GSM3723084: H3K27ac ChIP-seq [77_WT_Th0_IL6_H3K27ac]; Mus musculus; ChIP-Seq
Cells (10 x 10e6) were cross-linked for 10 min with 1% formaldehyde, quenched with Glycine, and washed. Cell pellets were resuspended in lysis buffer, and homogenized. Nuclei were resuspended in RIPA buffer and sonicated to a size range of 200-500 bp using Bioruptor. Cells were pelleted, RIPA buffer was added to supernatant to 1 ml, and appropriate antibody coupled to Dynabeads M-280 Sheep anti-rabbit IgG  was added. The immunocomplexes were washed, supernatants were collected, incubated at 65oC overnight, and immunoprecipiated DNA were purified using MinElute columns (Qiagen) [Antibodies] rabbit anti-BATF (Schraml et al., 2010), rabbit anti-CTCF (Active Motif, 61311), rabbit anti-Med12 (Bethyl, A300-774A),  rabbit anti-Ets1 (Santa Cruz, sc 350X), rabbit anti-IRF4 (Santa Cruz, sc-28696 X),  rabbit anti-H3K27me3 (Millipore, 07-449), rabbit anti-H3K4me3 (Millipore, 07-473), rabbit anti-H3K27ac (abcam, ab4729), and normal rabbit IgG (Millipore, 12-370) Immunoprecipitated DNA was used to prepare library using NEBNext Ultra II DNA library prep kit and NEBNext Multiplex Oligos for Illumina (NEB). ChIP-seq libraries were sequenced with single-end 50 bp using Illumina HiSeq 2500.