Massively parallel sequencing of the mouse exome to accurately identify rare, induced mutations: an immediate source for thousands of new mouse models
Identifieur interne : 005319 ( Main/Exploration ); précédent : 005318; suivant : 005320Massively parallel sequencing of the mouse exome to accurately identify rare, induced mutations: an immediate source for thousands of new mouse models
Auteurs : T. D. Andrews [Australie] ; B. Whittle [Australie] ; M. A. Field [Australie] ; B. Balakishnan [Australie] ; Y. Zhang [Australie] ; Y. Shao [Australie] ; V. Cho [Australie] ; M. Kirk [Australie] ; M. Singh [Australie] ; Y. Xia [États-Unis] ; J. Hager [France] ; S. Winslade [Australie] ; G. Sjollema [Australie] ; B. Beutler [États-Unis] ; A. Enders [Australie] ; C. C. Goodnow [Australie]Source :
- Open Biology [ 2046-2441 ] ; 2012.
Abstract
Accurate identification of sparse heterozygous single-nucleotide variants (SNVs) is a critical challenge for identifying the causative mutations in mouse genetic screens, human genetic diseases and cancer. When seeking to identify causal DNA variants that occur at such low rates, they are overwhelmed by false-positive calls that arise from a range of technical and biological sources. We describe a strategy using whole-exome capture, massively parallel DNA sequencing and computational analysis, which identifies with a low false-positive rate the majority of heterozygous and homozygous SNVs arising de novo with a frequency of one nucleotide substitution per megabase in progeny of
Url:
DOI: 10.1098/rsob.120061
PubMed: 22724066
PubMed Central: 3376740
Affiliations:
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<author><name sortKey="Shao, Y" sort="Shao, Y" uniqKey="Shao Y" first="Y." last="Shao">Y. Shao</name>
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<author><name sortKey="Cho, V" sort="Cho, V" uniqKey="Cho V" first="V." last="Cho">V. Cho</name>
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<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
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<country xml:lang="fr">Australie</country>
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<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Hager, J" sort="Hager, J" uniqKey="Hager J" first="J." last="Hager">J. Hager</name>
<affiliation wicri:level="1"><nlm:aff id="af6"><addr-line>Centre National de Génotypage, 2 rue Gaston Crémieux, CP5721, 91057, Evry Cedex</addr-line>
,<country>France</country>
</nlm:aff>
<country xml:lang="fr">France</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Winslade, S" sort="Winslade, S" uniqKey="Winslade S" first="S." last="Winslade">S. Winslade</name>
<affiliation wicri:level="1"><nlm:aff id="af3"><addr-line>Australian Phenomics Facility</addr-line>
,<institution>Australian National University</institution>
,<addr-line>Hugh Ennor Building, Building 117, Garran Road, Canberra City, Australian Capital Territory, 0200</addr-line>
,<country>Australia</country>
</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Sjollema, G" sort="Sjollema, G" uniqKey="Sjollema G" first="G." last="Sjollema">G. Sjollema</name>
<affiliation wicri:level="1"><nlm:aff id="af3"><addr-line>Australian Phenomics Facility</addr-line>
,<institution>Australian National University</institution>
,<addr-line>Hugh Ennor Building, Building 117, Garran Road, Canberra City, Australian Capital Territory, 0200</addr-line>
,<country>Australia</country>
</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Beutler, B" sort="Beutler, B" uniqKey="Beutler B" first="B." last="Beutler">B. Beutler</name>
<affiliation wicri:level="1"><nlm:aff id="af4"><addr-line>Center for the Genetics of Host Defense</addr-line>
,<institution>University of Texas Southwestern</institution>
,<addr-line>6000 Harry Hines Boulevard, Dallas, TX 75930-8505</addr-line>
,<country>USA</country>
</nlm:aff>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
<affiliation wicri:level="1"><nlm:aff id="af5"><addr-line>Department of Genetics</addr-line>
,<institution>Scripps Research Institute</institution>
,<addr-line>10550 North Torrey Pines Road, La Jolla, CA 92037</addr-line>
,<country>USA</country>
</nlm:aff>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Enders, A" sort="Enders, A" uniqKey="Enders A" first="A." last="Enders">A. Enders</name>
<affiliation wicri:level="1"><nlm:aff id="af2"><addr-line>Ramaciotti Immunisation Genomics Laboratory, John Curtin School of Medical Research</addr-line>
,<institution>Australian National University</institution>
,<addr-line>GPO Box 334, Canberra City, Australian Capital Territory, 2601</addr-line>
,<country>Australia</country>
