GSM6964886: WGBS_DNMT_WT_EpiLC_D1_rep2; Mus musculus; Bisulfite-Seq
As an input for WGBS 400 ng of genomic DNA was fragmented to between 300 and 400 bp using a Covaris E220 water bath sonicator following manufacturer's instructions. The DNA ends were repaired using NEBNext Ultra II Library Prep (New England Biolabs, E7645) reagents and methylated adapters were annealed according to the manufacturer's instructions. Fragments were purified using Agencourt Ampure XP beads (Fisher Scientific, 10136224) and bisulfite converted using a Zymo EZ Methylation Gold kit (Zymo, D5005) according to the manufacturer's instructions and eluting the converted DNA in 15µL of provided elution buffer. Bisulfite treated DNA was amplified using NEBNext index primers (NEB, E7335) and KAPA HiFi Uracil+ mastermix (Roche, KK2801/2) using the following protocol: 1 minute at 98ºC, 10 cycles of 15 seconds at 98ºC, 30 seconds at 65ºC, 30 seconds at 72ºC followed by a final extension for 1 minute at 72ºC. Libraries purified using 0.8x volumes of Agencourt Ampure XP beads (Fisher Scientific, 10136224) to remove contaminating adapters.