title: GSM5747902: H3K27me3 empty vector control rep 2; Homo sapiens; OTHER
library_construction_protocol: 1.5x105 nuclei extracted from each cell lines were captured to Concanavalin A beads (Bangs Laboratories) and then incubated with anti-H3K27me3 antibody (1/100, 07-449, EMD Millipore). After washing, beads were incubated with 700 ng/ml pA-MNase and the MNase reaction was carried out by incubating with CaCl2 on 0 °C cold metal block. After stopping this reaction, DNA-protein complex was released by incubation at 37°C and treated with Proteinase K. DNA were extracted with Phenol-Chloroform and purified by ethanol precipitation. For library preparation, end repair was conducted for 15 min at 12°C and 15 min at 37°C, followed by dA-tailing for 20 min at 72°C. After adaptor ligation using TruSeq DNA single indexes (Illumina) for 15 min at 20°C, adapter-ligated DNA was cleaned up by Ampure XP beads (Beckman Coulter). Libraries were PCR amplified for 14 cycle using KAPA HS HIFI polymerase. After cleaning up of PCR product by Ampure XP beads, DNA amount of library was measured by Qubit and size distribution of DNA was determined by Agilent TapeStation. CUT&RUN-seq