title: GSM7101056: MOLM-13, DMSO, H3K4me1 rep 2; Homo sapiens; OTHER
library_construction_protocol: Cells were washed in Wash Buffer (20 mmol/L HEPES pH 7.5, 150 mmol/L NaCl, 0.5 mmol/L Spermidine, protease inhibitor cocktail) and bound to Concanavalin A beads.  Cells were resuspended  in  50μL  Digitonin Wash  Buffer (20 mmol/L HEPES pH 7.5, 150 mmol/L NaCl, 0.5 mmol/L Spermidine, protease inhibitor cocktail, 2 mmol/L EDTA, 0.05% Digitonin) and incubated with anti-H3K4me3, -H3K4me1, -H3K27ac, -and H3K27me3 antibody at a 1:100 dilution overnight at 4°C. Beads were resuspended  in  100μL  Digitonin Wash  Buffer  with  a  secondary  antibody  diluted  1:100  and  incubated  for 30min at  room  temperature.  Cells were washed three times in Digitonin Wash Buffer and resuspended in Digitonin-300 Buffer (0.05% Digitonin, 20 mmol/L HEPES, pH 7.5, 300 mmol/L NaCl, 0.5 mmol/L Spermidine, protease inhibitor cocktail) containing 1:250 pA-Tn5 transposase and incubated at room temperature for 1 hour. Subsequently, cells were washed three times in Digitonin-300 Buffer and resuspended in 100 μL Tagmentation Buffer (10 mmol/L MgCl2 in Digitonin-300 Buffer) and incubated at 37°C for 1 hour. Tagmentation was stopped by adding 10μL of 0.5 M EDTA, 3 μL of 10% SDS, and 2.5 μL of 20 mg/mL Proteinase K, and samples were incubated 37°C overnight.  Tagmented DNA was purified using AMPureXP-beads from Beckman Coulter. Cut&Tag libraries were amplified from tagmented DNA isolates using 14 cycles on the thermocycler. Post amplification libraries were size selected using  AMPureXP-beads. Library fragment lenght was checked by HSD5000 Tape and DNA concentration by Qubit.