title: GSM9102100: Day 14, constant sgRNA, MOI20, imatinib, rep2; Homo sapiens; OTHER
library_construction_protocol: gDNA of live cells is extacted with the Quick-DNA 96 Kit (Zymo). gDNA of fixed cells is extacted by Phenol/Chloroform extraction. Cell pellets are resuspended and incubated overnight at 65 °C in 400µl of ChIP lysis buffer (1% SDS, 50 mM Tris, pH 8, 10 mM EDTA) with 16 µl 5 M NaCl. 8µl of RNAse A (10mg/ml - Thermo Fisher Scientific) is added the next day and incubated at 37 °C for 1 h followed by adding 8µl of Proteinase K (20mg/ml - Thermo Fisher Scientific) and a 1 h 55 °C incubation step. Genomic DNA was then isolated with phenol:chloroform:isoamyl alcohol, ethanol precipitated and washed with 70% ethanol. DNA was resuspended in water. Genomic DNA is PCR amplified with NEBNext Ultra II Q5 polymerase with primers amplifying the sgRNA cassette.