GSM7990608: WGBS_LOS_#2; Bos indicus x Bos taurus; Bisulfite-Seq
Bovine fibroblast cells were lysed in lysis buffer (0.05 M Tris-HCl (pH 8.0), 0.1 M EDTA, and 0.5% (w/v) SDS) with proteinase K (Fisher BioReagents, BP1700) at 55°C for four hours. Genomic DNA was extracted with Phenol:Chloroform:Isoamyl Alcohol (SIGMA, P3803) following the manufacturer's instructions. The concentration of DNA was measured by using a NanoDrop® ND-1000 Spectrophotometer and DNA integrity was confirmed by electrophoresis on a 0.7% agarose gel. Genomic DNA samples were stored at -20°C. WGBS for fibroblast cells was conducted by CD Genomics (Shirley, NY, USA). Information on library preparation and sequencing obtained from the company is as follows: Genomic DNA was fragmented by sonication to a mean size of approximately 200-400 bp. Fragmented DNA was end-repaired, 5'-phosphorylated, 3'-dA-tailed and then ligated to methylated adapters. The methylated adapter-ligated DNAs were purified using 0.8× Agencourt AMPure XP magnetic beads and subjected to bisulfite conversion by ZYMO EZ DNA Methylation-Gold Kit (Zymo Research). The converted DNAs then have the addition of 8-bp index primers to create the final cDNA library. Equimolar pooling of libraries was performed based on QC values and sequenced on Novaseq6000 sequencer (Illumina) with a read length configuration of 150 PE for 300M PE reads per sample (300M in each direction). 0.1-1% lambda DNA were added during the library preparation to monitor bisulfite conversion rate.