GSM7806714: rep1_HSC_2_M_H3K36M; Homo sapiens; OTHER
Cut&Run assays were performed with CUTANATM ChIC/CUT&RUN kit according to the manufacturer's protocol (EpiCypher, #14-1048). Briefly, fresh 500,000 cells collected carefully with a cell lifter (Corning, #3008) weredissolved in wash buffer (Pre-wash buffer (EpiCypher, #21-1002), protease inhibitor, and 0.5 mM spermidine)and bound to activated Concanavalin A beads. The cells bound to the beads were dissolved in Antibody buffer (Cell permeabilization buffer and 2 mM EDTA) and incubated with antibodies at 4°C overnight. After extensive washing with Cell permeabilization buffer (Wash buffer and 0.01% digitonin), the cells were incubated with pAG-MNase (EpiCypher, #15-1016) for 10 min at room temperature, and then chromatin digestion was performed with calcium chloride for 2 h, at 4°C. After chromatin digestion, the stop buffer and E. coli spike-in DNA (EpiCypher, #18-1401) were added and incubatedfor 10 min at 37°C. After discarding beads, chromatin fragments were purified with DNA cleanup columns. Immunoprecipitated DNA was used to generate libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs cat# 7370) following the manufacturer's instructions.