外显子组测序
再现性
计算生物学
外显子组
生物
基因组
DNA测序
全基因组测序
突变
遗传学
癌症基因组测序
基因
数学
统计
作者
Wenming Xiao,Luyao Ren,Zhong Chen,Li Tai Fang,Yongmei Zhao,Justin Lack,Meijian Guan,Bin Zhu,Erich Jaeger,Liz Kerrigan,Thomas Blomquist,Tiffany Hung,Marc Sultan,Kenneth B. Idler,Charles Lu,Andreas Scherer,Rebecca Kusko,Malcolm Moos,Chunlin Xiao,Stephen T. Sherry
标识
DOI:10.1038/s41587-021-00994-5
摘要
Clinical applications of precision oncology require accurate tests that can distinguish true cancer-specific mutations from errors introduced at each step of next-generation sequencing (NGS). To date, no bulk sequencing study has addressed the effects of cross-site reproducibility, nor the biological, technical and computational factors that influence variant identification. Here we report a systematic interrogation of somatic mutations in paired tumor–normal cell lines to identify factors affecting detection reproducibility and accuracy at six different centers. Using whole-genome sequencing (WGS) and whole-exome sequencing (WES), we evaluated the reproducibility of different sample types with varying input amount and tumor purity, and multiple library construction protocols, followed by processing with nine bioinformatics pipelines. We found that read coverage and callers affected both WGS and WES reproducibility, but WES performance was influenced by insert fragment size, genomic copy content and the global imbalance score (GIV; G > T/C > A). Finally, taking into account library preparation protocol, tumor content, read coverage and bioinformatics processes concomitantly, we recommend actionable practices to improve the reproducibility and accuracy of NGS experiments for cancer mutation detection. Recommendations are given on optimal read coverage and selection of calling algorithm to maximize the reproducibility of cancer mutation detection in whole-genome or whole-exome sequencing.
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