Bulk ATAC-seq of IL27 treated OT-1 cells
Bulk ATAC-seq of IL27 treated OT-1 cells
CD8+ T cells were purified from spleens of OT-I mice by negative selection with magnetic beads (EasySep, Stemcell Technologies). After purification, cells were >95% CD8+ T cells. In each well of a 96-well plate, 0.5x106 of the purified CD8+ T cells/ml were cultured in complete media (RPMI 1640, 10% FBS (Gibco), 1% 2 mM L-glutamine (Life Technologies), 1% HEPES (Life Technologies), 1 nM Sodium Pyruvate (Life Technologies), 1% non-essential amino acids (Life Technologies), 100 U/ml penicillin (Gibco) and 100 μg/ml streptomycin-sulfate (Gibco), 0.05 mM 2-mercaptoethanol (Sigma) with recombinant human IL-2 (10 ng/ml, Roche) with or without 10 ng/ml OVA(257–264) (SIINFEKL) peptide in the presence or absence of 25 ng/ml murine IL-27 (R&D Systems). For repeated peptide stimulation, 10 ng/ml OVA(257–264) peptide was added daily for 4–6 days. Cells were checked daily, passaged, and cultured with fresh complete media containing cytokines as required. After day 5 cells were washed and stained for flow cytometry. ATAC-seq libraries were generated from fresh cells using the ATAC-Seq Kit (Active Motif, catalog# 53150).  Libraries were quantified using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and profiled using the D5000 ScreenTape on TapeStation 4200 (Agilent Technologies).