DNA sequencing of serial diluted reference genomes with replicates in three Next Generation sequencing platforms. Homo sapiens

Accurate detection of somatic mutations with low variant allele fraction (VAF, 1) Serially diluted gDNA mixture dataGenomic DNA from two independent blood samples was mixed to mimic somatic mutations at four different VAFs: 0.5%, 1%, 5%, and 10%. Mixtures are then sequenced with three distinct platforms: hybridization-capture-based Illumina sequencing (ILH) and amplicon-based Illumina and Ion-Torrent sequencing (ILA and ITA, respectively). Sequencing libraries from each level were sequenced twice to generate data of sequencing replicates (designated as X12). For low-level mixtures with the VAF of 0.5% and 1%, two library-level replications were additionally prepared and sequenced (X21 and X31) to compare the effect of sequencing and library-level replication.2) Commercial reference standard dataTru-Q7 reference standard (1.3% Tier, HD734, Horizon Dx, Cambridge, UK) was sequenced with two different platforms: hybridization-capture-based Illumina sequencing (ILH) and amplicon-based Ion-Torrent sequencing (ITA). Widely-used cancer panels were used (SureSelect custom panel and Ion AmpliSeq cancer hotspot panel v2) to generate sequence data. Triplicate of library-level replication were made and sequenced with matched control (wild-type reference, HD752). Each library-level replicate data is designated as rep1, rep2, and, rep3, respectively.