GSM8868945: WT, 25kPa substrate, 72hr culture, rep3; Homo sapiens; ATAC-seq
Cells were trypsinized, collected in a 1.5mL DNA Lobind (Eppendorf) tube, and washed with ice cold ATAC-seq resuspension buffer (ATAC-RSB). After washing cells were lysed with ATAC-RSB containing .005% digitonin, .05% NP-40, and .05% Tween-20. Transposition was performed using TDE1 Tn5 tranposase (Nextera) in TD buffer (Nextera) for 45 minutes on a thermoshaker set to 37C with 1000 RPM mixing. The resulting DNA products were purified using a Qiagen MinElute purification kit. Samples were double-barcoded using Nextera i7 and i5 indexes (Illumina), amplified on a thermocycler, size selected to exclude fragments >600 bp using the SPRIselect system (Beckman-Coulter) and pooled to a final concentration of 20nM. Proper fragment size distribution was validated prior to sequencing via TapeStation (Agilent) as preliminary quality control (QC) to confirm DNA fragment sizes followed the anticipated pattern of nucleosomal periodicity.