Instrument: Illumina HiSeq 1500
Strategy: ChIP-Seq
Source: GENOMIC
Selection: ChIP
Layout: SINGLE
Construction protocol: Chromatin was prepared from 150 mm culture dishes first by crosslinking in 1% formaldehyde in PBS (10 minutes, RT). Cross-linking was stopped by adding glycine to a final concentration of 0.125 M (10 minutes, RT). The cells were washed twice in ice-cold PBS, and harvested in ice-cold lysis buffer (0.1% SDS, 1% Triton X-100, 0.15 M NaCl, 1 mM EDTA, 20 mM Tris pH=8) and sonicated at high setting in a Bioruptor-Twin (Diagenode) at a volume of 1.5 ml in 15-mL tubes for 40 cycles of 30 seconds on and 30 seconds off. The chromatin IPs were performed as described in (Nielsen R, et al. 2014. Methods in Enzymology 537: 261-279). Chromatin used for ChIP-seq of MED1 was also cross-linked in 2 mM disuccinimidyl glutarate (DSG) for 20 minutes at RT prior to cross-linking with formaldehyde. DNase-seq and ChIP-seq libraries were constructed from 10 to 20 ng of genomic DNA according to the manufacturer's instructions (Illumina) as described in (Nielsen R, et al. 2014. Methods in Enzymology 537: 261-279).