Multiplex Target Enrichment Using DNA Indexing for Ultra-High Throughput SNP Detection
Multiplex Target Enrichment Using DNA Indexing for Ultra-High Throughput SNP Detection
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DOI：
10.1093/dnares/dsq029
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发表时间：
2011-02-01
期刊：
影响因子：
4.1
通讯作者：
Morris, Derek W.
中科院分区：
文献类型：
--
作者：
Kenny, Elaine M.;Cormican, Paul;Morris, Derek W.
在多个DNA样品中筛选大量靶区域的序列变异是下一代测序的重要应用，但平行富集样品的有效方法尚未报道。我们描述了一种先进的方法，该方法在目标富集之前使用索引或条形码结合DNA样品，以促进这种类型的实验。将用于多个单独DNA样品的测序文库（每个都掺入独特的6-bp索引）以等量组合，使用单个溶液中靶富集测定富集并在单个反应中测序。基于索引解析序列读数，从而允许对单个样品进行序列分析。我们表明，使用索引的样品不影响富集反应的效率。对于三索引和九索引的HapMap DNA样品，发现该方法对于SNP鉴定是高度准确的。即使序列覆盖率低至8x，99%的序列SNP调用与已知基因型一致。在一个单一的实验中，这种方法可以测序的外显子区域的数百个基因在几十个样品的序列和结构变异，使用少至1 μ g的输入DNA每个样品。
Screening large numbers of target regions in multiple DNA samples for sequence variation is an important application of next-generation sequencing but an efficient method to enrich the samples in parallel has yet to be reported. We describe an advanced method that combines DNA samples using indexes or barcodes prior to target enrichment to facilitate this type of experiment. Sequencing libraries for multiple individual DNA samples, each incorporating a unique 6-bp index, are combined in equal quantities, enriched using a single in-solution target enrichment assay and sequenced in a single reaction. Sequence reads are parsed based on the index, allowing sequence analysis of individual samples. We show that the use of indexed samples does not impact on the efficiency of the enrichment reaction. For three- and nine-indexed HapMap DNA samples, the method was found to be highly accurate for SNP identification. Even with sequence coverage as low as 8x, 99% of sequence SNP calls were concordant with known genotypes. Within a single experiment, this method can sequence the exonic regions of hundreds of genes in tens of samples for sequence and structural variation using as little as 1 mu g of input DNA per sample.