title: GSM4088840: HCT116_DNMT3B_KO; Homo sapiens; Bisulfite-Seq
library_construction_protocol: Genomic DNA was extracted from cells using DNeasy Blood & Tissue Kit according to the manufacturer's protocol with some modifications. RNase A/T1 Cocktail (Ambion AM2286) was added to proteinase K and samples were incubated at 37°C for 1hr to remove RNA. At the final step, DNA was eluted with ddH2O instead of AE buffer. DNA was quantified by Nanodrop and Qubit. 200ng of purified DNA samples were processed using the Ovation RRBS Methyl-Seq system kit (NuGen Technologies) according to instructions with modifications. Briefly, 0.5ng unmethylated phage λ DNA (NEB) was spiked into each sample to allow assessment of bisulfite conversion efficiency. The methylation-insensitive restriction enzyme MspI was then used to digest the genomic DNA, and digested fragments were ligated to adapters. Adapter-ligated fragments were then repaired before bisulfite conversion with the Qiagen Epitect Fast Bisulfite Conversion kit. Bisulfite-treated adapter-ligated fragments were amplified by 9 cycles of PCR and purified using Agencourt RNAClean XP beads. Libraries were quantified using the Qubit dsDNA HS assay and assessed for size and quality using the Agilent Bioanalyzer DNA HS kit. Library preparation and sequencing was performed by the Edinburgh Clinical Research Facility.
attributes.tag: GEO Accession
attributes.value: GSM4088840