Assessment of the latest NGS enrichment capture methods in clinical context.
Assessment of the latest NGS enrichment capture methods in clinical context.
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DOI：
10.1038/srep20948
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发表时间：
2016-02-11
影响因子：
4.6
通讯作者：
Roux AF
中科院分区：
文献类型：
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作者：
García-García G;Baux D;Faugère V;Moclyn M;Koenig M;Claustres M;Roux AF
NGS的富集捕获方法得到了广泛使用，但是，它们发展迅速，在转移到诊断服务之前，有必要定期衡量它们的优缺点。我们评估了最近发布的两种用于NGS的定制DNA溶液捕获富集方法，即Illumina NRCCE和Agilent SureSelectQXT，与参考方法NimbleGen SeqCap EZ Choice在相似的基因面板上，共享678 kb和110个基因。对于三种方法中的每一种，使用相同的24名患者（患有感觉神经性疾病）进行了两次Illumina MiSeq运行，每次运行12个样品。对技术成果进行了计算和比较，包括覆盖的深度和均匀度、目标区域的富集、在GC丰富区域的性能以及生成一致的变异数据集的能力。虽然我们表明这三种方法产生了用于标准DNA变体发现的合适数据集，但我们描述了上述参数的结果之间的显著差异。NimbleGen提供了最好的覆盖深度和均匀度，而NRCCE显示出最高的目标水平，但重复率很高。SureSelectQXT的整体质量接近NimbleGen。新方法显示出减少的制备时间，但表现不同。这些发现将指导实验室选择文库富集方法。
Enrichment capture methods for NGS are widely used, however, they evolve rapidly and it is necessary to periodically measure their strengths and weaknesses before transfer to diagnostic services. We assessed two recently released custom DNA solution-capture enrichment methods for NGS, namely Illumina NRCCE and Agilent SureSelectQXT, against a reference method NimbleGen SeqCap EZ Choice on a similar gene panel, sharing 678 kb and 110 genes. Two Illumina MiSeq runs of 12 samples each have been performed, for each of the three methods, using the same 24 patients (affected with sensorineural disorders). Technical outcomes have been computed and compared, including depth and evenness of coverage, enrichment in targeted regions, performance in GC-rich regions and ability to generate consistent variant datasets. While we show that the three methods resulted in suitable datasets for standard DNA variant discovery, we describe significant differences between the results for the above parameters. NimbleGen offered the best depth of coverage and evenness, while NRCCE showed the highest on target levels but high duplicate rates. SureSelectQXT showed an overall quality close to that of NimbleGen. The new methods exhibit reduced preparation time but behave differently. These findings will guide laboratories in their choice of library enrichment approach.