Here, we compared the Twist exome capture kit’s coding sequence coverage and SNV detection sensitivity to other widely used exome kits as well as to SR- and LR-WGS. In the last few years, new exome capture and sequencing technologies, particularly the Twist exome capture kit and long read sequencing (LRS) technologies, have been applied in clinical sequencing studies. However, lower sequencing and storage costs as well as the demonstration of diagnostic yield of CNV detection have led WES to be proposed as a first-tier diagnostic test in recent studies. Short- and long-read whole genome sequencing (SR-WGS and LR-WGS, respectively) approaches generally yield more uniform and complete coverage profiles than exome sequencing, and the gapless nature of WGS data enables more accurate detection of CNVs and structural variants (SVs). Therefore, over- and underrepresentation of target regions due to extreme GC content and mappability issues can dramatically affect the robustness of CNV calling from exome data. Large sets of reference samples are typically required in order to robustly compare CNV coverage profiles in exome data. CNV detection from WES data particularly fully depends on the analysis of read depth variations at sequencing targets. While CNVs are not routinely detected from WES in each laboratory or pipeline, their additional clinical utility urges for reliable CNV detection from exomes, especially when patient cohorts are not routinely pre-screened by CNV-microarrays. Sufficient, uniform and reproducible/consistent sequence coverage is required for robust and sensitive single-nucleotide variant (SNV) and copy number variant (CNV) detection in exome data. In these comparative studies, extreme GC content and mappability issues are shown to be the major sources of coverage bias. Several studies that compared exome capture technologies have shown that there are major differences in their performance and that high average read depth does not guarantee coverage for individual targets. These characteristics may give rise to differences in the overall coverage uniformity and capture efficiency of specific targets, resulting in decreased variant calling sensitivity. Selection of target genomic regions, sequence features, length of probes and exome capture mechanisms are the major differences among these kits. There are various exome capture kits with different target enrichment strategies. With the improvements in targeted sequencing approaches, whole exome sequencing (WES) has become a standard tool in clinical diagnostics. Next-generation sequencing (NGS) techniques are widely used across clinical and research applications in genetics. We conclude that exome sequencing with Twist represents a significant improvement and could be performed at lower sequence coverage compared to other exome capture techniques. Additionally, we show that even at a reduced average coverage of 70× there is only minimal loss in sensitivity for SNV and CNV detection. Twist performance is comparable to that of both short- and long-read whole genome sequencing. We show that the Twist exome capture significantly improves complete coverage and coverage uniformity across coding regions compared to other exome capture kits. We compared three different widely used enrichment kits (Agilent SureSelect Human All Exon V5, Agilent SureSelect Human All Exon V7 and Twist Bioscience) as well as short-read and long-read WGS. Here we compared the ability to obtain comprehensive exome coverage for recent exome capture kits and genome sequencing techniques. Sufficient, uniform and reproducible/consistent sequence coverage is a main determinant for the sensitivity to detect single-nucleotide (SNVs) and copy number variants (CNVs). Exome and genome sequencing are the predominant techniques in the diagnosis and research of genetic disorders.
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