GSM9064093: Rv1, control, SMARCA1, rep2; Homo sapiens; ChIP-Seq
Cell nuclei were extracted using nuclear extraction buffer (20 mM HEPES, pH 7.9, 10 mM KCl, 0.1 % Triton X-100, 20% glycerol, 1X protease cocktail) and incubated with ConA Beads to absorb nuclei to beads for 10 min at RT. The nuclei-beads were placed in magnet and the supernatant was removed. Antibody buffer and 0.5 μg antibody was added to each sample and incubate overnight at 4°C. After washing the nuclei-beads, 2.5 μl of pAG-NMase was added and incubated for 10 min at RT. After washing, 1 μl of Chromatin Digest additive was added and incubate for 2 hrs at 4°C. Then, stop buffer and 0.5 ng of Spike-in DNA were added and incubate for 10 min at 37 °C. We purified DNA from this using DNA purification kit (EpiCypher). Using 5 ng of purified CUT&RUN-enriched DNA, we prepared sequencing library using library prep kit for illumine sequencing (E7645, NEB). The library was purified with 1.0 X AMPure beads (Beckman) and library fragment size was checked with Agilent Bioanalyzer.