GSM7949866: T21 V 1; Homo sapiens; OTHER
45 minutes after vehicle treatment, cells were collected, spun down, washed once in PBS,  then nuclei were isolated using a lysis buffer containing 10 mM Tris pH 7.4, 2 mM MgCl2, 3 mM CaCl2, 0.5% NP-40, 10% glycerol, 1 mM DTT, 1 uL/10 mL SUPERase-In, and protease/phosphatase inhibitor cocktail. Nuclei were snap-frozen in freezing buffer consisting of 50 mM Tris pH 8.3, 5 mM MgCl2, 40% glycerol. 0.1 mM EDTA, and SUPERase-In until library preparation 10 million human nuclei/replicate were mixed with 100,000 Drosophila melanogaster nuceli per replicate. Nuclear run-ons were performed in the presence of biotinylated CTP and nascent RNA was isolated using strepdavidin beads. Library preparation was performed as described in Fant et al. Mol Cell 2021 (PMID 32229306)