GSM2340146: PF negative 5; Homo sapiens; Bisulfite-Seq
Genomic DNA was extracted from the cell lines using standard phenol:chloroform extraction followed by ethanol precipitation. ERRBS libraries were prepared by digesting 75 ng of DNA with restriction enzyme MspI, followed by post end-repair, A-tailing, and adapter ligation with Illumina TruSeq adapters. The region corresponding to 84-334bp was size-selected as two fractions, and each fraction was subjected to overnight bisulfite conversion (55 cycles of 95 °C for 30 sec, 50 °C for 15 min) using EZ DNA methylation kit (Cat # D5002, Zymo Research, Irvine CA). Purified bisulfite converted DNA was PCR amplified (using TruSeq primers, Illumina Inc. San Diego, CA) for 18 cycles of denaturing, annealing and extension/elongation steps using Roche FastStart (cat # 03 553 361 001):  94 °C for 20 secs, 65 °C for 30 secs, 72 °C for 1 min, followed by 72 °C for 3 min. The resulting libraries were normalized to 2nM and pooled at the same molar ratio.
GEO Accession
GSM2340146