title: GSM8778065: ATAC-seq of CD8 Tex sgIcos cells, replicate 1; Mus musculus; ATAC-seq
library_construction_protocol: 70,000 Tpex or Tex cells per replicate were sorted into 1.5 ml tubes precoated with MACS buffer and processed for ATAC-seq as described in 1 with modifications as follows: centrifugation was performed at 800 x g for 8 min at 4ºC in a swing-bucket rotor; the transposition mix omitted RNAse inhibitor and a volume of 50 μl was used per sample; transposed DNA was purified using a Monarch DNA cleanup kit (NEB) and eluted in 22 μl. The optimal number of PCR cycles was determined on 2 μl of eluate and libraries were amplified and barcoded according to 2. NEBNext Ultra II Q5 mastermix (NEB) was preheated at 98ºC for 30 sec to restore its nick translation activity, then reactions were assembled on ice and amplified for 9 cycles. Libraries were cleaned up with SPRIselect beads (Beckman-Coulter) at 0.6X (right) and 1.5X (left) ratios for two-sided size selection and sequenced on the NextSeq 2000 system (Illumina) in 60bp paired-end mode at a target depth of 40M read pairs per replicate.