Histone h3k9me3

ChIP was performed as described previously (22 (link)), with the following modifications. Briefly, HCT116 cells were exposed to doxorubicin or 5-FU for 24 h prior to formaldehyde fixation. Fixed and lysed cells were sonicated using a Bioruptor Plus Sonication System (Diagenode) set at high power, 30 s ON, 90 s OFF, 60 cycles. Approximately 15–25 μg of sonicated chromatin was incubated overnight at 4°C with 2 μg of p53 (Santa Cruz Biotechnology), histone H3 (Abcam), histone H3K4me3 (Abcam), histone H3K9me3 (Abcam), JMJD2B (Cell Signaling) antibodies, followed by precipitation with protein A/G Dynabeads (Invitrogen). Normal mouse or rabbit IgG (Santa Cruz Biothechnology) was used as a non-specific IgG control. Approximately 5% of the sample from each immunoprecipitation was reserved for input control. Immunoprecipitated complexes were washed, eluted and reverse crosslinked. DNA was purified with QIAquick PCR purification kit following the manufacturer's protocol (Qiagen). Relative enrichment of the samples was measured by qPCR using a titration of pooled input samples as a standard curve, and normalized to input after subtraction of IgG signal. Relative occupancy is presented as percentage of input. For histone ChIPs, H3K4me3 and H3K9me3 enrichments were normalized to bulk histone H3 signal. Primer sequences are listed in Supplementary Table S3.