RNA immunoprecipitation (RIP), chromatin immunoprecipitation (ChIP) and chromatin RNA-immunoprecipitation (ChRIP)

RIP experiments were performed using total cell extracts and antibody against KDM4B or control IgG using RIP assay kit (MBL international, Woburn, MA, USA). RNAs associated with IgG- or KDM4B-immunoprecipitate were extracted using Trizol, treated with DNase I, converted to cDNA, and used for qRT-PCR. For ChIP and ChRIP, chromatin fractions were crosslinked with 1% paraformaldehyde, fragmented to ∼500 bp via sonication, and immunoprecipitated with anti-KDM4B antibody. Chromatin precipitates were split into two parts after reversing the cross-link. One part was treated with RNase and remaining DNA fragments were quantified by SYBR-based qPCR, normalized using the percent input method (Invitrogen) (ChIP-qPCR). The other part was extracted with Trizol for RNA, followed by DNase I treatment, cDNA synthesis and qPCR (ChRIP-qPCR).