them. Using standard PCR, we amplified Abp transcripts from cDNA libraries of C57BL/6 mice and found fifteen Abp gene transcripts in the lacrimal gland and five in the submandibular gland. Proteomic analyses identified proteins corresponding to eleven of the lacrimal gland transcripts, all of them different from the three salivary ABPs reported previously. Our qPCR results showed that five of the six transcripts that lacked corresponding proteins are expressed at very low levels compared to those transcripts with proteins. We found 1) no overlap in the repertoires of expressed Abp paralogs in lacrimal gland/tears and salivary glands/saliva; 2) substantial sex-limited expression of lacrimal gland/tear expressed-paralogs in males but no sex-limited expression in females; and 3) that the lacrimal gland/tear expressed-paralogs are found exclusively in ancestral clades 1, 2 and 3 of the five clades described previously while the salivary glands/saliva expressed-paralogs are found only in clade 5. The number of instances of extremely low levels of transcription without corresponding protein production in paralogs specific to tears and saliva suggested the role of subfunctionalization, a derived condition wherein genes that may have been expressed highly in both glands ancestrally were down-regulated subsequent to duplication. Thus, evidence for subfunctionalization can be seen in our data and we argue that the partitioning of paralog expression between lacrimal and salivary glands that we report here occurred as the result of adaptive evolution. PMID:25531410

  10. Did androgen-binding protein paralogs undergo neo- and/or Subfunctionalization as the Abp gene region expanded in the mouse genome?

    PubMed

    Karn, Robert C; Chung, Amanda G; Laukaitis, Christina M

    2014-01-01

    The Androgen-binding protein (Abp) region of the mouse genome contains 30 Abpa genes encoding alpha subunits and 34 Abpbg genes encoding betagamma subunits, their products forming dimers composed of an alpha and a betagamma subunit. We endeavored to determine how many Abp genes are expressed as proteins in tears and saliva, and as transcripts in the exocrine glands producing them. Using standard PCR, we amplified Abp transcripts from cDNA libraries of C57BL/6 mice and found fifteen Abp gene transcripts in the lacrimal gland and five in the submandibular gland. Proteomic analyses identified proteins corresponding to eleven of the lacrimal gland transcripts, all of them different from the three salivary ABPs reported previously. Our qPCR results showed that five of the six transcripts that lacked corresponding proteins are expressed at very low levels compared to those transcripts with proteins. We found 1) no overlap in the repertoires of expressed Abp paralogs in lacrimal gland/tears and salivary glands/saliva; 2) substantial sex-limited expression of lacrimal gland/tear expressed-paralogs in males but no sex-limited expression in females; and 3) that the lacrimal gland/tear expressed-paralogs are found exclusively in ancestral clades 1, 2 and 3 of the five clades described previously while the salivary glands/saliva expressed-paralogs are found only in clade 5. The number of instances of extremely low levels of transcription without corresponding protein production in paralogs specific to tears and saliva suggested the role of subfunctionalization, a derived condition wherein genes that may have been expressed highly in both glands ancestrally were down-regulated subsequent to duplication. Thus, evidence for subfunctionalization can be seen in our data and we argue that the partitioning of paralog expression between lacrimal and salivary glands that we report here occurred as the result of adaptive evolution.

  11. Concentration- and time-dependent genomic changes in the mouse urinary bladder following exposure to arsenate in drinking water for up to 12 weeks.

    PubMed

    Clewell, H J; Thomas, R S; Kenyon, E M; Hughes, M F; Adair, B M; Gentry, P R; Yager, J W

    2011-10-01

    Inorganic arsenic (As(i)) is a known human bladder carcinogen. The objective of this study was to examine the concentration dependence of the genomic response to As(i) in the urinary bladders of mice. C57BL/6J mice were exposed for 1 or 12 weeks to arsenate in drinking water at concentrations of 0.5, 2, 10, and 50 mg As/l. Urinary bladders were analyzed using gene expression microarrays. A consistent reversal was observed in the direction of gene expression change: from predominantly decreased expression at 1 week to predominantly increased expression at 12 weeks. These results are consistent with evidence from in vitro studies of an acute adaptive response that is suppressed on longer exposure due to downregulation of Fos. Pathways with the highest enrichment in gene expression changes were associated with epithelial-to-mesenchymal transition, inflammation, and proliferation. Benchmark dose (BMD) analysis determined that the lowest median BMD values for pathways were above 5 mg As/l, despite the fact that pathway enrichment was observed at the 0.5 mg As/l exposure concentration. This disparity may result from the nonmonotonic nature of the concentration-responses for the expression changes of a number of genes, as evidenced by the much fewer gene expression changes at 2 mg As/l compared with lower or higher concentrations. Pathway categories with concentration-related gene expression changes included cellular morphogenesis, inflammation, apoptosis/survival, cell cycle control, and DNA damage response. The results of this study provide evidence of a concentration-dependent transition in the mode of action for the subchronic effects of As(i) in mouse bladder cells in the vicinity of 2 mg As(i)/l.

  12. Highly and moderately aggressive mouse ovarian cancer cell lines exhibit differential gene expression

    PubMed Central

    Zhang, Wensheng; Kale, Shubha P.; McFerrin, Harris; Davenport, Ian; Wang, Guangdi; Skripnikova, Elena; Li, Xiao-Lin; Bowen, Nathan J.; McDaniels, Leticia B; Meng, Yuan-Xiang; Polk, Paula; Liu, Yong-Yu; Zhang, Qian-Jin

    2017-01-01

    Patients with advanced epithelial ovarian cancer often experience disease recurrence after standard therapies, a critical factor in determining their five-year survival rate. Recent reports indicated that long-term or short-term survival is associated with varied gene expression of cancer cells. Thus, identification of novel prognostic biomarkers should be considered. Since the mouse genome is similar to the human genome, we explored potential prognostic biomarkers using two groups of mouse ovarian cancer cell lines (group 1: IG-10, IG-10pw, and IG-10pw/agar; group 2: IG-10 clones 2, 3, and 11) which display highly and moderately aggressive phenotypes in vivo. Mice injected with these cell lines have different survival time and rates, capacities of tumor, and ascites formations, reflecting different prognostic potentials. Using an Affymetrix Mouse Genome 430 2.0 Array, a total of 181 genes were differentially expressed (P<0.01) by at least twofold between two groups of the cell lines. Of the 181 genes, 109 and 72 genes were overexpressed in highly and moderately aggressive cell lines, respectively. Analysis of the 109 and 72 genes using Ingenuity Pathway Analysis (IPA) tool revealed two cancer-related gene networks. One was associated with the highly aggressive cell lines and affiliated with MYC gene, and another was associated with the moderately aggressive cell lines and affiliated with the androgen receptor (AR). Finally, the gene enrichment analysis indicated that the overexpressed 89 genes (out of 109 genes) in highly aggressive cell lines had a function annotation in the David database. The cancer-relevant significant gene ontology (GO) terms included Cell cycle, DNA metabolic process, and Programmed cell death. None of the genes from a set of the 72 genes overexpressed in the moderately aggressive cell lines had a function annotation in the David database. Our results suggested that the overexpressed MYC and 109 gene set represented highly aggressive ovarian

  13. Age affects gene expression in mouse spermatogonial stem/progenitor cells.

    PubMed

    Kokkinaki, Maria; Lee, Tin-Lap; He, Zuping; Jiang, Jiji; Golestaneh, Nady; Hofmann, Marie-Claude; Chan, Wai-Yee; Dym, Martin

    2010-06-01

    Spermatogenesis in man starts with spermatogonial stem cells (SSCs), and leads to the production of sperm in approximately 64 days, common to old and young men. Sperm from elderly men are functional and able to fertilize eggs and produce offspring, even though daily sperm production is more than 50% lower and damage to sperm DNA is significantly higher in older men than in those who are younger. Our hypothesis is that the SSC/spermatogonial progenitors themselves age. To test this hypothesis, we studied the gene expression profile of mouse SSC/progenitor cells at several ages using microarrays. After sequential enzyme dispersion, we purified the SSC/progenitors with immunomagnetic cell sorting using an antibody to GFRA1, a known SSC/progenitor cell marker. RNA was isolated and used for the in vitro synthesis of amplified and labeled cRNAs that were hybridized to the Affymetrix mouse genome microarrays. The experiments were repeated twice with different cell preparations, and statistically significant results are presented. Quantitative RT-PCR analysis was used to confirm the microarray results. Comparison of four age groups (6 days, 21 days, 60 days, and 8 months old) showed a number of genes that were expressed specifically in the older mice. Two of them (i.e. Icam1 and Selp) have also been shown to mark aging hematopoietic stem cells. On the other hand, the expression levels of the genes encoding the SSC markers Gfra1 and Plzf did not seem to be significantly altered by age, indicating that age affects only certain SSC/progenitor properties.

  14. Methods comparison for high-resolution transcriptional analysis of archival material on Affymetrix Plus 2.0 and Exon 1.0 microarrays.

    PubMed

    Linton, Kim; Hey, Yvonne; Dibben, Sian; Miller, Crispin; Freemont, Anthony; Radford, John; Pepper, Stuart

    2009-07-01

    Microarray gene expression profiling of formalin-fixed paraffin-embedded (FFPE) tissues is a new and evolving technique. This report compares transcript detection rates on Affymetrix U133 Plus 2.0 and Human Exon 1.0 ST GeneChips across several RNA extraction and target labeling protocols, using routinely collected archival FFPE samples. All RNA extraction protocols tested (Ambion-Optimum, Ambion-RecoverAll, and Qiagen-RNeasy FFPE) provided extracts suitable for microarray hybridization. Compared with Affymetrix One-Cycle labeled extracts, NuGEN system protocols utilizing oligo(dT) and random hexamer primers, and cDNA target preparations instead of cRNA, achieved percent present rates up to 55% on Plus 2.0 arrays. Based on two paired-sample analyses, at 90% specificity this equalled an average 30 percentage-point increase (from 50% to 80%) in FFPE transcript sensitivity relative to fresh frozen tissues, which we have assumed to have 100% sensitivity and specificity. The high content of Exon arrays, with multiple probe sets per exon, improved FFPE sensitivity to 92% at 96% specificity, corresponding to an absolute increase of ~600 genes over Plus 2.0 arrays. While larger series are needed to confirm high correspondence between fresh-frozen and FFPE expression patterns, these data suggest that both Plus 2.0 and Exon arrays are suitable platforms for FFPE microarray expression analyses.

  15. 9. international mouse genome conference

    SciTech Connect

    1995-12-31

    This conference was held November 12--16, 1995 in Ann Arbor, Michigan. The purpose of this conference was to provide a multidisciplinary forum for exchange of state-of-the-art information on genetic mapping in mice. This report contains abstracts of presentations, focusing on the following areas: mutation identification; comparative mapping; informatics and complex traits; mutagenesis; gene identification and new technology; and genetic and physical mapping.

  16. Computational Integration of Structural and Functional Genomics Data Across Species to Develop Information on Porcine Inflammatory Gene Regulatory Pathway

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Comparative integration of structural and functional genomic data across species holds great promise in finding genes controlling disease resistance. We are investigating the porcine gut immune response to infection through gene expression profiling. We have collected porcine Affymetrix GeneChip da...

  17. Genome-wide and parental allele-specific analysis of CTCF and cohesin DNA binding in mouse brain reveals a tissue-specific binding pattern and an association with imprinted differentially methylated regions.

    PubMed

    Prickett, Adam R; Barkas, Nikolaos; McCole, Ruth B; Hughes, Siobhan; Amante, Samuele M; Schulz, Reiner; Oakey, Rebecca J

    2013-10-01

    DNA binding factors are essential for regulating gene expression. CTCF and cohesin are DNA binding factors with central roles in chromatin organization and gene expression. We determined the sites of CTCF and cohesin binding to DNA in mouse brain, genome wide and in an allele-specific manner with high read-depth ChIP-seq. By comparing our results with existing data for mouse liver and embryonic stem (ES) cells, we investigated the tissue specificity of CTCF binding sites. ES cells have fewer unique CTCF binding sites occupied than liver and brain, consistent with a ground-state pattern of CTCF binding that is elaborated during differentiation. CTCF binding sites without the canonical consensus motif were highly tissue specific. In brain, a third of CTCF and cohesin binding sites coincide, consistent with the potential for many interactions between cohesin and CTCF but also many instances of independent action. In the context of genomic imprinting, CTCF and/or cohesin bind to a majority but not all differentially methylated regions, with preferential binding to the unmethylated parental allele. Whether the parental allele-specific methylation was established in the parental germlines or post-fertilization in the embryo is not a determinant in CTCF or cohesin binding. These findings link CTCF and cohesin with the control regions of a subset of imprinted genes, supporting the notion that imprinting control is mechanistically diverse.

  18. Functional annotation of mammalian genomic DNA sequence by chemical mutagenesis: a fine-structure genetic mutation map of a 1- to 2-cM segment of mouse chromosome 7 corresponding to human chromosome 11p14-p15.

    PubMed

    Rinchik, Eugene M; Carpenter, Donald A; Johnson, Dabney K

    2002-01-22

    Eleven independent, recessive, N-ethyl-N-nitrosourea-induced mutations that map to a approximately 1- to 2-cM region of mouse chromosome (Chr) 7 homologous to human Chr 11p14-p15 were recovered from a screen of 1,218 gametes. These mutations were initially identified in a hemizygous state opposite a large p-locus deletion and subsequently were mapped to finer genomic intervals by crosses to a panel of smaller p deletions. The 11 mutations also were classified into seven complementation groups by pairwise crosses. Four complementation groups were defined by seven prenatally lethal mutations, including a group (l7R3) comprised of two alleles of obvious differing severity. Two allelic mutations (at the psrt locus) result in a severe seizure and runting syndrome, but one mutation (at the fit2 locus) results in a more benign runting phenotype. This experiment has added seven loci, defined by phenotypes of presumed point mutations, to the genetic map of a small (1-2 cM) region of mouse Chr 7 and will facilitate the task of functional annotation of DNA sequence and transcription maps both in the mouse and the corresponding human 11p14-p15 homology region.

  19. Comparison of TCDD-elicited genome-wide hepatic gene expression in Sprague–Dawley rats and C57BL/6 mice

    SciTech Connect

    Nault, Rance; Kim, Suntae; Zacharewski, Timothy R.

    2013-03-01

    Although the structure and function of the AhR are conserved, emerging evidence suggests that downstream effects are species-specific. In this study, rat hepatic gene expression data from the DrugMatrix database (National Toxicology Program) were compared to mouse hepatic whole-genome gene expression data following treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). For the DrugMatrix study, male Sprague–Dawley rats were gavaged daily with 20 μg/kg TCDD for 1, 3 and 5 days, while female C57BL/6 ovariectomized mice were examined 1, 3 and 7 days after a single oral gavage of 30 μg/kg TCDD. A total of 649 rat and 1386 mouse genes (|fold change| ≥ 1.5, P1(t) ≥ 0.99) were differentially expressed following treatment. HomoloGene identified 11,708 orthologs represented across the rat Affymetrix 230 2.0 GeneChip (12,310 total orthologs), and the mouse 4 × 44K v.1 Agilent oligonucleotide array (17,578 total orthologs). Comparative analysis found 563 and 922 orthologs differentially expressed in response to TCDD in the rat and mouse, respectively, with 70 responses associated with immune function and lipid metabolism in common to both. Moreover, QRTPCR analysis of Ceacam1, showed divergent expression (induced in rat; repressed in mouse) functionally consistent with TCDD-elicited hepatic steatosis in the mouse but not the rat. Functional analysis identified orthologs involved in nucleotide binding and acetyltransferase activity in rat, while mouse-specific responses were associated with steroid, phospholipid, fatty acid, and carbohydrate metabolism. These results provide further evidence that TCDD elicits species-specific regulation of distinct gene networks, and outlines considerations for future comparisons of publicly available microarray datasets. - Highlights: ► We performed a whole-genome comparison of TCDD-regulated genes in mice and rats. ► Previous species comparisons were extended using data from the DrugMatrix database. ► Less than 15% of TCDD

  20. Aging and chronic alcohol consumption are determinants of p16 gene expression, genomic DNA methylation and p16 promoter methylation in the mouse colon

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Elder age and chronic alcohol consumption are important risk factors for the development of colon cancer. Each factor can alter genomic and gene-specific DNA methylation. This study examined the effects of aging and chronic alcohol consumption on genomic and p16-specific methylation, and p16 express...

  1. Ageing, chronic alcohol consumption and folate are determinants of genomic DNA methylation, p16 promoter methylation and the expression of p16 in the mouse colon

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Elder age and chronic alcohol consumption are important risk factors for the development of colon cancer. Each factor can alter genomic and gene-specific DNA methylation. This study examined the effects of aging and chronic alcohol consumption on genomic and p16-specific methylation, and p16 express...

  2. Gene Expression Analysis of Cultured Rat-Endothelial Cells after Nd:YAG Laser Irradiation by Affymetrix GeneChip Array

    PubMed Central

    MASUDA, YOSHIKO; YOKOSE, SATOSHI; SAKAGAMI, HIROSHI

    2017-01-01

    Endothelial cells and dental pulp cells enhance osteo-/odontogenic and angiogenic differentiation. In our previous study, rat pulp cells migrated to Nd:YAG laser-irradiated endothelial cells in an insert cell culture system. The purpose of this study was to examine the possible changes in the gene expression of cultured rat aortic endothelial cells after Nd:YAG laser irradiation using affymetrix GeneChip Array. Total RNA was extracted from the cells at 5 h after laser irradiation. Gene expressions were evaluated by DNA array chip. Up-regulated genes were related to cell migration and cell structure (membrane stretch, actin regulation and junctional complexes), neurotransmission and inflammation. Heat-shock 70 kDa protein (Hsp70) was related to the development of tooth germ. This study offers candidate genes for understanding the relationship between the laser-stimulated endothelial cells and dental pulp cells. PMID:28064220

  3. Genome-Wide Analysis in Swine Associates Corneal Graft Rejection with Donor-Recipient Mismatches in Three Novel Histocompatibility Regions and One Locus Homologous to the Mouse H-3 Locus.

    PubMed

    Nicholls, Susan; Pong-Wong, Ricardo; Mitchard, Louisa; Harley, Ross; Archibald, Alan; Dick, Andrew; Bailey, Michael

    2016-01-01

    In rodents, immune responses to minor histocompatibility antigens are the most important drivers of corneal graft rejection. However, this has not been confirmed in humans or in a large animal model and the genetic loci are poorly characterised, even in mice. The gene sequence data now available for a range of relevant species permits the use of genome-wide association (GWA) techniques to identify minor antigens associated with transplant rejection. We have used this technique in a pre-clinical model of corneal transplantation in semi-inbred NIH minipigs and Babraham swine to search for novel minor histocompatibility loci and to determine whether rodent findings have wider applicability. DNA from a cohort of MHC-matched and MHC-mismatched donors and recipients was analysed for single nucleotide polymorphisms (SNPs). The level of SNP homozygosity for each line was assessed. Genome-wide analysis of the association of SNP disparities with rejection was performed using log-likelihood ratios. Four genomic blocks containing four or more SNPs significantly linked to rejection were identified (on chromosomes 1, 4, 6 and 9), none at the location of the MHC. One block of 36 SNPs spanned a region that exhibits conservation of synteny with the mouse H-3 histocompatibility locus and contains the pig homologue of the mouse Zfp106 gene, which encodes peptide epitopes known to mediate corneal graft rejection. The other three regions are novel minor histocompatibility loci. The results suggest that rejection can be predicted from SNP analysis prior to transplant in this model and that a similar GWA analysis is merited in humans.

  4. Quantitative LC-MS Provides No Evidence for m(6) dA or m(4) dC in the Genome of Mouse Embryonic Stem Cells and Tissues.

    PubMed

    Schiffers, Sarah; Ebert, Charlotte; Rahimoff, René; Kosmatchev, Olesea; Steinbacher, Jessica; Bohne, Alexandra-Viola; Spada, Fabio; Michalakis, Stylianos; Nickelsen, Jörg; Müller, Markus; Carell, Thomas

    2017-03-30

    Until recently, it was believed that the genomes of higher organisms contain, in addition to the four canonical DNA bases, only 5-methyl-dC (m(5) dC) as a modified base to control epigenetic processes. In recent years, this view has changed dramatically with the discovery of 5-hydroxymethyl-dC (hmdC), 5-formyl-dC (fdC), and 5-carboxy-dC (cadC) in DNA from stem cells and brain tissue. N(6) -methyldeoxyadenosine (m(6) dA) is the most recent base reported to be present in the genome of various eukaryotic organisms. This base, together with N(4) -methyldeoxycytidine (m(4) dC), was first reported to be a component of bacterial genomes. In this work, we investigated the levels and distribution of these potentially epigenetically relevant DNA bases by using a novel ultrasensitive UHPLC-MS method. We further report quantitative data for m(5) dC, hmdC, fdC, and cadC, but we were unable to detect either m(4) dC or m(6) dA in DNA isolated from mouse embryonic stem cells or brain and liver tissue, which calls into question their epigenetic relevance.

  5. Biology of breast cancer during pregnancy using genomic profiling.

    PubMed

    Azim, Hatem A; Brohée, Sylvain; Peccatori, Fedro A; Desmedt, Christine; Loi, Sherene; Lambrechts, Diether; Dell'Orto, Patrizia; Majjaj, Samira; Jose, Vinu; Rotmensz, Nicole; Ignatiadis, Michail; Pruneri, Giancarlo; Piccart, Martine; Viale, Giuseppe; Sotiriou, Christos

    2014-08-01

    Breast cancer during pregnancy is rare and is associated with relatively poor prognosis. No information is available on its biological features at the genomic level. Using a dataset of 54 pregnant and 113 non-pregnant breast cancer patients, we evaluated the pattern of hot spot somatic mutations and did transcriptomic profiling using Sequenom and Affymetrix respectively. We performed gene set enrichment analysis to evaluate the pathways associated with diagnosis during pregnancy. We also evaluated the expression of selected cancer-related genes in pregnant and non-pregnant patients and correlated the results with changes occurring in the normal breast using a pregnant murine model. We finally investigated aberrations associated with disease-free survival (DFS). No significant differences in mutations were observed. Of the total number of patients, 18.6% of pregnant and 23% of non-pregnant patients had a PIK3CA mutation. Around 30% of tumors were basal, with no differences in the distribution of breast cancer molecular subtypes between pregnant and non-pregnant patients. Two pathways were enriched in tumors diagnosed during pregnancy: the G protein-coupled receptor pathway and the serotonin receptor pathway (FDR <0.0001). Tumors diagnosed during pregnancy had higher expression of PD1 (PDCD1; P=0.015), PDL1 (CD274; P=0.014), and gene sets related to SRC (P=0.004), IGF1 (P=0.032), and β-catenin (P=0.019). Their expression increased almost linearly throughout gestation when evaluated on the normal breast using a pregnant mouse model underscoring the potential effect of the breast microenvironment on tumor phenotype. No genes were associated with DFS in a multivariate model, which could be due to low statistical power. Diagnosis during pregnancy impacts the breast cancer transcriptome including potential cancer targets.

  6. Mouse genetics: catalogue and scissors.

    PubMed

    Sung, Young Hoon; Baek, In-Jeoung; Seong, Je Kyung; Kim, Jin Soo; Lee, Han-Woong

    2012-12-01

    Phenotypic analysis of gene-specific knockout (KO) mice has revolutionized our understanding of in vivo gene functions. As the use of mouse embryonic stem (ES) cells is inevitable for conventional gene targeting, the generation of knockout mice remains a very time-consuming and expensive process. To accelerate the large-scale production and phenotype analyses of KO mice, international efforts have organized global consortia such as the International Knockout Mouse Consortium (IKMC) and International Mouse Phenotype Consortium (IMPC), and they are persistently expanding the KO mouse catalogue that is publicly available for the researches studying specific genes of interests in vivo. However, new technologies, adopting zinc-finger nucleases (ZFNs) or Transcription Activator-Like Effector (TALE) Nucleases (TALENs) to edit the mouse genome, are now emerging as valuable and effective shortcuts alternative for the conventional gene targeting using ES cells. Here, we introduce the recent achievement of IKMC, and evaluate the significance of ZFN/TALEN technology in mouse genetics.

  7. Genome databases

    SciTech Connect

    Courteau, J.

    1991-10-11

    Since the Genome Project began several years ago, a plethora of databases have been developed or are in the works. They range from the massive Genome Data Base at Johns Hopkins University, the central repository of all gene mapping information, to small databases focusing on single chromosomes or organisms. Some are publicly available, others are essentially private electronic lab notebooks. Still others limit access to a consortium of researchers working on, say, a single human chromosome. An increasing number incorporate sophisticated search and analytical software, while others operate as little more than data lists. In consultation with numerous experts in the field, a list has been compiled of some key genome-related databases. The list was not limited to map and sequence databases but also included the tools investigators use to interpret and elucidate genetic data, such as protein sequence and protein structure databases. Because a major goal of the Genome Project is to map and sequence the genomes of several experimental animals, including E. coli, yeast, fruit fly, nematode, and mouse, the available databases for those organisms are listed as well. The author also includes several databases that are still under development - including some ambitious efforts that go beyond data compilation to create what are being called electronic research communities, enabling many users, rather than just one or a few curators, to add or edit the data and tag it as raw or confirmed.

  8. Mouse Models of Genomic Syndromes as Tools for Understanding the Basis of Complex Traits: An Example with the Smith-Magenis and the Potocki-Lupski Syndromes

    PubMed Central

    Carmona-Mora, P; Molina, J; Encina, C.A; Walz, K

    2009-01-01

    Each human's genome is distinguished by extra and missing DNA that can be “benign” or powerfully impact everything from development to disease. In the case of genomic disorders DNA rearrangements, such as deletions or duplications, correlate with a clinical specific phenotype. The clinical presentations of genomic disorders were thought to result from altered gene copy number of physically linked dosage sensitive genes. Genomic disorders are frequent diseases (~1 per 1,000 births). Smith-Magenis syndrome (SMS) and Potocki-Lupski syndrome (PTLS) are genomic disorders, associated with a deletion and a duplication, of 3.7 Mb respectively, within chromosome 17 band p11.2. This region includes 23 genes. Both syndromes have complex and distinctive phenotypes including multiple congenital and neurobehavioral abnormalities. Human chromosome 17p11.2 is syntenic to the 32-34 cM region of murine chromosome 11. The number and order of the genes are highly conserved. In this review, we will exemplify how genomic disorders can be modeled in mice and the advantages that such models can give in the study of genomic disorders in particular and gene copy number variation (CNV) in general. The contributions of the SMS and PTLS animal models in several aspects ranging from more specific ones, as the definition of the clinical aspects of the human clinical spectrum, the identification of dosage sensitive genes related to the human syndromes, to the more general contributions as the definition of genetic locus impacting obesity and behavior and the elucidation of general mechanisms related to the pathogenesis of gene CNV are discussed. PMID:19949547

  9. Mouse models of genomic syndromes as tools for understanding the basis of complex traits: an example with the smith-magenis and the potocki-lupski syndromes.

    PubMed

    Carmona-Mora, P; Molina, J; Encina, C A; Walz, K

    2009-06-01

    Each human's genome is distinguished by extra and missing DNA that can be "benign" or powerfully impact everything from development to disease. In the case of genomic disorders DNA rearrangements, such as deletions or duplications, correlate with a clinical specific phenotype. The clinical presentations of genomic disorders were thought to result from altered gene copy number of physically linked dosage sensitive genes. Genomic disorders are frequent diseases (~1 per 1,000 births). Smith-Magenis syndrome (SMS) and Potocki-Lupski syndrome (PTLS) are genomic disorders, associated with a deletion and a duplication, of 3.7 Mb respectively, within chromosome 17 band p11.2. This region includes 23 genes. Both syndromes have complex and distinctive phenotypes including multiple congenital and neurobehavioral abnormalities. Human chromosome 17p11.2 is syntenic to the 32-34 cM region of murine chromosome 11. The number and order of the genes are highly conserved. In this review, we will exemplify how genomic disorders can be modeled in mice and the advantages that such models can give in the study of genomic disorders in particular and gene copy number variation (CNV) in general. The contributions of the SMS and PTLS animal models in several aspects ranging from more specific ones, as the definition of the clinical aspects of the human clinical spectrum, the identification of dosage sensitive genes related to the human syndromes, to the more general contributions as the definition of genetic locus impacting obesity and behavior and the elucidation of general mechanisms related to the pathogenesis of gene CNV are discussed.

  10. A comprehensive proteomics and genomics analysis reveals novel transmembrane proteins in human platelets and mouse megakaryocytes including G6b-B, a novel immunoreceptor tyrosine-based inhibitory motif protein.

    PubMed

    Senis, Yotis A; Tomlinson, Michael G; García, Angel; Dumon, Stephanie; Heath, Victoria L; Herbert, John; Cobbold, Stephen P; Spalton, Jennifer C; Ayman, Sinem; Antrobus, Robin; Zitzmann, Nicole; Bicknell, Roy; Frampton, Jon; Authi, Kalwant S; Martin, Ashley; Wakelam, Michael J O; Watson, Stephen P

    2007-03-01

    The platelet surface is poorly characterized due to the low abundance of many membrane proteins and the lack of specialist tools for their investigation. In this study we identified novel human platelet and mouse megakaryocyte membrane proteins using specialist proteomics and genomics approaches. Three separate methods were used to enrich platelet surface proteins prior to identification by liquid chromatography and tandem mass spectrometry: lectin affinity chromatography, biotin/NeutrAvidin affinity chromatography, and free flow electrophoresis. Many known, abundant platelet surface transmembrane proteins and several novel proteins were identified using each receptor enrichment strategy. In total, two or more unique peptides were identified for 46, 68, and 22 surface membrane, intracellular membrane, and membrane proteins of unknown subcellular localization, respectively. The majority of these were single transmembrane proteins. To complement the proteomics studies, we analyzed the transcriptome of a highly purified preparation of mature primary mouse megakaryocytes using serial analysis of gene expression in view of the increasing importance of mutant mouse models in establishing protein function in platelets. This approach identified all of the major classes of platelet transmembrane receptors, including multitransmembrane proteins. Strikingly 17 of the 25 most megakaryocyte-specific genes (relative to 30 other serial analysis of gene expression libraries) were transmembrane proteins, illustrating the unique nature of the megakaryocyte/platelet surface. The list of novel plasma membrane proteins identified using proteomics includes the immunoglobulin superfamily member G6b, which undergoes extensive alternate splicing. Specific antibodies were used to demonstrate expression of the G6b-B isoform, which contains an immunoreceptor tyrosine-based inhibition motif. G6b-B undergoes tyrosine phosphorylation and association with the SH2 domain-containing phosphatase

  11. Effects of L1 retrotransposon insertion on transcript processing, localization and accumulation: lessons from the retinal degeneration 7 mouse and implications for the genomic ecology of L1 elements.

    PubMed

    Chen, Jichao; Rattner, Amir; Nathans, Jeremy

    2006-07-01

    The retinal degeneration 7 (rd7) mouse is a naturally occurring model of enhanced S-cone syndrome, Goldman-Favre syndrome and clumped pigmentary retinopathy in humans, allelic disorders caused by inactivation of a photoreceptor-specific nuclear hormone receptor, NR2E3. We show here that the rd7 mutation arose from the antisense insertion of a long interspersed nuclear element (LINE-1) (or L1) into exon 5 of the mouse Nr2e3 gene. L1 insertion blocks splicing of Nr2e3 intron 5 by separating an inefficient splice donor from essential splicing enhancers within exon 5, with the result that incompletely spliced transcripts accumulate to high levels at the mutant Nr2e3 locus in photoreceptor nuclei. The high efficiency of transcription through the 7 kb L1 was unexpected and led us to compare the effect on transcript abundance of sense or antisense L1 insertions in transfected cells. In a variety of sequence contexts antisense L1 insertions had little or no effect on transcript levels or the production of full-length transcripts, whereas sense L1 insertions reduced transcript levels from several-fold to more than 10-fold. A bioinformatic analysis of all mouse L1s shows a approximately 2-fold under-representation of L1s in introns when compared with bulk genomic DNA, and, within introns, a further approximately 2-fold under-representation of sense when compared with antisense L1s. Interestingly, there is no evidence for orientation-specific positive or negative selection within any subregions of the L1 element. These data suggest that L1s have evolved to present the host transcriptional machinery with a minimally disruptive profile in the antisense orientation such that antisense intronic L1s often escape purifying negative selection.

