Supplementary data: Quantifying sequencing error and effective sequencing depth of liquid biopsy NGS with UMI error correction
Supplementary Figure 1. Ratio of forward-to-reverse reads is no useful quality criterion for UMI-libraries after error-correction bioinformatics. The density plots show the position balance of forward reads to reverse reads at each genomic position, from targeted sequencing of the PIK3CA coding regions. The position balance was computed with GenSearchNGS, with 1 being optimal and 0 being worst. (A) Position balance in the four non-UMI-libraries (TruSeq 776-1, 776-2, 779-1, 779-2). The majority of genomic positions have a near-optimal read balance. (B) Position balance in the two UMI-libraries (Oncology 776, 779). Nearly all positions are covered by error-corrected reads oriented in just one genomic direction. The conventional quality check of forward/reverse balance is therefore no longer useful after UMI-based error-correction has been performed.
Supplementary Table 3: Deduplication (removal of redundant sequences) increases the allele frequency of signal noise (PCR or sequencing errors) and reduces the effective sequencing depth
Supplementary Table 2: Allelic frequency of PIK3CA p.E545K mutation for non-UMI libraries vs UMI libraries and bioinformatic non-filtering vs filtering, in wild-type standard HD776 (0.00%) and mutated standard HD779 (0.13%)
Supplementary Table 1: Bioinformatic tools, versions, and settings that were used