GSM8128669: IgG_TELOEC_flagSTAT1_sko-STAT1_CUTandRUN_SYN+A485_rep2; Homo sapiens; OTHER
ATAC-seq:Followed protocol from Grandi et al 2022, Nature Protocols, https://doi.org/10.1038/s41596-022-00692-9 CUT&RUN: CUT&RUN was performed as previously described with a few modifications. 1xe6 cells were fixed in 0.1% Formaldyhde for 1 minute before fixation was quenched with 1.5M glycine for 1 minute. Cells were bound to 10uL of activated My-One Concavalin beads per IP. Cells and beads were washed with buffers containing (20mM HEPES pH 7.5, 150mM NaCl, 0.5mM Spermidine, 0.1% BSA Protease and Phosphatase Inhibitors (Thermo Scientific, 78440). Cells were permeabilized in the buffer above plus 0.01% digitonin and 0.7ug of each respective antibody was added to the buffer above plus 0.01% digitonin and 250mM EDTA. Samples then incubated overnight on a nutator at 4C. Samples were washed in the buffer above plus 0.01% digitonin and then pAG-MNase was added to the samples and bound for 1 hour at 4C, after which 100mM CaCL2 was added and pAG-MNase cleavage went for 30 minutes with samples submerged in ice. DNA was purified via a phenol-chloroform isolation after which the purified DNA was utilized to generate sequencing libraries. ATAC-seq:Followed protocol from Grandi et al 2022, Nature Protocols, https://doi.org/10.1038/s41596-022-00692-9 CUT&RUN:DNA libraries were generated using the NEBNext Ultra II DNA Library Prep Kit (NEB #E7645) with the following PCR cycling adjustments for the PCR Enrichment of adaptor ligated DNA  a. 45 sec at 98ºC  b. 15 sec at 98ºC  c. 10 sec at 60ºC  d. Repeat Steps b-c for a total of 14 cycles e. 1 min at 72ºC  final extension.