Chromatin immunoprecipitation (ChIP) assay
ChIP assays were performed using the ChIP Assay Kit (ChIP-IT High
Sensitive Kit, Active Motif, CA, USA) according to the manufacturer’s
instructions. Briefly, cells were cross-linked and lysed and the
chromatin was then sheared by sonication to produce fragments from 100
to 500 bps. The complexes were immunoprecipitated using anti-IgG control
or anti-Brd4 antibody, followed by incubation with protein G-agarose
beads. Cross-links were reversed and immunoprecipitated DNA was
purified. The presence of potential Brd4 binding sites was measured by
qPCR using the primers listed in supplemental Table S4.