  12. Concentration-and time-dependent genomic changes in the mouse urinary bladder following exposure to arsenate in drinking water for up to twelve weeks

    EPA Science Inventory

    Inorganic arsenic (AsD is a known human bladder carcinogen. The objective of this study was to examine the concentration dependence of the genomic response to ASi in the urinary bladders of mice. C57BL/6J mice were exposed for 1 or 12 weeks to arsenate in drinking water at concen...

  13. Mouse models for neurological disease.

    PubMed

    Hafezparast, Majid; Ahmad-Annuar, Azlina; Wood, Nicholas W; Tabrizi, Sarah J; Fisher, Elizabeth M C

    2002-08-01

    The mouse has many advantages over human beings for the study of genetics, including the unique property that genetic manipulation can be routinely carried out in the mouse genome. Most importantly, mice and human beings share the same mammalian genes, have many similar biochemical pathways, and have the same diseases. In the minority of cases where these features do not apply, we can still often gain new insights into mouse and human biology. In addition to existing mouse models, several major programmes have been set up to generate new mouse models of disease. Alongside these efforts are new initiatives for the clinical, behavioural, and physiological testing of mice. Molecular genetics has had a major influence on our understanding of the causes of neurological disorders in human beings, and much of this has come from work in mice.

  14. University of California San Francisco (UCSF-2): Gene Expression Profiling of Normal Mouse Skin, Hras WT and Hras -/- | Office of Cancer Genomics

    Cancer.gov

    University of California San Francisco (UCSF-2): Gene Expression Profiling of Normal Mouse Skin, Hras WT and Hras -/- This data set contains the transcriptional profiles of 20 dorsal skin samples from eight-week-old mice. Mice were generated by crossing FVB/N to Mus spretus mice to generate F1 mice, and then crossing F1 mice back to the FVB/N strain. 10  FVB/N mice lacking Hras1 (aka HrasKO, Hras-/-) and 10  FVB/N mice with wild-type Hras1 were generated. Read the abstract.

  15. Strategies and tools for whole genome alignments

    SciTech Connect

    Couronne, Olivier; Poliakov, Alexander; Bray, Nicolas; Ishkhanov,Tigran; Ryaboy, Dmitriy; Rubin, Edward; Pachter, Lior; Dubchak, Inna

    2002-11-25

    The availability of the assembled mouse genome makespossible, for the first time, an alignment and comparison of two largevertebrate genomes. We have investigated different strategies ofalignment for the subsequent analysis of conservation of genomes that areeffective for different quality assemblies. These strategies were appliedto the comparison of the working draft of the human genome with the MouseGenome Sequencing Consortium assembly, as well as other intermediatemouse assemblies. Our methods are fast and the resulting alignmentsexhibit a high degree of sensitivity, covering more than 90 percent ofknown coding exons in the human genome. We have obtained such coveragewhile preserving specificity. With a view towards the end user, we havedeveloped a suite of tools and websites for automatically aligning, andsubsequently browsing and working with whole genome comparisons. Wedescribe the use of these tools to identify conserved non-coding regionsbetween the human and mouse genomes, some of which have not beenidentified by other methods.

  16. Increased 4R tau expression and behavioural changes in a novel MAPT-N296H genomic mouse model of tauopathy

    PubMed Central

    Wobst, Heike J.; Denk, Franziska; Oliver, Peter L.; Livieratos, Achilleas; Taylor, Tonya N.; Knudsen, Maria H.; Bengoa-Vergniory, Nora; Bannerman, David; Wade-Martins, Richard

    2017-01-01

    The microtubule-associated protein tau is implicated in various neurodegenerative diseases including Alzheimer’s disease, progressive supranuclear palsy and corticobasal degeneration, which are characterized by intracellular accumulation of hyperphosphorylated tau. Mutations in the tau gene MAPT cause frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17). In the human central nervous system, six tau isoforms are expressed, and imbalances in tau isoform ratios are associated with pathology. To date, few animal models of tauopathy allow for the potential influence of these protein isoforms, relying instead on cDNA-based transgene expression. Using the P1-derived artificial chromosome (PAC) technology, we created mouse lines expressing all six tau isoforms from the human MAPT locus, harbouring either the wild-type sequence or the disease-associated N296H mutation on an endogenous Mapt−/− background. Animals expressing N296H mutant tau recapitulated early key features of tauopathic disease, including a tau isoform imbalance and tau hyperphosphorylation in the absence of somatodendritic tau inclusions. Furthermore, N296H animals displayed behavioural anomalies such as hyperactivity, increased time in the open arms of the elevated plus maze and increased immobility during the tail suspension test. The mouse models described provide an excellent model to study the function of wild-type or mutant tau in a highly physiological setting. PMID:28233851

  17. Genomics-based screening of differentially expressed genes in the brains of mice exposed to silver nanoparticles via inhalation

    NASA Astrophysics Data System (ADS)

    Lee, Hye-Young; Choi, You-Jin; Jung, Eun-Jung; Yin, Hu-Quan; Kwon, Jung-Taek; Kim, Ji-Eun; Im, Hwang-Tae; Cho, Myung-Haing; Kim, Ju-Han; Kim, Hyun-Young; Lee, Byung-Hoon

    2010-06-01

    Silver nanoparticles (AgNP) are among the fastest growing product categories in the nanotechnology industry. Despite the importance of AgNP in consumer products and clinical applications, relatively little is known regarding AgNP toxicity and its associated risks. We investigated the effects of AgNP on gene expression in the mouse brain using Affymetrix Mouse Genome Arrays. C57BL/6 mice were exposed to AgNP (geometric mean diameter, 22.18 ± 1.72 nm; 1.91 × 107 particles/cm3) for 6 h/day, 5 days/week using the nose-only exposure system for 2 weeks. Total RNA isolated from the cerebrum and cerebellum was subjected to hybridization. From over 39,000 probe sets, 468 genes in the cerebrum and 952 genes in the cerebellum were identified as AgNP-responsive (one-way analysis of variance; p < 0.05). The largest groups of gene products affected by AgNP exposure included 73 genes in the cerebrum and 144 genes in the cerebellum. AgNP exposure modulated the expression of several genes associated with motor neuron disorders, neurodegenerative disease, and immune cell function, indicating potential neurotoxicity and immunotoxicity associated with AgNP exposure. Real-time PCR data for five genes analyzed from whole blood showed good correlation with the observed changes in the brain. Following rigorous validation and substantiation, these genes may assist in the development of surrogate markers for AgNP exposure and/or toxicity.

  18. Molecular changes during neurodevelopment following second-trimester binge ethanol exposure in a mouse model of fetal alcohol spectrum disorder: from immediate effects to long-term adaptation.

    PubMed

    Mantha, Katarzyna; Laufer, Benjamin I; Singh, Shiva M

    2014-01-01

    Fetal alcohol spectrum disorder (FASD) is an umbrella term that refers to a wide range of behavioral and cognitive deficits resulting from prenatal alcohol exposure. It involves changes in brain gene expression that underlie lifelong FASD symptoms. How these changes are achieved from immediate to long-term effects, and how they are maintained, is unknown. We have used the C57BL/6J mouse to assess the dynamics of genomic alterations following binge alcohol exposure. Ethanol-exposed fetal (short-term effect) and adult (long-term effect) brains were assessed for gene expression and microRNA (miRNA) changes using Affymetrix mouse arrays. We identified 48 and 68 differentially expressed genes in short- and long-term groups, respectively. No gene was common between the 2 groups. Short-term (immediate) genes were involved in cellular compromise and apoptosis, which represent ethanol's toxic effects. Long-term genes were involved in various cellular functions, including epigenetics. Using quantitative RT-PCR, we confirmed the downregulation of long-term genes: Camk1g, Ccdc6, Egr3, Hspa5, and Xbp1. miRNA arrays identified 20 differentially expressed miRNAs, one of which (miR-302c) was confirmed. miR-302c was involved in an inverse relationship with Ccdc6. A network-based model involving altered genes illustrates the importance of cellular redox, stress and inflammation in FASD. Our results also support a critical role of apoptosis in FASD, and the potential involvement of miRNAs in the adaptation of gene expression following prenatal ethanol exposure. The ultimate molecular footprint involves inflammatory disease, neurological disease and skeletal and muscular disorders as major alterations in FASD. At the cellular level, these processes represent abnormalities in redox, stress and inflammation, with potential underpinnings to anxiety.

  19. Global gene expression analysis in the mouse brainstem after hyperalgesia induced by facial carrageenan injection--evidence for a form of neurovascular coupling?

    PubMed

    Poh, Kay-Wee; Lutfun, Nahar; Manikandan, Jayapal; Ong, Wei-Yi; Yeo, Jin-Fei

    2009-03-01

    The present study was carried out to examine global gene expression in the brainstem, in a mouse facial carrageenan injection model of orofacial pain. Mice that received facial carrageenan injection showed increased mechanical allodynia, demonstrated by increased responses to von Frey hair stimulation of the face. The brainstem was harvested at 3 days post-injection, corresponding to the time of peak responses, and analyzed by Affymetrix Mouse Genome 430 2.0 microarrays. We sought to identify common genes that are changed in the respective sides of the brainstem after either right- or left-sided facial carrageenan injection. The result is a relatively small list of genes (22 genes), which were then classified using DAVID software. Many of them fell into the categories of "response to stress", "defence response", "response to biotic stimulus", "cell adhesion" and "leukocyte adhesion". Of these, increased expression of P-selectin, ICAM-1 and CCL12 after carrageenan injection could be verified by real-time RT-PCR on both the right and left sides, and increased in P-selectin and ICAM-1 further verified by Western blot analysis. P-selectin and ICAM-1 were immunolocalized to endothelial cells, and were double labelled with von Willebrand factor. Intraperitoneal injection of the P-selectin inhibitor KF38789 significantly reduced mechanical allodynia in the facial carrageenan-injected mice. P-selectin mediates the capturing of leukocytes from the bloodstream and rolling of leukocytes along the endothelial surface. We hypothesize that increased nociceptive input to the brainstem could attract circulating macrophages into the brain, resulting in neuroinflammation and pain.

  20. Comparative transcriptomic profiling of Vitis vinifera under high light using a custom-made array and the Affymetrix GeneChip.

    PubMed

    Carvalho, Luísa C; Vilela, Belmiro J; Mullineaux, Phil M; Amâncio, Sara

    2011-11-01

    Understanding abiotic stress responses is one of the most important issues in plant research nowadays. Abiotic stress, including excess light, can promote the onset of oxidative stress through the accumulation of reactive oxygen species. Oxidative stress also arises when in vitro propagated plants are exposed to high light upon transfer to ex vitro. To determine whether the underlying pathways activated at the transfer of in vitro grapevine to ex vitro conditions reflect the processes occurring upon light stress, we used Vitis vinifera Affymetrix GeneChip (VvGA) and a custom array of genes responsive to light stress (LSCA) detected by real-time reverse transcriptase PCR (qRT-PCR). When gene-expression profiles were compared, 'protein metabolism and modification', 'signaling', and 'anti-oxidative' genes were more represented in LSCA, while, in VvGA, 'cell wall metabolism' and 'secondary metabolism' were the categories in which gene expression varied more significantly. The above functional categories confirm previous studies involving other types of abiotic stresses, enhancing the common attributes of abiotic stress defense pathways. The LSCA analysis of our experimental system detected strong response of heat shock genes, particularly the protein rescuing mechanism involving the cooperation of two ATP-dependent chaperone systems, Hsp100 and Hsp70, which showed an unusually late response during the recovery period, of extreme relevance to remove non-functional, potentially harmful polypeptides arising from misfolding, denaturation, or aggregation brought about by stress. The success of LSCA also proves the feasibility of a custom-made qRT-PCR approach, particularly for species for which no GeneChip is available and for researchers dealing with a specific and focused problem.

  1. The Bam repeats of the mouse genome belong in several superfamilies the longest of which is over 9 kb in size.

    PubMed Central

    Meunier-Rotival, M; Bernardi, G

    1984-01-01

    Mouse DNA contains two equally abundant, homologous subfamilies of MspI 3.6 and 5 kb repeated fragments. The first subfamily corresponds to the previously described (1) Bam 4 kb repeats, the second one to Bam repeated fragments of higher molecular weight. These subfamilies account for the vast majority of long Bam repeats and are linked with contiguous short Bam 0.5 kb repeats. A minority of these composite Bam repeats extend, on the 0.5 kb side, into R repeats. In turn, a fraction of the composite Bam/R repeats extend further, for at least 3 kb, into other repeated sequences contiguous to the R repeats. The long Bam repeats belong, therefore, in at least three superfamilies of repeats, the longest one being over 9 kb in size. Some general properties of these superfamilies are discussed. Images PMID:6322110

  2. Targeted Genomic Disruption of H-ras and N-ras, Individually or in Combination, Reveals the Dispensability of Both Loci for Mouse Growth and Development

    PubMed Central

    Esteban, Luis M.; Vicario-Abejón, Carlos; Fernández-Salguero, Pedro; Fernández-Medarde, Alberto; Swaminathan, Nalini; Yienger, Kate; Lopez, Eva; Malumbres, Marcos; McKay, Ron; Ward, Jerrold M.; Pellicer, Angel; Santos, Eugenio

    2001-01-01

    Mammalian cells harbor three highly homologous and widely expressed members of the ras family (H-ras, N-ras, and K-ras), but it remains unclear whether they play specific or overlapping cellular roles. To gain insight into such functional roles, here we generated and analyzed H-ras null mutant mice, which were then also bred with N-ras knockout animals to ascertain the viability and properties of potential double null mutations in both loci. Mating among heterozygous H-ras+/− mice produced H-ras−/− offspring with a normal Mendelian pattern of inheritance, indicating that the loss of H-ras did not interfere with embryonic and fetal viability in the uterus. Homozygous mutant H-ras−/− mice reached sexual maturity at the same age as their littermates, and both males and females were fertile. Characterization of lymphocyte subsets in the spleen and thymus showed no significant differences between wild-type and H-ras−/− mice. Analysis of neuronal markers in the brains of knockout and wild-type H-ras mice showed that disruption of this locus did not impair or alter neuronal development. Breeding between our H-ras mutant animals and previously available N-ras null mutants gave rise to viable double knockout (H-ras−/−/N-ras−/−) offspring expressing only K-ras genes which grew normally, were fertile, and did not show any obvious phenotype. Interestingly, however, lower-than-expected numbers of adult, double knockout animals were consistently obtained in Mendelian crosses between heterozygous N-ras/H-ras mice. Our results indicate that, as for N-ras, H-ras gene function is dispensable for normal mouse development, growth, fertility, and neuronal development. Additionally, of the three ras genes, K-ras appears to be not only essential but also sufficient for normal mouse development. PMID:11238881

  3. QuadBase: genome-wide database of G4 DNA--occurrence and conservation in human, chimpanzee, mouse and rat promoters and 146 microbes.

    PubMed

    Yadav, Vinod Kumar; Abraham, James Kappukalayil; Mani, Prithvi; Kulshrestha, Rashi; Chowdhury, Shantanu

    2008-01-01

    Emerging evidence indicates the importance of G-quadruplex motifs as drug targets. [Stuart A. Borman, Ascent of quadruplexes-nucleic acid structures become promising drug targets. Chem. Eng. News, 2007;85, 12-17], which stems from the fact that these motifs are present in a surprising number of promoters wherein their role in controlling gene expression has been demonstrated for a few. We present a compendium of quadruplex motifs, with particular focus on their occurrence and conservation in promoters-QuadBase. It is composed of two parts (EuQuad and ProQuad). EuQuad gives information on quadruplex motifs present within 10 kb of transcription starts sites in 99 980 human, chimpanzee, rat and mouse genes. ProQuad contains quadruplex information of 146 prokaryotes. Apart from gene-specific searches for quadruplex motifs, QuadBase has a number of other modules. 'Orthologs Analysis' queries for conserved motifs across species based on a selected reference organism; 'Pattern Search' can be used to fetch specific motifs of interest from a selected organism using user-defined criteria for quadruplex motifs, i.e. stem, loop size, etc. 'Pattern Finder' tool can search for motifs in any given sequence. QuadBase is freely available to users from non-profit organization at http://quadbase.igib.res.in/.

  4. BEAT: Bioinformatics Exon Array Tool to store, analyze and visualize Affymetrix GeneChip Human Exon Array data from disease experiments

    PubMed Central

    2012-01-01

    Background It is known from recent studies that more than 90% of human multi-exon genes are subject to Alternative Splicing (AS), a key molecular mechanism in which multiple transcripts may be generated from a single gene. It is widely recognized that a breakdown in AS mechanisms plays an important role in cellular differentiation and pathologies. Polymerase Chain Reactions, microarrays and sequencing technologies have been applied to the study of transcript diversity arising from alternative expression. Last generation Affymetrix GeneChip Human Exon 1.0 ST Arrays offer a more detailed view of the gene expression profile providing information on the AS patterns. The exon array technology, with more than five million data points, can detect approximately one million exons, and it allows performing analyses at both gene and exon level. In this paper we describe BEAT, an integrated user-friendly bioinformatics framework to store, analyze and visualize exon arrays datasets. It combines a data warehouse approach with some rigorous statistical methods for assessing the AS of genes involved in diseases. Meta statistics are proposed as a novel approach to explore the analysis results. BEAT is available at http://beat.ba.itb.cnr.it. Results BEAT is a web tool which allows uploading and analyzing exon array datasets using standard statistical methods and an easy-to-use graphical web front-end. BEAT has been tested on a dataset with 173 samples and tuned using new datasets of exon array experiments from 28 colorectal cancer and 26 renal cell cancer samples produced at the Medical Genetics Unit of IRCCS Casa Sollievo della Sofferenza. To highlight all possible AS events, alternative names, accession Ids, Gene Ontology terms and biochemical pathways annotations are integrated with exon and gene level expression plots. The user can customize the results choosing custom thresholds for the statistical parameters and exploiting the available clinical data of the samples for a

  5. Genome-wide gene expression profiling of SCID mice with T-cell-mediated Colitis.

    PubMed

    Brudzewsky, D; Pedersen, A E; Claesson, M H; Gad, M; Kristensen, N N; Lage, K; Jensen, T; Tommerup, N; Larsen, L A; Knudsen, S; Tümer, Z

    2009-05-01

    Inflammatory bowel disease (IBD) is a multifactorial disorder with an unknown aetiology. The aim of this study is to employ a murine model of IBD to identify pathways and genes, which may play a key role in the pathogenesis of IBD and could be important for discovery of new disease markers in human disease. Here, we have investigated severe combined immunodeficient (SCID) mice, which upon adoptive transfer with concanavalin A-activated CD4(+) T cells develop inflammation of the colon with predominance in rectum. Mice with increasing level of inflammation was studied. RNA from rectum of transplanted and non-transplanted SCID mice was investigated by a genome-wide gene expression analysis using the Affymetrix mouse expression array 430A (MOE430A) including 22,626 probe sets. A significant change in gene expression (P = 0.00001) is observed in 152 of the genes between the non-transplanted control mice and colitis mice, and among these genes there is an overrepresentation of genes involved in inflammatory processes. Some of the most significant genes showing higher expression encode S100A proteins and chemokines involved in trafficking of leucocytes in inflammatory areas. Classification by gene clustering based on the genes with the significantly altered gene expression corresponds to two different levels of inflammation as established by the histological scoring of the inflamed rectum. These data demonstrate that this SCID T-cell transfer model is a useful animal model for human IBD and can be used for suggesting candidate genes involved in the pathogenesis and for identifying new molecular markers of chronic inflammation in human IBD.

  6. ATR-FTIR spectroscopy reveals genomic loci regulating the tissue response in high fat diet fed BXD recombinant inbred mouse strains

    PubMed Central

    2013-01-01

    Background Obesity-associated organ-specific pathological states can be ensued from the dysregulation of the functions of the adipose tissues, liver and muscle. However, the influence of genetic differences underlying gross-compositional differences in these tissues is largely unknown. In the present study, the analytical method of ATR-FTIR spectroscopy has been combined with a genetic approach to identify genetic differences responsible for phenotypic alterations in adipose, liver and muscle tissues. Results Mice from 29 BXD recombinant inbred mouse strains were put on high fat diet and gross-compositional changes in adipose, liver and muscle tissues were measured by ATR-FTIR spectroscopy. The analysis of genotype-phenotype correlations revealed significant quantitative trait loci (QTL) on chromosome 12 for the content of fat and collagen, collagen integrity, and the lipid to protein ratio in adipose tissue and on chromosome 17 for lipid to protein ratio in liver. Using gene expression and sequence information, we suggest Rsad2 (viperin) and Colec11 (collectin-11) on chromosome 12 as potential quantitative trait candidate genes. Rsad2 may act as a modulator of lipid droplet contents and lipid biosynthesis; Colec11 might play a role in apoptopic cell clearance and maintenance of adipose tissue. An increased level of Rsad2 transcripts in adipose tissue of DBA/2J compared to C57BL/6J mice suggests a cis-acting genetic variant leading to differential gene activation. Conclusion The results demonstrate that the analytical method of ATR-FTIR spectroscopy effectively contributed to decompose the macromolecular composition of tissues that accumulate fat and to link this information with genetic determinants. The candidate genes in the QTL regions may contribute to obesity-related diseases in humans, in particular if the results can be verified in a bigger BXD cohort. PMID:23758785

  7. Proteasome Inhibitors Enhance Gene Delivery by AAV Virus Vectors Expressing Large Genomes in Hemophilia Mouse and Dog Models: A Strategy for Broad Clinical Application

    PubMed Central

    Monahan, Paul E; Lothrop, Clinton D; Sun, Junjiang; Hirsch, Matthew L; Kafri, Tal; Kantor, Boris; Sarkar, Rita; Tillson, D Michael; Elia, Joseph R; Samulski, R Jude

    2010-01-01

    Delivery of genes that are larger than the wild-type adeno-associated virus (AAV) 4,681 nucleotide genome is inefficient using AAV vectors. We previously demonstrated in vitro that concurrent proteasome inhibitor (PI) treatment improves transduction by AAV vectors encoding oversized transgenes. In this study, an AAV vector with a 5.6 kilobase (kb) factor VIII expression cassette was used to test the effect of an US Food and Drug Administration–approved PI (bortezomib) treatment concurrent with vector delivery in vivo. Intrahepatic vector delivery resulted in factor VIII expression that persisted for >1 year in hemophilia mice. Single-dose bortezomib given with AAV2 or AAV8 factor VIII vector enhanced expression on average ~600 and ~300%, respectively. Moreover, coadministration of AAV8.canineFVIII (1 × 1013 vg/kg) and bortezomib in hemophilia A dogs (n = 4) resulted in normalization of the whole blood clotting time (WBCT) and 90% reduction in hemorrhages for >32 months compared to untreated hemophilia A dogs (n = 3) or dogs administered vector alone (n = 3). Demonstration of long-term phenotypic correction of hemophilia A dogs with combination adjuvant bortezomib and AAV vector expressing the oversized transgene establishes preclinical studies that support testing in humans and provides a working paradigm to facilitate a significant expansion of therapeutic targets for human gene therapy. PMID:20700109

  8. Relationship between somatic mosaicism of Pax6 mutation and variable developmental eye abnormalities-an analysis of CRISPR genome-edited mouse embryos.

    PubMed

    Yasue, Akihiro; Kono, Hitomi; Habuta, Munenori; Bando, Tetsuya; Sato, Keita; Inoue, Junji; Oyadomari, Seiichi; Noji, Sumihare; Tanaka, Eiji; Ohuchi, Hideyo

    2017-12-01

    The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein (Cas) system is a rapid gene-targeting technology that does not require embryonic stem cells. To demonstrate dosage effects of the Pax6 gene on eye formation, we generated Pax6-deficient mice with the CRISPR/Cas system. Eyes of founder embryos at embryonic day (E) 16.5 were examined and categorized according to macroscopic phenotype as class 1 (small eye with distinct pigmentation), class 2 (pigmentation without eye globes), or class 3 (no pigmentation and no eyes). Histologically, class 1 eyes were abnormally small in size with lens still attached to the cornea at E16.5. Class 2 eyes had no lens and distorted convoluted retinas. Class 3 eyes had only rudimentary optic vesicle-like tissues or histological anophthalmia. Genotyping of neck tissue cells from the founder embryos revealed somatic mosaicism and allelic complexity for Pax6. Relationships between eye phenotype and genotype were developed. The present results demonstrated that development of the lens from the surface ectoderm requires a higher gene dose of Pax6 than development of the retina from the optic vesicle. We further anticipate that mice with somatic mosaicism in a targeted gene generated by CRISPR/Cas-mediated genome editing will give some insights for understanding the complexity in human congenital diseases that occur in mosaic form.

  9. Genome-wide analysis reveals methyl-CpG-binding protein 2-dependent regulation of microRNAs in a mouse model of Rett syndrome.

    PubMed

    Wu, Hao; Tao, Jifang; Chen, Pauline J; Shahab, Atif; Ge, Weihong; Hart, Ronald P; Ruan, Xiaoan; Ruan, Yijun; Sun, Yi E

    2010-10-19

    MicroRNAs (miRNAs) are a class of small, noncoding RNAs that function as posttranscriptional regulators of gene expression. Many miRNAs are expressed in the developing brain and regulate multiple aspects of neural development, including neurogenesis, dendritogenesis, and synapse formation. Rett syndrome (RTT) is a progressive neurodevelopmental disorder caused by mutations in the gene encoding methyl-CpG-binding protein 2 (MECP2). Although Mecp2 is known to act as a global transcriptional regulator, miRNAs that are directly regulated by Mecp2 in the brain are not known. Using massively parallel sequencing methods, we have identified miRNAs whose expression is altered in cerebella of Mecp2-null mice before and after the onset of severe neurological symptoms. In vivo genome-wide analyses indicate that promoter regions of a significant fraction of dysregulated miRNA transcripts, including a large polycistronic cluster of brain-specific miRNAs, are DNA-methylated and are bound directly by Mecp2. Functional analysis demonstrates that the 3' UTR of messenger RNA encoding Brain-derived neurotrophic factor (Bdnf) can be targeted by multiple miRNAs aberrantly up-regulated in the absence of Mecp2. Taken together, these results suggest that dysregulation of miRNAs may contribute to RTT pathoetiology and also may provide a valuable resource for further investigations of the role of miRNAs in RTT.

  10. Bioinformatics Pipelines for Targeted Resequencing and Whole-Exome Sequencing of Human and Mouse Genomes: A Virtual Appliance Approach for Instant Deployment

    PubMed Central

    Saeed, Isaam; Wong, Stephen Q.; Mar, Victoria; Goode, David L.; Caramia, Franco; Doig, Ken; Ryland, Georgina L.; Thompson, Ella R.; Hunter, Sally M.; Halgamuge, Saman K.; Ellul, Jason; Dobrovic, Alexander; Campbell, Ian G.; Papenfuss, Anthony T.; McArthur, Grant A.; Tothill, Richard W.

    2014-01-01

    Targeted resequencing by massively parallel sequencing has become an effective and affordable way to survey small to large portions of the genome for genetic variation. Despite the rapid development in open source software for analysis of such data, the practical implementation of these tools through construction of sequencing analysis pipelines still remains a challenging and laborious activity, and a major hurdle for many small research and clinical laboratories. We developed TREVA (Targeted REsequencing Virtual Appliance), making pre-built pipelines immediately available as a virtual appliance. Based on virtual machine technologies, TREVA is a solution for rapid and efficient deployment of complex bioinformatics pipelines to laboratories of all sizes, enabling reproducible results. The analyses that are supported in TREVA include: somatic and germline single-nucleotide and insertion/deletion variant calling, copy number analysis, and cohort-based analyses such as pathway and significantly mutated genes analyses. TREVA is flexible and easy to use, and can be customised by Linux-based extensions if required. TREVA can also be deployed on the cloud (cloud computing), enabling instant access without investment overheads for additional hardware. TREVA is available at http://bioinformatics.petermac.org/treva/. PMID:24752294

  11. Analysis of copy number variations in Mexican Holstein cattle using axiom genome-wide Bos 1 array

    PubMed Central

    Salomon-Torres, Ricardo; Villa-Angulo, Rafael; Villa-Angulo, Carlos

    2015-01-01

    Recently, for copy number variation (CNV) analysis, bovine researchers have focused mainly on the use of genome-wide SNP genotyping arrays. One of the highest densities commercially available SNPchips for cattle is the Affymetrix axiom genome-wide Bos 1, which assays 648,315 informative SNPs across the whole bovine genome. Here, we describe the microarray data, quality controls and validation implemented in a study published in Genetics and Molecular Research Journal in 2015 [1]. The microarray raw data has been deposited into Gene Expression Omnibus under accession #GSE54813. PMID:26981375

  12. Matrix metalloproteinase-9, -10, and -12, MDM2 and p53 expression in mouse liver during dimethylnitrosamine-induced oxidative stress and genomic injury.

    PubMed

    Syed, Ismail; Rathod, Jasmine; Parmar, Mayur; Corcoran, George B; Ray, Sidhartha D

    2012-06-01

    Treatment during early tumor development has greater success because tissue growth remains largely confined to its original locus. At later stages, malignant cells migrate from their original location, invade surrounding normal areas, and can disseminate widely throughout the body. Remodeling of the extracellular matrix (ECM) serves as a key facilitator of this dissemination. Proteolytic enzymes including plasmin and matrix metalloproteinases (MMPs) play an integral role in degrading the surrounding ECM proteins and clearing a path for tumor cell migration. Specific MMPs are highly expressed late during malignant tumor invasion. It is not understood whether early changes in MMPs influence apoptotic and necrotic cell death, processes known to govern the early stages of carcinogenesis. Similarly, the interaction between MDM2 and p53 is tightly controlled by a complex array of post-translational modifications, which in turn dictates the stability and activity of both p53 and MDM2. The present studies examine the hypothesis that model hepatotoxin dimethylnitrosamine (DMN), which is also a model carcinogen, will induce the MMP family of proteins after administration in hepatotoxic doses. Doses of 25, 50, and 100 mg/kg DMN were administered i.p. to male C3H mice. Changes in parameters associated with apoptotic and necrotic cell death, DNA damage, cell proliferation, and extracellular proteinases were examined in liver at 24 h. Serum ALT activity, oxidative stress [malondialdehyde], and caspase-activated DNAse mediated DNA laddering increased in a dose-dependent manner, as did the level of MDM2 protein. MMP-9, -10 and -12 (gelatinase-B, stromelysin-2, macrophage elastase), and p53 protein levels increased following 25 mg/kg DMN, but were successively decreased after higher DMN doses. The results of this study demonstrate changes in MDM2 and MMPs during DMN-induced acute liver injury and provide a plausible linkage between DMN-induced oxidative stress-mediated genomic

  13. Genome-wide Mechanosensitive MicroRNA (MechanomiR) Screen Uncovers Dysregulation of Their Regulatory Networks in the mdm Mouse Model of Muscular Dystrophy*

    PubMed Central

    Mohamed, Junaith S.; Hajira, Ameena; Lopez, Michael A.; Boriek, Aladin M.

