"Pre-treatment ctDNA ULP-WGS in a patient with small cell lung cancer, liver lesion"
"The DSP Circulating DNA kit from Qiagen was utilized to extract cell-free DNA from aliquots of plasma which were eluted into 40-80 uL of re-suspension buffer using the Qiagen Circulating DNA kit on the QIAsymphony liquid handling system. Library preparation utilized the Kapa Hyper Prep kit with custom adapters (IDT and Broad Institute). Samples were sequenced to meet a goal of 0.1x mean coverage utilizing the Laboratory Picard bioinformatics pipeline and Illumina instruments were used for all of ctDNA sequencing with 150 base pair and paired-end sequencing. Library construction was performed as previously described (Fisher et al. 2011). Kapa HyperPrep reagents in 96-reaction kit format were used for end repair/A-tailing, adapter ligation, and library enrichment PCR.  After library construction, hybridization and capture were performed using the relevant components of Illumina's Nextera Exome Kit and following the manufacturer's suggested protocol. Cluster amplification of DNA libraries was performed according to the manufacturer's protocol (Illumina) using exclusion amplification chemistry and flowcells. Flowcells were sequenced utilizing Sequencing-by-Synthesis chemistry. Each pool of whole exome libraries was sequenced on paired 76 cycle runs with two 8 cycle index reads across the number of lanes needed to meet coverage for all libraries in the pool."
NIH-phs002089
S3