GSM5138756: WGBS_EpiSC_WT_Rep2; Mus musculus; Bisulfite-Seq
Genomic DNA was extracted from cells using the DNeasy Blood and Tissue kit (QIAGEN, 69506) following the manufacturer's instructions. The whole-genome bisulfite-seq library was prepared starting from 5 μg of sonicated genomic DNA for 2 runs of 10 cycles [30 sec ''ON'', 30 sec ''OFF'']. In order to obtain 200 nt fragments. Sonicated DNA was then end-repaired twice, dA-tailed, and ligated to adapters, using the Illumina TruSeq DNA Sample Prep Kit, following manufacturer's instructions. Adapter-ligated DNA was loaded on an EGel Size select 2% agarose pre-cast gel (Invitrogen), and a fraction corresponding to fragments ranging from 25 0bp to 350bp was recovered. Purified DNA was then subjected to bisulfite conversion using the EpiTect Bisulfite Kit (QIAGEN) following the manufacturer's instruction except for the double desulfonation after the conversion. Bisulfite-converted DNA was finally enriched by 15 cycles of PCR using Pfu Turbo Cx. HotStart Taq (Agilent). Libraries were sequenced on Illumina Novaseq 6000 System.