    2015-01-01

    Muscular dystrophies (MDs) are a heterogeneous group of genetic and neuromuscular disorders, which result in severe loss of motor ability and skeletal muscle mass and function. Aberrant mechanotransduction and dysregulated-microRNA pathways are often associated with the progression of MD. Here, we hypothesized that dysregulation of mechanosensitive microRNAs (mechanomiRs) in dystrophic skeletal muscle plays a major role in the progression of MD. To test our hypothesis, we performed a genome-wide expression profile of anisotropically regulated mechanomiRs and bioinformatically analyzed their target gene networks. We assessed their functional roles in the advancement of MD using diaphragm muscles from mdm (MD with myositis) mice, an animal model of human tibial MD (titinopathy), and their wild-type littermates. We were able to show that ex vivo anisotropic mechanical stretch significantly alters the miRNA expression profile in diaphragm muscles from WT and mdm mice; as a result, some of the genes associated with MDs are dysregulated in mdm mice due to differential regulation of a distinct set of mechanomiRs. Interestingly, we found a contrasting expression pattern of the highly expressed let-7 family mechanomiRs, let-7e-5p and miR-98–5p, and their target genes associated with the extracellular matrix and TGF-β pathways, respectively, between WT and mdm mice. Gain- and loss-of-function analysis of let-7e-5p in myocytes isolated from the diaphragms of WT and mdm mice confirmed Col1a1, Col1a2, Col3a1, Col24a1, Col27a1, Itga1, Itga4, Scd1, and Thbs1 as target genes of let-7e-5p. Furthermore, we found that miR-98 negatively regulates myoblast differentiation. Our study therefore introduces additional biological players in the regulation of skeletal muscle structure and myogenesis that may contribute to unexplained disorders of MD. PMID:26272747

  14. Trace levels of mitomycin C disrupt genomic integrity and lead to DNA damage response defect in long-term-cultured human embryonic stem cells.

    PubMed

    Zhou, Di; Lin, Ge; Zeng, Si-Cong; Xiong, Bo; Xie, Ping-Yuan; Cheng, De-Hua; Zheng, Qing; Ouyang, Qi; Zhou, Xiao-Ying; Tang, Wei-Ling; Sun, Yi; Lu, Guang-Ying; Lu, Guang-Xiu

    2015-01-01

    How to maintain the genetic integrity of cultured human embryonic stem (hES) cells is raising crucial concerns for future clinical use in regenerative medicine. Mitomycin C(MMC), a DNA damage agent, is widely used for preparation of feeder cells in many laboratories. However, to what extent MMC affects the karyotypic stability of hES cells is not clear. Here, we measured residual MMC using High Performance Liquid Chromatography-Mass Spectrometry/Mass Spectrometry following each step of feeder preparation and found that 2.26 ± 0.77 and 3.50 ± 0.92 ng/ml remained in mouse feeder cells and human feeder cells, respectively. In addition, different amounts of MMC caused different chromosomal aberrations in hES cells. In particular, one abnormality, dup(1)(p32p36), was the same identical to one we previously reported in another hES cell line. Using Affymetrix SNP 6.0 arrays, the copy number variation changes of the hES cells maintained on MMC-inactivated feeders (MMC-feeder) were significantly more than those cultured on γ-inactivated feeder (IR-feeder) cells. Furthermore, DNA damage response (DDR) genes were down-regulated during long-term culture in the MMC-containing system, leading to DDR defect and shortened telomeres of hES cells, a sign of genomic instability. Therefore, MMC-feeder and MMC-induced genomic variation present an important safety problem that would limit such hES from being applied for future clinic use and drug screening.

  15. Mouse genetics: Catalogue and scissors

    PubMed Central

    Sung, Young Hoon; Baek, In-Jeoung; Seong, Je Kyung; Kim, Jin-Soo; Lee, Han-Woong

    2012-01-01

    Phenotypic analysis of gene-specific knockout (KO) mice has revolutionized our understanding of in vivo gene functions. As the use of mouse embryonic stem (ES) cells is inevitable for conventional gene targeting, the generation of knockout mice remains a very time-consuming and expensive process. To accelerate the large-scale production and phenotype analyses of KO mice, international efforts have organized global consortia such as the International Knockout Mouse Consortium (IKMC) and International Mouse Phenotype Consortium (IMPC), and they are persistently expanding the KO mouse catalogue that is publicly available for the researches studying specific genes of interests in vivo. However, new technologies, adopting zinc-finger nucleases (ZFNs) or Transcription Activator-Like Effector (TALE) Nucleases (TALENs) to edit the mouse genome, are now emerging as valuable and effective shortcuts alternative for the conventional gene targeting using ES cells. Here, we introduce the recent achievement of IKMC, and evaluate the significance of ZFN/TALEN technology in mouse genetics. [BMB Reports 2012; 45(12): 686-692] PMID:23261053

  16. Partnering for functional genomics research conference: Abstracts of poster presentations

    SciTech Connect

    1998-06-01

    This reports contains abstracts of poster presentations presented at the Functional Genomics Research Conference held April 16--17, 1998 in Oak Ridge, Tennessee. Attention is focused on the following areas: mouse mutagenesis and genomics; phenotype screening; gene expression analysis; DNA analysis technology development; bioinformatics; comparative analyses of mouse, human, and yeast sequences; and pilot projects to evaluate methodologies.

  17. Transcriptional signatures of BALB/c mouse macrophages housing multiplying Leishmania amazonensis amastigotes

    PubMed Central

    Fortéa, José Osorio y; de La Llave, Emilie; Regnault, Béatrice; Coppée, Jean-Yves; Milon, Geneviève; Lang, Thierry; Prina, Eric

    2009-01-01

    Background Mammal macrophages (MΦ) display a wide range of functions which contribute to surveying and maintaining tissue integrity. One such function is phagocytosis, a process known to be subverted by parasites like Leishmania (L). Indeed, the intracellular development of L. amazonensis amastigote relies on the biogenesis and dynamic remodelling of a phagolysosome, termed the parasitophorous vacuole, primarily within dermal MΦ. Results Using BALB/c mouse bone marrow-derived MΦ loaded or not with amastigotes, we analyzed the transcriptional signatures of MΦ 24 h later, when the amastigote population was growing. Total RNA from MΦ cultures were processed and hybridized onto Affymetrix Mouse430_2 GeneChips®, and some transcripts were also analyzed by Real-Time quantitative PCR (RTQPCR). A total of 1,248 probe-sets showed significant differential expression. Comparable fold-change values were obtained between the Affymetrix technology and the RTQPCR method. Ingenuity Pathway Analysis software® pinpointed the up-regulation of the sterol biosynthesis pathway (p-value = 1.31e-02) involving several genes (1.95 to 4.30 fold change values), and the modulation of various genes involved in polyamine synthesis and in pro/counter-inflammatory signalling. Conclusion Our findings suggest that the amastigote growth relies on early coordinated gene expression of the MΦ lipid and polyamine pathways. Moreover, these MΦ hosting multiplying L. amazonensis amastigotes display a transcriptional profile biased towards parasite-and host tissue-protective processes. PMID:19302708

  18. Prenatal stress-induced programming of genome-wide promoter DNA methylation in 5-HTT-deficient mice.

    PubMed

    Schraut, K G; Jakob, S B; Weidner, M T; Schmitt, A G; Scholz, C J; Strekalova, T; El Hajj, N; Eijssen, L M T; Domschke, K; Reif, A; Haaf, T; Ortega, G; Steinbusch, H W M; Lesch, K P; Van den Hove, D L

    2014-10-21

    The serotonin transporter gene (5-HTT/SLC6A4)-linked polymorphic region has been suggested to have a modulatory role in mediating effects of early-life stress exposure on psychopathology rendering carriers of the low-expression short (s)-variant more vulnerable to environmental adversity in later life. The underlying molecular mechanisms of this gene-by-environment interaction are not well understood, but epigenetic regulation including differential DNA methylation has been postulated to have a critical role. Recently, we used a maternal restraint stress paradigm of prenatal stress (PS) in 5-HTT-deficient mice and showed that the effects on behavior and gene expression were particularly marked in the hippocampus of female 5-Htt+/- offspring. Here, we examined to which extent these effects are mediated by differential methylation of DNA. For this purpose, we performed a genome-wide hippocampal DNA methylation screening using methylated-DNA immunoprecipitation (MeDIP) on Affymetrix GeneChip Mouse Promoter 1.0 R arrays. Using hippocampal DNA from the same mice as assessed before enabled us to correlate gene-specific DNA methylation, mRNA expression and behavior. We found that 5-Htt genotype, PS and their interaction differentially affected the DNA methylation signature of numerous genes, a subset of which showed overlap with the expression profiles of the corresponding transcripts. For example, a differentially methylated region in the gene encoding myelin basic protein (Mbp) was associated with its expression in a 5-Htt-, PS- and 5-Htt × PS-dependent manner. Subsequent fine-mapping of this Mbp locus linked the methylation status of two specific CpG sites to Mbp expression and anxiety-related behavior. In conclusion, hippocampal DNA methylation patterns and expression profiles of female prenatally stressed 5-Htt+/- mice suggest that distinct molecular mechanisms, some of which are promoter methylation-dependent, contribute to the behavioral effects of the 5-Htt

  19. Upstream—News in Genomics

    PubMed Central

    2002-01-01

    This report on the literature spans from May to July, highlighting breakthroughs on several important genomes, including mouse, zebrafish, Fugu and Plasmodium. Recent papers have reported on a mechanism for genome size reduction in Arabidopsis, comparisons and verifications of large-scale protein–protein interaction datasets, developments in RNA interference approaches for mammalian systems and a solidphase peptide tagging method for proteomics. PMID:18629049

  20. Genome-wide gene expression effects in B6C3F1 mouse intestinal epithelia following 7 and 90 days of exposure to hexavalent chromium in drinking water

    SciTech Connect

    Kopec, Anna K.; Kim, Suntae; Forgacs, Agnes L.; Zacharewski, Timothy R.; Proctor, Deborah M.; Harris, Mark A.; Haws, Laurie C.; Thompson, Chad M.

    2012-02-15

    Chronic administration of high doses of hexavalent chromium [Cr(VI)] as sodium dichromate dihydrate (SDD) elicits alimentary cancers in mice. To further elucidate key events underlying tumor formation, a 90-day drinking water study was conducted in B6C3F1 mice. Differential gene expression was examined in duodenal and jejunal epithelial samples following 7 or 90 days of exposure to 0, 0.3, 4, 14, 60, 170 or 520 mg/L SDD in drinking water. Genome-wide microarray analyses identified 6562 duodenal and 4448 jejunal unique differentially expressed genes at day 8, and 4630 and 4845 unique changes, respectively, in the duodenum and jejunum at day 91. Comparative analysis identified significant overlap in duodenal and jejunal differential gene expression. Automated dose–response modeling identified > 80% of the differentially expressed genes exhibited sigmoidal dose–response curves with EC{sub 50} values ranging from 10 to 100 mg/L SDD. Only 16 genes satisfying the dose-dependent differential expression criteria had EC{sub 50} values < 10 mg/L SDD, 3 of which were regulated by Nrf2, suggesting oxidative stress in response to SDD at low concentrations. Analyses of differentially expressed genes identified over-represented functions associated with oxidative stress, cell cycle, lipid metabolism, and immune responses consistent with the reported effects on redox status and histopathology at corresponding SDD drinking water concentrations. Collectively, these data are consistent with a mode of action involving oxidative stress and cytotoxicity as early key events. This suggests that the tumorigenic effects of chronic Cr(VI) oral exposure likely require chronic tissue damage and compensatory epithelial cell proliferation. Highlights: ► Mouse small intestine gene expression is highly responsive to hexavalent chromium [Cr(VI)]. ► Cr(VI) elicits more differential gene expression after 7 days of exposure than 90 days of exposure. ► Oral exposure to Cr(VI) leads to

  1. Comparative Analysis of Gene Regulation by the Transcription Factor PPARα between Mouse and Human

    PubMed Central

    Rakhshandehroo, Maryam; Hooiveld, Guido; Müller, Michael; Kersten, Sander

    2009-01-01

    Background Studies in mice have shown that PPARα is an important regulator of hepatic lipid metabolism and the acute phase response. However, little information is available on the role of PPARα in human liver. Here we set out to compare the function of PPARα in mouse and human hepatocytes via analysis of target gene regulation. Methodology/Principal Findings Primary hepatocytes from 6 human and 6 mouse donors were treated with PPARα agonist Wy14643 and gene expression profiling was performed using Affymetrix GeneChips followed by a systems biology analysis. Baseline PPARα expression was similar in human and mouse hepatocytes. Depending on species and time of exposure, Wy14643 significantly induced the expression of 362–672 genes. Surprisingly minor overlap was observed between the Wy14643-regulated genes from mouse and human, although more substantial overlap was observed at the pathway level. Xenobiotics metabolism and apolipoprotein synthesis were specifically regulated by PPARα in human hepatocytes, whereas glycolysis-gluconeogenesis was regulated specifically in mouse hepatocytes. Most of the genes commonly regulated in mouse and human were involved in lipid metabolism and many represented known PPARα targets, including CPT1A, HMGCS2, FABP1, ACSL1, and ADFP. Several genes were identified that were specifically induced by PPARα in human (MBL2, ALAS1, CYP1A1, TSKU) or mouse (Fbp2, lgals4, Cd36, Ucp2, Pxmp4). Furthermore, several putative novel PPARα targets were identified that were commonly regulated in both species, including CREB3L3, KLF10, KLF11 and MAP3K8. Conclusions/Significance Our results suggest that PPARα activation has a major impact on gene regulation in human hepatocytes. Importantly, the role of PPARα as master regulator of hepatic lipid metabolism is generally well-conserved between mouse and human. Overall, however, PPARα regulates a mostly divergent set of genes in mouse and human hepatocytes. PMID:19710929

  2. MouseMine: a new data warehouse for MGI.

    PubMed

    Motenko, H; Neuhauser, S B; O'Keefe, M; Richardson, J E

    2015-08-01

    MouseMine (www.mousemine.org) is a new data warehouse for accessing mouse data from Mouse Genome Informatics (MGI). Based on the InterMine software framework, MouseMine supports powerful query, reporting, and analysis capabilities, the ability to save and combine results from different queries, easy integration into larger workflows, and a comprehensive Web Services layer. Through MouseMine, users can access a significant portion of MGI data in new and useful ways. Importantly, MouseMine is also a member of a growing community of online data resources based on InterMine, including those established by other model organism databases. Adopting common interfaces and collaborating on data representation standards are critical to fostering cross-species data analysis. This paper presents a general introduction to MouseMine, presents examples of its use, and discusses the potential for further integration into the MGI interface.

  3. High-throughput mouse phenotyping.

    PubMed

    Gates, Hilary; Mallon, Ann-Marie; Brown, Steve D M

    2011-04-01

    Comprehensive phenotyping will be required to reveal the pleiotropic functions of a gene and to uncover the wider role of genetic loci within diverse biological systems. The challenge will be to devise phenotyping approaches to characterise the thousands of mutants that are being generated as part of international efforts to acquire a mutant for every gene in the mouse genome. In order to acquire robust datasets of broad based phenotypes from mouse mutants it is necessary to design and implement pipelines that incorporate standardised phenotyping platforms that are validated across diverse mouse genetics centres or mouse clinics. We describe here the rationale and methodology behind one phenotyping pipeline, EMPReSSslim, that was designed as part of the work of the EUMORPHIA and EUMODIC consortia, and which exemplifies some of the challenges facing large-scale phenotyping. EMPReSSslim captures a broad range of data on diverse biological systems, from biochemical to physiological amongst others. Data capture and dissemination is pivotal to the operation of large-scale phenotyping pipelines, including the definition of parameters integral to each phenotyping test and the associated ontological descriptions. EMPReSSslim data is displayed within the EuroPhenome database, where a variety of tools are available to allow the user to search for interesting biological or clinical phenotypes.

  4. Genome-wide analysis links NFATC2 with asparaginase hypersensitivity

    PubMed Central

    Fernandez, Christian A.; Smith, Colton; Yang, Wenjian; Mullighan, Charles G.; Qu, Chunxu; Larsen, Eric; Bowman, W. Paul; Liu, Chengcheng; Ramsey, Laura B.; Chang, Tamara; Karol, Seth E.; Loh, Mignon L.; Raetz, Elizabeth A.; Winick, Naomi J.; Hunger, Stephen P.; Carroll, William L.; Jeha, Sima; Pui, Ching-Hon; Evans, William E.; Devidas, Meenakshi

    2015-01-01

    Asparaginase is used to treat acute lymphoblastic leukemia (ALL); however, hypersensitivity reactions can lead to suboptimal asparaginase exposure. Our objective was to use a genome-wide approach to identify loci associated with asparaginase hypersensitivity in children with ALL enrolled on St. Jude Children’s Research Hospital (SJCRH) protocols Total XIIIA (n = 154), Total XV (n = 498), and Total XVI (n = 271), or Children’s Oncology Group protocols POG 9906 (n = 222) and AALL0232 (n = 2163). Germline DNA was genotyped using the Affymetrix 500K, Affymetrix 6.0, or the Illumina Exome BeadChip array. In multivariate logistic regression, the intronic rs6021191 variant in nuclear factor of activated T cells 2 (NFATC2) had the strongest association with hypersensitivity (P = 4.1 × 10−8; odds ratio [OR] = 3.11). RNA-seq data available from 65 SJCRH ALL tumor samples and 52 Yoruba HapMap samples showed that samples carrying the rs6021191 variant had higher NFATC2 expression compared with noncarriers (P = 1.1 × 10−3 and 0.03, respectively). The top ranked nonsynonymous polymorphism was rs17885382 in HLA-DRB1 (P = 3.2 × 10−6; OR = 1.63), which is in near complete linkage disequilibrium with the HLA-DRB1*07:01 allele we previously observed in a candidate gene study. The strongest risk factors for asparaginase allergy are variants within genes regulating the immune response. PMID:25987655

  5. Integration of Mouse Phenome Data Resources

    SciTech Connect

    Hancock, John M; Adams, Neils; Aidinis, Vassilis; Blake, Judith A; Bogue, Molly; Brown, Steve D M; Chesler, Elissa J; Davidson, Duncan; Duran, Christopher; Eppig, Janan T; Gailus-Durner, Valerie; Gkoutos, Georgios V; Greenaway, Simon; Angelis, Martin Hrabe de; Kollias, George; Leblanc, Sophie; Lee, Kirsty; Lengger, Christoph; Maier, Holger; Mallon, Ann-Marie; Masuya, Hiroshi; Melvin, David; Muller, Werner; Parkinson, Helen; Proctor, Glenn; Reuveni, Eli; Schofield, Paul; Shukla, Aadya; Smith, Cynthia; Toyoda, Tetsuro; Vasseur, Laurent; Wakana, Shigeharu; Walling, Alison; White, Jacqui; Wood, Joe; Zouberakis, Michalis

    2008-01-01

    Understanding the functions encoded in the mouse genome will be central to an understanding of the genetic basis of human disease. To achieve this it will be essential to be able to characterise the phenotypic consequences of variation and alterations in individual genes. Data on the phenotypes of mouse strains are currently held in a number of different forms (detailed descriptions of mouse lines, first line phenotyping data on novel mutations, data on the normal features of inbred lines, etc.) at many sites worldwide. For the most efficient use of these data sets, we have set in train a process to develop standards for the description of phenotypes (using ontologies), and file formats for the description of phenotyping protocols and phenotype data sets. This process is ongoing, and needs to be supported by the wider mouse genetics and phenotyping communities to succeed. We invite interested parties to contact us as we develop this process further.

  6. Mouse models of sickle cell disease.

    PubMed

    Beuzard, Y

    2008-01-01

    In the absence of a natural animal model for sickle cell disease, transgenic mouse models have been generated to better understand the complex pathophysiology of the disease and to evaluate potential specific therapies. In the early nineties, the simple addition of human globin genes induced the expression of hemoglobin S (HbS) or HbS-related human hemoglobins in mice still expressing mouse hemoglobin. To increase the proportion of human hemoglobin and the severity of the mouse sickle cell syndrome, the proportion of mouse hemoglobin could be decreased by a combination of mouse alpha- and beta-thalassemic defects, leading to complex genotypes and mild disease. Following the discovery of gene targeting in the mouse embryonic stem cells (ES cells), it was made possible to knock out all mouse adult globin genes (2alpha and 2beta) and to add the human homologous genes elsewhere in the mouse genome. In addition, the human gamma gene of fetal hemoglobin was protecting the fetus from HbS polymer formation. Accordingly, the resulting adult mouse models obtained in 1997, expressing human HbS-only, had a very severe anemia (Hb=5-6 g/dL). In order to survive, these "HbS-only mice" had to reduce the HbS concentration within the red blood cells. The phenotype could be less severe by adding modified human gamma genes, still expressed in adult mice. In 2006, a last "S-only" model was obtained by homologous knock in, replacing the mouse globin genes by human genes. This array of models contributes to better understand the role of different interacting factors in the complexity of sickle cell events, such as red cell defects, changes in blood flow and vaso-occlusion, hyperhemolysis, vascular tone dysregulation, oxidations, inflammation, activation and adhesion of cells, ischemia, reperfusion... In addition, each model has an appropriate usefulness to evaluate experimental therapies in vivo and to perform preclinical studies.

  7. The UCSC Genome Browser database: 2014 update

    PubMed Central

    Karolchik, Donna; Barber, Galt P.; Casper, Jonathan; Clawson, Hiram; Cline, Melissa S.; Diekhans, Mark; Dreszer, Timothy R.; Fujita, Pauline A.; Guruvadoo, Luvina; Haeussler, Maximilian; Harte, Rachel A.; Heitner, Steve; Hinrichs, Angie S.; Learned, Katrina; Lee, Brian T.; Li, Chin H.; Raney, Brian J.; Rhead, Brooke; Rosenbloom, Kate R.; Sloan, Cricket A.; Speir, Matthew L.; Zweig, Ann S.; Haussler, David; Kuhn, Robert M.; Kent, W. James

    2014-01-01

    The University of California Santa Cruz (UCSC) Genome Browser (http://genome.ucsc.edu) offers online public access to a growing database of genomic sequence and annotations for a large collection of organisms, primarily vertebrates, with an emphasis on the human and mouse genomes. The Browser’s web-based tools provide an integrated environment for visualizing, comparing, analysing and sharing both publicly available and user-generated genomic data sets. As of September 2013, the database contained genomic sequence and a basic set of annotation ‘tracks’ for ∼90 organisms. Significant new annotations include a 60-species multiple alignment conservation track on the mouse, updated UCSC Genes tracks for human and mouse, and several new sets of variation and ENCODE data. New software tools include a Variant Annotation Integrator that returns predicted functional effects of a set of variants uploaded as a custom track, an extension to UCSC Genes that displays haplotype alleles for protein-coding genes and an expansion of data hubs that includes the capability to display remotely hosted user-provided assembly sequence in addition to annotation data. To improve European access, we have added a Genome Browser mirror (http://genome-euro.ucsc.edu) hosted at Bielefeld University in Germany. PMID:24270787

  8. The UCSC Genome Browser database: 2014 update.

    PubMed

    Karolchik, Donna; Barber, Galt P; Casper, Jonathan; Clawson, Hiram; Cline, Melissa S; Diekhans, Mark; Dreszer, Timothy R; Fujita, Pauline A; Guruvadoo, Luvina; Haeussler, Maximilian; Harte, Rachel A; Heitner, Steve; Hinrichs, Angie S; Learned, Katrina; Lee, Brian T; Li, Chin H; Raney, Brian J; Rhead, Brooke; Rosenbloom, Kate R; Sloan, Cricket A; Speir, Matthew L; Zweig, Ann S; Haussler, David; Kuhn, Robert M; Kent, W James

    2014-01-01

    The University of California Santa Cruz (UCSC) Genome Browser (http://genome.ucsc.edu) offers online public access to a growing database of genomic sequence and annotations for a large collection of organisms, primarily vertebrates, with an emphasis on the human and mouse genomes. The Browser's web-based tools provide an integrated environment for visualizing, comparing, analysing and sharing both publicly available and user-generated genomic data sets. As of September 2013, the database contained genomic sequence and a basic set of annotation 'tracks' for ∼90 organisms. Significant new annotations include a 60-species multiple alignment conservation track on the mouse, updated UCSC Genes tracks for human and mouse, and several new sets of variation and ENCODE data. New software tools include a Variant Annotation Integrator that returns predicted functional effects of a set of variants uploaded as a custom track, an extension to UCSC Genes that displays haplotype alleles for protein-coding genes and an expansion of data hubs that includes the capability to display remotely hosted user-provided assembly sequence in addition to annotation data. To improve European access, we have added a Genome Browser mirror (http://genome-euro.ucsc.edu) hosted at Bielefeld University in Germany.

  9. An experimental validation of genomic selection in octoploid strawberry

    PubMed Central

    Gezan, Salvador A; Osorio, Luis F; Verma, Sujeet; Whitaker, Vance M

    2017-01-01

    The primary goal of genomic selection is to increase genetic gains for complex traits by predicting performance of individuals for which phenotypic data are not available. The objective of this study was to experimentally evaluate the potential of genomic selection in strawberry breeding and to define a strategy for its implementation. Four clonally replicated field trials, two in each of 2 years comprised of a total of 1628 individuals, were established in 2013–2014 and 2014–2015. Five complex yield and fruit quality traits with moderate to low heritability were assessed in each trial. High-density genotyping was performed with the Affymetrix Axiom IStraw90 single-nucleotide polymorphism array, and 17 479 polymorphic markers were chosen for analysis. Several methods were compared, including Genomic BLUP, Bayes B, Bayes C, Bayesian LASSO Regression, Bayesian Ridge Regression and Reproducing Kernel Hilbert Spaces. Cross-validation within training populations resulted in higher values than for true validations across trials. For true validations, Bayes B gave the highest predictive abilities on average and also the highest selection efficiencies, particularly for yield traits that were the lowest heritability traits. Selection efficiencies using Bayes B for parent selection ranged from 74% for average fruit weight to 34% for early marketable yield. A breeding strategy is proposed in which advanced selection trials are utilized as training populations and in which genomic selection can reduce the breeding cycle from 3 to 2 years for a subset of untested parents based on their predicted genomic breeding values. PMID:28090334

  10. Genome-wide analysis correlates Ayurveda Prakriti

    PubMed Central

    Govindaraj, Periyasamy; Nizamuddin, Sheikh; Sharath, Anugula; Jyothi, Vuskamalla; Rotti, Harish; Raval, Ritu; Nayak, Jayakrishna; Bhat, Balakrishna K.; Prasanna, B. V.; Shintre, Pooja; Sule, Mayura; Joshi, Kalpana S.; Dedge, Amrish P.; Bharadwaj, Ramachandra; Gangadharan, G. G.; Nair, Sreekumaran; Gopinath, Puthiya M.; Patwardhan, Bhushan; Kondaiah, Paturu; Satyamoorthy, Kapaettu; Valiathan, Marthanda Varma Sankaran; Thangaraj, Kumarasamy

    2015-01-01

    The practice of Ayurveda, the traditional medicine of India, is based on the concept of three major constitutional types (Vata, Pitta and Kapha) defined as “Prakriti”. To the best of our knowledge, no study has convincingly correlated genomic variations with the classification of Prakriti. In the present study, we performed genome-wide SNP (single nucleotide polymorphism) analysis (Affymetrix, 6.0) of 262 well-classified male individuals (after screening 3416 subjects) belonging to three Prakritis. We found 52 SNPs (p ≤ 1 × 10−5) were significantly different between Prakritis, without any confounding effect of stratification, after 106 permutations. Principal component analysis (PCA) of these SNPs classified 262 individuals into their respective groups (Vata, Pitta and Kapha) irrespective of their ancestry, which represent its power in categorization. We further validated our finding with 297 Indian population samples with known ancestry. Subsequently, we found that PGM1 correlates with phenotype of Pitta as described in the ancient text of Caraka Samhita, suggesting that the phenotypic classification of India’s traditional medicine has a genetic basis; and its Prakriti-based practice in vogue for many centuries resonates with personalized medicine. PMID:26511157

  11. Two thyroid hormone regulated genes, the beta-subunits of nerve growth factor (NGFB) and thyroid stimulating hormone (TSHB), are located less than 310 kb apart in both human and mouse genomes.

    PubMed

    Dracopoli, N C; Rose, E; Whitfield, G K; Guidon, P T; Bale, S J; Chance, P A; Kourides, I A; Housman, D E

    1988-08-01

    Two thyroid hormone regulated genes, the beta-subunits of nerve growth factor (NGFB) and thyroid stimulating hormone (TSHB), have been assigned to mouse chromosome 3 and human chromosome 1p22. We have used the techniques of linkage analysis and pulsed field gel electrophoresis to determine the proximity of these two antithetically regulated genes in this conserved linkage group. Four novel restriction fragment length polymorphisms were identified at the human TSHB gene. Two-point linkage analysis between TSHB and NGFB in 46 families, including the Centre d'Etude du Polymorphisme Humain (CEPH) reference panel, demonstrated no recombination (theta = 0.00, Z = 42.8). Analysis of this region by pulsed field gel electrophoresis showed that the genes for TSHB and NGFB are located less than 310 kb apart in man and 220 kb in the mouse.

  12. Single-strand DNA-mediated targeted mutagenesis of genomic DNA in early mouse embryos is stimulated by Rad51/54 and by Ku70/86 inhibition.

    PubMed

    Morozov, V; Wawrousek, E F

    2008-03-01

    Low and variable efficiency is a major problem in targeted gene alteration, which is used as a primary tool in gene therapy and animal model studies. We tested several types of constructs alone, or in combination with other factors, to introduce a point mutation into the alphaB-crystallin gene in one-celled mouse embryos. We found that co-injection of ssDNA along with antibodies against Ku70/86, or supplementing the system with hRad51/hRad54, increases efficiency of targeted mutagenesis. These findings suggest that proteins in the homologous recombination DNA repair pathway contribute, and that proteins involved in the alternative nonhomologous end-joining pathway inhibit, ssDNA-mediated targeted mutagenesis. This is the first successful demonstration of targeted mutation in early mouse embryos. This novel methodology of supplying protein factors to stimulate gene modification in the nucleus has not been previously reported.

  13. A genomic approach to myoblast fusion in Drosophila

    PubMed Central

    Estrada, Beatriz; Michelson, Alan M.

