Discovery of candidate disease genes in ENU-induced mouse mutants by large-scale sequencing, including a splice-site mutation in nucleoredoxin.
Discovery of candidate disease genes in ENU-induced mouse mutants by large-scale sequencing, including a splice-site mutation in nucleoredoxin.
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DOI：
10.1371/journal.pgen.1000759
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发表时间：
2009-12
期刊：
影响因子：
4.5
通讯作者：
Justice MJ
中科院分区：
文献类型：
--
作者：
Boles MK;Wilkinson BM;Wilming LG;Liu B;Probst FJ;Harrow J;Grafham D;Hentges KE;Woodward LP;Maxwell A;Mitchell K;Risley MD;Johnson R;Hirschi K;Lupski JR;Funato Y;Miki H;Marin-Garcia P;Matthews L;Coffey AJ;Parker A;Hubbard TJ;Rogers J;Bradley A;Adams DJ;Justice MJ
准确和精确注释的基因组组装是功能基因组分析的基本要求。在此，使用小鼠11号染色体的完整DNA序列和基因注释来测试大规模测序用于突变鉴定的功效。我们重新测序了来自41个隐性突变小鼠品系中900多个基因的14，000个注释外显子和边界，这些小鼠品系是在针对小鼠11号染色体的N-乙基-N-亚硝基脲（ENU）突变筛选中分离的。在31个突变株系的55个基因中鉴定出59个序列变异。39%的病变位于编码序列中，主要产生错义突变。其他61%位于非编码区，其中许多是高度保守的序列。在围产期致死线l11 Jus 13的病变改变了核氧还蛋白（Nxn）的共识剪接位点，插入10个氨基酸到所得的蛋白质。我们的结论是，点突变可以准确和灵敏地恢复大规模测序，保守的非编码区应包括疾病突变鉴定。我们报告的候选基因中只有7个先前在小鼠或大鼠中被突变靶向，这表明尽管正在努力对哺乳动物基因组中的基因进行功能注释，但表型和功能之间仍然存在巨大差距。我们的数据表明，疾病突变识别的经典位置映射方法可以使用高通量测序扩展到大的靶区域。在这里，我们表明可以在大量DNA序列数据中发现微小的DNA损伤，类似于大海捞针。这些病变确定了许多与出生缺陷、不育和生长相关的疾病基因的新候选者。此外，我们的数据表明，我们对哺乳动物基因的作用知之甚少。测序方法变得更便宜和更快。因此，我们在这里首次展示的战略将变得司空见惯。
An accurate and precisely annotated genome assembly is a fundamental requirement for functional genomic analysis. Here, the complete DNA sequence and gene annotation of mouse Chromosome 11 was used to test the efficacy of large-scale sequencing for mutation identification. We re-sequenced the 14,000 annotated exons and boundaries from over 900 genes in 41 recessive mutant mouse lines that were isolated in an N-ethyl-N-nitrosourea (ENU) mutation screen targeted to mouse Chromosome 11. Fifty-nine sequence variants were identified in 55 genes from 31 mutant lines. 39% of the lesions lie in coding sequences and create primarily missense mutations. The other 61% lie in noncoding regions, many of them in highly conserved sequences. A lesion in the perinatal lethal line l11Jus13 alters a consensus splice site of nucleoredoxin (Nxn), inserting 10 amino acids into the resulting protein. We conclude that point mutations can be accurately and sensitively recovered by large-scale sequencing, and that conserved noncoding regions should be included for disease mutation identification. Only seven of the candidate genes we report have been previously targeted by mutation in mice or rats, showing that despite ongoing efforts to functionally annotate genes in the mammalian genome, an enormous gap remains between phenotype and function. Our data show that the classical positional mapping approach of disease mutation identification can be extended to large target regions using high-throughput sequencing. Here we show that tiny DNA lesions can be found in huge amounts of DNA sequence data, similar to finding a needle in a haystack. These lesions identify many new candidates for disease genes associated with birth defects, infertility, and growth. Further, our data suggest that we know very little about what mammalian genes do. Sequencing methods are becoming cheaper and faster. Therefore, our strategy, shown here for the first time, will become commonplace.
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影响因子：
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作者：
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