Seqcap ez human exome library v2.0 kit

Genomic DNA was isolated from 24 tissues using the Puregene™ DNA purification kit (Qiagen, Venlo, Netherlands). Library construction and targeted exome enrichment were performed using the Illumina TruSeq DNA Sample Prep Kit (San Diego, CA, USA) and the SeqCap EZ Human Exome Library v2.0 kit (Roche NimbleGen, WI, USA), respectively. Next, paired-end sequencing was performed on the Illumina HiSeq 2000 sequencing instrument, according to the manufacturer's instructions, yielding ~100 bp-sized short sequencing reads. The sequencing reads were aligned on human reference genome 19 using Burrows Wheelers Aligner [34 (link)], and duplicates reads were removed using Picard (Broad Institute). Then, the remaining reads were calibrated and realigned using Genome Analysis Toolkit [35 (link)]. The realigned BAM files were analyzed using MuTect [29 (link)] and Strelka [36 (link)] to detect somatic single-nucleotide variants and insertions/deletions, respectively. Coding variants were selected by dbNSFP annotation [37 (link)] and germline variants were filtered out by dbSNP database (dbSNP version 132). SciClone was used to infer tumor clonality [38 (link)], and ASCAT v2.1 was used to estimate tumor purity [39 (link)]. We run all these programs under the default parameter settings.