Acetyl histone h3 antibody

ChIP assays were performed using a SimpleChIP Enzymatic Chromatin IP Kit (#9003, Cell Signaling Technology, USA) following the manufacturer’s instructions. The protein-RNA interaction was first cross-linked using 1% formaldehyde. After cell lysis, chromatin was partially digested using micrococcal nuclease and sonication. The digested chromatin was then incubated with HIF-1α antibody (#36169, Cell Signaling Technology, USA) or acetyl-histone H3 antibody (#9649, Cell Signaling Technology, USA) overnight at 4 °C. After pulldown by ChIP-Grade Protein G Magnetic Beads (#9006, Cell Signaling Technology, USA), the immune complex was eluted from the beads, and DNA was purified. The presence of TUG1 was examined using quantitative real-time PCR (qRT-PCR) with the primer pair forwards 5′-GGCACCCAGTGTAAAGCA-3′ and reverse 5′-AAGCAGCAGATAACAGAGTTGA-3′ (GenePharma, Shanghai, China). Rabbit immunoglobulin G (IgG) (#8726, Cell Signaling Technology, USA) was used as a negative control. PCR products were then separated using gel electrophoresis.