GSM3178274: U937_ATAC-seq; Homo sapiens; ATAC-seq
Cells were washed with ice cold FACS Buffer and kept on ice until cell sorting. 25,000 live cells from each condition were sorted in to FACS Buffer and pelleted by centrifugation at 500 RCF for 5 minutes at 4C in a pre-cooled fixed angle centrifuge. Cell lines were then tagmented according to the previously described Fast-ATAC protocol (92). Briefly, all supernatant was removed being careful to not disturb the not visible cell pellet. 50 ul transposase mixture (25 ul of 2x TD, 2.5 ul of TDE1, 0.5 ul of 1% digitonin, 22 ul of nuclease-free water) (Cat# FC-121-1030, Illumina; Cat# G9441, Promega) was added to the cells, the pellet was dissociated by pipetting. Transposition reactions were incubated at 37C for 30 minutes in an Eppendorf ThermoMixer with agitation at 300 RPM. Transposed DNA was purified using a QIAgen MinElute Reaction Cleanup kit (Cat# 28204) and purified DNA was eluted in 12 ul elution buffer (10 mM Tris-HCl, pH 8). Transposed fragments were amplified and purified as described previously (93) with modified primers (94). Libraries were quantified using qPCR prior to sequencing. All Fast-ATAC libraries were sequenced using paired-end, dual-index sequencing on a NextSeq with 76x8x8x76 cycle reads at an average read depth of 30 million reads per sample.