Circulating tumor DNA (ctDNA) is emerging as a promising biomarker for cancer diagnosis. However, the system to detect gene mutations with very low frequencies from plasma remains to be established in terms of technical aspects of sequencing technologies and cost for universal use. One strategy is to employ a cancer sequencing panel to detect mutations in a primary tumor in a time- and cost-effective manner, and subsequently assess these mutations with a digital PCR technology from plasma ctDNA. This strategy enables the accurate detection of low frequency mutations (i.e., less than 1% allele frequency) from ctDNA, since both comprehensive coverage of genes and quantitative mutation detection with very low frequencies are required for cancer diagnosis from plasma samples. Here, we present a pipeline can be used to detect mutations from plasma ctDNA with very low allele frequencies using a next-generation sequencing technology for comprehensive coverage of primary tumors and droplet digital PCR for sensitive detection from plasma ctDNA.
Keywords: Allele frequency; Circulating tumor DNA; Colorectal cancer; Digital PCR; Gastrointestinal tract cancer; Next-generation sequencer; Sequencing panel.