</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
<author><name sortKey="Goodnow, C C" sort="Goodnow, C C" uniqKey="Goodnow C" first="C. C." last="Goodnow">C. C. Goodnow</name>
<affiliation wicri:level="1"><nlm:aff id="af1"><addr-line>Immunogenomics Laboratory</addr-line>
,<institution>Australian National University</institution>
,<addr-line>GPO Box 334, Canberra City, Australian Capital Territory, 2601</addr-line>
,<country>Australia</country>
</nlm:aff>
<country xml:lang="fr">Australie</country>
<wicri:regionArea># see nlm:aff country strict</wicri:regionArea>
</affiliation>
</author>
</analytic>
<series><title level="j">Open Biology</title>
<idno type="eISSN">2046-2441</idno>
<imprint><date when="2012">2012</date>
</imprint>
</series>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc><textClass></textClass>
</profileDesc>
</teiHeader>
<front><div type="abstract" xml:lang="en"><title>Summary</title>
<p>Accurate identification of sparse heterozygous single-nucleotide variants (SNVs) is a critical challenge for identifying the causative mutations in mouse genetic screens, human genetic diseases and cancer. When seeking to identify causal DNA variants that occur at such low rates, they are overwhelmed by false-positive calls that arise from a range of technical and biological sources. We describe a strategy using whole-exome capture, massively parallel DNA sequencing and computational analysis, which identifies with a low false-positive rate the majority of heterozygous and homozygous SNVs arising de novo with a frequency of one nucleotide substitution per megabase in progeny of <italic>N</italic>
-ethyl-<italic>N</italic>
-nitrosourea (ENU)-mutated C57BL/6j mice. We found that by applying a strategy of filtering raw SNV calls against known and platform-specific variants we could call true SNVs with a false-positive rate of 19.4 per cent and an estimated false-negative rate of 21.3 per cent. These error rates are small enough to enable calling a causative mutation from both homozygous and heterozygous candidate mutation lists with little or no further experimental validation. The efficacy of this approach is demonstrated by identifying the causative mutation in the <italic>Ptprc</italic>
gene in a lymphocyte-deficient strain and in 11 other strains with immune disorders or obesity, without the need for meiotic mapping. Exome sequencing of first-generation mutant mice revealed hundreds of unphenotyped protein-changing mutations, 52 per cent of which are predicted to be deleterious, which now become available for breeding and experimental analysis. We show that exome sequencing data alone are sufficient to identify induced mutations. This approach transforms genetic screens in mice, establishes a general strategy for analysing rare DNA variants and opens up a large new source for experimental models of human disease.</p>
</div>
</front>
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</back>
</TEI>
<affiliations><list><country><li>Australie</li>
<li>France</li>
<li>États-Unis</li>
</country>
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<name sortKey="Field, M A" sort="Field, M A" uniqKey="Field M" first="M. A." last="Field">M. A. Field</name>
<name sortKey="Goodnow, C C" sort="Goodnow, C C" uniqKey="Goodnow C" first="C. C." last="Goodnow">C. C. Goodnow</name>
<name sortKey="Kirk, M" sort="Kirk, M" uniqKey="Kirk M" first="M." last="Kirk">M. Kirk</name>
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<name sortKey="Shao, Y" sort="Shao, Y" uniqKey="Shao Y" first="Y." last="Shao">Y. Shao</name>
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<name sortKey="Singh, M" sort="Singh, M" uniqKey="Singh M" first="M." last="Singh">M. Singh</name>
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<name sortKey="Whittle, B" sort="Whittle, B" uniqKey="Whittle B" first="B." last="Whittle">B. Whittle</name>
<name sortKey="Winslade, S" sort="Winslade, S" uniqKey="Winslade S" first="S." last="Winslade">S. Winslade</name>
<name sortKey="Zhang, Y" sort="Zhang, Y" uniqKey="Zhang Y" first="Y." last="Zhang">Y. Zhang</name>
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<country name="États-Unis"><noRegion><name sortKey="Xia, Y" sort="Xia, Y" uniqKey="Xia Y" first="Y." last="Xia">Y. Xia</name>
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<name sortKey="Xia, Y" sort="Xia, Y" uniqKey="Xia Y" first="Y." last="Xia">Y. Xia</name>
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<country name="France"><noRegion><name sortKey="Hager, J" sort="Hager, J" uniqKey="Hager J" first="J." last="Hager">J. Hager</name>
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