    2009-01-01

    Summary We have developed an integrated genetic, genomic and computational approach to identify and characterize genes involved in myoblast fusion in Drosophila. We first used fluorescence activated cell sorting to purify mesodermal cells both from wild-type embryos and from twelve variant genotypes in which muscle development is perturbed in known ways. Then, we obtained gene expression profiles for the purified cells by hybridizing isolated mesodermal RNA to Affymetrix GeneChip arrays. These data were subsequently compounded into a statistical meta-analysis that predicts myoblast subtype-specific gene expression signatures that were later validated by in situ hybridization experiments. Finally, we analyzed the myogenic functions of a subset of these myoblast genes using a double-stranded RNA interference assay in living embryos expressing green fluorescent protein under control of a muscle-specific promoter. This experimental strategy led to the identification of several previously uncharacterized genes required for myoblast fusion in Drosophila. PMID:18979251

  14. The transcriptional landscape of the mammalian genome.

    PubMed

    Carninci, P; Kasukawa, T; Katayama, S; Gough, J; Frith, M C; Maeda, N; Oyama, R; Ravasi, T; Lenhard, B; Wells, C; Kodzius, R; Shimokawa, K; Bajic, V B; Brenner, S E; Batalov, S; Forrest, A R R; Zavolan, M; Davis, M J; Wilming, L G; Aidinis, V; Allen, J E; Ambesi-Impiombato, A; Apweiler, R; Aturaliya, R N; Bailey, T L; Bansal, M; Baxter, L; Beisel, K W; Bersano, T; Bono, H; Chalk, A M; Chiu, K P; Choudhary, V; Christoffels, A; Clutterbuck, D R; Crowe, M L; Dalla, E; Dalrymple, B P; de Bono, B; Della Gatta, G; di Bernardo, D; Down, T; Engstrom, P; Fagiolini, M; Faulkner, G; Fletcher, C F; Fukushima, T; Furuno, M; Futaki, S; Gariboldi, M; Georgii-Hemming, P; Gingeras, T R; Gojobori, T; Green, R E; Gustincich, S; Harbers, M; Hayashi, Y; Hensch, T K; Hirokawa, N; Hill, D; Huminiecki, L; Iacono, M; Ikeo, K; Iwama, A; Ishikawa, T; Jakt, M; Kanapin, A; Katoh, M; Kawasawa, Y; Kelso, J; Kitamura, H; Kitano, H; Kollias, G; Krishnan, S P T; Kruger, A; Kummerfeld, S K; Kurochkin, I V; Lareau, L F; Lazarevic, D; Lipovich, L; Liu, J; Liuni, S; McWilliam, S; Madan Babu, M; Madera, M; Marchionni, L; Matsuda, H; Matsuzawa, S; Miki, H; Mignone, F; Miyake, S; Morris, K; Mottagui-Tabar, S; Mulder, N; Nakano, N; Nakauchi, H; Ng, P; Nilsson, R; Nishiguchi, S; Nishikawa, S; Nori, F; Ohara, O; Okazaki, Y; Orlando, V; Pang, K C; Pavan, W J; Pavesi, G; Pesole, G; Petrovsky, N; Piazza, S; Reed, J; Reid, J F; Ring, B Z; Ringwald, M; Rost, B; Ruan, Y; Salzberg, S L; Sandelin, A; Schneider, C; Schönbach, C; Sekiguchi, K; Semple, C A M; Seno, S; Sessa, L; Sheng, Y; Shibata, Y; Shimada, H; Shimada, K; Silva, D; Sinclair, B; Sperling, S; Stupka, E; Sugiura, K; Sultana, R; Takenaka, Y; Taki, K; Tammoja, K; Tan, S L; Tang, S; Taylor, M S; Tegner, J; Teichmann, S A; Ueda, H R; van Nimwegen, E; Verardo, R; Wei, C L; Yagi, K; Yamanishi, H; Zabarovsky, E; Zhu, S; Zimmer, A; Hide, W; Bult, C; Grimmond, S M; Teasdale, R D; Liu, E T; Brusic, V; Quackenbush, J; Wahlestedt, C; Mattick, J S; Hume, D A; Kai, C; Sasaki, D; Tomaru, Y; Fukuda, S; Kanamori-Katayama, M; Suzuki, M; Aoki, J; Arakawa, T; Iida, J; Imamura, K; Itoh, M; Kato, T; Kawaji, H; Kawagashira, N; Kawashima, T; Kojima, M; Kondo, S; Konno, H; Nakano, K; Ninomiya, N; Nishio, T; Okada, M; Plessy, C; Shibata, K; Shiraki, T; Suzuki, S; Tagami, M; Waki, K; Watahiki, A; Okamura-Oho, Y; Suzuki, H; Kawai, J; Hayashizaki, Y

    2005-09-02

    This study describes comprehensive polling of transcription start and termination sites and analysis of previously unidentified full-length complementary DNAs derived from the mouse genome. We identify the 5' and 3' boundaries of 181,047 transcripts with extensive variation in transcripts arising from alternative promoter usage, splicing, and polyadenylation. There are 16,247 new mouse protein-coding transcripts, including 5154 encoding previously unidentified proteins. Genomic mapping of the transcriptome reveals transcriptional forests, with overlapping transcription on both strands, separated by deserts in which few transcripts are observed. The data provide a comprehensive platform for the comparative analysis of mammalian transcriptional regulation in differentiation and development.

  15. SEARCH FOR GENOMIC ALTERATIONS IN MONOZYGOTIC TWINS DISCORDANT FOR CLEFT LIP AND/OR PALATE

    PubMed Central

    Kimani, Jane W.; Yoshiura, Koh-ichiro; Shi, Min; Jugessur, Astanand; Moretti-Ferreira, Danilo; Christensen, Kaare; Murray, Jeffrey C.

    2010-01-01

    Phenotypically discordant monozygotic twins offer the possibility of gene discovery through delineation of molecular abnormalities in one member of the twin pair. One proposed mechanism of discordance is postzygotically occurring genomic alterations resulting from mitotic recombination and other somatic changes. Detection of altered genomic fragments can reveal candidate gene loci that can be verified through additional analyses. We investigated this hypothesis using array comparative genomic hybridization; the 50K and 250K Affymetrix GeneChip® SNP arrays and an Illumina custom array consisting of 1,536 SNPs, to scan for genomic alterations in a sample of monozygotic twin pairs with discordant cleft lip and/or palate phenotypes. Paired analysis for deletions, amplifications and loss of heterozygosity, along with sequence verification of SNPs with discordant genotype calls did not reveal any genomic discordance between twin pairs in lymphocyte DNA samples. Our results demonstrate that postzygotic genomic alterations are not a common cause of monozygotic twin discordance for isolated cleft lip and/or palate. However, rare or balanced genomic alterations, tissue-specific events and small aberrations beyond the detection level of our experimental approach cannot be ruled out. The stability of genomes we observed in our study samples also suggests that detection of discordant events in other monozygotic twin pairs would be remarkable and of potential disease significance. PMID:19803774

  16. Genome Editing in Mice Using TALE Nucleases.

    PubMed

    Wefers, Benedikt; Brandl, Christina; Ortiz, Oskar; Wurst, Wolfgang; Kühn, Ralf

    2016-01-01

    Gene engineering for generating targeted mouse mutants is a key technology for biomedical research. Using TALENs as sequence-specific nucleases to induce targeted double-strand breaks, the mouse genome can be directly modified in zygotes in a single step without the need for embryonic stem cells. By embryo microinjection of TALEN mRNAs and targeting vectors, knockout and knock-in alleles can be generated fast and efficiently. In this chapter we provide protocols for the application of TALENs in mouse zygotes.

  17. Sense-antisense gene pairs: sequence, transcription, and structure are not conserved between human and mouse

    PubMed Central

    Wood, Emily J.; Chin-Inmanu, Kwanrutai; Jia, Hui; Lipovich, Leonard

    2013-01-01

    Previous efforts to characterize conservation between the human and mouse genomes focused largely on sequence comparisons. These studies are inherently limited because they don't account for gene structure differences, which may exist despite genomic sequence conservation. Recent high-throughput transcriptome studies have revealed widespread and extensive overlaps between genes, and transcripts, encoded on both strands of the genomic sequence. This overlapping gene organization, which produces sense-antisense (SAS) gene pairs, is capable of effecting regulatory cascades through established mechanisms. We present an evolutionary conservation assessment of SAS pairs, on three levels: genomic, transcriptomic, and structural. From a genome-wide dataset of human SAS pairs, we first identified orthologous loci in the mouse genome, then assessed their transcription in the mouse, and finally compared the genomic structures of SAS pairs expressed in both species. We found that approximately half of human SAS loci have single orthologous locations in the mouse genome; however, only half of those orthologous locations have SAS transcriptional activity in the mouse. This suggests that high human-mouse gene conservation overlooks widespread distinctions in SAS pair incidence and expression. We compared gene structures at orthologous SAS loci, finding frequent differences in gene structure between human and orthologous mouse SAS pair members. Our categorization of human SAS pairs with respect to mouse conservation of expression as well as structure points to limitations of mouse models. Gene structure differences, including at SAS loci, may account for some of the phenotypic distinctions between primates and rodents. Genes in non-conserved SAS pairs may contribute to evolutionary lineage-specific regulatory outcomes. PMID:24133500

  18. Mouse Curve Biometrics

    SciTech Connect

    Schulz, Douglas A.

    2007-10-08

    A biometric system suitable for validating user identity using only mouse movements and no specialized equipment is presented. Mouse curves (mouse movements with little or no pause between them) are individually classied and used to develop classication histograms, which are representative of an individual's typical mouse use. These classication histograms can then be compared to validate identity. This classication approach is suitable for providing continuous identity validation during an entire user session.

  19. Mouse homologues of human hereditary disease.

    PubMed Central

    Searle, A G; Edwards, J H; Hall, J G

    1994-01-01

    Details are given of 214 loci known to be associated with human hereditary disease, which have been mapped on both human and mouse chromosomes. Forty two of these have pathological variants in both species; in general the mouse variants are similar in their effects to the corresponding human ones, but exceptions include the Dmd/DMD and Hprt/HPRT mutations which cause little, if any, harm in mice. Possible reasons for phenotypic differences are discussed. In most pathological variants the gene product seems to be absent or greatly reduced in both species. The extensive data on conserved segments between human and mouse chromosomes are used to predict locations in the mouse of over 50 loci of medical interest which are mapped so far only on human chromosomes. In about 80% of these a fairly confident prediction can be made. Some likely homologies between mapped mouse loci and unmapped human ones are also given. Sixty six human and mouse proto-oncogene and growth factor gene homologies are also listed; those of confirmed location are all in known conserved segments. A survey of 18 mapped human disease loci and chromosome regions in which the manifestation or severity of pathological effects is thought to be the result of genomic imprinting shows that most of the homologous regions in the mouse are also associated with imprinting, especially those with homologues on human chromosomes 11p and 15q. Useful methods of accelerating the production of mouse models of human hereditary disease include (1) use of a supermutagen, such as ethylnitrosourea (ENU), (2) targeted mutagenesis involving ES cells, and (3) use of gene transfer techniques, with production of 'knockout mutations'. PMID:8151633

  20. Building a Brainier Mouse.

    ERIC Educational Resources Information Center

    Tsien, Joe Z.

    2000-01-01

    Describes a genetic engineering project to build an intelligent mouse. Cites understanding the molecular basis of learning and memory as a very important step. Concludes that while science will never create a genius mouse that plays the stock market, it can turn a mouse into a quick learner with a better memory. (YDS)

  1. An Affymetrix Microarray Design for Microbial Genotyping

    DTIC Science & Technology

    2009-10-01

    Clostridium botulinum APRT Okra 5 Clostridium botulinum A str. ATCC 19397 5 Clostridium botulinum ATCC 3502 40 Clostridium botulinum B str. Eklund 17B 5...Clostridium botulinum SNP B1 str. Okra plasmid pCLD 20 Clostridium botulinum B1 str. Okra plasmid pCLD 5 Clostridium botulinum Bf 5 Clostridium...botulinum HPT Eklund 17B 10 Clostridium botulinum HPT Loch Maree 20 Clostridium botulinum HPT Okra 5 Clostridium botulinum A3 str. Loch Maree 5

  2. Mouse Tumor Biology (MTB): a database of mouse models for human cancer.

    PubMed

    Bult, Carol J; Krupke, Debra M; Begley, Dale A; Richardson, Joel E; Neuhauser, Steven B; Sundberg, John P; Eppig, Janan T

    2015-01-01

    The Mouse Tumor Biology (MTB; http://tumor.informatics.jax.org) database is a unique online compendium of mouse models for human cancer. MTB provides online access to expertly curated information on diverse mouse models for human cancer and interfaces for searching and visualizing data associated with these models. The information in MTB is designed to facilitate the selection of strains for cancer research and is a platform for mining data on tumor development and patterns of metastases. MTB curators acquire data through manual curation of peer-reviewed scientific literature and from direct submissions by researchers. Data in MTB are also obtained from other bioinformatics resources including PathBase, the Gene Expression Omnibus and ArrayExpress. Recent enhancements to MTB improve the association between mouse models and human genes commonly mutated in a variety of cancers as identified in large-scale cancer genomics studies, provide new interfaces for exploring regions of the mouse genome associated with cancer phenotypes and incorporate data and information related to Patient-Derived Xenograft models of human cancers.

  3. Preterm Birth Genome Project (PGP) -- validation of resources for preterm birth genome-wide studies.

    PubMed

    Pennell, Craig E; Vadillo-Ortega, Felipe; Olson, David M; Ha, Eun-Hee; Williams, Scott; Frayling, Tim M; Dolan, Siobhan; Katz, Michael; Merialdi, Mario; Menon, Ramkumar

    2013-01-01

    We determined a series of quality control (QC) analyses to assess the usability of DNA collected and processed from different countries utilizing different DNA extraction techniques prior to genome-wide association studies (GWAS). The quality of DNA collected utilizing four different DNA extraction techniques and the impact of shipping DNA at different temperatures on array performance were evaluated. Fifteen maternal-fetal pairs were used from four countries. DNA was extracted using four approaches: whole blood, blood spots with whole genome amplification (WGA), saliva and buccal swab. Samples were sent to a genotyping facility, either on dry ice or at room temperature and genotyped using Affymetrix SNP array 6.0. QC measured included extraction techniques, effect of shipping temperatures, accuracy and Mendelian concordance. Significantly fewer (50 % ) single nucleotide polymorphisms (SNPs) passed QC metrics for buccal swab DNA (P < 0.0001) due to missing genotype data (P < 0.0001). Whole blood or saliva DNA had the highest call rates (99.2 0.4 % and 99.3 0.2 % , respectively) and Mendelian concordance. Shipment temperature had no effect. DNA from blood or saliva had the highest call rate accuracy, and buccal swabs had the lowest. DNA extracted from blood, saliva and blood spots were found suitable for GWAS in our study.

  4. Structure of the mouse IL-10 gene and chromosomal localization of the mouse and human genes

    SciTech Connect

    Kim, J.M.; Khan, T.A.; Moore, K.W. ); Brannan, C.I.; Copeland, N.G.; Jenkins, N.A. )

    1992-06-01

    The nucleotide sequence of a 7.2-kb segment containing the mouse IL-10 (mIL-10) gene was determined. Comparison to the mIL-10 cDNA sequence revealed the presence of five exons that span [approximately]5.1 kb of genomic DNA. The noncoding regions of the mIL-10 gene contain sequences that have been associated with transcriptional regulation of several cytokine genes. The mIL-10 gene was mapped to mouse chromosome 1 and the human IL-10 gene was also mapped to human chromosome 1. 35 refs., 4 figs., 3 tabs.

  5. Generation of targeted mouse mutants by embryo microinjection of TALENs.

    PubMed

    Wefers, Benedikt; Ortiz, Oskar; Wurst, Wolfgang; Kühn, Ralf

    2014-08-15

    Gene engineering for generating targeted mouse mutants is a key technology for biomedical research. Using TALENs as nucleases to induce targeted double-strand breaks, the mouse genome can be directly modified in zygotes in a single step, without the need for embryonic stem cells. Thereby, knockout and knockin alleles can be generated fast and efficiently by embryo microinjection of TALEN mRNAs and targeting vectors. In this article we present an introduction into the TALEN technology and provide protocols for the application of TALENs in mouse zygotes.

  6. Microarray-based genomic profiling reveals novel genomic aberrations in follicular lymphoma which associate with patient survival and gene expression status.

    PubMed

    Schwaenen, Carsten; Viardot, Andreas; Berger, Hilmar; Barth, Thomas F E; Bentink, Stefan; Döhner, Hartmut; Enz, Martina; Feller, Alfred C; Hansmann, Martin-Leo; Hummel, Michael; Kestler, Hans A; Klapper, Wolfram; Kreuz, Markus; Lenze, Dido; Loeffler, Markus; Möller, Peter; Müller-Hermelink, Hans-Konrad; Ott, German; Rosolowski, Maciej; Rosenwald, Andreas; Ruf, Sandra; Siebert, Reiner; Spang, Rainer; Stein, Harald; Truemper, Lorenz; Lichter, Peter; Bentz, Martin; Wessendorf, Swen

    2009-01-01

    Follicular lymphoma (FL) is characterized by a large number of chromosomal aberrations. However, their exact genomic extension and involved target genes remain to be determined. For this purpose, we used array-based intermediate-high resolution genomic profiling in combination with Affymetrix gene expression analysis. Tumor specimens from 128 FL patients were analyzed for the presence of genomic aberrations and the results were correlated to clinical data sets and mRNA expression levels. In 114 (89%) of the 128 analyzed cases, a total of 688 genomic aberrations (384 gains/amplifications and 304 losses) were detected. Frequent genomic aberrations were: -1p36 (18%), +2p15 (24%), -3q (14%), -6q (25%), +7p (19%), +7q (23%), +8q (14%), -9p (16%), -11q (15%), +12q (20%), -13q (11%), -17p (16%), +18p (18%), and +18q (28%). Critical segments of these imbalances were delineated to genomic fragments with a minimum size down to 0.2 Mb. By comparison of these with mRNA gene expression data, putative candidate genes were identified. Moreover, we found that deletions affecting the tumor suppressor gene CDKN2A/B on 9p21 were detected in nontransformed FL grade I-II. For this aberration as well as for -6q25 and -6q26, an association with inferior survival was observed.

  7. The gene for the serpin thrombin inhibitor (P17), protease nexin I, is located on human chromosome 2q33-q35 and on syntenic regions in the mouse and sheep genomes

    SciTech Connect

    Carter, R.E.; Burkin, D.J.; Fournier, R.E.K.

    1995-05-01

    Protease nexin I (PNI) is the most important physiologic regulator of {alpha}-thrombin in tissues. PNI is highly expressed and developmentally regulated in the nervous system where it is concentrated at neuromuscular junctions and also central synapses in the hippocampus and striatum. Approximately 10% of identified proteins at mammalian neuromuscular junctions are serine protease inhibitors, consistent with their central role in balancing serine protease activity to develop, maintain, and remodel synapses. Southern blot hybridization of PNI cDNA to somatic cell hybrids placed the structural gene for PNI (locus PI7) on human chromosome 2q33-q35 and to syntenic chromosomes in the mouse (chromosome 1) and sheep (chromosome 2). 30 refs., 2 figs.

  8. Mermaid, a family of short interspersed repetitive elements, is useful for zebrafish genome mapping.

    PubMed

    Shimoda, N; Chevrette, M; Ekker, M; Kikuchi, Y; Hotta, Y; Okamoto, H

    1996-03-07

    A family of short interspersed repetitive elements (SINEs), designated mermaid, is present in the genomes of fish, amphibian and primates, but absent in the mouse genome. We have demonstrated that the sequences of the mermaid family are highly polymorphic in the zebrafish genome as in the human genome. We have also shown that the mermaid sequence can be used to recover zebrafish specific DNA from zebrafish-mouse cell hybrids by using mermaid-specific oligonucleotides as PCR primers. Thus, the mermaid family serves as a valuable genetic tool for the zebrafish genome mapping.

  9. Genetic identification of a novel hantavirus of the harvest mouse Reithrodontomys megalotis.

    PubMed Central

    Hjelle, B; Chavez-Giles, F; Torrez-Martinez, N; Yates, T; Sarisky, J; Webb, J; Ascher, M

    1994-01-01

    We have cloned the S genomic segment of a novel hantavirus of the harvest mouse Reithrodontomys megalotis. The virus is phylogenetically distinct from other hantaviruses. The new hantavirus was identified in harvest mice separated by approximately 1,000 km. A wood rat (Neotoma mexicana) was found to be infected with the harvest mouse hantavirus. PMID:8084007

  10. Genetics and genomics of Drosophila mating behavior

    PubMed Central

    Mackay, Trudy F. C.; Heinsohn, Stefanie L.; Lyman, Richard F.; Moehring, Amanda J.; Morgan, Theodore J.; Rollmann, Stephanie M.

    2005-01-01

    The first steps of animal speciation are thought to be the development of sexual isolating mechanisms. In contrast to recent progress in understanding the genetic basis of postzygotic isolating mechanisms, little is known about the genetic architecture of sexual isolation. Here, we have subjected Drosophila melanogaster to 29 generations of replicated divergent artificial selection for mating speed. The phenotypic response to selection was highly asymmetrical in the direction of reduced mating speed, with estimates of realized heritability averaging 7%. The selection response was largely attributable to a reduction in female receptivity. We assessed the whole genome transcriptional response to selection for mating speed using Affymetrix GeneChips and a rigorous statistical analysis. Remarkably, >3,700 probe sets (21% of the array elements) exhibited a divergence in message levels between the Fast and Slow replicate lines. Genes with altered transcriptional abundance in response to selection fell into many different biological process and molecular function Gene Ontology categories, indicating substantial pleiotropy for this complex behavior. Future functional studies are necessary to test the extent to which transcript profiling of divergent selection lines accurately predicts genes that directly affect the selected trait. PMID:15851659

  11. Phylogenomic Insights into Mouse Evolution Using a Pseudoreference Approach

    PubMed Central

    Sarver, Brice A.J.; Keeble, Sara; Cosart, Ted; Tucker, Priscilla K.; Dean, Matthew D.

    2017-01-01

    Comparative genomic studies are now possible across a broad range of evolutionary timescales, but the generation and analysis of genomic data across many different species still present a number of challenges. The most sophisticated genotyping and down-stream analytical frameworks are still predominantly based on comparisons to high-quality reference genomes. However, established genomic resources are often limited within a given group of species, necessitating comparisons to divergent reference genomes that could restrict or bias comparisons across a phylogenetic sample. Here, we develop a scalable pseudoreference approach to iteratively incorporate sample-specific variation into a genome reference and reduce the effects of systematic mapping bias in downstream analyses. To characterize this framework, we used targeted capture to sequence whole exomes (∼54 Mbp) in 12 lineages (ten species) of mice spanning the Mus radiation. We generated whole exome pseudoreferences for all species and show that this iterative reference-based approach improved basic genomic analyses that depend on mapping accuracy while preserving the associated annotations of the mouse reference genome. We then use these pseudoreferences to resolve evolutionary relationships among these lineages while accounting for phylogenetic discordance across the genome, contributing an important resource for comparative studies in the mouse system. We also describe patterns of genomic introgression among lineages and compare our results to previous studies. Our general approach can be applied to whole or partitioned genomic data and is easily portable to any system with sufficient genomic resources, providing a useful framework for phylogenomic studies in mice and other taxa. PMID:28338821

  12. 76 FR 3642 - National Human Genome Research Institute; Notice of Closed Meetings

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-01-20

    ... HUMAN SERVICES National Institutes of Health National Human Genome Research Institute; Notice of Closed... of Committee: National Human Genome Research Institute Special Emphasis Panel, KOMP (KNOCK-OUT MOUSE..., MD 20814, 301-594- 4280, mckenneyk@mail.nih.gov . Name of Committee: National Human Genome...

  13. A Pooled Genome-Wide Association Study of Asperger Syndrome.

    PubMed

    Warrier, Varun; Chakrabarti, Bhismadev; Murphy, Laura; Chan, Allen; Craig, Ian; Mallya, Uma; Lakatošová, Silvia; Rehnstrom, Karola; Peltonen, Leena; Wheelwright, Sally; Allison, Carrie; Fisher, Simon E; Baron-Cohen, Simon

    2015-01-01

    Asperger Syndrome (AS) is a neurodevelopmental condition characterized by impairments in social interaction and communication, alongside the presence of unusually repetitive, restricted interests and stereotyped behaviour. Individuals with AS have no delay in cognitive and language development. It is a subset of Autism Spectrum Conditions (ASC), which are highly heritable and has a population prevalence of approximately 1%. Few studies have investigated the genetic basis of AS. To address this gap in the literature, we performed a genome-wide pooled DNA association study to identify candidate loci in 612 individuals (294 cases and 318 controls) of Caucasian ancestry, using the Affymetrix GeneChip Human Mapping version 6.0 array. We identified 11 SNPs that had a p-value below 1x10-5. These SNPs were independently genotyped in the same sample. Three of the SNPs (rs1268055, rs7785891 and rs2782448) were nominally significant, though none remained significant after Bonferroni correction. Two of our top three SNPs (rs7785891 and rs2782448) lie in loci previously implicated in ASC. However, investigation of the three SNPs in the ASC genome-wide association dataset from the Psychiatric Genomics Consortium indicated that these three SNPs were not significantly associated with ASC. The effect sizes of the variants were modest, indicating that our study was not sufficiently powered to identify causal variants with precision.

  14. A Pooled Genome-Wide Association Study of Asperger Syndrome

    PubMed Central

    Warrier, Varun; Chakrabarti, Bhismadev; Murphy, Laura; Chan, Allen; Craig, Ian; Mallya, Uma; Lakatošová, Silvia; Rehnstrom, Karola; Wheelwright, Sally; Allison, Carrie; Fisher, Simon E.; Baron-Cohen, Simon

    2015-01-01

    Asperger Syndrome (AS) is a neurodevelopmental condition characterized by impairments in social interaction and communication, alongside the presence of unusually repetitive, restricted interests and stereotyped behaviour. Individuals with AS have no delay in cognitive and language development. It is a subset of Autism Spectrum Conditions (ASC), which are highly heritable and has a population prevalence of approximately 1%. Few studies have investigated the genetic basis of AS. To address this gap in the literature, we performed a genome-wide pooled DNA association study to identify candidate loci in 612 individuals (294 cases and 318 controls) of Caucasian ancestry, using the Affymetrix GeneChip Human Mapping version 6.0 array. We identified 11 SNPs that had a p-value below 1x10-5. These SNPs were independently genotyped in the same sample. Three of the SNPs (rs1268055, rs7785891 and rs2782448) were nominally significant, though none remained significant after Bonferroni correction. Two of our top three SNPs (rs7785891 and rs2782448) lie in loci previously implicated in ASC. However, investigation of the three SNPs in the ASC genome-wide association dataset from the Psychiatric Genomics Consortium indicated that these three SNPs were not significantly associated with ASC. The effect sizes of the variants were modest, indicating that our study was not sufficiently powered to identify causal variants with precision. PMID:26176695

  15. Antarctic Genomics

    PubMed Central

    Clarke, Andrew; Cockell, Charles S.; Convey, Peter; Detrich III, H. William; Fraser, Keiron P. P.; Johnston, Ian A.; Methe, Barbara A.; Murray, Alison E.; Peck, Lloyd S.; Römisch, Karin; Rogers, Alex D.

    2004-01-01

    With the development of genomic science and its battery of technologies, polar biology stands on the threshold of a revolution, one that will enable the investigation of important questions of unprecedented scope and with extraordinary depth and precision. The exotic organisms of polar ecosystems are ideal candidates for genomic analysis. Through such analyses, it will be possible to learn not only the novel features that enable polar organisms to survive, and indeed thrive, in their extreme environments, but also fundamental biological principles that are common to most, if not all, organisms. This article aims to review recent developments in Antarctic genomics and to demonstrate the global context of such studies. PMID:18629155

  16. Aberrant Splicing in Transgenes Containing Introns, Exons, and V5 Epitopes: Lessons from Developing an FSHD Mouse Model Expressing a D4Z4 Repeat with Flanking Genomic Sequences

    PubMed Central

    Ansseau, Eugénie; Domire, Jacqueline S.; Wallace, Lindsay M.; Eidahl, Jocelyn O.; Guckes, Susan M.; Giesige, Carlee R.; Pyne, Nettie K.; Belayew, Alexandra; Harper, Scott Q.

    2015-01-01

    The DUX4 gene, encoded within D4Z4 repeats on human chromosome 4q35, has recently emerged as a key factor in the pathogenic mechanisms underlying Facioscapulohumeral muscular dystrophy (FSHD). This recognition prompted development of animal models expressing the DUX4 open reading frame (ORF) alone or embedded within D4Z4 repeats. In the first published model, we used adeno-associated viral vectors (AAV) and strong viral control elements (CMV promoter, SV40 poly A) to demonstrate that the DUX4 cDNA caused dose-dependent toxicity in mouse muscles. As a follow-up, we designed a second generation of DUX4-expressing AAV vectors to more faithfully genocopy the FSHD-permissive D4Z4 repeat region located at 4q35. This new vector (called AAV.D4Z4.V5.pLAM) contained the D4Z4/DUX4 promoter region, a V5 epitope-tagged DUX4 ORF, and the natural 3’ untranslated region (pLAM) harboring two small introns, DUX4 exons 2 and 3, and the non-canonical poly A signal required for stabilizing DUX4 mRNA in FSHD. AAV.D4Z4.V5.pLAM failed to recapitulate the robust pathology of our first generation vectors following delivery to mouse muscle. We found that the DUX4.V5 junction sequence created an unexpected splice donor in the pre-mRNA that was preferentially utilized to remove the V5 coding sequence and DUX4 stop codon, yielding non-functional DUX4 protein with 55 additional residues on its carboxyl-terminus. Importantly, we further found that aberrant splicing could occur in any expression construct containing a functional splice acceptor and sequences resembling minimal splice donors. Our findings represent an interesting case study with respect to AAV.D4Z4.V5.pLAM, but more broadly serve as a note of caution for designing constructs containing V5 epitope tags and/or transgenes with downstream introns and exons. PMID:25742305

  17. Genomic Testing

    MedlinePlus

    ... Services released a report identifying gaps in the regulation, oversight, and usefulness of genetic testing. They expressed ... December 20, 2016 Content source: Center for Surveillance, Epidemiology and Laboratory Services (CSELS) , Public Health Genomics Email ...

  18. Genome-wide association study and mouse expression data identify a highly conserved 32 kb intergenic region between WNT3 and WNT9b as possible susceptibility locus for isolated classic exstrophy of the bladder.

    PubMed

    Reutter, Heiko; Draaken, Markus; Pennimpede, Tracie; Wittler, Lars; Brockschmidt, Felix F; Ebert, Anne-Karolin; Bartels, Enrika; Rösch, Wolfgang; Boemers, Thomas M; Hirsch, Karin; Schmiedeke, Eberhard; Meesters, Christian; Becker, Tim; Stein, Raimund; Utsch, Boris; Mangold, Elisabeth; Nordenskjöld, Agneta; Barker, Gillian; Kockum, Christina Clementsson; Zwink, Nadine; Holmdahl, Gundula; Läckgren, Göran; Jenetzky, Ekkehart; Feitz, Wouter F J; Marcelis, Carlo; Wijers, Charlotte H W; Van Rooij, Iris A L M; Gearhart, John P; Herrmann, Bernhard G; Ludwig, Michael; Boyadjiev, Simeon A; Nöthen, Markus M; Mattheisen, Manuel

    2014-10-15

    Bladder exstrophy-epispadias complex (BEEC), the severe end of the urorectal malformation spectrum, has a profound impact on continence as well as sexual and renal functions. It is widely accepted that for the majority of cases the genetic basis appears to be multifactorial. Here, we report the first study which utilizes genome-wide association methods to analyze a cohort comprising patients presenting the most common BEEC form, classic bladder exstrophy (CBE), to identify common variation associated with risk for isolated CBE. We employed discovery and follow-up samples comprising 218 cases/865 controls and 78 trios in total, all of European descent. Our discovery sample identified a marker near SALL1, showing genome-wide significant association with CBE. However, analyses performed on follow-up samples did not add further support to these findings. We were also able to identify an association with CBE across our study samples (discovery: P = 8.88 × 10(-5); follow-up: P = 0.0025; combined: 1.09 × 10(-6)) in a highly conserved 32 kb intergenic region containing regulatory elements between WNT3 and WNT9B. Subsequent analyses in mice revealed expression for both genes in the genital region during stages relevant to the development of CBE in humans. Unfortunately, we were not able to replicate the suggestive signal for WNT3 and WNT9B in a sample that was enriched for non-CBE BEEC cases (P = 0.51). Our suggestive findings support the hypothesis that larger samples are warranted to identify association of common variation with CBE.

  19. Genome-Wide Association Study of Copy Number Variations in Patients with Familial Neurocardiogenic Syncope.

    PubMed

    Demir, Emre; Hasdemir, Can; Ak, Handan; Atay, Sevcan; Aydin, Hikmet Hakan

    2016-08-01

    Neurocardiogenic syncope (NCS) is the most frequent type of syncope characterized by a self-limited episode of systemic hypotension. In this study, we conducted the first genome-wide association study testing copy number variations for association with NCS. Study population consisted of 107 consecutive patients with recurrent syncope and positive head-up tilt table testing. Four families with NCS were selected for CNV analysis. Affymetrix GeneChip(®) SNP 6.0 array was used for CNV analysis. Data and statistical analysis were performed with Affymetrix genotyping console 4.0 and GraphPad Prism v6. Positive family history of NCS was present in 19.6 % (n = 21) in our study population (n = 107). Twenty-six CNV regions were found to be significantly altered in families with NCS (P < 0.05). Several CNVs were identified in families with NCS. Further studies comprising wider study population are required to determine the effect of these variations on NCS development.

