title: GSM4162860: NPC1-ExpA GRO-seq; Homo sapiens; OTHER
library_construction_protocol: For the run-on assay, re-suspended nuclear extract was mixed with an equal volume of run-on reaction buffer (10 mM Tris-HCl pH 8.0, 5 mM MgCl2, 300 mM KCl, 1 mM DTT, 200 U/ml RNaseOut, 1% Sarkosyl, 500 uM ATP, 500 uM GTP, 500uM CTP, and 500 uM Br-UTP) and incubated for 5 min at 30C. The reaction was stopped by adding 1mL TRIzol LS reagent (Thermo Fisher). The total nuclear RNA was extracted by acidic phenol/chloroform and precipitated by ethanol.  The library construct was done according to GRO-seq protocol described in Core, Leighton J., Joshua J. Waterfall, and John T. Lis. "Nascent RNA sequencing reveals widespread pausing and divergent initiation at human promoters." Science 322.5909 (2008): 1845-1848.
attributes.tag: GEO Accession
attributes.value: GSM4162860