GSM8973938: PU knockdown replicate 3; Mus musculus; ATAC-seq
Microglia were isolated using cold mechanical homogenization followed by FACS sorting. T cells were obtained from the spleen with cold mechanical homogenization followed by RBC lysis. ATAC-seq libraries were prepared using the OMNI-ATAC protocol (Buenrostro et al., 2015). Briefly, nuclei were isolated from 40,000 FACS-sorted eYFP+ cells. Transposition was performed using Tn5 transposase (Illumina) at 37°C for 30 min, followed by DNA purification with the Zymo DNA Clean & Concentrator kit. Libraries were pre-amplified for five cycles using NEBNext High-Fidelity 2X PCR Master Mix. Additional amplification cycles were determined via qPCR. Double-sided size selection was performed using AMPure XP beads. Libraries were quantified via Qubit and analyzed by Bioanalyzer or TapeStation. Final libraries were pooled equimolarly and sequenced on an Illumina NovaSeq (2 × 150 bp), targeting ~40 million reads per sample.