  20. The Functional Genomics Initiative at Oak Ridge National Laboratory

    SciTech Connect

    Johnson, Dabney; Justice, Monica; Beattle, Ken; Buchanan, Michelle; Ramsey, Michael; Ramsey, Rose; Paulus, Michael; Ericson, Nance; Allison, David; Kress, Reid; Mural, Richard; Uberbacher, Ed; Mann, Reinhold

    1997-12-31

    The Functional Genomics Initiative at the Oak Ridge National Laboratory integrates outstanding capabilities in mouse genetics, bioinformatics, and instrumentation. The 50 year investment by the DOE in mouse genetics/mutagenesis has created a one-of-a-kind resource for generating mutations and understanding their biological consequences. It is generally accepted that, through the mouse as a surrogate for human biology, we will come to understand the function of human genes. In addition to this world class program in mammalian genetics, ORNL has also been a world leader in developing bioinformatics tools for the analysis, management and visualization of genomic data. Combining this expertise with new instrumentation technologies will provide a unique capability to understand the consequences of mutations in the mouse at both the organism and molecular levels. The goal of the Functional Genomics Initiative is to develop the technology and methodology necessary to understand gene function on a genomic scale and apply these technologies to megabase regions of the human genome. The effort is scoped so as to create an effective and powerful resource for functional genomics. ORNL is partnering with the Joint Genome Institute and other large scale sequencing centers to sequence several multimegabase regions of both human and mouse genomic DNA, to identify all the genes in these regions, and to conduct fundamental surveys to examine gene function at the molecular and organism level. The Initiative is designed to be a pilot for larger scale deployment in the post-genome era. Technologies will be applied to the examination of gene expression and regulation, metabolism, gene networks, physiology and development.

  1. INFRAFRONTIER—providing mutant mouse resources as research tools for the international scientific community

    PubMed Central

    2015-01-01

    The laboratory mouse is a key model organism to investigate mechanism and therapeutics of human disease. The number of targeted genetic mouse models of disease is growing rapidly due to high-throughput production strategies employed by the International Mouse Phenotyping Consortium (IMPC) and the development of new, more efficient genome engineering techniques such as CRISPR based systems. We have previously described the European Mouse Mutant Archive (EMMA) resource and how this international infrastructure provides archiving and distribution worldwide for mutant mouse strains. EMMA has since evolved into INFRAFRONTIER (http://www.infrafrontier.eu), the pan-European research infrastructure for the systemic phenotyping, archiving and distribution of mouse disease models. Here we describe new features including improved search for mouse strains, support for new embryonic stem cell resources, access to training materials via a comprehensive knowledgebase and the promotion of innovative analytical and diagnostic techniques. PMID:25414328

  2. Molecular analysis of heritable mouse mutations.

    PubMed

    Rinchik, E M

    1987-10-01

    Germ-line mutations of the mouse have for years comprised one class of biological markers for mammalian reproductive and developmental toxicology. Understanding the molecular nature of mutations and the mechanisms by which mutations are translated into specific (and often complex) phenotypes, however, still looms as a major goal of mammalian biology. Molecular genetic analysis of heritable mouse mutations constitutes a significant, experimentally malleable strategy for relating genomic DNA structure to genic expression and function in mammals. The integrated use of recombinant DNA technology, which allows both the identification and analysis of expression of single genes, and classical genetic and cytogenetic analysis, which allow the important correlation between basic DNA defects and the organismic consequences of such defects, has been crucial to this strategy. Some of the approaches (e.g., specific-gene cloning, random-clone analysis of genomic regions, insertional mutagenesis) for studying the nature and effect of both mutations and their wild-type counterparts that have resulted from this integration of genetic analysis and molecular biology have been applied to many loci within the murine genome. Studies of the nature and effects of a complex set of radiation-induced mutations at the dilute-short ear (d-se) region of chromosome 9, a specific example of this type of integrated analysis, are discussed.

  3. Identification and Applications of Repetitive Probes for Gene Mapping in the Mouse

    PubMed Central

    Siracusa, L. D.; Jenkins, N. A.; Copeland, N. G.

    1991-01-01

    Interspecific mouse hybrids that are viable and fertile provide a wealth of genetic variation that is useful for gene mapping. We are using this genetic variation to develop multilocus linkage maps of the mouse genome. As an outgrowth of this work, we have identified three repetitive probes that collectively identify 28 loci dispersed on 16 of the 19 mouse autosomes and the X chromosome. These loci establish a skeleton linkage map that can be used to detect linkage over much of the mouse genome. The molecular probes are derived from the mouse mammary tumor virus envelope gene, the ornithine decarboxylase gene, and the triose phosphate isomerase gene. The ability to scan the mouse genome quickly and efficiently in an interspecific cross using these three repetitive probes makes this system a powerful tool for identifying the chromosomal location of mutations that have yet to be cloned, mapping multigenic traits, and identifying recessive protooncogene loci associated with murine neoplastic disease. Ultimately, interspecific hybrids in conjunction with repetitive and single-copy probes will provide a rapid means to access virtually any gene of interest in the mouse genome at the molecular level. PMID:1673105

  4. The Vertebrate Genome Annotation browser 10 years on.

    PubMed

    Harrow, Jennifer L; Steward, Charles A; Frankish, Adam; Gilbert, James G; Gonzalez, Jose M; Loveland, Jane E; Mudge, Jonathan; Sheppard, Dan; Thomas, Mark; Trevanion, Stephen; Wilming, Laurens G

    2014-01-01

    The Vertebrate Genome Annotation (VEGA) database (http://vega.sanger.ac.uk), initially designed as a community resource for browsing manual annotation of the human genome project, now contains five reference genomes (human, mouse, zebrafish, pig and rat). Its introduction pages have been redesigned to enable the user to easily navigate between whole genomes and smaller multi-species haplotypic regions of interest such as the major histocompatibility complex. The VEGA browser is unique in that annotation is updated via the Human And Vertebrate Analysis aNd Annotation (HAVANA) update track every 2 weeks, allowing single gene updates to be made publicly available to the research community quickly. The user can now access different haplotypic subregions more easily, such as those from the non-obese diabetic mouse, and display them in a more intuitive way using the comparative tools. We also highlight how the user can browse manually annotated updated patches from the Genome Reference Consortium (GRC).

  5. Genome Sequencing.

    PubMed

    Verma, Mansi; Kulshrestha, Samarth; Puri, Ayush

    2017-01-01

    Genome sequencing is an important step toward correlating genotypes with phenotypic characters. Sequencing technologies are important in many fields in the life sciences, including functional genomics, transcriptomics, oncology, evolutionary biology, forensic sciences, and many more. The era of sequencing has been divided into three generations. First generation sequencing involved sequencing by synthesis (Sanger sequencing) and sequencing by cleavage (Maxam-Gilbert sequencing). Sanger sequencing led to the completion of various genome sequences (including human) and provided the foundation for development of other sequencing technologies. Since then, various techniques have been developed which can overcome some of the limitations of Sanger sequencing. These techniques are collectively known as "Next-generation sequencing" (NGS), and are further classified into second and third generation technologies. Although NGS methods have many advantages in terms of speed, cost, and parallelism, the accuracy and read length of Sanger sequencing is still superior and has confined the use of NGS mainly to resequencing genomes. Consequently, there is a continuing need to develop improved real time sequencing techniques. This chapter reviews some of the options currently available and provides a generic workflow for sequencing a genome.

  6. Isolation of the mouse homologue of BRCA1 and genetic mapping to mouse chromosome 11

    SciTech Connect

    Bennett, L.M.; Haugen-Strano, A.; Cochran, C.

    1995-10-10

    The BRCA1 gene is in large part responsible for hereditary human breast and ovarian cancer. Here we report the isolation of the murine Brca1 homologue cDNA clones. In addition, we identified genomic P1 clones that contain most, if not all, of the mouse Brca1 locus. DNA sequence analysis revealed that the mouse and human coding regions are 75% identical at the nucleotide level while the predicted amino acid identity is only 58%. A DNA sequence variant in the Brcal locus was identified and used to map this gene on a (Mus m. musculus Czech II x C57BL/KsJ)F1 x C57BL/KsJ intersubspecific backcross to distal mouse chromosome 11. The mapping of this gene to a region highly syntenic with human chromosome 17, coupled with Southern and Northern analyses, confirms that we isolated the murine Brcal homologue rather than a related RING finger gene. The isolation of the mouse Brca1 homologue will facilitate the creation of mouse models for germline BRCA1 defects. 12 refs., 3 figs.

  7. Localization and regulation of mouse pantothenate kinase 2.

    PubMed

    Leonardi, Roberta; Zhang, Yong-Mei; Lykidis, Athanasios; Rock, Charles O; Jackowski, Suzanne

    2007-10-02

    Coenzyme A (CoA) biosynthesis is initiated by pantothenate kinase (PanK) and CoA levels are controlled through differential expression and feedback regulation of PanK isoforms. PanK2 is a mitochondrial protein in humans, but comparative genomics revealed that acquisition of a mitochondrial targeting signal was limited to primates. Human and mouse PanK2 possessed similar biochemical properties, with inhibition by acetyl-CoA and activation by palmitoylcarnitine. Mouse PanK2 localized in the cytosol, and the expression of PanK2 was higher in human brain compared to mouse brain. Differences in expression and subcellular localization should be considered in developing a mouse model for human PanK2 deficiency.

  8. Identification of transcriptional regulators in the mouse immune system.

    PubMed

    Jojic, Vladimir; Shay, Tal; Sylvia, Katelyn; Zuk, Or; Sun, Xin; Kang, Joonsoo; Regev, Aviv; Koller, Daphne; Best, Adam J; Knell, Jamie; Goldrath, Ananda; Joic, Vladimir; Koller, Daphne; Shay, Tal; Regev, Aviv; Cohen, Nadia; Brennan, Patrick; Brenner, Michael; Kim, Francis; Rao, Tata Nageswara; Wagers, Amy; Heng, Tracy; Ericson, Jeffrey; Rothamel, Katherine; Ortiz-Lopez, Adriana; Mathis, Diane; Benoist, Christophe; Bezman, Natalie A; Sun, Joseph C; Min-Oo, Gundula; Kim, Charlie C; Lanier, Lewis L; Miller, Jennifer; Brown, Brian; Merad, Miriam; Gautier, Emmanuel L; Jakubzick, Claudia; Randolph, Gwendalyn J; Monach, Paul; Blair, David A; Dustin, Michael L; Shinton, Susan A; Hardy, Richard R; Laidlaw, David; Collins, Jim; Gazit, Roi; Rossi, Derrick J; Malhotra, Nidhi; Sylvia, Katelyn; Kang, Joonsoo; Kreslavsky, Taras; Fletcher, Anne; Elpek, Kutlu; Bellemarte-Pelletier, Angelique; Malhotra, Deepali; Turley, Shannon

    2013-06-01

    The differentiation of hematopoietic stem cells into cells of the immune system has been studied extensively in mammals, but the transcriptional circuitry that controls it is still only partially understood. Here, the Immunological Genome Project gene-expression profiles across mouse immune lineages allowed us to systematically analyze these circuits. To analyze this data set we developed Ontogenet, an algorithm for reconstructing lineage-specific regulation from gene-expression profiles across lineages. Using Ontogenet, we found differentiation stage-specific regulators of mouse hematopoiesis and identified many known hematopoietic regulators and 175 previously unknown candidate regulators, as well as their target genes and the cell types in which they act. Among the previously unknown regulators, we emphasize the role of ETV5 in the differentiation of γδ T cells. As the transcriptional programs of human and mouse cells are highly conserved, it is likely that many lessons learned from the mouse model apply to humans.

  9. Immunologic applications of conditional gene modification technology in the mouse.

    PubMed

    Sharma, Suveena; Zhu, Jinfang

    2014-04-02

    Since the success of homologous recombination in altering mouse genome and the discovery of Cre-loxP system, the combination of these two breakthroughs has created important applications for studying the immune system in the mouse. Here, we briefly summarize the general principles of this technology and its applications in studying immune cell development and responses; such implications include conditional gene knockout and inducible and/or tissue-specific gene over-expression, as well as lineage fate mapping. We then discuss the pros and cons of a few commonly used Cre-expressing mouse lines for studying lymphocyte development and functions. We also raise several general issues, such as efficiency of gene deletion, leaky activity of Cre, and Cre toxicity, all of which may have profound impacts on data interpretation. Finally, we selectively list some useful links to the Web sites as valuable mouse resources.

  10. Immunologic Applications of Conditional Gene Modification Technology in the Mouse

    PubMed Central

    Sharma, Suveena; Zhu, Jinfang

    2014-01-01

    Since the success of homologous recombination in altering mouse genome and the discovery of Cre-loxP system, the combination of these two breakthroughs has created important applications for studying the immune system in the mouse. Here, we briefly summarize the general principles of this technology and its applications in studying immune cell development and responses; such implications include conditional gene knockout and inducible and/or tissue-specific gene over-expression, as well as lineage fate mapping. We then discuss the pros and cons of a few commonly used Cre-expressing mouse lines for studying lymphocyte development and functions. We also raise several general issues, such as efficiency of gene deletion, leaky activity of Cre, and Cre toxicity, all of which may have profound impacts on data interpretation. Finally, we selectively list some useful links to the Web sites as valuable mouse resources. PMID:24700321

  11. Listeria Genomics

    NASA Astrophysics Data System (ADS)

    Cabanes, Didier; Sousa, Sandra; Cossart, Pascale

    The opportunistic intracellular foodborne pathogen Listeria monocytogenes has become a paradigm for the study of host-pathogen interactions and bacterial adaptation to mammalian hosts. Analysis of L. monocytogenes infection has provided considerable insight into how bacteria invade cells, move intracellularly, and disseminate in tissues, as well as tools to address fundamental processes in cell biology. Moreover, the vast amount of knowledge that has been gathered through in-depth comparative genomic analyses and in vivo studies makes L. monocytogenes one of the most well-studied bacterial pathogens. This chapter provides an overview of progress in the exploration of genomic, transcriptomic, and proteomic data in Listeria spp. to understand genome evolution and diversity, as well as physiological aspects of metabolism used by bacteria when growing in diverse environments, in particular in infected hosts.

  12. Bovine Genome Database: new tools for gleaning function from the Bos taurus genome.

    PubMed

    Elsik, Christine G; Unni, Deepak R; Diesh, Colin M; Tayal, Aditi; Emery, Marianne L; Nguyen, Hung N; Hagen, Darren E

    2016-01-04

    We report an update of the Bovine Genome Database (BGD) (http://BovineGenome.org). The goal of BGD is to support bovine genomics research by providing genome annotation and data mining tools. We have developed new genome and annotation browsers using JBrowse and WebApollo for two Bos taurus genome assemblies, the reference genome assembly (UMD3.1.1) and the alternate genome assembly (Btau_4.6.1). Annotation tools have been customized to highlight priority genes for annotation, and to aid annotators in selecting gene evidence tracks from 91 tissue specific RNAseq datasets. We have also developed BovineMine, based on the InterMine data warehousing system, to integrate the bovine genome, annotation, QTL, SNP and expression data with external sources of orthology, gene ontology, gene interaction and pathway information. BovineMine provides powerful query building tools, as well as customized query templates, and allows users to analyze and download genome-wide datasets. With BovineMine, bovine researchers can use orthology to leverage the curated gene pathways of model organisms, such as human, mouse and rat. BovineMine will be especially useful for gene ontology and pathway analyses in conjunction with GWAS and QTL studies.

  13. Impact of copy number variations burden on coding genome in humans using integrated high resolution arrays.

    PubMed

    Veerappa, Avinash M; Lingaiah, Kusuma; Vishweswaraiah, Sangeetha; Murthy, Megha N; Suresh, Raviraj V; Manjegowda, Dinesh S; Ramachandra, Nallur B

    2014-12-16

    Copy number variations (CNVs) alter the transcriptional and translational levels of genes by disrupting the coding structure and this burden of CNVs seems to be a significant contributor to phenotypic variations. Therefore it was necessary to assess the complexities of CNV burden on the coding genome. A total of 1715 individuals from 12 populations were used for CNV analysis in the present investigation. Analysis was performed using Affymetrix Genome-Wide Human SNP Array 6·0 chip and CytoScan High-Density arrays. CNVs were more frequently observed in the coding region than in the non-coding region. CNVs were observed vastly more frequently in the coding region than the non-coding region. CNVs were found to be enriched in the regions containing functional genes (83-96%) compared with the regions containing pseudogenes (4-17%). CNVs across the genome of an individual showed multiple hits across many genes, whose proteins interact physically and function under the same pathway. We identified varying numbers of proteins and degrees of interactions within protein complexes of single individual genomes. This study represents the first draft of a population-specific CNV genes map as well as a cross-populational map. The complex relationship of CNVs on genes and their physically interacting partners unravels many complexities involved in phenotype expression. This study identifies four mechanisms contributing to the complexities caused by the presence of multiple CNVs across many genes in the coding part of the genome.

  14. Genome-wide gene expression effects in B6C3F1 mouse intestinal epithelia following 7 and 90days of exposure to hexavalent chromium in drinking water.

    PubMed

    Kopec, Anna K; Kim, Suntae; Forgacs, Agnes L; Zacharewski, Timothy R; Proctor, Deborah M; Harris, Mark A; Haws, Laurie C; Thompson, Chad M

    2012-02-15

    Chronic administration of high doses of hexavalent chromium [Cr(VI)] as sodium dichromate dihydrate (SDD) elicits alimentary cancers in mice. To further elucidate key events underlying tumor formation, a 90-day drinking water study was conducted in B6C3F1 mice. Differential gene expression was examined in duodenal and jejunal epithelial samples following 7 or 90days of exposure to 0, 0.3, 4, 14, 60, 170 or 520mg/L SDD in drinking water. Genome-wide microarray analyses identified 6562 duodenal and 4448 jejunal unique differentially expressed genes at day 8, and 4630 and 4845 unique changes, respectively, in the duodenum and jejunum at day 91. Comparative analysis identified significant overlap in duodenal and jejunal differential gene expression. Automated dose-response modeling identified >80% of the differentially expressed genes exhibited sigmoidal dose-response curves with EC(50) values ranging from 10 to 100mg/L SDD. Only 16 genes satisfying the dose-dependent differential expression criteria had EC(50) values <10mg/L SDD, 3 of which were regulated by Nrf2, suggesting oxidative stress in response to SDD at low concentrations. Analyses of differentially expressed genes identified over-represented functions associated with oxidative stress, cell cycle, lipid metabolism, and immune responses consistent with the reported effects on redox status and histopathology at corresponding SDD drinking water concentrations. Collectively, these data are consistent with a mode of action involving oxidative stress and cytotoxicity as early key events. This suggests that the tumorigenic effects of chronic Cr(VI) oral exposure likely require chronic tissue damage and compensatory epithelial cell proliferation.

  15. Chromosomal localization of the human and mouse hyaluronan synthase genes

    SciTech Connect

    Spicer, A.P.; McDonald, J.A.; Seldin, M.F.

    1997-05-01

    We have recently identified a new vertebrate gene family encoding putative hyaluronan (HA) synthases. Three highly conserved related genes have been identified, designated HAS1, HAS2, and HAS3 in humans and Has1, Has2, and Has3 in the mouse. All three genes encode predicted plasma membrane proteins with multiple transmembrane domains and approximately 25% amino acid sequence identity to the Streptococcus pyogenes HA synthase, HasA. Furthermore, expression of any one HAS gene in transfected mammalian cells leads to high levels of HA biosynthesis. We now report the chromosomal localization of the three HAS genes in human and in mouse. The genes localized to three different positions within both the human and the mouse genomes. HAS1 was localized to the human chromosome 19q13.3-q13.4 boundary and Has1 to mouse Chr 17. HAS2 was localized to human chromosome 8q24.12 and Has2 to mouse Chr 15. HAS3 was localized to human chromosome 16q22.1 and Has3 to mouse Chr 8. The map position for HAS1 reinforces the recently reported relationship between a small region of human chromosome 19q and proximal mouse chromosome 17. HAS2 mapped outside the predicted critical region delineated for the Langer-Giedion syndrome and can thus be excluded as a candidate gene for this genetic syndrome. 33 refs., 2 figs.

  16. Structural Genomics of Protein Phosphatases

    SciTech Connect

    Almo,S.; Bonanno, J.; Sauder, J.; Emtage, S.; Dilorenzo, T.; Malashkevich, V.; Wasserman, S.; Swaminathan, S.; Eswaramoorthy, S.; et al

    2007-01-01

    The New York SGX Research Center for Structural Genomics (NYSGXRC) of the NIGMS Protein Structure Initiative (PSI) has applied its high-throughput X-ray crystallographic structure determination platform to systematic studies of all human protein phosphatases and protein phosphatases from biomedically-relevant pathogens. To date, the NYSGXRC has determined structures of 21 distinct protein phosphatases: 14 from human, 2 from mouse, 2 from the pathogen Toxoplasma gondii, 1 from Trypanosoma brucei, the parasite responsible for African sleeping sickness, and 2 from the principal mosquito vector of malaria in Africa, Anopheles gambiae. These structures provide insights into both normal and pathophysiologic processes, including transcriptional regulation, regulation of major signaling pathways, neural development, and type 1 diabetes. In conjunction with the contributions of other international structural genomics consortia, these efforts promise to provide an unprecedented database and materials repository for structure-guided experimental and computational discovery of inhibitors for all classes of protein phosphatases.

  17. Admixture mapping identifies introgressed genomic regions in North American canids.

    PubMed

    vonHoldt, Bridgett M; Kays, Roland; Pollinger, John P; Wayne, Robert K

    2016-06-01

    Hybrid zones typically contain novel gene combinations that can be tested by natural selection in a unique genetic context. Parental haplotypes that increase fitness can introgress beyond the hybrid zone, into the range of parental species. We used the Affymetrix canine SNP genotyping array to identify genomic regions tagged by multiple ancestry informative markers that are more frequent in an admixed population than expected. We surveyed a hybrid zone formed in the last 100 years as coyotes expanded their range into eastern North America. Concomitant with expansion, coyotes hybridized with wolves and some populations became more wolflike, such that coyotes in the northeast have the largest body size of any coyote population. Using a set of 3102 ancestry informative markers, we identified 60 differentially introgressed regions in 44 canines across this admixture zone. These regions are characterized by an excess of exogenous ancestry and, in northeastern coyotes, are enriched for genes affecting body size and skeletal proportions. Further, introgressed wolf-derived alleles have penetrated into Southern US coyote populations. Because no wolves currently exist in this area, these alleles are unlikely to have originated from recent hybridization. Instead, they probably originated from intraspecific gene flow or ancient admixture. We show that grey wolf and coyote admixture has far-reaching effects and, in addition to phenotypically transforming admixed populations, allows for the differential movement of alleles from different parental species to be tested in new genomic backgrounds.

  18. Genome mapping

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genome maps can be thought of much like road maps except that, instead of traversing across land, they traverse across the chromosomes of an organism. Genetic markers serve as landmarks along the chromosome and provide researchers information as to how close they may be to a gene or region of inter...

  19. Comparison of Comparative Genomic Hybridization Technologies across Microarray Platforms

    EPA Science Inventory

    In the 2007 Association of Biomolecular Resource Facilities (ABRF) Microarray Research Group (MARG) project, we analyzed HL-60 DNA with five platforms: Agilent, Affymetrix 500K, Affymetrix U133 Plus 2.0, Illumina, and RPCI 19K BAC arrays. Copy number variation (CNV) was analyzed ...

  20. Stabilin-1 is expressed in human breast cancer and supports tumor growth in mammary adenocarcinoma mouse model

    PubMed Central

    Riabov, Vladimir; Yin, Shuiping; Song, Bin; Avdic, Aida; Schledzewski, Kai; Ovsiy, Ilja; Gratchev, Alexei; Verdiell, Maria Llopis; Sticht, Carsten; Schmuttermaier, Christina; Schönhaber, Hiltrud; Weiss, Christel; Fields, Alan P.; Simon-Keller, Katja; Pfister, Frederick; Berlit, Sebastian; Marx, Alexander; Arnold, Bernd; Goerdt, Sergij; Kzhyshkowska, Julia

    2016-01-01

    Stabilin-1 is a multifunctional scavenger receptor expressed on alternatively-activated macrophages. Stabilin-1 mediates phagocytosis of “unwanted-self” components, intracellular sorting, and endocytic clearance of extracellular ligands including SPARC that modulates breast cancer growth. The expression of stabilin-1 was found on tumor-associated macrophages (TAM) in mouse and human cancers including melanoma, lymphoma, glioblastoma, and pancreatic insulinoma. Despite its tumor-promoting role in mouse models of melanoma and lymphoma the expression and functional role of stabilin-1 in breast cancer was unknown. Here, we demonstrate that stabilin-1 is expressed on TAM in human breast cancer, and its expression is most pronounced on stage I disease. Using stabilin-1 knockout (ko) mice we show that stabilin-1 facilitates growth of mouse TS/A mammary adenocarcinoma. Endocytosis assay on stabilin-1 ko TAM demonstrated impaired clearance of stabilin-1 ligands including SPARC that was capable of inducing cell death in TS/A cells. Affymetrix microarray analysis on purified TAM and reporter assays in stabilin-1 expressing cell lines demonstrated no influence of stabilin-1 expression on intracellular signalling. Our results suggest stabilin-1 mediated silent clearance of extracellular tumor growth-inhibiting factors (e.g. SPARC) as a mechanism of stabilin-1 induced tumor growth. Silent clearance function of stabilin-1 makes it an attractive candidate for delivery of immunomodulatory anti-cancer therapeutic drugs to TAM. PMID:27105498

  1. [Echocardiography in mouse].

    PubMed

    Fayssoil, A

    2008-06-01

    Assessing cardiac phenotype requires invasive or noninvasive techniques in mouse. Echocardiography is a noninvasive technique for evaluating cardiac function. The purpose of this paper is to underline echocardiography modalities and new tools Doppler applications like tissue Doppler imaging.

  2. Mouse Cleaning Apparatus and Method

    NASA Technical Reports Server (NTRS)

    Williams, Glenn L. (Inventor)

    2005-01-01

    The method of using the mouse pad cleaning apparatus is disclosed and claimed. The method comprises the steps of uncovering the mouse cleaning surface, applying the mouse and ball of the mouse to the cleaning surface, moving the mouse in a rotational pattern on the mouse cleaning surface, removing the mouse form the mouse cleaning surface, washing the cleaning surface, and covering the mouse cleaning surface. A mouse pad cleaning apparatus comprising a plurality of substrates, each said substrate having adhesive thereon, said plurality of substrates residing in and affixed to a receptacle. A single substrate having adhesive, which may be washable or non-washable, thereon may be employed. The washable adhesive may be an organopolysiloxane or gelatinous elastomer.

  3. Characterization of the mouse lymphotoxin-beta gene.

    PubMed

    Lawton, P; Nelson, J; Tizard, R; Browning, J L

    1995-01-01

    Lymphotoxin-beta (LT-beta) is a member of the TNF family of ligands which when expressed with lymphotoxin-alpha (LT-alpha, i.e., the original LT or TNF-beta) forms a heteromeric complex with LT-alpha on the cell surface. The mouse gene structure was determined by both cDNA cloning and analysis of a genomic DNA fragment encompassing the TNF/LT locus in the H-2 region of chromosome 17. The mouse and human genomic structures were found to be similar in terms of location in the class III region of the MHC; however, the mouse gene lacks one intron found in most members of the family. Both the cDNA and the genomic sequences revealed an altered splice donor in the conventional intron 2 position, rendering it nonfunctional. The altered gene retains an open reading frame such that an additional 66 amino acids are inserted into the stalk region connecting the transmembrane domain with the receptor binding domain encoded by exon 4 in this type II membrane protein. Northern analysis showed that this gene is expressed predominantly in lymphoid organs. The outlining of the complete mouse TNF locus will further studies of the relationship between these genes and immune function.

  4. Genome cartography: charting the apicomplexan genome.

    PubMed

    Kissinger, Jessica C; DeBarry, Jeremy

    2011-08-01

    Genes reside in particular genomic contexts that can be mapped at many levels. Historically, 'genetic maps' were used primarily to locate genes. Recent technological advances in the determination of genome sequences have made the analysis and comparison of whole genomes possible and increasingly tractable. What do we see if we shift our focus from gene content (the 'inventory' of genes contained within a genome) to the composition and organization of a genome? This review examines what has been learned about the evolution of the apicomplexan genome as well as the significance and impact of genomic location on our understanding of the eukaryotic genome and parasite biology.

  5. Whole-genome linkage analysis in mapping alcoholism genes using single-nucleotide polymorphisms and microsatellites.

    PubMed

    Wang, Shuang; Huang, Song; Liu, Nianjun; Chen, Liang; Oh, Cheongeun; Zhao, Hongyu

    2005-12-30

    There is currently a great interest in using single-nucleotide polymorphisms (SNPs) in genetic linkage and association studies because of the abundance of SNPs as well as the availability of high-throughput genotyping technologies. In this study, we compared the performance of whole-genome scans using SNPs with microsatellites on 143 pedigrees from the Collaborative Studies on Genetics of Alcoholism provided by Genetic Analysis Workshop 14. A total of 315 microsatellites and 10,081 SNPs from Affymetrix on 22 autosomal chromosomes were used in our analyses. We found that the results from the two scans had good overall concordance. One region on chromosome 2 and two regions on chromosome 7 showed significant linkage signals (i.e., NPL >or= 2) for alcoholism from both the SNP and microsatellite scans. The different results observed between the two scans may be explained by the difference observed in information content between the SNPs and the microsatellites.

  6. Personal genomics services: whose genomes?

    PubMed Central

    Gurwitz, David; Bregman-Eschet, Yael

    2009-01-01

    New companies offering personal whole-genome information services over the internet are dynamic and highly visible players in the personal genomics field. For fees currently ranging from US$399 to US$2500 and a vial of saliva, individuals can now purchase online access to their individual genetic information regarding susceptibility to a range of chronic diseases and phenotypic traits based on a genome-wide SNP scan. Most of the companies offering such services are based in the United States, but their clients may come from nearly anywhere in the world. Although the scientific validity, clinical utility and potential future implications of such services are being hotly debated, several ethical and regulatory questions related to direct-to-consumer (DTC) marketing strategies of genetic tests have not yet received sufficient attention. For example, how can we minimize the risk of unauthorized third parties from submitting other people's DNA for testing? Another pressing question concerns the ownership of (genotypic and phenotypic) information, as well as the unclear legal status of customers regarding their own personal information. Current legislation in the US and Europe falls short of providing clear answers to these questions. Until the regulation of personal genomics services catches up with the technology, we call upon commercial providers to self-regulate and coordinate their activities to minimize potential risks to individual privacy. We also point out some specific steps, along the trustee model, that providers of DTC personal genomics services as well as regulators and policy makers could consider for addressing some of the concerns raised below. PMID:19259127

  7. Making the Mouse Blastocyst: Past, Present, and Future.

    PubMed

    Rossant, Janet

    2016-01-01

    The study of the preimplantation mouse embryo has progressed over the past 50 years from descriptive biology through experimental embryology to molecular biology and genetics. Along the way, the molecular pathways that lead to the establishment of the three cell lineages of the blastocyst have become more clearly understood but the fundamental questions of lineage commitment remain the same as those laid out in early studies. With new tools of genome manipulation, in vivo imaging and single-cell analysis, the mouse blastocyst is an excellent model system to understand how organized cell fate decisions are made in a self-organizing developmental context.

  8. Citrus Genomics

    PubMed Central

    Talon, Manuel; Gmitter Jr., Fred G.

