GSM5397981: BMSC_T7E_IP_MC; Homo sapiens; MeDIP-Seq
Aliquots of purified genomic DNA from each situation of MSCs were first sonicated to ∼250 bp using Bioruptor (Diagenode, Seraing, Belgium), then end-repaired and ligated to Illumina PE adaptors using NEB Next DNA Library Prep Master Mix Set (New England Biolabs, Beverly, MA, USA). In DIP assays, 10 μg of the adaptor-ligated genomic DNA was used as input, and 5 μl of anti-5-methylcytosine antibody、anti-TET1 or anti-PARP1 antibodies was added to immunoprecipitate modified DNA. Each of the immunoprecipitated DNA was amplified with forward primer and reverse primers in a 50 μl PCR reaction with 0.2× Sybr-Green I and 1× Phusion High-fidelity PCR Master Mix (New England Biolabs, Beverly, MA, USA). PCR products that contain barcodes were sequenced after the standard first read using N2IndSeq primer. After base-calling, each sequence was decoded using an in-house Perl script which required at least five out of six matching positions in the barcode sequence. All sequencing reads were trimmed from 5' end to obtain final 37 bases in length for further analysis.