cfDNA extraction, library construction, and targeted sequencing
tumor
BLOOD, TUMOR
Patient blood samples were collected and centrifuged to separate peripheral blood lymphocytes from plasma. Genomic DNA from peripheral blood lymphocytes and cell-free DNA (cfDNA) from plasma were extracted using the RelaxGene Blood DNA System (TianGen Biotech Co., Ltd., Beijing, China) and the QIAamp Circulating Nucleic Acid Kit (Qiagen, Hilden, Germany), respectively. Circulating tumor DNA (ctDNA) was included in cfDNA.Targeted capture was achieved using a panel of biotinylated DNA probes which cover 605 cancer-related genes (HapOncoCDx™, Roche). Amplified sample libraries and the SeqCap EZ Library were hybridized according to the manufacturer’s protocol. The captured libraries were sequenced using PE150 paired-end sequencing on the NovaSeq 6000 system (Illumina, Inc., San Diego, CA, USA).