Optimizing Cancer Genome Sequencing and Analysis
Optimizing Cancer Genome Sequencing and Analysis
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DOI：
10.1016/j.cels.2015.08.015
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发表时间：
2015-09-23
期刊：
影响因子：
9.3
通讯作者：
Wilson, Richard K.
中科院分区：
文献类型：
--
作者：
Griffith, Malachi;Miller, Christopher A.;Wilson, Richard K.
肿瘤通常被测序到75 x-100 x（外显子组）或30 x-50 x（全基因组）的深度。我们证明，目前的测序模式是不充分的肿瘤是不纯的，非整倍体，或克隆异质性。为了重新评估最佳测序策略，我们对原发性急性髓性白血病、其后续复发和匹配的正常皮肤样本进行了超深（高达312倍）全基因组测序和外显子组捕获（高达433倍）。我们测试了多种比对和变异识别算法，并通过对200，000个推定SNV进行测序至类似于1，000 x的深度来验证它们。额外的靶向测序提供了超过10，000 x的覆盖率，ddPCR测定提供了高达250，000 x的选定位点采样。我们评估了不同文库生成方法、测序深度和分析策略对有效表征复杂肿瘤的能力的影响。该数据集代表了迄今为止描述的最全面的肿瘤测序，将作为宝贵的社区资源（dbGaP：phs 000159）。
Tumors are typically sequenced to depths of 75x-100x (exome) or 30x-50x (whole genome). We demonstrate that current sequencing paradigms are inadequate for tumors that are impure, aneuploid, or clonally heterogeneous. To reassess optimal sequencing strategies, we performed ultra-deep (up to similar to 312x) whole genome sequencing and exome capture (up to similar to 433x) of a primary acute myeloid leukemia, its subsequent relapse, and a matched normal skin sample. We tested multiple alignment and variant calling algorithms and validated similar to 200,000 putative SNVs by sequencing them to depths of similar to 1,000x. Additional targeted sequencing provided over 10,000x coverage and ddPCR assays provided up to similar to 250,000x sampling of selected sites. We evaluated the effects of different library generation approaches, depth of sequencing, and analysis strategies on the ability to effectively characterize a complex tumor. This dataset, representing the most comprehensively sequenced tumor described to date, will serve as an invaluable community resource (dbGaP: phs000159).