title: GSM8333112: HEK293T_WT_WGBS; Homo sapiens; Bisulfite-Seq
library_construction_protocol: DNA was isolated using the Cell Culture DNA Midi Kit from QIAGEN (Qiagen, 13343). Prior to library construction, 1µg sample genome DNA mixed with unmethylated lambda DNA was fragmented using a Bioruptor system to a mean size of approximately 250bp After fragmentation, the purified randomly fragmented DNA was subsequently treated with a mix of T4 DNA polymerase, Klenow Fragment and T4 polynucleotide kinase to repair, blunt and phosphorylate the ends. The blunt DNA fragments were subsequently 3' adenylated using Klenow Fragment (3'-5' exo-) (Enzymatics, P7010-LC-L) and following with ligation to adaptors synthesized with 5'-methylcytosine instead of cytosine using T4 DNA Ligase. Then, the DNA was purified using QIAquick PCR purification kit (Qiagen, 28104). After that, ZYMO EZ DNA Methylation-Gold Kit™ (ZYMO, D5005) was employed to convert unmethylated cytosine into uracil according to the instructions. Finally, the WGBS libraries were generated by PCR amplification using JumpStartTM Taq DNA Polymerase (Sigma, D4184-50UN) and purification using QIAquick PCR purification kit (Qiagen, 28104).