Exome-wide DNA capture and next generation sequencing in domestic and wild species.
Exome-wide DNA capture and next generation sequencing in domestic and wild species.
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DOI：
10.1186/1471-2164-12-347
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发表时间：
2011-07-05
期刊：
影响因子：
4.4
通讯作者：
Luikart G
中科院分区：
文献类型：
--
作者：
Cosart T;Beja-Pereira A;Chen S;Ng SB;Shendure J;Luikart G
基因靶向标记和全基因组标记通过提高我们对适应和物种形成的遗传过程的理解，对推进进化生物学、农业和生物多样性保护至关重要。不幸的是，对于具有大基因组的真核生物物种，获得基因组序列和开发基因组资源如全基因组SNP仍然是昂贵的。需要一种方法来允许基因靶向的下一代测序，其足够灵活以包括任何基因或任何数量的基因，而不像转录组测序。这种方法将允许对许多个体进行测序，避免在随后的群体遗传分析中的确定偏差。我们证明了最近的技术，外显子捕获，在没有基因组资源的物种全基因组，基因靶向标记发现的有用性。我们使用来自家养奶牛基因组序列（Bos taurus）的编码基因序列来捕获（富集）B的数千个外显子，并随后对其进行测序。taurus，B. B. indicus和野牛野牛（野生野牛）。我们的捕获阵列具有针对2，570个基因中的16，131个外显子的探针，其中包括203个具有已知功能的候选基因，以及与疾病和其他健康性状相关的感兴趣的基因。我们成功地对B中29条常染色体和X染色体的外显子序列进行了测序和作图。金牛座基因组序列外显子捕获和高通量测序鉴定了所有三个个体中均匀分布在所有参考染色体上的数千个推定SNP，包括我们靶向候选基因中的数百个SNP。这项研究表明，外显子捕获可以在许多个体和没有基因组资源的非模式物种中定制SNP发现。我们捕获的外显子组子集足够小，可以进行负担得起的下一代测序，并使用国内奶牛基因组作为参考，成功捕获了来自不同野生物种的外显子。
Gene-targeted and genome-wide markers are crucial to advance evolutionary biology, agriculture, and biodiversity conservation by improving our understanding of genetic processes underlying adaptation and speciation. Unfortunately, for eukaryotic species with large genomes it remains costly to obtain genome sequences and to develop genome resources such as genome-wide SNPs. A method is needed to allow gene-targeted, next-generation sequencing that is flexible enough to include any gene or number of genes, unlike transcriptome sequencing. Such a method would allow sequencing of many individuals, avoiding ascertainment bias in subsequent population genetic analyses. We demonstrate the usefulness of a recent technology, exon capture, for genome-wide, gene-targeted marker discovery in species with no genome resources. We use coding gene sequences from the domestic cow genome sequence (Bos taurus) to capture (enrich for), and subsequently sequence, thousands of exons of B. taurus, B. indicus, and Bison bison (wild bison). Our capture array has probes for 16,131 exons in 2,570 genes, including 203 candidate genes with known function and of interest for their association with disease and other fitness traits. We successfully sequenced and mapped exon sequences from across the 29 autosomes and X chromosome in the B. taurus genome sequence. Exon capture and high-throughput sequencing identified thousands of putative SNPs spread evenly across all reference chromosomes, in all three individuals, including hundreds of SNPs in our targeted candidate genes. This study shows exon capture can be customized for SNP discovery in many individuals and for non-model species without genomic resources. Our captured exome subset was small enough for affordable next-generation sequencing, and successfully captured exons from a divergent wild species using the domestic cow genome as reference.
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