Characterisation and Validation of Insertions and Deletions in 173 Patient Exomes

Recent advances in genomics technologies have spurred unprecedented efforts in genome and exome re-sequencing aiming to unravel the genetic component of rare and complex disorders. While in rare disorders this allowed the identification of novel causal genes, the missing heritability paradox in complex diseases remains so far elusive. Despite rapid advances of next-generation sequencing, both the technology and the analysis of the data it produces are in its infancy. At present there is abundant knowledge pertaining to the role of rare single nucleotide variants (SNVs) in rare disorders and of common SNVs in common disorders. Although the 1,000 genome project has clearly highlighted the prevalence of rare variants and more complex variants (e.g. insertions, deletions), their role in disease is as yet far from elucidated. We set out to analyse the properties of sequence variants identified in a comprehensive collection of exome re-sequencing studies performed on samples from patients affected by a broad range of complex and rare diseases (N = 173). Given the known potential for Loss of Function (LoF) variants to be false positive, we performed an extensive validation of the common, rare and private LoF variants identified, which indicated that most of the private and rare variants identified were indeed true, while common novel variants had a significantly higher false positive rate. Our results indicated a strong enrichment of very low-frequency insertion/deletion variants, so far under-investigated, which might be difficult to capture with low coverage and imputation approaches and for which most of study designs would be under-powered. These insertions and deletions might play a significant role in disease genetics, contributing specifically to the underlining rare and private variation predicted to be discovered through next generation sequencing.近年来，基因组学技术的飞速发展推动了基因组与外显子组重测序领域的空前研究热潮，其核心目标是阐明罕见病与复杂疾病的遗传机制。尽管针对罕见病的此类研究已成功鉴定出多个全新的致病基因，但复杂疾病的缺失遗传度悖论至今仍未得到破解。尽管下一代测序（next-generation sequencing）技术发展迅猛，但其本身及所产生数据的分析方法仍处于起步阶段。目前学界已对罕见病中罕见单核苷酸变异（single nucleotide variants, SNVs）以及常见病中常见SNVs的致病作用形成了较为充分的认知。尽管千人基因组计划（1,000 Genome Project）已明确证实了罕见变异及更复杂变异（如插入、缺失序列）的普遍性，但这类变异在疾病发生中的作用至今仍未被完全阐明。本研究旨在对一项综合外显子组重测序研究队列中的序列变异特征进行分析，该队列纳入了173名（N=173）罹患多种复杂疾病与罕见病的患者样本。鉴于功能丧失变异（Loss of Function, LoF）存在较高的假阳性检出风险，我们对所鉴定出的常见、罕见及私有LoF变异进行了大规模验证。结果显示，绝大多数私有与罕见变异均为真实存在的变异，而新发常见变异的假阳性率则显著更高。本研究结果显示，极低频率插入/缺失变异存在显著富集现象——这类变异迄今尚未得到充分研究，且低覆盖度测序与基因型填充方法难以有效捕获，多数研究设计对其检测效力亦显不足。这些插入与缺失序列可能在疾病遗传学中发挥重要作用，特别是针对下一代测序技术有望发现的潜在罕见与私有变异，其贡献尤为突出。