PCR Free Whole Genome of COLO829 an immortal cell line derived from a metastasis from a cutaneous melanoma male patient: Sample COLO829_GSC
Library preparation and sequencing. To minimize genome library bias and coverage gaps associated with PCR amplification of high GC or AT-rich regions, we implemented an automated version of the TruSeq DNA PCR-free kit (FC-121-1002, Illumina Inc.). Briefly, one microgram of high molecular weight genomic DNA was arrayed in a 96-well microtitre plate and subjected to shearing to 300-600bp by sonication for 30 seconds (Covaris). Sheared DNA was end-repaired and size selected using AMPure XP beads targeting a 300-400bp fraction. After 3’ A-tailing, full length TruSeq adapters were ligated. Libraries were purified using AMPure XP beads and fragment sizes were assessed using an aliquot of PCR amplified library DNA on an Agilent 2100 Bioanalyzer DNA1000 chip, or Caliper GX DNA1000 chip. The PCR-free library concentration was quantified using a qPCR Library Quantification kit (KAPA, KK4824). Tumor and normal genome libraries were sequenced with paired 125bp reads using Illumina HiSeq2500 V4 chemistry running HCS2.2 controller software and RTA 1.18.61. The five lanes of each library generated 2.819 billion reads for the tumor and 2.968 billion reads for the normal.