title: GSM4724885: VAT CD4+ (MD-scATAC_5); Mus musculus; ATAC-seq
library_construction_protocol: T cells were isolated and pre-enriched as described in the associated publication. Afterwards, they were stained with live/dead dye and surface antibodies as described. Cells were then sorted on an BDTM FACSAriaTM IIu or FACSAriaTM FUSION cell sorter using 85 µm nozzle size and 4-way-purity sort mode into 1.5 ml tubes filled with FACS buffer. Sample and post-sort quality control were recorded where applicable and are displayed in supplementary figures of the manuscript. Cells were pelleted by centrifugation. Supernatant was removed and cells were resuspended in 0.04%BSA-PBS buffer. Cells were centrifuged, supernatant was completely removed and chilled nuclei lysis buffer was added. Lysis was performed for 2 minutes at 4°C, followed by addition of 50 µl washing buffer. After centrifugation, supernatant was removed and 45 µl chilled diluted nuclei buffer was added. After centrifugation, supernatant was removed and 7µl chilled diluted nuclei buffer was added. Nuclei were counted (if applicable) and transposition mix was prepared with 5 µl of nuclei suspension based on manufacturer's protocol. Transposition was performed for one hour at 37°C, followed by supplementation of master mix and beads, loading on 10X Chromium Next GEM Chip H and processing on 10X Chromium Controller. GEM incubation was performed with 12 cycles of PCR. Library was prepared according to protocol with cycle numbers dependent on input nuclei concentration. Libraries were sequenced on an Illumina NextSeqTM 550 with NextSeqTM 500/550 High Output Kit v2.5 (75 cycles).