GSM9258148: HEC-1A THC 20uM 72h Biol rep 2; Homo sapiens; RNA-Seq
RNA-seq: Total RNA was purified using the RNeasy Mini Kit (QIAgen, 74104) following manufacturer's instructions. ATAC-seq: 50x10^3 HEC-1A cells per condition were pelleted (500xg, 4°C, 5min), washed with PBS, and lysed in buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3mM MgCl2, 0.1% Igepal CA-630) for 20min at 4°C. Nuclei were isolated by centrifugation and resuspended in 2x transposition reaction mix (Diagenode, C01070012) for 1h at 37°C. DNA was purified with cleanup buffer containing 5% SDS and Proteinase K, followed by 2x SPRI bead cleanup and elution in 10mM Tris-HCl pH 8. Cut&run: 10x10^4 HEC-1A cells per condition were isolated with concanavalin A magnetic beads, permeabilized with digitonin, and incubated with anti-H3K4me3 antibody or IgG control. Cut&Run was performed using pAG-MNase digestion following manufacturer's protocol (Cell Signaling Technology, 86652). RNA-seq libraries were prepared using the Illumina TruSeq Stranded Total RNA library prep kit according to manufacturer's protocol; ATAC-seq Genomic libraries were prepared using Buenrostro et al. 2013 primers with PCR program: 72°C 5min, 98°C 30sec, 9 cycles of (98°C 10sec, 63°C 30sec, 72°C 1min). Libraries purified with SPRI beads, eluted in 21μL 10mM Tris-HCl pH 8.0. DNA quantified using Qubit dsDNA BR assay kit (ThermoFisher Scientific, Q32850).;  Cut&run DNA libraries were purified and prepared with standard end repair/adapter ligation, and sequenced on NextSeq550 (150bp paired-end)