title: GSM2448910: MCF-7 B7 TamS E2; Homo sapiens; OTHER
library_construction_protocol: Nuclei were isolated as described in previously (Core et al., 2008). Briefly, cells were rinsed three times with ice-cold PBS and lysed using lysis buffer (10 mM Tris-HCl pH 7.4, 2 mM MgCl2, 3 mM CaCl2, 0.5% NP-40, 10% Glycerol, 1 mM DTT, 1X PIC (Roche; Cat# 11836153001), and 1 ml/10 mL SUPERase-In (ThermoFisher; Cat# AM2694) dissolved in DEPC water). Cells were homogenized by gently pipetting at least 30 times and the nuclei were harvested by centrifugation at 1000g for five minutes at 4°C. The isolated nuclei were washed twice with lysis buffer and were resuspended in 100 mL freezing buffer (50 mM Tris HCl pH 8.3, 5 mM MgCl2, 40% Glycerol, 0.1 mM EDTA pH 8.0, and 4 U/mL SUPERase-In). The isolated nuclei were used for nuclear run-on and precision nuclear run-on sequencing (PRO-seq) library preparation. We prepared PRO-seq libraries as described in Kwak et. al. (2013) Science.
attributes.tag: GEO Accession
attributes.value: GSM2448910