GSM6189353: CT27, EVT, p20, rep1; Homo sapiens; ATAC-seq
The Omni-ATAC library preparation was performed on freshly isolated cells according to the established protocol (Corces et al. 2017) until the post-transposition cleanup step. After the cleanup of the transposed DNA was complete, samples were stored at -20°C until library amplification. The samples underwent 5 cycles of PCR before an aliquot was used in a quantitative PCR (qPCR) reaction to determine whether additional amplification was required. An Applied Biosciences Viia7 Real-Time PCR System (Applied Biosciences, Beverly Hills, CA) was used for qPCR and the number of additional PCR cycles needed was determined from the multicomponent plot, which plots fluorescence versus the number of PCR cycles. The maximum fluorescence for each sample was found from the plot, ¼ of the maximum fluorescence value was calculated, and the number of additional PCR cycles needed was determined by determining the cycle number that reached ¼ maximum fluorescence on the graph. In cases where the ¼ maximum fluorescence value was halfway between two cycle numbers, the lower number of cycles was selected since slight under-amplification was more favorable than overamplification. The original samples were then PCR amplified with their determined number of additional cycles. Cleanup of the samples was performed after PCR amplification according to the published protocol (Corces et al. 2017). Libraries were quantified using a Qubit dsDNA BR Assay Kit (Q32853, Thermo-Fisher) and the size was determined with a High Sensitivity DNA Bioanalyzer Kit (5067-4626, Agilent, Santa Clara, CA).