GSM2341634: IMR90-GFP-DamID; Homo sapiens; OTHER
For DamID-Seq,  cells were harvested and gDNA was isolated using the Qiagen DNeasy Blood & Tissue Kit. gDNA was  digested with DpnI (NEB, 20 U/ μl) and ligated with the adaptor AdR, which was made by mixing and slowly annealing AdR-top (5’ CTAATACGACTCACTATAGGGCAGCGTGGTCGCGGCCGAGGA 3’) and AdR-bottom (5’ TCCTCGGCCG 3’). This ligation was completed using 1uL T4 Ligase (Roche, 5U/μl) for 2 hours at 16°C. The regions flanked by adaptors were amplified by PCR using the Bio-Adr-primer (5’ Bio-GGTCGCGGCCGAGGATC 3’) and purified using the Qiagen MinElute PCR purification kit. Amplified products were sonicated using a Bioruptor (Diagenode) to obtain DNA fragments of sizes between 100-500bp. MyOneC1 Streptavidin dynabeads (Invitrogen) were used to pull down the biotinylated ends of the sonicated PCR products. The bound DNA was removed off the beads by digestion with DpnII for 1 hour. ChIP-Seq and DamID-Seq libraries were generated using the TruSeq ChIP sample Prep kit (Illumina) or the Kapa Hyper Prep Kit for Illumina Platforms followed by Illumina sequencing.
GEO Accession
GSM2341634