The Centre for Applied Genomics, Genetics and Genome Biology, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada.
The Centre for Applied Genomics, Genetics and Genome Biology, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
Am J Hum Genet. 2018 Jan 4;102(1):142-155. doi: 10.1016/j.ajhg.2017.12.007.
A remaining hurdle to whole-genome sequencing (WGS) becoming a first-tier genetic test has been accurate detection of copy-number variations (CNVs). Here, we used several datasets to empirically develop a detailed workflow for identifying germline CNVs >1 kb from short-read WGS data using read depth-based algorithms. Our workflow is comprehensive in that it addresses all stages of the CNV-detection process, including DNA library preparation, sequencing, quality control, reference mapping, and computational CNV identification. We used our workflow to detect rare, genic CNVs in individuals with autism spectrum disorder (ASD), and 120/120 such CNVs tested using orthogonal methods were successfully confirmed. We also identified 71 putative genic de novo CNVs in this cohort, which had a confirmation rate of 70%; the remainder were incorrectly identified as de novo due to false positives in the proband (7%) or parental false negatives (23%). In individuals with an ASD diagnosis in which both microarray and WGS experiments were performed, our workflow detected all clinically relevant CNVs identified by microarrays, as well as additional potentially pathogenic CNVs < 20 kb. Thus, CNVs of clinical relevance can be discovered from WGS with a detection rate exceeding microarrays, positioning WGS as a single assay for genetic variation detection.
全基因组测序（WGS）成为一线遗传检测方法的一个遗留障碍是准确检测拷贝数变异（CNVs）。在这里，我们使用了几个数据集，通过基于读取深度的算法，从短读 WGS 数据中经验性地开发了一个用于识别种系 CNVs>1 kb 的详细工作流程。我们的工作流程是全面的，它解决了 CNV 检测过程的所有阶段，包括 DNA 文库制备、测序、质量控制、参考映射和计算 CNV 识别。我们使用该工作流程来检测自闭症谱系障碍（ASD）个体中的罕见基因 CNVs，使用正交方法测试的 120/120 个此类 CNVs 均成功得到确认。我们还在该队列中鉴定了 71 个推定基因从头 CNVs，其确认率为 70%；其余的由于先证者中的假阳性（7%）或父母中的假阴性（23%）而被错误地鉴定为从头。在进行了 ASD 诊断的个体中，同时进行了微阵列和 WGS 实验，我们的工作流程检测到了微阵列识别的所有临床相关 CNVs，以及额外的<20 kb 的潜在致病性 CNVs。因此，具有临床相关性的 CNVs 可以通过 WGS 以超过微阵列的检测率发现，使 WGS 成为一种用于遗传变异检测的单一检测方法。