    2008-01-01

    Citrus is one of the most widespread fruit crops globally, with great economic and health value. It is among the most difficult plants to improve through traditional breeding approaches. Currently, there is risk of devastation by diseases threatening to limit production and future availability to the human population. As technologies rapidly advance in genomic science, they are quickly adapted to address the biological challenges of the citrus plant system and the world's industries. The historical developments of linkage mapping, markers and breeding, EST projects, physical mapping, an international citrus genome sequencing project, and critical functional analysis are described. Despite the challenges of working with citrus, there has been substantial progress. Citrus researchers engaged in international collaborations provide optimism about future productivity and contributions to the benefit of citrus industries worldwide and to the human population who can rely on future widespread availability of this health-promoting and aesthetically pleasing fruit crop. PMID:18509486

  9. Sequence, molecular properties, and chromosomal mapping of mouse lumican

    NASA Technical Reports Server (NTRS)

    Funderburgh, J. L.; Funderburgh, M. L.; Hevelone, N. D.; Stech, M. E.; Justice, M. J.; Liu, C. Y.; Kao, W. W.; Conrad, G. W.; Spooner, B. S. (Principal Investigator)

    1995-01-01

    PURPOSE. Lumican is a major proteoglycan of vertebrate cornea. This study characterizes mouse lumican, its molecular form, cDNA sequence, and chromosomal localization. METHODS. Lumican sequence was determined from cDNA clones selected from a mouse corneal cDNA expression library using a bovine lumican cDNA probe. Tissue expression and size of lumican mRNA were determined using Northern hybridization. Glycosidase digestion followed by Western blot analysis provided characterization of molecular properties of purified mouse corneal lumican. Chromosomal mapping of the lumican gene (Lcn) used Southern hybridization of a panel of genomic DNAs from an interspecific murine backcross. RESULTS. Mouse lumican is a 338-amino acid protein with high-sequence identity to bovine and chicken lumican proteins. The N-terminus of the lumican protein contains consensus sequences for tyrosine sulfation. A 1.9-kb lumican mRNA is present in cornea and several other tissues. Antibody against bovine lumican reacted with recombinant mouse lumican expressed in Escherichia coli and also detected high molecular weight proteoglycans in extracts of mouse cornea. Keratanase digestion of corneal proteoglycans released lumican protein, demonstrating the presence of sulfated keratan sulfate chains on mouse corneal lumican in vivo. The lumican gene (Lcn) was mapped to the distal region of mouse chromosome 10. The Lcn map site is in the region of a previously identified developmental mutant, eye blebs, affecting corneal morphology. CONCLUSIONS. This study demonstrates sulfated keratan sulfate proteoglycan in mouse cornea and describes the tools (antibodies and cDNA) necessary to investigate the functional role of this important corneal molecule using naturally occurring and induced mutants of the murine lumican gene.

  10. Ancient genomics

    PubMed Central

    Der Sarkissian, Clio; Allentoft, Morten E.; Ávila-Arcos, María C.; Barnett, Ross; Campos, Paula F.; Cappellini, Enrico; Ermini, Luca; Fernández, Ruth; da Fonseca, Rute; Ginolhac, Aurélien; Hansen, Anders J.; Jónsson, Hákon; Korneliussen, Thorfinn; Margaryan, Ashot; Martin, Michael D.; Moreno-Mayar, J. Víctor; Raghavan, Maanasa; Rasmussen, Morten; Velasco, Marcela Sandoval; Schroeder, Hannes; Schubert, Mikkel; Seguin-Orlando, Andaine; Wales, Nathan; Gilbert, M. Thomas P.; Willerslev, Eske; Orlando, Ludovic

    2015-01-01

    The past decade has witnessed a revolution in ancient DNA (aDNA) research. Although the field's focus was previously limited to mitochondrial DNA and a few nuclear markers, whole genome sequences from the deep past can now be retrieved. This breakthrough is tightly connected to the massive sequence throughput of next generation sequencing platforms and the ability to target short and degraded DNA molecules. Many ancient specimens previously unsuitable for DNA analyses because of extensive degradation can now successfully be used as source materials. Additionally, the analytical power obtained by increasing the number of sequence reads to billions effectively means that contamination issues that have haunted aDNA research for decades, particularly in human studies, can now be efficiently and confidently quantified. At present, whole genomes have been sequenced from ancient anatomically modern humans, archaic hominins, ancient pathogens and megafaunal species. Those have revealed important functional and phenotypic information, as well as unexpected adaptation, migration and admixture patterns. As such, the field of aDNA has entered the new era of genomics and has provided valuable information when testing specific hypotheses related to the past. PMID:25487338

  11. Ancient genomics.

    PubMed

    Der Sarkissian, Clio; Allentoft, Morten E; Ávila-Arcos, María C; Barnett, Ross; Campos, Paula F; Cappellini, Enrico; Ermini, Luca; Fernández, Ruth; da Fonseca, Rute; Ginolhac, Aurélien; Hansen, Anders J; Jónsson, Hákon; Korneliussen, Thorfinn; Margaryan, Ashot; Martin, Michael D; Moreno-Mayar, J Víctor; Raghavan, Maanasa; Rasmussen, Morten; Velasco, Marcela Sandoval; Schroeder, Hannes; Schubert, Mikkel; Seguin-Orlando, Andaine; Wales, Nathan; Gilbert, M Thomas P; Willerslev, Eske; Orlando, Ludovic

    2015-01-19

    The past decade has witnessed a revolution in ancient DNA (aDNA) research. Although the field's focus was previously limited to mitochondrial DNA and a few nuclear markers, whole genome sequences from the deep past can now be retrieved. This breakthrough is tightly connected to the massive sequence throughput of next generation sequencing platforms and the ability to target short and degraded DNA molecules. Many ancient specimens previously unsuitable for DNA analyses because of extensive degradation can now successfully be used as source materials. Additionally, the analytical power obtained by increasing the number of sequence reads to billions effectively means that contamination issues that have haunted aDNA research for decades, particularly in human studies, can now be efficiently and confidently quantified. At present, whole genomes have been sequenced from ancient anatomically modern humans, archaic hominins, ancient pathogens and megafaunal species. Those have revealed important functional and phenotypic information, as well as unexpected adaptation, migration and admixture patterns. As such, the field of aDNA has entered the new era of genomics and has provided valuable information when testing specific hypotheses related to the past.

  12. Identification of evolutionary hotspots in the rodent genomes.

    PubMed

    Yap, Von Bing; Pachter, Lior

    2004-04-01

    We describe a whole-genome comparative analysis of the human, mouse, and rat genomes to describe the average substitution patterns of four genomic regions: ancient repeats, rodent-specific DNA, exons, and conserved (coding and noncoding) regions, and to identify rodent evolutionary hotspots. In all types of regions, except the rodent-specific DNA, the rat branch is slightly longer than the mouse branch. Moreover, the mouse-rat distance is longer in the rodent-specific DNA than in the ancient repeats. Analysis of individual conserved regions with different substitution models yielded the conclusion that the Jukes-Cantor model is inadequate, and the Hasegawa-Kishino-Yano model is almost as good as the REV model. Using human as an outgroup, we identified 5055 evolutionary hotspots, which are highly conserved subalignment blocks (each consisting of at least 100 aligned sites and a small fraction of gaps) with a large and statistically significant difference in the branch lengths of the rodent species. The cutoffs used to identify the hotspots are partially based on estimates of the average rates of substitution. The fractions of hotspots overlapping with the rodent RefSeq genes, RefSeq exons, and ESTs are all higher than expected. Still, more than half of the hotspots lie in noncoding regions of the mouse genome. We believe that the hotspots represent biologically interesting regions in the rodent genomes.

  13. A Fast Implementation of a Scan Statistic for Identifying Chromosomal Patterns of Genome Wide Association Studies.

    PubMed

    Sun, Yan V; Jacobsen, Douglas M; Turner, Stephen T; Boerwinkle, Eric; Kardia, Sharon L R

    2009-03-15

    In order to take into account the complex genomic distribution of SNP variations when identifying chromosomal regions with significant SNP effects, a single nucleotide polymorphism (SNP) association scan statistic was developed. To address the computational needs of genome wide association (GWA) studies, a fast Java application, which combines single-locus SNP tests and a scan statistic for identifying chromosomal regions with significant clusters of significant SNP effects, was developed and implemented. To illustrate this application, SNP associations were analyzed in a pharmacogenomic study of the blood pressure lowering effect of thiazide-diuretics (N=195) using the Affymetrix Human Mapping 100K Set. 55,335 tagSNPs (pair-wise linkage disequilibrium R(2)<0.5) were selected to reduce the frequency correlation between SNPs. A typical workstation can complete the whole genome scan including 10,000 permutation tests within 3 hours. The most significant regions locate on chromosome 3, 6, 13 and 16, two of which contain candidate genes that may be involved in the underlying drug response mechanism. The computational performance of ChromoScan-GWA and its scalability were tested with up to 1,000,000 SNPs and up to 4,000 subjects. Using 10,000 permutations, the computation time grew linearly in these datasets. This scan statistic application provides a robust statistical and computational foundation for identifying genomic regions associated with disease and provides a method to compare GWA results even across different platforms.

  14. New types of mouse centromeric satellite DNAs.

    PubMed

    Kuznetsova, Inna S; Prusov, Andrey N; Enukashvily, Natella I; Podgornaya, Olga I

    2005-01-01

    Genomic databases do not contain complete sequences of the centromeric regions. We created a pUC19-based library of DNA fragments from isolated chromocentres of interphase nuclei. In this library we have found major satellite (MaSat) and two new satellite sequences - MS3 and MS4. The computer analysis of MS3 and MS4 sequences by alignment, fragment curved state and search for MAR motifs in comparison with the mouse major and minor satellite (MiSat) DNA has shown them to be new satellite fragments. Southern blot of MS3 and MS4 with total DNA digested by restriction enzymes shows the ladder characteristic of satellite DNA. 2.2% of the total DNA consists of MS3, the monomer of which is 150 bp long. The MS4 monomer is 300 bp long and accounts for 1.6% of the total DNA. On metaphase chromosomes MS3 and MS4 are located at the centromeric region. FISH analysis of L929 nuclei during the cell cycle showed relative positions of MaSat, MiSat, MS3, and MS4. All mapped satDNA fragments except MaSat belong to the outer layer of the chromocentres in the G0/G1 phase. MS3 is likely to be involved in the centromere formation. The mouse genome contains at least four satDNA types: AT-rich (MaSat and MiSat), and CG-rich (MS3 and MS4).

  15. Integration and Fixation Preferences of Human and Mouse Endogenous Retroviruses Uncovered with Functional Data Analysis

    PubMed Central

    Pini, Alessia; Chiaromonte, Francesca; Makova, Kateryna D.

    2016-01-01

    Endogenous retroviruses (ERVs), the remnants of retroviral infections in the germ line, occupy ~8% and ~10% of the human and mouse genomes, respectively, and affect their structure, evolution, and function. Yet we still have a limited understanding of how the genomic landscape influences integration and fixation of ERVs. Here we conducted a genome-wide study of the most recently active ERVs in the human and mouse genome. We investigated 826 fixed and 1,065 in vitro HERV-Ks in human, and 1,624 fixed and 242 polymorphic ETns, as well as 3,964 fixed and 1,986 polymorphic IAPs, in mouse. We quantitated >40 human and mouse genomic features (e.g., non-B DNA structure, recombination rates, and histone modifications) in ±32 kb of these ERVs’ integration sites and in control regions, and analyzed them using Functional Data Analysis (FDA) methodology. In one of the first applications of FDA in genomics, we identified genomic scales and locations at which these features display their influence, and how they work in concert, to provide signals essential for integration and fixation of ERVs. The investigation of ERVs of different evolutionary ages (young in vitro and polymorphic ERVs, older fixed ERVs) allowed us to disentangle integration vs. fixation preferences. As a result of these analyses, we built a comprehensive model explaining the uneven distribution of ERVs along the genome. We found that ERVs integrate in late-replicating AT-rich regions with abundant microsatellites, mirror repeats, and repressive histone marks. Regions favoring fixation are depleted of genes and evolutionarily conserved elements, and have low recombination rates, reflecting the effects of purifying selection and ectopic recombination removing ERVs from the genome. In addition to providing these biological insights, our study demonstrates the power of exploiting multiple scales and localization with FDA. These powerful techniques are expected to be applicable to many other genomic investigations

  16. Genome editing systems in novel therapies.

    PubMed

    Jang, Yoon-Young; Cai, Liuhong; Ye, Zhaohui

    2016-01-01

    Genome editing is the process in which DNA sequences at precise genomic locations are modified. In the past three decades, genome editing by homologous recombination has been successfully performed in mouse for generating genetic models. The low efficiency of this process in human cells, however, had prevented its clinical application until the recent advancements in designer endonuclease technologies. The significantly improved genome editing efficiencies aided by ZFN, TALEN, and CRISPR systems provide unprecedented opportunities not only for biomedical research, but also for developing novel therapies. Applications based on these genome editing tools to disrupt deleterious genes, correct genetic mutations, deliver functional transgenes more effectively or even modify the epigenetic landscape are being actively investigated for gene and cell therapy purposes. Encouraging results have been obtained in limited clinical trials in the past two years. While most of the applications are still in proof-of-principle or preclinical development stages, it is anticipated that the coming years will see increasing clinical success in novel therapies based on the modern genome editing technologies. It should be noted that critical issues still remain before the technologies can be translated into more reliable therapies. These key issues include off-target evaluation, establishing appropriate preclinical models and improving the currently low efficiency of homology-based precise gene replacement. In this review we discuss the preclinical and clinical studies aiming at translating the genome editing technologies as well as the issues that are important for more successful translation.

  17. Genomic Islands of Speciation in Anopheles gambiae

    PubMed Central

    Hahn, Matthew W; Nuzhdin, Sergey V

    2005-01-01

    The African malaria mosquito, Anopheles gambiae sensu stricto (A. gambiae), provides a unique opportunity to study the evolution of reproductive isolation because it is divided into two sympatric, partially isolated subtaxa known as M form and S form. With the annotated genome of this species now available, high-throughput techniques can be applied to locate and characterize the genomic regions contributing to reproductive isolation. In order to quantify patterns of differentiation within A. gambiae, we hybridized population samples of genomic DNA from each form to Affymetrix GeneChip microarrays. We found that three regions, together encompassing less than 2.8 Mb, are the only locations where the M and S forms are significantly differentiated. Two of these regions are adjacent to centromeres, on Chromosomes 2L and X, and contain 50 and 12 predicted genes, respectively. Sequenced loci in these regions contain fixed differences between forms and no shared polymorphisms, while no fixed differences were found at nearby control loci. The third region, on Chromosome 2R, contains only five predicted genes; fixed differences in this region were also verified by direct sequencing. These “speciation islands” remain differentiated despite considerable gene flow, and are therefore expected to contain the genes responsible for reproductive isolation. Much effort has recently been applied to locating the genes and genetic changes responsible for reproductive isolation between species. Though much can be inferred about speciation by studying taxa that have diverged for millions of years, studying differentiation between taxa that are in the early stages of isolation will lead to a clearer view of the number and size of regions involved in the genetics of speciation. Despite appreciable levels of gene flow between the M and S forms of A. gambiae, we were able to isolate three small regions of differentiation where genes responsible for ecological and behavioral isolation are

  18. The Mouse Isolated Perfused Kidney Technique.

    PubMed

    Czogalla, Jan; Schweda, Frank; Loffing, Johannes

    2016-11-17

    The mouse isolated perfused kidney (MIPK) is a technique for keeping a mouse kidney under ex vivo conditions perfused and functional for 1 hr. This is a prerequisite for studying the physiology of the isolated organ and for many innovative applications that may be possible in the future, including perfusion decellularization for kidney bioengineering or the administration of anti-rejection or genome-editing drugs in high doses to prime the kidney for transplantation. During the time of the perfusion, the kidney can be manipulated, renal function can be assessed, and various pharmaceuticals administered. After the procedure, the kidney can be transplanted or processed for molecular biology, biochemical analysis, or microscopy. This paper describes the perfusate and the surgical technique needed for the ex vivo perfusion of mouse kidneys. Details of the perfusion apparatus are given and data are presented showing the viability of the kidney's preparation: renal blood flow, vascular resistance, and urine data as functional, transmission electron micrographs of different nephron segments as morphological readouts, and western blots of transport proteins of different nephron segments as molecular readout.

  19. Cloning the mouse homologue of the human lysosomal acid {alpha}-glucosidase gene

    SciTech Connect

    Ding, J.H.; Yang, B.Z.; Liu, H.M.

    1994-09-01

    Pompe disease (GSD II) is an autosomal recessive disorder caused by a deficiency of lysosomal acid {alpha}-glucosidase (GAA). In an attempt to create a mouse model for Pompe disease, we isolated and characterized the gene encoding the mouse homologue of the human GAA. Twenty clones that extend from exon 2 to the poly(A) tail were isolated from a mouse liver cDNA library, but the remainder of the mRNA proved difficult to obtain by conventional cDNA library screening. Sequences spanning exons 1-2 were cloned by RACE from mouse liver RNA. The full-length liver GAA cDNA contains 3365 nucleotides with a coding region of 2859 nucleotides and a 394 base pair 3{prime}-nontranslated region. The deduced amino acid sequence of the mouse GAA shows 84% identity to the human GAA. Southern blot analysis demonstrated that the mouse GAA was encoded by a single copy gene. Then six bacteriophages containing DNA from the GAA gene were isolated by screening 10{sup 6} phage plaques of a mouse 129 genomic library using a mouse GAA cDNA as a probe. From one of these bacteriophages, an 11-kilobase EcoRI fragment containing exons 3 to 15 was subcloned and sequenced. Work is in progress using this genomic clone to disrupt the GAA gene in murine embryonic stem cells in order to create GSD II mice.

  20. Mouse bladder wall injection.

    PubMed

    Fu, Chi-Ling; Apelo, Charity A; Torres, Baldemar; Thai, Kim H; Hsieh, Michael H

    2011-07-12

    Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. Before starting injections, the surgical area must be cleaned with soap and water and antiseptic solution. Surgical equipment must be sterilized before use and between each animal. Each mouse is placed under inhaled isoflurane anesthesia (2-5% for induction, 1-3% for maintenance) and its bladder exposed by making a midline abdominal incision with scissors. If the bladder is full, it is partially decompressed by gentle squeezing between two fingers. The cell suspension of interest is intramurally injected into the wall of the bladder dome using a 29 or 30 gauge needle and 1 cc or smaller syringe. The wound is then closed using wound clips and the mouse allowed to recover on a warming pad. Bladder wall injection is a delicate microsurgical technique that can be mastered with practice.

  1. Genome-wide DNA polymorphism analyses using VariScan

    PubMed Central

    Hutter, Stephan; Vilella, Albert J; Rozas, Julio

    2006-01-01

    Background DNA sequence polymorphisms analysis can provide valuable information on the evolutionary forces shaping nucleotide variation, and provides an insight into the functional significance of genomic regions. The recent ongoing genome projects will radically improve our capabilities to detect specific genomic regions shaped by natural selection. Current available methods and software, however, are unsatisfactory for such genome-wide analysis. Results We have developed methods for the analysis of DNA sequence polymorphisms at the genome-wide scale. These methods, which have been tested on a coalescent-simulated and actual data files from mouse and human, have been implemented in the VariScan software package version 2.0. Additionally, we have also incorporated a graphical-user interface. The main features of this software are: i) exhaustive population-genetic analyses including those based on the coalescent theory; ii) analysis adapted to the shallow data generated by the high-throughput genome projects; iii) use of genome annotations to conduct a comprehensive analyses separately for different functional regions; iv) identification of relevant genomic regions by the sliding-window and wavelet-multiresolution approaches; v) visualization of the results integrated with current genome annotations in commonly available genome browsers. Conclusion VariScan is a powerful and flexible suite of software for the analysis of DNA polymorphisms. The current version implements new algorithms, methods, and capabilities, providing an important tool for an exhaustive exploratory analysis of genome-wide DNA polymorphism data. PMID:16968531

  2. Researchers Create Artificial Mouse 'Embryo'

    MedlinePlus

    ... news/fullstory_163881.html Researchers Create Artificial Mouse 'Embryo' Experiment used two types of gene-modified stem ... they've created a kind of artificial mouse embryo using stem cells, which can be coaxed to ...

  3. Step-specific Sorting of Mouse Spermatids by Flow Cytometry.

    PubMed

    Simard, Olivier; Leduc, Frédéric; Acteau, Geneviève; Arguin, Mélina; Grégoire, Marie-Chantal; Brazeau, Marc-André; Marois, Isabelle; Richter, Martin V; Boissonneault, Guylain

    2015-12-31

    The differentiation of mouse spermatids is one critical process for the production of a functional male gamete with an intact genome to be transmitted to the next generation. So far, molecular studies of this morphological transition have been hampered by the lack of a method allowing adequate separation of these important steps of spermatid differentiation for subsequent analyses. Earlier attempts at proper gating of these cells using flow cytometry may have been difficult because of a peculiar increase in DNA fluorescence in spermatids undergoing chromatin remodeling. Based on this observation, we provide details of a simple flow cytometry scheme, allowing reproducible purification of four populations of mouse spermatids fixed with ethanol, each representing a different state in the nuclear remodeling process. Population enrichment is confirmed using step-specific markers and morphological criterions. The purified spermatids can be used for genomic and proteomic analyses.

  4. Genomic arrays in chronic lymphocytic leukemia routine clinical practice: are we ready to substitute conventional cytogenetics and fluorescence in situ hybridization techniques?

    PubMed

    Puiggros, Anna; Puigdecanet, Eulàlia; Salido, Marta; Ferrer, Ana; Abella, Eugènia; Gimeno, Eva; Nonell, Lara; Herranz, María José; Galván, Ana Belén; Rodríguez-Rivera, María; Melero, Carme; Pairet, Silvia; Bellosillo, Beatriz; Serrano, Sergi; Florensa, Lourdes; Solé, Francesc; Espinet, Blanca

    2013-05-01

    Chronic lymphocytic leukemia (CLL) is characterized by a highly variable clinical course. Del(11q) and del(17p), routinely studied by conventional G-banding cytogenetics (CGC) and fluorescence in situ hybridization (FISH), have been related to progression and shorter overall survival. Recently, array-based karyotyping has gained acceptance as a high-resolution new tool for detecting genomic imbalances. The aim of the present study was to compare genomic arrays with CGC and FISH to ascertain whether the current techniques could be substituted in routine procedures. We analyzed 70 patients with CLL using the Cytogenetics Whole-Genome 2.7M Array and CytoScan HD Array (Affymetrix), CGC and FISH with the classical CLL panel. Whereas 31.4% and 68.6% of patients presented abnormalities when studied by CGC and FISH, respectively, these rates increased when arrays were also analyzed (78.6% and 80%). Although abnormality detection is higher when arrays are applied, one case with del(11q) and three with del(17p) were missed by genomic arrays due to their limited sensitivity. We consider that the complete substitution of CGC and FISH by genomic arrays in routine laboratories could negatively affect the management of some patients harboring 11q or 17p deletions. In conclusion, genomic arrays are valid to detect known and novel genomic imbalances in CLL, but should be maintained as a complementary tool to the current techniques.

  5. Complexity, polymorphism, and connectivity of mouse Vk gene families.

    PubMed

    Kofler, R; Duchosal, M A; Dixon, F J

    1989-01-01

    To define the polymorphism and extent of the mouse immunoglobulin kappa (Igk) gene complex, we have analyzed restriction-enzyme digested genomic DNA from 33 inbred strains of mice with labeled DNA probes corresponding to 16 Vk protein groups (1 of them previously undescribed) and the Jk/Ck region (V, variable; J, joining; C, constant). These probes detected between 1 and 25 distinct restriction enzyme fragments (REF) that appeared in up to eight polymorphic patterns, thus defining eight mouse Igk haplotypes. The investigated portion of the Vk repertoire was estimated to encompass between 60 and 120 discernable Vk gene-containing REFs. In contrast to mouse VH gene families, several Vk gene families defined by these probes appeared to overlap. This observation has implications for Vk gene analyses by nucleic acid hybridization and raises the possibility that the Vk gene complex is a continuum of related sequences.

  6. Mouse Models of Thyroid Cancer: A 2015 Update

    PubMed Central

    Kirschner, Lawrence S.; Qamri, Zahida; Kari, Suresh; Ashtekar, Amruta

    2015-01-01

    Thyroid cancer is the most common endocrine neoplasm, and its rate is rising at an alarming pace. Thus, there is a compelling need to develop in vivo models which will not only enable the confirmation of the oncogenic potential of driver genes, but also point the way towards the development of new therapeutics. Over the past 20 years, techniques for the generation of mouse models of human diseases have progressed substantially, accompanied by parallel advances in the genetics and genomics of human tumors. This convergence has enabled the development of mouse lines carrying mutations in the genes that cause thyroid cancers of all subtypes, including differentiated papillary and follicular thyroid cancers, poorly differentiated/anaplastic cancers, and medullary thyroid cancers. In this review, we will discuss the state of the art of mouse modeling of thyroid cancer, with the eventual goal of providing insight into tumor biology and treatment. PMID:26123589

  7. Genomic dark matter: the reliability of short read mapping illustrated by the genome mappability score

    PubMed Central

    Lee, Hayan; Schatz, Michael C.

    2012-01-01

    Motivation: Genome resequencing and short read mapping are two of the primary tools of genomics and are used for many important applications. The current state-of-the-art in mapping uses the quality values and mapping quality scores to evaluate the reliability of the mapping. These attributes, however, are assigned to individual reads and do not directly measure the problematic repeats across the genome. Here, we present the Genome Mappability Score (GMS) as a novel measure of the complexity of resequencing a genome. The GMS is a weighted probability that any read could be unambiguously mapped to a given position and thus measures the overall composition of the genome itself. Results: We have developed the Genome Mappability Analyzer to compute the GMS of every position in a genome. It leverages the parallelism of cloud computing to analyze large genomes, and enabled us to identify the 5–14% of the human, mouse, fly and yeast genomes that are difficult to analyze with short reads. We examined the accuracy of the widely used BWA/SAMtools polymorphism discovery pipeline in the context of the GMS, and found discovery errors are dominated by false negatives, especially in regions with poor GMS. These errors are fundamental to the mapping process and cannot be overcome by increasing coverage. As such, the GMS should be considered in every resequencing project to pinpoint the ‘dark matter’ of the genome, including of known clinically relevant variations in these regions. Availability: The source code and profiles of several model organisms are available at http://gma-bio.sourceforge.net Contact: hlee@cshl.edu Supplementary Information: Supplementary data are available at Bioinformatics online. PMID:22668792

  8. Genome engineering in cattle: recent technological advancements.

    PubMed

    Wang, Zhongde

    2015-02-01

    Great strides in technological advancements have been made in the past decade in cattle genome engineering. First, the success of cloning cattle by somatic cell nuclear transfer (SCNT) or chromatin transfer (CT) is a significant advancement that has made obsolete the need for using embryonic stem (ES) cells to conduct cell-mediated genome engineering, whereby site-specific genetic modifications can be conducted in bovine somatic cells via DNA homologous recombination (HR) and whereby genetically engineered cattle can subsequently be produced by animal cloning from the genetically modified cells. With this approach, a chosen bovine genomic locus can be precisely modified in somatic cells, such as to knock out (KO) or knock in (KI) a gene via HR, a gene-targeting strategy that had almost exclusively been used in mouse ES cells. Furthermore, by the creative application of embryonic cloning to rejuvenate somatic cells, cattle genome can be sequentially modified in the same line of somatic cells and complex genetic modifications have been achieved in cattle. Very recently, the development of designer nucleases-such as zinc finger nucleases (ZFNs) and transcription activator-like effector nuclease (TALENs), and clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9)-has enabled highly efficient and more facile genome engineering in cattle. Most notably, by employing such designer nucleases, genomes can be engineered at single-nucleotide precision; this process is now often referred to as genome or gene editing. The above achievements are a drastic departure from the traditional methods of creating genetically modified cattle, where foreign DNAs are randomly integrated into the animal genome, most often along with the integrations of bacterial or viral DNAs. Here, I review the most recent technological developments in cattle genome engineering by highlighting some of the major achievements in creating genetically engineered

  9. The 3D folding of metazoan genomes correlates with the association of similar repetitive elements

    PubMed Central

    Cournac, Axel; Koszul, Romain; Mozziconacci, Julien

    2016-01-01

    The potential roles of the numerous repetitive elements found in the genomes of multi-cellular organisms remain speculative. Several studies have suggested a role in stabilizing specific 3D genomic contacts. To test this hypothesis, we exploited inter-chromosomal contacts frequencies obtained from Hi-C experiments and show that the folding of the human, mouse and Drosophila genomes is associated with a significant co-localization of several specific repetitive elements, notably many elements of the SINE family. These repeats tend to be the oldest ones and are enriched in transcription factor binding sites. We propose that the co-localization of these repetitive elements may explain the global conservation of genome folding observed between homologous regions of the human and mouse genome. Taken together, these results support a contribution of specific repetitive elements in maintaining and/or reshaping genome architecture over evolutionary times. PMID:26609133

  10. The platypus genome unraveled.

    PubMed

    O'Brien, Stephen J

    2008-06-13

    The genome of the platypus has been sequenced, assembled, and annotated by an international genomics team. Like the animal itself the platypus genome contains an amalgam of mammal, reptile, and bird-like features.

  11. Ultraconserved elements in the human genome.

    PubMed

    Bejerano, Gill; Pheasant, Michael; Makunin, Igor; Stephen, Stuart; Kent, W James; Mattick, John S; Haussler, David

    2004-05-28

    There are 481 segments longer than 200 base pairs (bp) that are absolutely conserved (100% identity with no insertions or deletions) between orthologous regions of the human, rat, and mouse genomes. Nearly all of these segments are also conserved in the chicken and dog genomes, with an average of 95 and 99% identity, respectively. Many are also significantly conserved in fish. These ultraconserved elements of the human genome are most often located either overlapping exons in genes involved in RNA processing or in introns or nearby genes involved in the regulation of transcription and development. Along with more than 5000 sequences of over 100 bp that are absolutely conserved among the three sequenced mammals, these represent a class of genetic elements whose functions and evolutionary origins are yet to be determined, but which are more highly conserved between these species than are proteins and appear to be essential for the ontogeny of mammals and other vertebrates.

  12. Mouse ENU Mutagenesis to Understand Immunity to Infection: Methods, Selected Examples, and Perspectives

    PubMed Central

    Caignard, Grégory; Eva, Megan M.; van Bruggen, Rebekah; Eveleigh, Robert; Bourque, Guillaume; Malo, Danielle; Gros, Philippe; Vidal, Silvia M.

    2014-01-01

    Infectious diseases are responsible for over 25% of deaths globally, but many more individuals are exposed to deadly pathogens. The outcome of infection results from a set of diverse factors including pathogen virulence factors, the environment, and the genetic make-up of the host. The completion of the human reference genome sequence in 2004 along with technological advances have tremendously accelerated and renovated the tools to study the genetic etiology of infectious diseases in humans and its best characterized mammalian model, the mouse. Advancements in mouse genomic resources have accelerated genome-wide functional approaches, such as gene-driven and phenotype-driven mutagenesis, bringing to the fore the use of mouse models that reproduce accurately many aspects of the pathogenesis of human infectious diseases. Treatment with the mutagen N-ethyl-N-nitrosourea (ENU) has become the most popular phenotype-driven approach. Our team and others have employed mouse ENU mutagenesis to identify host genes that directly impact susceptibility to pathogens of global significance. In this review, we first describe the strategies and tools used in mouse genetics to understand immunity to infection with special emphasis on chemical mutagenesis of the mouse germ-line together with current strategies to efficiently identify functional mutations using next generation sequencing. Then, we highlight illustrative examples of genes, proteins, and cellular signatures that have been revealed by ENU screens and have been shown to be involved in susceptibility or resistance to infectious diseases caused by parasites, bacteria, and viruses. PMID:25268389

  13. WGE: a CRISPR database for genome engineering

    PubMed Central

    Hodgkins, Alex; Farne, Anna; Perera, Sajith; Grego, Tiago; Parry-Smith, David J.; Skarnes, William C.; Iyer, Vivek

    2015-01-01

    Summary: The rapid development of CRISPR-Cas9 mediated genome editing techniques has given rise to a number of online and stand-alone tools to find and score CRISPR sites for whole genomes. Here we describe the Wellcome Trust Sanger Institute Genome Editing database (WGE), which uses novel methods to compute, visualize and select optimal CRISPR sites in a genome browser environment. The WGE database currently stores single and paired CRISPR sites and pre-calculated off-target information for CRISPRs located in the mouse and human exomes. Scoring and display of off-target sites is simple, and intuitive, and filters can be applied to identify high-quality CRISPR sites rapidly. WGE also provides a tool for the design and display of gene targeting vectors in the same genome browser, along with gene models, protein translation and variation tracks. WGE is open, extensible and can be set up to compute and present CRISPR sites for any genome. Availability and implementation: The WGE database is freely available at www.sanger.ac.uk/htgt/wge Contact: vvi@sanger.ac.uk or skarnes@sanger.ac.uk Supplementary information: Supplementary data are available at Bioinformatics online. PMID:25979474

  14. Genomic imprinting--insights from studies in mice.

    PubMed

    Ferguson-Smith, Anne; Lin, Shau-Ping; Tsai, Chen-En; Youngson, Neil; Tevendale, Maxine

    2003-02-01

    A subset of mammalian genes is controlled by genomic imprinting. This process causes a gene to be expressed from only one chromosome homologue depending on whether it originally came from the egg or the sperm. Parental origin-specific gene regulation is controlled by epigenetic modifications to DNA and chromatin. Genomic imprinting is therefore a useful model system to study the epigenetic control of genome function. Here we consider the value of the mouse as an experimental organism to address questions about the role of imprinted genes, about the regulation of mono-allelic gene expression and about the evolution of the imprinting function and mechanism.

