GSM4569625: NRVM_Veh_WCE; Rattus norvegicus; ChIP-Seq
NRVM were plated in 15 cm dishes at 4 x 10^6 cells/dish. Total of 15 million cells were used for each immunoprecipitation. After indicated treatments, cells were crosslinked with 1% formaldehyde for 10 min at room temperature followed by quenching with 0.125M glycine for 5 min. Cells were then lysed and chromatin was extracted. Isolated chromatin was subjected to shearing with Bioruptor (Diagenode) for 16 cycles (30sec on and 30 sec off/cycle) at high intensity. Small volume of sheared chromatin (50 uL) from each was stored at -80 °C as input. The rest of sheared chromatin was then incubated with 5mg RNAPII antibody (clone 8WG16, BioLegend 920102 lot B217159) with 50uL Dynabeads (Invitrogen) overnight at 4 °C. Chromatin was washed, eluted and reverse crosslinked overnight at 65 °C. Genomic DNA was purified from both immunoprecipitated samples and input samples. The library was prepared with TrueSeq ChIP kit (Illumina)