  15. Metabolism, Genomics, and DNA Repair in the Mouse Aging Liver

    PubMed Central

    Lebel, Michel; de Souza-Pinto, Nadja C.; Bohr, Vilhelm A.

    2011-01-01

    The liver plays a pivotal role in the metabolism of nutrients, drugs, hormones, and metabolic waste products, thereby maintaining body homeostasis. The liver undergoes substantial changes in structure and function within old age. Such changes are associated with significant impairment of many hepatic metabolic and detoxification activities, with implications for systemic aging and age-related disease. It has become clear, using rodent models as biological tools, that genetic instability in the form of gross DNA rearrangements or point mutations accumulate in the liver with age. DNA lesions, such as oxidized bases or persistent breaks, increase with age and correlate well with the presence of senescent hepatocytes. The level of DNA damage and/or mutation can be affected by changes in carcinogen activation, decreased ability to repair DNA, or a combination of these factors. This paper covers some of the DNA repair pathways affecting liver homeostasis with age using rodents as model systems. PMID:21559242

  16. Genome evolution: the dynamics of static genomes.

    PubMed

    Stechmann, Alexandra

    2004-06-22

    A random survey of a microsporidian genome has revealed some striking features. Although the genomes of microsporidians are among the smallest known for eukaryotes, their organisation appears to be well conserved.

  17. Plant Genome Duplication Database.

    PubMed

    Lee, Tae-Ho; Kim, Junah; Robertson, Jon S; Paterson, Andrew H

    2017-01-01

    Genome duplication, widespread in flowering plants, is a driving force in evolution. Genome alignments between/within genomes facilitate identification of homologous regions and individual genes to investigate evolutionary consequences of genome duplication. PGDD (the Plant Genome Duplication Database), a public web service database, provides intra- or interplant genome alignment information. At present, PGDD contains information for 47 plants whose genome sequences have been released. Here, we describe methods for identification and estimation of dates of genome duplication and speciation by functions of PGDD.The database is freely available at http://chibba.agtec.uga.edu/duplication/.

  18. Rice-arsenate interactions in hydroponics: whole genome transcriptional analysis.

    PubMed

    Norton, Gareth J; Lou-Hing, Daniel E; Meharg, Andrew A; Price, Adam H

    2008-01-01

    Rice (Oryza sativa) varieties that are arsenate-tolerant (Bala) and -sensitive (Azucena) were used to conduct a transcriptome analysis of the response of rice seedlings to sodium arsenate (AsV) in hydroponic solution. RNA extracted from the roots of three replicate experiments of plants grown for 1 week in phosphate-free nutrient with or without 13.3 muM AsV was used to challenge the Affymetrix (52K) GeneChip Rice Genome array. A total of 576 probe sets were significantly up-regulated at least 2-fold in both varieties, whereas 622 were down-regulated. Ontological classification is presented. As expected, a large number of transcription factors, stress proteins, and transporters demonstrated differential expression. Striking is the lack of response of classic oxidative stress-responsive genes or phytochelatin synthases/synthatases. However, the large number of responses from genes involved in glutathione synthesis, metabolism, and transport suggests that glutathione conjugation and arsenate methylation may be important biochemical responses to arsenate challenge. In this report, no attempt is made to dissect differences in the response of the tolerant and sensitive variety, but analysis in a companion article will link gene expression to the known tolerance loci available in the BalaxAzucena mapping population.

  19. Genomic analysis of gum disease and hypertrichosis in foxes.

    PubMed

    Clark, J-A B J; Whalen, D; Marshall, H D

    2016-05-20

    Since the 1940s, a proliferative gingival disease called hereditary hyperplastic gingivitis (HHG) has been described in the farmed silver fox, Vulpes vulpes (Dyrendahl and Henricson 1960). HHG displays an autosomal recessive transmission and has a pleiotropic relationship with superior fur quality in terms of length and thickness of guard hairs. An analogous human disease, hereditary gingival fibromatosis (HGF), is characterized by a predominantly autosomal dominant transmission and a complex etiology, occurring either as an isolated condition or as a part of a syndrome. Similar to HHG, the symptom most commonly associated with syndromic HGF is hypertrichosis. Here we explore potential mechanisms involved in HHG by comparison to known genetic information about hypertrichosis co-occurring with HGF, using an Affymetrix canine genome microarray platform, quantitative PCR, and candidate gene sequencing. We conclude that the mitogen-activated protein kinase pathway is involved in HHG, however despite involvement of the mitogen-activated protein kinase kinase 6 gene in congenital hypertrichosis with gingival fibromatosis in humans, this gene did not contain any fixed mutations in exons or exon-intron boundaries in HHG-affected foxes, suggesting that it is not causative of HHG in the farmed silver fox population. Differential up-regulation of MAP2K6 gene in HHG-affected foxes does implicate this gene in the HHG phenotype.

  20. Brain Perihematoma Genomic Profile Following Spontaneous Human Intracerebral Hemorrhage

    PubMed Central

    Rosell, Anna; Vilalta, Anna; García-Berrocoso, Teresa; Fernández-Cadenas, Israel; Domingues-Montanari, Sophie; Cuadrado, Eloy; Delgado, Pilar; Ribó, Marc; Martínez-Sáez, Elena; Ortega-Aznar, Arantxa; Montaner, Joan

    2011-01-01

    Background Spontaneous intracerebral hemorrhage (ICH) represents about 15% of all strokes and is associated with high mortality rates. Our aim was to identify the gene expression changes and biological pathways altered in the brain following ICH. Methodology/Principal Findings Twelve brain samples were obtained from four deceased patients who suffered an ICH including perihematomal tissue (PH) and the corresponding contralateral white (CW) and grey (CG) matter. Affymetrix GeneChip platform for analysis of over 47,000 transcripts was conducted. Microarray Analysis Suite 5.0 was used to process array images and the Ingenuity Pathway Analysis System was used to analyze biological mechanisms and functions of the genes. We identified 468 genes in the PH areas displaying a different expression pattern with a fold change between −3.74 and +5.16 when compared to the contralateral areas (291 overexpressed and 177 underexpressed). The top genes which appeared most significantly overexpressed in the PH areas codify for cytokines, chemokines, coagulation factors, cell growth and proliferation factors while the underexpressed codify for proteins involved in cell cycle or neurotrophins. Validation and replication studies at gene and protein level in brain samples confirmed microarray results. Conclusions The genomic responses identified in this study provide valuable information about potential biomarkers and target molecules altered in the perihematomal regions. PMID:21311749

  1. Genome Scans for Transmission Ratio Distortion Regions in Mice

    PubMed Central

    Casellas, Joaquim; Gularte, Rodrigo J.; Farber, Charles R.; Varona, Luis; Mehrabian, Margarete; Schadt, Eric E.; Lusis, Aldon J.; Attie, Alan D.; Yandell, Brian S.; Medrano, Juan F.

    2012-01-01

    Transmission ratio distortion (TRD) is the departure from the expected genotypic frequencies under Mendelian inheritance. This departure can be due to multiple physiological mechanisms during gametogenesis, fertilization, fetal and embryonic development, and early neonatal life. Although a few TRD loci have been reported in mouse, inheritance patterns have never been evaluated for TRD. In this article, we developed a Bayesian binomial model accounting for additive and dominant deviation TRD mechanisms. Moreover, this model was used to perform genome-wide scans for TRD quantitative trait loci (QTL) on six F2 mouse crosses involving between 296 and 541 mice and between 72 and 1854 genetic markers. Statistical significance of each model was checked at each genetic marker with Bayes factors. Genome scans revealed overdominance TRD QTL located in mouse chromosomes 1, 2, 12, 13, and 14 and additive TRD QTL in mouse chromosomes 2, 3, and 15, although these results did not replicate across mouse crosses. This research contributes new statistical tools for the analysis of specific genetic patterns involved in TRD in F2 populations, our results suggesting a relevant incidence of TRD phenomena in mouse with important implications for both statistical analyses and biological research. PMID:22367040

  2. Generation of mouse ES cell lines engineered for the forced induction of transcription factors

    PubMed Central

    Correa-Cerro, Lina S.; Piao, Yulan; Sharov, Alexei A.; Nishiyama, Akira; Cadet, Jean S.; Yu, Hong; Sharova, Lioudmila V.; Xin, Li; Hoang, Hien G.; Thomas, Marshall; Qian, Yong; Dudekula, Dawood B.; Meyers, Emily; Binder, Bernard Y.; Mowrer, Gregory; Bassey, Uwem; Longo, Dan L.; Schlessinger, David; Ko, Minoru S. H.

    2011-01-01

    Here we report the generation and characterization of 84 mouse ES cell lines with doxycycline-controllable transcription factors (TFs) which, together with the previous 53 lines, cover 7–10% of all TFs encoded in the mouse genome. Global gene expression profiles of all 137 lines after the induction of TFs for 48 hrs can associate each TF with the direction of ES cell differentiation, regulatory pathways, and mouse phenotypes. These cell lines and microarray data provide building blocks for a variety of future biomedical research applications as a community resource. PMID:22355682

  3. Ensembl genomes 2016: more genomes, more complexity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ensembl Genomes (http://www.ensemblgenomes.org) is an integrating resource for genome-scale data from non-vertebrate species, complementing the resources for vertebrate genomics developed in the context of the Ensembl project (http://www.ensembl.org). Together, the two resources provide a consistent...

  4. Ensembl Genomes 2016: more genomes, more complexity

    PubMed Central

    Kersey, Paul Julian; Allen, James E.; Armean, Irina; Boddu, Sanjay; Bolt, Bruce J.; Carvalho-Silva, Denise; Christensen, Mikkel; Davis, Paul; Falin, Lee J.; Grabmueller, Christoph; Humphrey, Jay; Kerhornou, Arnaud; Khobova, Julia; Aranganathan, Naveen K.; Langridge, Nicholas; Lowy, Ernesto; McDowall, Mark D.; Maheswari, Uma; Nuhn, Michael; Ong, Chuang Kee; Overduin, Bert; Paulini, Michael; Pedro, Helder; Perry, Emily; Spudich, Giulietta; Tapanari, Electra; Walts, Brandon; Williams, Gareth; Tello–Ruiz, Marcela; Stein, Joshua; Wei, Sharon; Ware, Doreen; Bolser, Daniel M.; Howe, Kevin L.; Kulesha, Eugene; Lawson, Daniel; Maslen, Gareth; Staines, Daniel M.

    2016-01-01

    Ensembl Genomes (http://www.ensemblgenomes.org) is an integrating resource for genome-scale data from non-vertebrate species, complementing the resources for vertebrate genomics developed in the context of the Ensembl project (http://www.ensembl.org). Together, the two resources provide a consistent set of programmatic and interactive interfaces to a rich range of data including reference sequence, gene models, transcriptional data, genetic variation and comparative analysis. This paper provides an update to the previous publications about the resource, with a focus on recent developments. These include the development of new analyses and views to represent polyploid genomes (of which bread wheat is the primary exemplar); and the continued up-scaling of the resource, which now includes over 23 000 bacterial genomes, 400 fungal genomes and 100 protist genomes, in addition to 55 genomes from invertebrate metazoa and 39 genomes from plants. This dramatic increase in the number of included genomes is one part of a broader effort to automate the integration of archival data (genome sequence, but also associated RNA sequence data and variant calls) within the context of reference genomes and make it available through the Ensembl user interfaces. PMID:26578574

  5. Ensembl Genomes 2016: more genomes, more complexity.

    PubMed

    Kersey, Paul Julian; Allen, James E; Armean, Irina; Boddu, Sanjay; Bolt, Bruce J; Carvalho-Silva, Denise; Christensen, Mikkel; Davis, Paul; Falin, Lee J; Grabmueller, Christoph; Humphrey, Jay; Kerhornou, Arnaud; Khobova, Julia; Aranganathan, Naveen K; Langridge, Nicholas; Lowy, Ernesto; McDowall, Mark D; Maheswari, Uma; Nuhn, Michael; Ong, Chuang Kee; Overduin, Bert; Paulini, Michael; Pedro, Helder; Perry, Emily; Spudich, Giulietta; Tapanari, Electra; Walts, Brandon; Williams, Gareth; Tello-Ruiz, Marcela; Stein, Joshua; Wei, Sharon; Ware, Doreen; Bolser, Daniel M; Howe, Kevin L; Kulesha, Eugene; Lawson, Daniel; Maslen, Gareth; Staines, Daniel M

    2016-01-04

    Ensembl Genomes (http://www.ensemblgenomes.org) is an integrating resource for genome-scale data from non-vertebrate species, complementing the resources for vertebrate genomics developed in the context of the Ensembl project (http://www.ensembl.org). Together, the two resources provide a consistent set of programmatic and interactive interfaces to a rich range of data including reference sequence, gene models, transcriptional data, genetic variation and comparative analysis. This paper provides an update to the previous publications about the resource, with a focus on recent developments. These include the development of new analyses and views to represent polyploid genomes (of which bread wheat is the primary exemplar); and the continued up-scaling of the resource, which now includes over 23 000 bacterial genomes, 400 fungal genomes and 100 protist genomes, in addition to 55 genomes from invertebrate metazoa and 39 genomes from plants. This dramatic increase in the number of included genomes is one part of a broader effort to automate the integration of archival data (genome sequence, but also associated RNA sequence data and variant calls) within the context of reference genomes and make it available through the Ensembl user interfaces.

  6. Microarray Data Processing Techniques for Genome-Scale Network Inference from Large Public Repositories.

    PubMed

    Chockalingam, Sriram; Aluru, Maneesha; Aluru, Srinivas

    2016-09-19

    Pre-processing of microarray data is a well-studied problem. Furthermore, all popular platforms come with their own recommended best practices for differential analysis of genes. However, for genome-scale network inference using microarray data collected from large public repositories, these methods filter out a considerable number of genes. This is primarily due to the effects of aggregating a diverse array of experiments with different technical and biological scenarios. Here we introduce a pre-processing pipeline suitable for inferring genome-scale gene networks from large microarray datasets. We show that partitioning of the available microarray datasets according to biological relevance into tissue- and process-specific categories significantly extends the limits of downstream network construction. We demonstrate the effectiveness of our pre-processing pipeline by inferring genome-scale networks for the model plant Arabidopsis thaliana using two different construction methods and a collection of 11,760 Affymetrix ATH1 microarray chips. Our pre-processing pipeline and the datasets used in this paper are made available at http://alurulab.cc.gatech.edu/microarray-pp.

  7. Soybean Knowledge Base (SoyKB): a Web Resource for Soybean Translational Genomics

    SciTech Connect

    Joshi, Trupti; Patil, Kapil; Fitzpatrick, Michael R.; Franklin, Levi D.; Yao, Qiuming; Cook, Jeffrey R.; Wang, Zhem; Libault, Marc; Brechenmacher, Laurent; Valliyodan, Babu; Wu, Xiaolei; Cheng, Jianlin; Stacey, Gary; Nguyen, Henry T.; Xu, Dong

    2012-01-17

    Background: Soybean Knowledge Base (SoyKB) is a comprehensive all-inclusive web resource for soybean translational genomics. SoyKB is designed to handle the management and integration of soybean genomics, transcriptomics, proteomics and metabolomics data along with annotation of gene function and biological pathway. It contains information on four entities, namely genes, microRNAs, metabolites and single nucleotide polymorphisms (SNPs). Methods: SoyKB has many useful tools such as Affymetrix probe ID search, gene family search, multiple gene/ metabolite search supporting co-expression analysis, and protein 3D structure viewer as well as download and upload capacity for experimental data and annotations. It has four tiers of registration, which control different levels of access to public and private data. It allows users of certain levels to share their expertise by adding comments to the data. It has a user-friendly web interface together with genome browser and pathway viewer, which display data in an intuitive manner to the soybean researchers, producers and consumers. Conclusions: SoyKB addresses the increasing need of the soybean research community to have a one-stop-shop functional and translational omics web resource for information retrieval and analysis in a user-friendly way. SoyKB can be publicly accessed at http://soykb.org/.

  8. Rapid Identification of Potential Drugs for Diabetic Nephropathy Using Whole-Genome Expression Profiles of Glomeruli

    PubMed Central

    Shi, Jingsong; Jiang, Song; Qiu, Dandan; Le, Weibo; Wang, Xiao; Lu, Yinhui; Liu, Zhihong

    2016-01-01

    Objective. To investigate potential drugs for diabetic nephropathy (DN) using whole-genome expression profiles and the Connectivity Map (CMAP). Methodology. Eighteen Chinese Han DN patients and six normal controls were included in this study. Whole-genome expression profiles of microdissected glomeruli were measured using the Affymetrix human U133 plus 2.0 chip. Differentially expressed genes (DEGs) between late stage and early stage DN samples and the CMAP database were used to identify potential drugs for DN using bioinformatics methods. Results. (1) A total of 1065 DEGs (FDR < 0.05 and fold change > 1.5) were found in late stage DN patients compared with early stage DN patients. (2) Piperlongumine, 15d-PGJ2 (15-delta prostaglandin J2), vorinostat, and trichostatin A were predicted to be the most promising potential drugs for DN, acting as NF-κB inhibitors, histone deacetylase inhibitors (HDACIs), PI3K pathway inhibitors, or PPARγ agonists, respectively. Conclusion. Using whole-genome expression profiles and the CMAP database, we rapidly predicted potential DN drugs, and therapeutic potential was confirmed by previously published studies. Animal experiments and clinical trials are needed to confirm both the safety and efficacy of these drugs in the treatment of DN. PMID:27069916

  9. The Mouse That Soared

    NASA Astrophysics Data System (ADS)

    2004-09-01

    Astronomers have used an X-ray image to make the first detailed study of the behavior of high-energy particles around a fast moving pulsar. The image, from NASA's Chandra X-ray Observatory, shows the shock wave created as a pulsar plows supersonically through interstellar space. These results will provide insight into theories for the production of powerful winds of matter and antimatter by pulsars. Chandra's image of the glowing cloud, known as the Mouse, shows a stubby bright column of high-energy particles, about four light years in length, swept back by the pulsar's interaction with interstellar gas. The intense source at the head of the X-ray column is the pulsar, estimated to be moving through space at about 1.3 million miles per hour. VLA Radio Image of the Mouse, Full Field VLA Radio Image of the Mouse, Full Field A cone-shaped cloud of radio-wave-emitting particles envelopes the X-ray column. The Mouse, a.k.a. G359.23-0.82, was discovered in 1987 by radio astronomers using the National Science Foundation's Very Large Array in New Mexico. It gets its name from its appearance in radio images that show a compact snout, a bulbous body, and a remarkable long, narrow, tail that extends for about 55 light years. "A few dozen pulsar wind nebulae are known, including the spectacular Crab Nebula, but none have the Mouse's combination of relatively young age and incredibly rapid motion through interstellar space," said Bryan Gaensler of the Harvard-Smithsonian Center for Astrophysics and lead author of a paper on the Mouse that will appear in an upcoming issue of The Astrophysical Journal. "We effectively are seeing a supersonic cosmic wind tunnel, in which we can study the effects of a pulsar's motion on its pulsar wind nebula, and test current theories." Illustration of the Mouse System Illustration of the Mouse System Pulsars are known to be rapidly spinning, highly magnetized neutron stars -- objects so dense that a mass equal to that of the Sun is packed into a

  10. Genome-wide patterns of population structure and admixture in West Africans and African Americans.

    PubMed

    Bryc, Katarzyna; Auton, Adam; Nelson, Matthew R; Oksenberg, Jorge R; Hauser, Stephen L; Williams, Scott; Froment, Alain; Bodo, Jean-Marie; Wambebe, Charles; Tishkoff, Sarah A; Bustamante, Carlos D

    2010-01-12

    Quantifying patterns of population structure in Africans and African Americans illuminates the history of human populations and is critical for undertaking medical genomic studies on a global scale. To obtain a fine-scale genome-wide perspective of ancestry, we analyze Affymetrix GeneChip 500K genotype data from African Americans (n = 365) and individuals with ancestry from West Africa (n = 203 from 12 populations) and Europe (n = 400 from 42 countries). We find that population structure within the West African sample reflects primarily language and secondarily geographical distance, echoing the Bantu expansion. Among African Americans, analysis of genomic admixture by a principal component-based approach indicates that the median proportion of European ancestry is 18.5% (25th-75th percentiles: 11.6-27.7%), with very large variation among individuals. In the African-American sample as a whole, few autosomal regions showed exceptionally high or low mean African ancestry, but the X chromosome showed elevated levels of African ancestry, consistent with a sex-biased pattern of gene flow with an excess of European male and African female ancestry. We also find that genomic profiles of individual African Americans afford personalized ancestry reconstructions differentiating ancient vs. recent European and African ancestry. Finally, patterns of genetic similarity among inferred African segments of African-American genomes and genomes of contemporary African populations included in this study suggest African ancestry is most similar to non-Bantu Niger-Kordofanian-speaking populations, consistent with historical documents of the African Diaspora and trans-Atlantic slave trade.

  11. High-resolution genomic profiling of chronic lymphocytic leukemia reveals new recurrent genomic alterations.

    PubMed

    Edelmann, Jennifer; Holzmann, Karlheinz; Miller, Florian; Winkler, Dirk; Bühler, Andreas; Zenz, Thorsten; Bullinger, Lars; Kühn, Michael W M; Gerhardinger, Andreas; Bloehdorn, Johannes; Radtke, Ina; Su, Xiaoping; Ma, Jing; Pounds, Stanley; Hallek, Michael; Lichter, Peter; Korbel, Jan; Busch, Raymonde; Mertens, Daniel; Downing, James R; Stilgenbauer, Stephan; Döhner, Hartmut

    2012-12-06

    To identify genomic alterations in chronic lymphocytic leukemia (CLL), we performed single-nucleotide polymorphism-array analysis using Affymetrix Version 6.0 on 353 samples from untreated patients entered in the CLL8 treatment trial. Based on paired-sample analysis (n = 144), a mean of 1.8 copy number alterations per patient were identified; approximately 60% of patients carried no copy number alterations other than those detected by fluorescence in situ hybridization analysis. Copy-neutral loss-of-heterozygosity was detected in 6% of CLL patients and was found most frequently on 13q, 17p, and 11q. Minimally deleted regions were refined on 13q14 (deleted in 61% of patients) to the DLEU1 and DLEU2 genes, on 11q22.3 (27% of patients) to ATM, on 2p16.1-2p15 (gained in 7% of patients) to a 1.9-Mb fragment containing 9 genes, and on 8q24.21 (5% of patients) to a segment 486 kb proximal to the MYC locus. 13q deletions exhibited proximal and distal breakpoint cluster regions. Among the most common novel lesions were deletions at 15q15.1 (4% of patients), with the smallest deletion (70.48 kb) found in the MGA locus. Sequence analysis of MGA in 59 samples revealed a truncating mutation in one CLL patient lacking a 15q deletion. MNT at 17p13.3, which in addition to MGA and MYC encodes for the network of MAX-interacting proteins, was also deleted recurrently.

  12. Effect of chronic valproic Acid treatment on hepatic gene expression profile in wfs1 knockout mouse.

    PubMed

    Punapart, Marite; Eltermaa, Mall; Oflijan, Julia; Sütt, Silva; Must, Anne; Kõks, Sulev; Schalkwyk, Leonard C; Fernandes, Catherine; Vasar, Eero; Soomets, Ursel; Terasmaa, Anton

    2014-01-01

    Valproic acid (VPA) is a widely used anticonvulsant and mood-stabilizing drug whose use is often associated with drug-induced weight gain. Treatment with VPA has been shown to upregulate Wfs1 expression in vitro. Aim of the present study was to compare the effect of chronic VPA treatment in wild type (WT) and Wfs1 knockout (KO) mice on hepatic gene expression profile. Wild type, Wfs1 heterozygous, and homozygous mice were treated with VPA for three months (300 mg/kg i.p. daily) and gene expression profiles in liver were evaluated using Affymetrix Mouse GeneChip 1.0 ST array. We identified 42 genes affected by Wfs1 genotype, 10 genes regulated by VPA treatment, and 9 genes whose regulation by VPA was dependent on genotype. Among the genes that were regulated differentially by VPA depending on genotype was peroxisome proliferator-activated receptor delta (Ppard), whose expression was upregulated in response to VPA treatment in WT, but not in Wfs1 KO mice. Thus, regulation of Ppard by VPA is dependent on Wfs1 genotype.

  13. A Provisional Gene Regulatory Atlas for Mouse Heart Development

    PubMed Central

    Chen, Hailin; VanBuren, Vincent

    2014-01-01

    Congenital Heart Disease (CHD) is one of the most common birth defects. Elucidating the molecular mechanisms underlying normal cardiac development is an important step towards early identification of abnormalities during the developmental program and towards the creation of early intervention strategies. We developed a novel computational strategy for leveraging high-content data sets, including a large selection of microarray data associated with mouse cardiac development, mouse genome sequence, ChIP-seq data of selected mouse transcription factors and Y2H data of mouse protein-protein interactions, to infer the active transcriptional regulatory network of mouse cardiac development. We identified phase-specific expression activity for 765 overlapping gene co-expression modules that were defined for obtained cardiac lineage microarray data. For each co-expression module, we identified the phase of cardiac development where gene expression for that module was higher than other phases. Co-expression modules were found to be consistent with biological pathway knowledge in Wikipathways, and met expectations for enrichment of pathways involved in heart lineage development. Over 359,000 transcription factor-target relationships were inferred by analyzing the promoter sequences within each gene module for overrepresentation against the JASPAR database of Transcription Factor Binding Site (TFBS) motifs. The provisional regulatory network will provide a framework of studying the genetic basis of CHD. PMID:24421884

  14. Mouse models in oncoimmunology.

    PubMed

    Zitvogel, Laurence; Pitt, Jonathan M; Daillère, Romain; Smyth, Mark J; Kroemer, Guido

    2016-12-01

    Fundamental cancer research and the development of efficacious antineoplastic treatments both rely on experimental systems in which the relationship between malignant cells and immune cells can be studied. Mouse models of transplantable, carcinogen-induced or genetically engineered malignancies - each with their specific advantages and difficulties - have laid the foundations of oncoimmunology. These models have guided the immunosurveillance theory that postulates that evasion from immune control is an essential feature of cancer, the concept that the long-term effects of conventional cancer treatments mostly rely on the reinstatement of anticancer immune responses and the preclinical development of immunotherapies, including currently approved immune checkpoint blockers. Specific aspects of pharmacological development, as well as attempts to personalize cancer treatments using patient-derived xenografts, require the development of mouse models in which murine genes and cells are replaced with their human equivalents. Such 'humanized' mouse models are being progressively refined to characterize the leukocyte subpopulations that belong to the innate and acquired arms of the immune system as they infiltrate human cancers that are subjected to experimental therapies. We surmise that the ever-advancing refinement of murine preclinical models will accelerate the pace of therapeutic optimization in patients.

  15. Funding Opportunity: Genomic Data Centers

    Cancer.gov

    Funding Opportunity CCG, Funding Opportunity Center for Cancer Genomics, CCG, Center for Cancer Genomics, CCG RFA, Center for cancer genomics rfa, genomic data analysis network, genomic data analysis network centers,

  16. Mechanical Unloading of Mouse Bone in Microgravity Significantly Alters Cell Cycle Gene Set Expression

    NASA Astrophysics Data System (ADS)

    Blaber, Elizabeth; Dvorochkin, Natalya; Almeida, Eduardo; Kaplan, Warren; Burns, Brnedan

    2012-07-01

    unloading in spaceflight, we conducted genome wide microarray analysis of total RNA isolated from the mouse pelvis. Specifically, 16 week old mice were subjected to 15 days spaceflight onboard NASA's STS-131 space shuttle mission. The pelvis of the mice was dissected, the bone marrow was flushed and the bones were briefly stored in RNAlater. The pelvii were then homogenized, and RNA was isolated using TRIzol. RNA concentration and quality was measured using a Nanodrop spectrometer, and 0.8% agarose gel electrophoresis. Samples of cDNA were analyzed using an Affymetrix GeneChip\\S Gene 1.0 ST (Sense Target) Array System for Mouse and GenePattern Software. We normalized the ST gene arrays using Robust Multichip Average (RMA) normalization, which summarizes perfectly matched spots on the array through the median polish algorithm, rather than normalizing according to mismatched spots. We also used Limma for statistical analysis, using the BioConductor Limma Library by Gordon Smyth, and differential expression analysis to identify genes with significant changes in expression between the two experimental conditions. Finally we used GSEApreRanked for Gene Set Enrichment Analysis (GSEA), with Kolmogorov-Smirnov style statistics to identify groups of genes that are regulated together using the t-statistics derived from Limma. Preliminary results show that 6,603 genes expressed in pelvic bone had statistically significant alterations in spaceflight compared to ground controls. These prominently included cell cycle arrest molecules p21, and p18, cell survival molecule Crbp1, and cell cycle molecules cyclin D1, and Cdk1. Additionally, GSEA results indicated alterations in molecular targets of cyclin D1 and Cdk4, senescence pathways resulting from abnormal laminin maturation, cell-cell contacts via E-cadherin, and several pathways relating to protein translation and metabolism. In total 111 gene sets out of 2,488, about 4%, showed statistically significant set alterations. These

  17. REVIEW: Zebrafish: A Renewed Model System For Functional Genomics

    NASA Astrophysics Data System (ADS)

    Wen, Xiao-Yan

    2008-01-01

    In the post genome era, a major goal in molecular biology is to determine the function of the many thousands of genes present in the vertebrate genome. The zebrafish (Danio rerio) provides an almost ideal genetic model to identify the biological roles of these novel genes, in part because their embryos are transparent and develop rapidly. The zebrafish has many advantages over mouse for genome-wide mutagenesis studies, allowing for easier, cheaper and faster functional characterization of novel genes in the vertebrate genome. Many molecular research tools such as chemical mutagenesis, transgenesis, gene trapping, gene knockdown, TILLING, gene targeting, RNAi and chemical genetic screen are now available in zebrafish. Combining all the forward, reverse, and chemical genetic tools, it is expected that zebrafish will make invaluable contribution to vertebrate functional genomics in functional annotation of the genes, modeling human diseases and drug discoveries.

  18. Ontology for Genome Comparison and Genomic Rearrangements

    PubMed Central

    Flanagan, Keith; Stevens, Robert; Pocock, Matthew; Lee, Pete

    2004-01-01

    We present an ontology for describing genomes, genome comparisons, their evolution and biological function. This ontology will support the development of novel genome comparison algorithms and aid the community in discussing genomic evolution. It provides a framework for communication about comparative genomics, and a basis upon which further automated analysis can be built. The nomenclature defined by the ontology will foster clearer communication between biologists, and also standardize terms used by data publishers in the results of analysis programs. The overriding aim of this ontology is the facilitation of consistent annotation of genomes through computational methods, rather than human annotators. To this end, the ontology includes definitions that support computer analysis and automated transfer of annotations between genomes, rather than relying upon human mediation. PMID:18629137

  19. Enabling functional genomics with genome engineering.

    PubMed

    Hilton, Isaac B; Gersbach, Charles A

    2015-10-01

    Advances in genome engineering technologies have made the precise control over genome sequence and regulation possible across a variety of disciplines. These tools can expand our understanding of fundamental biological processes and create new opportunities for therapeutic designs. The rapid evolution of these methods has also catalyzed a new era of genomics that includes multiple approaches to functionally characterize and manipulate the regulation of genomic information. Here, we review the recent advances of the most widely adopted genome engineering platforms and their application to functional genomics. This includes engineered zinc finger proteins, TALEs/TALENs, and the CRISPR/Cas9 system as nucleases for genome editing, transcription factors for epigenome editing, and other emerging applications. We also present current and potential future applications of these tools, as well as their current limitations and areas for future advances.

  20. Enabling functional genomics with genome engineering

    PubMed Central

    Hilton, Isaac B.; Gersbach, Charles A.

    2015-01-01

    Advances in genome engineering technologies have made the precise control over genome sequence and regulation possible across a variety of disciplines. These tools can expand our understanding of fundamental biological processes and create new opportunities for therapeutic designs. The rapid evolution of these methods has also catalyzed a new era of genomics that includes multiple approaches to functionally characterize and manipulate the regulation of genomic information. Here, we review the recent advances of the most widely adopted genome engineering platforms and their application to functional genomics. This includes engineered zinc finger proteins, TALEs/TALENs, and the CRISPR/Cas9 system as nucleases for genome editing, transcription factors for epigenome editing, and other emerging applications. We also present current and potential future applications of these tools, as well as their current limitations and areas for future advances. PMID:26430154

  1. ECR Browser: A Tool For Visualizing And Accessing Data From Comparisons Of Multiple Vertebrate Genomes

    SciTech Connect

    Loots, G G; Ovcharenko, I; Stubbs, L; Nobrega, M A

    2004-01-06

    The increasing number of vertebrate genomes being sequenced in draft or finished form provide a unique opportunity to study and decode the language of DNA sequence through comparative genome alignments. However, novel tools and strategies are required to accommodate this increasing volume of genomic information and to facilitate experimental annotation of genome function. Here we present the ECR Browser, a tool that provides an easy and dynamic access to whole genome alignments of human, mouse, rat and fish sequences. This web-based tool (http://ecrbrowser.dcode.org) provides the starting point for discovery of novel genes, identification of distant gene regulatory elements and prediction of transcription factor binding sites. The genome alignment portal of the ECR Browser also permits fast and automated alignment of any user-submitted sequence to the genome of choice. The interconnection of the ECR browser with other DNA sequence analysis tools creates a unique portal for studying and exploring vertebrate genomes.

  2. Genome Sequence Alterations Detected upon Passage of Burkholderia mallei ATCC 23344 in Culture and in Mammalian Hosts

    DTIC Science & Technology

    2006-09-05

    genome sequence of B. mallei ATCC 23344: nine in the laboratory culture passaged isolate, eight in the mouse spleen isolate, eight in the horse lung...mouse spleen isolate and 8 in the lab culture and indels 3 in the human liver isolate and 3 in the human blood isolate, were different. Intergenic...8 CTGTCGTG 21 22 no no BMAA0376 Transporter 9 GTGCGAT 19 20 no no BMAA1878 Transcriptional regulator Mouse Spleen 1 GAGGCGT 26 25 no no BMA2774

  3. Construction of a mouse model of factor VIII deficiency by gene targeting

    SciTech Connect

    Bi, L.; Lawler, A.; Gearhart, J.

    1994-09-01

    To develop a small animal model of hemophilia A for gene therapy experiments, we set out to construct a mouse model for factor VIII deficiency by gene targeting. First, we screened a mouse liver cDNA library using a human FVIII cDNA probe. We cloned a 2.6 Kb partial mouse factor VIII cDNA which extends from 800 base pairs of the 3{prime} end of exon 14 to the 5{prime} end of exon 26. A mouse genomic library made from strain 129 was then screened to obtain genomic fragments covering the exons desired for homologous recombination. Two genomic clones were obtained, and one covering exon 15 through 22 was used for gene targeting. To make gene targeting constructs, a 5.8 Kb genomic DNA fragment covering exons 15 to 19 of the mouse FVIII gene was subcloned, and the neo expression cassette was inserted into exons 16 and 17 separately by different strategies. These two constructs were named MFVIIIC-16 and MFVIIIC-17. The constructs were linearized and transfected into strain 129 mouse ES cells by electroporation. Factor VIII gene-knockout ES cell lines were selected by G-418 and screened by genomic Southern blots. Eight exon 16 targeted cell lines and five exon 17 targeted cell lines were obtained. Three cell lines from each construct were injected into blastocysts and surgically transferred into foster mothers. Multiple chimeric mice with 70-90% hair color derived from the ES-cell genotype were seen with both constructs. Germ line transmission of the ES-cell genotype has been obtained for the MFVIIIC-16 construct, and multiple hemophilia A carrier females have been identified. Factor VIII-deficient males will be conceived soon.

  4. Navigating yeast genome maintenance with functional genomics.

    PubMed

    Measday, Vivien; Stirling, Peter C

    2016-03-01

    Maintenance of genome integrity is a fundamental requirement of all organisms. To address this, organisms have evolved extremely faithful modes of replication, DNA repair and chromosome segregation to combat the deleterious effects of an unstable genome. Nonetheless, a small amount of genome instability is the driver of evolutionary change and adaptation, and thus a low level of instability is permitted in populations. While defects in genome maintenance almost invariably reduce fitness in the short term, they can create an environment where beneficial mutations are more likely to occur. The importance of this fact is clearest in the development of human cancer, where genome instability is a well-established enabling characteristic of carcinogenesis. This raises the crucial question: what are the cellular pathways that promote genome maintenance and what are their mechanisms? Work in model organisms, in particular the yeast Saccharomyces cerevisiae, has provided the global foundations of genome maintenance mechanisms in eukaryotes. The development of pioneering genomic tools inS. cerevisiae, such as the systematic creation of mutants in all nonessential and essential genes, has enabled whole-genome approaches to identifying genes with roles in genome maintenance. Here, we review the extensive whole-genome approaches taken in yeast, with an emphasis on functional genomic screens, to understand the genetic basis of genome instability, highlighting a range of genetic and cytological screening modalities. By revealing the biological pathways and processes regulating genome integrity, these analyses contribute to the systems-level map of the yeast cell and inform studies of human disease, especially cancer.

  5. Culex genome is not just another genome for comparative genomics.

    PubMed

    Reddy, B P Niranjan; Labbé, Pierrick; Corbel, Vincent

    2012-03-30

    Formal publication of the Culex genome sequence has closed the human disease vector triangle by meeting the Anopheles gambiae and Aedes aegypti genome sequences. Compared to these other mosquitoes, Culex quinquefasciatus possesses many specific hallmark characteristics, and may thus provide different angles for research which ultimately leads to a practical solution for controlling the ever increasing burden of insect-vector-borne diseases around the globe. We argue the special importance of the cosmopolitan species- Culex genome sequence by invoking many interesting questions and the possible of potential of the Culex genome to answer those.

  6. Exploring Other Genomes: Bacteria.

    ERIC Educational Resources Information Center

    Flannery, Maura C.

    2001-01-01

    Points out the importance of genomes other than the human genome project and provides information on the identified bacterial genomes Pseudomonas aeuroginosa, Leprosy, Cholera, Meningitis, Tuberculosis, Bubonic Plague, and plant pathogens. Considers the computer's use in genome studies. (Contains 14 references.) (YDS)

  7. Revealing Transcriptome Landscape of Mouse Spermatogonial Cells by Tiling Microarray

    PubMed Central

    Lee, Tin-Lap.; Rennert, Owen M.; Chan, Wai-Yee.

    2014-01-01

    Summary Spermatogenesis is a highly regulated developmental process by which spermatogonia develop into mature spermatozoa. This process involves many testis- or male germ cell-specific events through tightly regulated gene expression programs. In the past decade the advent of microarray technologies has allowed functional genomic studies of male germ cell development, resulting in the identification of genes governing various processes. A major limitation with conventional gene expression microarray is that there is a bias from gene probe design. The gene probes for expression microarrays are usually represented by a small number probes located at the 3’ end of a transcirpt. Tiling microarrays eliminate such issue by interrogating the genome in an unbiased fashion through probes tiled for the entire genome. These arrays provide a higher genomic resolution and allow identification of novel transcripts. To reveal the complexity of the genomic landscape of developing male germ cells, we applied tiling microarray to evaluate the transcriptome in spermatogonial cells. Over 50% of the mouse and rat genome are expressed during testicular development. More than 47% of transcripts are uncharacterized. The results suggested the transcription machinery in spermaotogonial cells are more complex than previously envisioned. PMID:22144238

  8. Manipulation of Mouse Embryonic Stem Cells for Knockout Mouse Production

    PubMed Central

    Limaye, Advait; Hall, Bradford; Kulkarni, Ashok B

    2009-01-01

    The establishment of mouse embryonic stem (ES) cell liness has allowed for the generation of the knockout mouse. ES cells that are genetically altered in culture can then be manipulated to derive a whole mouse containing the desired mutation. To successfully generate a knockout mouse, however, the ES cells must be carefully cultivated in a pluripotent state throughout the gene targeting experiment. This unit describes detailed step-by-step protocols, reagents, equipment, and strategies needed for the successful generation of gene knockout embryonic stem cells using homologous recombination technologies. PMID:19731225

  9. Functional analysis of limb transcriptional enhancers in the mouse

    PubMed Central

    Nolte, Mark J.; Wang, Ying; Deng, Jian Min; Swinton, Paul G.; Wei, Caimiao; Guindani, Michele; Schwartz, Robert J.; Behringer, Richard R.

    2014-01-01

    Transcriptional enhancers are genomic sequences bound by transcription factors that act together with basal transcriptional machinery to regulate gene transcription. Several high-throughput methods have generated large datasets of tissue-specific enhancer sequences with putative roles in developmental processes. However, few enhancers have been deleted from the genome to determine their roles in development. To understand the roles of two enhancers active in the mouse embryonic limb bud we deleted them from the genome. Although the genes regulated by these enhancers are unknown, they were selected because they were identified in a screen for putative limb bud-specific enhancers associated with p300, an acetyltransferase that participates in protein complexes that promote active transcription, and because the orthologous human enhancers (H1442 and H280) drive distinct lacZ expression patterns in limb buds of embryonic day (E) 11.5 transgenic mice. We show that the orthologous mouse sequences, M280 and M1442, regulate dynamic expression in the developing limb. Although significant transcriptional differences in enhancer-proximal genes in embryonic limb buds accompany the deletion of M280 and M1442 no gross limb malformations during embryonic development were observed, demonstrating that M280 and M1442 are not required for mouse limb development. However, M280 is required for the development and/or maintenance of body size; M280 mice are significantly smaller than controls. M280 also harbors an “ultraconserved” sequence that is identical between human, rat, and mouse. This is the first report of a phenotype resulting from the deletion of an ultraconserved element. These studies highlight the importance of determining enhancer regulatory function by experiments that manipulate them in situ and suggest that some of an enhancer's regulatory capacities may be developmentally tolerated rather than developmentally required. PMID:24920384

  10. Exploiting the Genome

    DTIC Science & Technology

    1998-09-11

    complete human genome sequence . 14. SUBJECT TERMS 15. NUMBER OF PAGES 16. PRICE CODE 17. SECURITY CLASSIFICATION OF REPORT Unclassified 18. SECURITY...goal of the project is to ob- tain the complete sequence of the human genome by the year 2005. The genome contains approximately 3.3 Gb (billion base...and second, to consider possible roles for the DOE in the "post- genomic " era, following acquisition of the complete human genome

  11. Genome Maps, a new generation genome browser

    PubMed Central

    Medina, Ignacio; Salavert, Francisco; Sanchez, Rubén; de Maria, Alejandro; Alonso, Roberto; Escobar, Pablo; Bleda, Marta; Dopazo, Joaquín

    2013-01-01

    Genome browsers have gained importance as more genomes and related genomic information become available. However, the increase of information brought about by new generation sequencing technologies is, at the same time, causing a subtle but continuous decrease in the efficiency of conventional genome browsers. Here, we present Genome Maps, a genome browser that implements an innovative model of data transfer and management. The program uses highly efficient technologies from the new HTML5 standard, such as scalable vector graphics, that optimize workloads at both server and client sides and ensure future scalability. Thus, data management and representation are entirely carried out by the browser, without the need of any Java Applet, Flash or other plug-in technology installation. Relevant biological data on genes, transcripts, exons, regulatory features, single-nucleotide polymorphisms, karyotype and so forth, are imported from web services and are available as tracks. In addition, several DAS servers are already included in Genome Maps. As a novelty, this web-based genome browser allows the local upload of huge genomic data files (e.g. VCF or BAM) that can be dynamically visualized in real time at the client side, thus facilitating the management of medical data affected by privacy restrictions. Finally, Genome Maps can easily be integrated in any web application by including only a few lines of code. Genome Maps is an open source collaborative initiative available in the GitHub repository (https://github.com/compbio-bigdata-viz/genome-maps). Genome Maps is available at: http://www.genomemaps.org. PMID:23748955

  12. Genome Maps, a new generation genome browser.

    PubMed

    Medina, Ignacio; Salavert, Francisco; Sanchez, Rubén; de Maria, Alejandro; Alonso, Roberto; Escobar, Pablo; Bleda, Marta; Dopazo, Joaquín

    2013-07-01

    Genome browsers have gained importance as more genomes and related genomic information become available. However, the increase of information brought about by new generation sequencing technologies is, at the same time, causing a subtle but continuous decrease in the efficiency of conventional genome browsers. Here, we present Genome Maps, a genome browser that implements an innovative model of data transfer and management. The program uses highly efficient technologies from the new HTML5 standard, such as scalable vector graphics, that optimize workloads at both server and client sides and ensure future scalability. Thus, data management and representation are entirely carried out by the browser, without the need of any Java Applet, Flash or other plug-in technology installation. Relevant biological data on genes, transcripts, exons, regulatory features, single-nucleotide polymorphisms, karyotype and so forth, are imported from web services and are available as tracks. In addition, several DAS servers are already included in Genome Maps. As a novelty, this web-based genome browser allows the local upload of huge genomic data files (e.g. VCF or BAM) that can be dynamically visualized in real time at the client side, thus facilitating the management of medical data affected by privacy restrictions. Finally, Genome Maps can easily be integrated in any web application by including only a few lines of code. Genome Maps is an open source collaborative initiative available in the GitHub repository (https://github.com/compbio-bigdata-viz/genome-maps). Genome Maps is available at: http://www.genomemaps.org.

  13. Genomic disorders on 22q11.

    PubMed

    McDermid, Heather E; Morrow, Bernice E

    2002-05-01

    The 22q11 region is involved in chromosomal rearrangements that lead to altered gene dosage, resulting in genomic disorders that are characterized by mental retardation and/or congenital malformations. Three such disorders-cat-eye syndrome (CES), der(22) syndrome, and velocardiofacial syndrome/DiGeorge syndrome (VCFS/DGS)-are associated with four, three, and one dose, respectively, of parts of 22q11. The critical region for CES lies centromeric to the deletion region of VCFS/DGS, although, in some cases, the extra material in CES extends across the VCFS/DGS region. The der(22) syndrome region overlaps both the CES region and the VCFS/DGS region. Molecular approaches have revealed a set of common chromosome breakpoints that are shared between the three disorders, implicating specific mechanisms that cause these rearrangements. Most VCFS/DGS and CES rearrangements are likely to occur by homologous recombination events between blocks of low-copy repeats (e.g., LCR22), whereas nonhomologous recombination mechanisms lead to the constitutional t(11;22) translocation. Meiotic nondisjunction events in carriers of the t(11;22) translocation can then lead to offspring with der(22) syndrome. The molecular basis of the clinical phenotype of these genomic disorders has also begun to be addressed. Analysis of both the genomic sequence for the 22q11 interval and the orthologous regions in the mouse has identified >24 genes that are shared between VCFS/DGS and der(22) syndrome and has identified 14 putative genes that are shared between CES and der(22) syndrome. The ability to manipulate the mouse genome aids in the identification of candidate genes in these three syndromes. Research on genomic disorders on 22q11 will continue to expand our knowledge of the mechanisms of chromosomal rearrangements and the molecular basis of their phenotypic consequences.

  14. Genetic Networks in Mouse Retinal Ganglion Cells

    PubMed Central

    Struebing, Felix L.; Lee, Richard K.; Williams, Robert W.; Geisert, Eldon E.

    2016-01-01

    Retinal ganglion cells (RGCs) are the output neuron of the eye, transmitting visual information from the retina through the optic nerve to the brain. The importance of RGCs for vision is demonstrated in blinding diseases where RGCs are lost, such as in glaucoma or after optic nerve injury. In the present study, we hypothesize that normal RGC function is transcriptionally regulated. To test our hypothesis, we examine large retinal expression microarray datasets from recombinant inbred mouse strains in GeneNetwork and define transcriptional networks of RGCs and their subtypes. Two major and functionally distinct transcriptional networks centering around Thy1 and Tubb3 (Class III beta-tubulin) were identified. Each network is independently regulated and modulated by unique genomic loci. Meta-analysis of publically available data confirms that RGC subtypes are differentially susceptible to death, with alpha-RGCs and intrinsically photosensitive RGCs (ipRGCs) being less sensitive to cell death than other RGC subtypes in a mouse model of glaucoma. PMID:27733864

  15. A humanoid mouse model of autism.

    PubMed

    Takumi, Toru

    2010-10-01

    Even now fruit of the human genome project is available, we have difficulties to approach neuropsychiatric disorders at the molecular level. Autism is a complex psychiatric illness but has received considerable attention as a developmental brain disorder not only from basic researchers but also from society. Substantial evidence suggests that chromosomal abnormalities contribute to autism risk. The duplication of human chromosome 15q11-13 is known to be the most frequent cytogenetic abnormality in autism. We succeeded to generate mice with a 6.3-Mb-wide interstitial duplication in mouse chromosome 7c that is highly syntenic to human 15q11-13 by using a Cre-loxP-based chromosome-engineering technique. The only paternally duplicated mice display autistic behavioral features such as poor social interaction and stereotypical behavior, and exhibit a developmental abnormality in ultrasonic vocalizations as well as anxiety. The detailed analysis focusing on a non-coding small nucleolar RNA, MBII52, within the duplicated region, revealed that the paternally duplicated mice alter the editing ratio of serotonin (5-HT) 2c receptor pre-mRNA and intracellular calcium responses by a 5-HT2c receptor specific agonist are changed in neurons. This result may explain one of molecular mechanisms of abnormal behaviors in the paternal duplicated mice. The first chromosome-engineered mouse model for human chromosome 15q11-13 duplication fulfills not only face validity of human autistic phenotypes but also construct validity based on human chromosome abnormality. This model will be a founder mouse for forward genetics of autistic disease and an invaluable tool for its therapeutic development.

  16. Genomics: Looking at Life in New Ways

    SciTech Connect

    Adams, Mark D.

    2003-10-22

    The availability of complete or nearly complete mouse, human, and rat genomes (in addition to those from many other species) has resulted in a series of new and powerful opportunities to apply the technologies and approaches developed for large-scale genome sequencing to the study of disease. New approaches to biological problems are being explored that involve concepts from computer science such as systems theory and modern large scale computing techniques. A recent project at Celera Genomics involved sequencing protein coding regions from several humans and a chimpanzee. Computational models of evolutionary divergence enabled us to identify genes with unique evolutionary signatures. These genes give us some insight into features that may be uniquely human. The laboratory mouse and rat have long been favorite mammalian models of human disease. Integrated approaches to the study of disease that combine genetics, DNA sequence analysis, and careful analysis of phenotype at a molecular level are becoming more common and powerful. In addition, evaluation of the variation inherent in normal populations is now being used to build networks to describe heart function based on the interaction of multiple phenotypes in randomized populations using a factorial design.

  17. Genome-Wide Association Studies for Comb Traits in Chickens

    PubMed Central

    Ma, Meng; Dou, Taocun; Lu, Jian; Guo, Jun; Hu, Yuping; Yi, Guoqiang; Yuan, Jingwei; Sun, Congjiao; Wang, Kehua; Yang, Ning

    2016-01-01

    The comb, as a secondary sexual character, is an important trait in chicken. Indicators of comb length (CL), comb height (CH), and comb weight (CW) are often selected in production. DNA-based marker-assisted selection could help chicken breeders to accelerate genetic improvement for comb or related economic characters by early selection. Although a number of quantitative trait loci (QTL) and candidate genes have been identified with advances in molecular genetics, candidate genes underlying comb traits are limited. The aim of the study was to use genome-wide association (GWA) studies by 600 K Affymetrix chicken SNP arrays to detect genes that are related to comb, using an F2 resource population. For all comb characters, comb exhibited high SNP-based heritability estimates (0.61–0.69). Chromosome 1 explained 20.80% genetic variance, while chromosome 4 explained 6.89%. Independent univariate genome-wide screens for each character identified 127, 197, and 268 novel significant SNPs with CL, CH, and CW, respectively. Three candidate genes, VPS36, AR, and WNT11B, were determined to have a plausible function in all comb characters. These genes are important to the initiation of follicle development, gonadal growth, and dermal development, respectively. The current study provides the first GWA analysis for comb traits. Identification of the genetic basis as well as promising candidate genes will help us understand the underlying genetic architecture of comb development and has practical significance in breeding programs for the selection of comb as an index for sexual maturity or reproduction. PMID:27427764

  18. A Whole Genome Association Study on Meat Palatability in Hanwoo

    PubMed Central

    Hyeong, K.-E.; Lee, Y.-M.; Kim, Y.-S.; Nam, K. C.; Jo, C.; Lee, K.-H.; Lee, J.-E.; Kim, J.-J.

    2014-01-01

    A whole genome association (WGA) study was carried out to find quantitative trait loci (QTL) for sensory evaluation traits in Hanwoo. Carcass samples of 250 Hanwoo steers were collected from National Agricultural Cooperative Livestock Research Institute, Ansung, Gyeonggi province, Korea, between 2011 and 2012 and genotyped with the Affymetrix Bovine Axiom Array 640K single nucleotide polymorphism (SNP) chip. Among the SNPs in the chip, a total of 322,160 SNPs were chosen after quality control tests. After adjusting for the effects of age, slaughter-year-season, and polygenic effects using genome relationship matrix, the corrected phenotypes for the sensory evaluation measurements were regressed on each SNP using a simple linear regression additive based model. A total of 1,631 SNPs were detected for color, aroma, tenderness, juiciness and palatability at 0.1% comparison-wise level. Among the significant SNPs, the best set of 52 SNP markers were chosen using a forward regression procedure at 0.05 level, among which the sets of 8, 14, 11, 10, and 9 SNPs were determined for the respectively sensory evaluation traits. The sets of significant SNPs explained 18% to 31% of phenotypic variance. Three SNPs were pleiotropic, i.e. AX-26703353 and AX-26742891 that were located at 101 and 110 Mb of BTA6, respectively, influencing tenderness, juiciness and palatability, while AX-18624743 at 3 Mb of BTA10 affected tenderness and palatability. Our results suggest that some QTL for sensory measures are segregating in a Hanwoo steer population. Additional WGA studies on fatty acid and nutritional components as well as the sensory panels are in process to characterize genetic architecture of meat quality and palatability in Hanwoo. PMID:25178363

  19. Genomic Analysis of Stress Response against Arsenic in Caenorhabditis elegans

    PubMed Central

    Sahu, Surasri N.; Lewis, Jada; Patel, Isha; Bozdag, Serdar; Lee, Jeong H.; Sprando, Robert; Cinar, Hediye Nese

    2013-01-01

    Arsenic, a known human carcinogen, is widely distributed around the world and found in particularly high concentrations in certain regions including Southwestern US, Eastern Europe, India, China, Taiwan and Mexico. Chronic arsenic poisoning affects millions of people worldwide and is associated with increased risk of many diseases including arthrosclerosis, diabetes and cancer. In this study, we explored genome level global responses to high and low levels of arsenic exposure in Caenorhabditis elegans using Affymetrix expression microarrays. This experimental design allows us to do microarray analysis of dose-response relationships of global gene expression patterns. High dose (0.03%) exposure caused stronger global gene expression changes in comparison with low dose (0.003%) exposure, suggesting a positive dose-response correlation. Biological processes such as oxidative stress, and iron metabolism, which were previously reported to be involved in arsenic toxicity studies using cultured cells, experimental animals, and humans, were found to be affected in C. elegans. We performed genome-wide gene expression comparisons between our microarray data and publicly available C. elegans microarray datasets of cadmium, and sediment exposure samples of German rivers Rhine and Elbe. Bioinformatics analysis of arsenic-responsive regulatory networks were done using FastMEDUSA program. FastMEDUSA analysis identified cancer-related genes, particularly genes associated with leukemia, such as dnj-11, which encodes a protein orthologous to the mammalian ZRF1/MIDA1/MPP11/DNAJC2 family of ribosome-associated molecular chaperones. We analyzed the protective functions of several of the identified genes using RNAi. Our study indicates that C. elegans could be a substitute model to study the mechanism of metal toxicity using high-throughput expression data and bioinformatics tools such as FastMEDUSA. PMID:23894281

  20. Gene expression profiling in the lung and liver of PFOA-exposed mouse fetuses.

    PubMed

    Rosen, Mitchell B; Thibodeaux, Julie R; Wood, Carmen R; Zehr, Robert D; Schmid, Judith E; Lau, Christopher

    2007-09-24

    Perfluorooctanoic acid (PFOA) is a stable perfluoroalkyl acid used to synthesize fluoropolymers during the manufacture of a wide variety of products. Concerns have been raised over the potential health effects of PFOA because it is persistent in the environment and can be detected in blood and other tissues of many animal species, including humans. PFOA has also been shown to induce growth deficits and mortality in murine neonates. To better understand the mechanism of PFOA induced developmental toxicity, lung and liver gene expression profiling was conducted in PFOA-exposed full-term mouse fetuses. Thirty timed-pregnant CD-1 mice were orally dosed from gestation days 1-17 with either 0, 1, 3, 5, or 10mg/(kgday) PFOA in water. At term, fetal lung and liver were collected, total RNA prepared, and samples pooled from three fetuses per litter. Five biological replicates consisting of individual litter samples were then evaluated for each treatment group using Affymetrix mouse 430_2 microarrays. The expression of genes related to fatty acid catabolism was altered in both the fetal liver and lung. In the fetal liver, the effects of PFOA were robust and also included genes associated with lipid transport, ketogenesis, glucose metabolism, lipoprotein metabolism, cholesterol biosynthesis, steroid metabolism, bile acid biosynthesis, phospholipid metabolism, retinol metabolism, proteosome activation, and inflammation. These changes are consistent with transactivation of PPARalpha, although, with regard to bile acid biosynthesis and glucose metabolism, non-PPARalpha related effects were suggested as well. Additional studies will be needed to more thoroughly address the role of PPARalpha, and other nuclear receptors, in PFOA mediated developmental toxicity.

  1. The Mouse House: a brief history of the ORNL mouse-genetics program, 1947-2009.

    PubMed

    Russell, Liane B

    2013-01-01

    The large mouse genetics program at the Oak Ridge National Laboratory (ORNL) is often remembered chiefly for the germ-cell mutation-rate data it generated and their uses in estimating the risk of heritable radiation damage. In fact, it soon became a multi-faceted research effort that, over a period of almost 60 years, generated a wealth of information in the areas of mammalian mutagenesis, basic genetics (later enriched by molecular techniques), cytogenetics, reproductive biology, biochemistry of germ cells, and teratology. Research in the area of germ-cell mutagenesis explored the important physical and biological factors that affect the frequency and nature of induced mutations and made several unexpected discoveries, such as the major importance of the perigametic interval (the zygote stage) for the origin of spontaneous mutations and for the sensitivity to induced genetic change. Of practical value was the discovery that ethylnitrosourea was a supermutagen for point mutations, making high-efficiency mutagenesis in the mouse feasible worldwide. Teratogenesis findings resulted in recommendations still generally accepted in radiological practice. Studies supporting the mutagenesis research added whole bodies of information about mammalian germ-cell development and about molecular targets in germ cells. The early decision to not merely count but propagate genetic variants of all sorts made possible further discoveries, such as the Y-chromosome's importance in mammalian sex determination and the identification of rare X-autosome translocations, which, in turn, led to the formulation of the single-active-X hypothesis and provided tools for studies of functional mosaicism for autosomal genes, male sterility, and chromosome-pairing mechanism. Extensive genetic and then molecular analyses of large numbers of induced specific-locus mutants resulted in fine-structure physical and correlated functional mapping of significant portions of the mouse genome and constituted a

  2. The Mouse House: A brief history of the ORNL mouse-genetics program, 1947–2009

    SciTech Connect

    Russell, Liane B.

    2013-10-01

    The large mouse genetics program at the Oak Ridge National Lab is often re-membered chiefly for the germ-cell mutation-rate data it generated and their uses in estimating the risk of heritable radiation damage. In fact, it soon became a multi-faceted research effort that, over a period of almost 60 years, generated a wealth of information in the areas of mammalian mutagenesis, basic genetics (later enriched by molecular techniques), cytogenetics, reproductive biology, biochemistry of germ cells, and teratology. Research in the area of germ-cell mutagenesis explored the important physical and biological factors that affect the frequency and nature of induced mutations and made several unexpected discoveries, such as the major importance of the perigametic interval (the zygote stage) for the origin of spontaneous mutations and for the sensitivity to induced genetic change. Of practical value was the discovery that ethylnitrosourea was a supermutagen for point mutations, making high-efficiency mutagenesis in the mouse feasible worldwide. Teratogenesis findings resulted in recommendations still generally accepted in radiological practice. Studies supporting the mutagenesis research added whole bodies of information about mammalian germ-cell development and about molecular targets in germ cells. The early decision to not merely count but propagate genetic variants of all sorts made possible further discoveries, such as the Y-Chromosome s importance in mammalian sex determination and the identification of rare X-autosome translocations, which, in turn, led to the formulation of the single-active-X hypothesis and provided tools for studies of functional mosaicism for autosomal genes, male sterility, and chromosome-pairing mechanism. Extensive genetic and then molecular analyses of large numbers of induced specific-locus mutants resulted in fine-structure physical and correlated functional mapping of significant portions of the mouse genome and constituted a valuable

  3. Diversity and Complexity of the Mouse Saa1 and Saa2 genes

    PubMed Central

    Mori, Masayuki; Tian, Geng; Ishikawa, Akira; Higuchi, Keiichi

    2014-01-01

    Mouse strains show polymorphisms in the amino acid sequences of serum amyloid A 1 (SAA1) and serum amyloid A 2 (SAA2). Major laboratory mouse strains are classified based on the sequence as carrying the A haplotype (e.g., BALB/c) or B haplotype (e.g., SJL/J) of the Saa1 and Saa2 gene unit. We attempted to elucidate the diversity of the mouse Saa1 and Saa2 family genes at the nucleotide sequence level by a systematic survey of 6 inbred mouse strains from 4 Mus subspecies, including Mus musculus domesticus, Mus musculus musculus, Mus musculus castaneus, and Mus spretus. Saa1 and Saa2 genes were obtained from the mouse genome by PCR amplification, and each full-length nucleotide sequence was determined. We found that Mus musculus castaneus mice uniquely possess 2 divergent Saa1 genes linked on chromosome 7. Overall, the mouse strains had distinct composite patterns of amino acid substitutions at 9 positions in SAA1 and SAA2 isoforms. The mouse strains also had distinct composite patterns of 2 polymorphic upstream regulatory elements that influenced gene transcription in in vitro reporter assays. B haplotype mice were revealed to possess an LTR insertion in the downstream region of Saa1. Collectively, these results indicate that the mouse Saa genes hold broader diversity and greater complexity than previously known, and these characteristics were likely attained through gene duplication and repeated gene conversion events in the Mus lineage. PMID:24521869

  4. "Replicated" genome wide association for dependence on illegal substances: genomic regions identified by overlapping clusters of nominally positive SNPs.

    PubMed

    Drgon, Tomas; Johnson, Catherine A; Nino, Michelle; Drgonova, Jana; Walther, Donna M; Uhl, George R

    2011-03-01

    Declaring "replication" from results of genome wide association (GWA) studies is straightforward when major gene effects provide genome-wide significance for association of the same allele of the same SNP in each of multiple independent samples. However, such unambiguous replication may be unlikely when phenotypes display polygenic genetic architecture, allelic heterogeneity, locus heterogeneity, and when different samples display linkage disequilibria with different fine structures. We seek chromosomal regions that are tagged by clustered SNPs that display nominally significant association in each of several independent samples. This approach provides one "nontemplate" approach to identifying overall replication of groups of GWA results in the face of diffi