GSM5033141: THP-1_WT0hIP_rep2_ChIP-seq; Homo sapiens; ChIP-Seq
Untreated and IFN treated cells were crosslinked by adding 1% paraformaldehyde to the culture media and incubating at room temperature for 10 minutes. The crosslinked cells were quenched by adding ice-cold glycine to a final concentration of 125mM and incubating for 5 minutes. Cells were then washed with PBS, resuspended in Supplement Cell Lysis buffer (5 mM PIPES [pH 8.0], 85 mM KCl, 0.5% NP-40) supplemented with 1 X proteinase inhibitor cocktail (Roche), and incubated on ice for 10 minutes. Nuclear pellets were lysed with SDS lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris-HCl, pH 8.1) containing 1 X proteinase inhibitor cocktail for 15 minutes prior to sonication. Cells were sonicated using the Bioruptor plus (Diagenode) on the High setting: 30 seconds On, 30 seconds Off for 20 cycles. SDS was removed from the chromatin solution by centrifugation for 10 minutes at 4°C. The sheared chromatin was measured, diluted with ChIP Dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-HCl, pH 8.1, 167 mM NaCl), and incubated with Protein A agarose beads for 1 hour at 4°C. An aliquot was removed as input from the pre-cleared chromatin and 300 ug of each sample was then incubated with H3K27Ac antibody (Active Motif #91193) overnight at 4°C. Then, Protein A agarose beads were added for 1 hour at 4 °C. The immune complexes were washed once with low salt wash buffer (0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20 mM Tris-HCl, pH 8.1, 150 mM NaCl), once with high salt wash buffer (0.1% SDS, 1% Triton X-100, 2 mM EDTA, 20 mM Tris-HCl, pH 8.1, 500 mM NaCl), once with LiCl wash buffer (0.25 M LiCl, 1% NP-40, 1% deoxycholate, 1 mM EDTA, 10 mM Tris-HCl, pH 8.1), and twice with 1x TE buffer (10 mM Tris-HCl, 1 mM EDTA [pH 8.0]). The complexes were eluted with elution buffer (1% SDS, 0.1 M NaHCO3). The eluates (200 µl) were reverse crosslinked at 65°C overnight by adding 8 µl of 5 M NaCl, followed by RNase A treatment for one hour at 37°C. The samples were mixed with 4 µl of 0.5 M EDTA, 8 µl of 1 M Tris-HCl pH 8.0, and 1 µl of Proteinase K for two hours at 45°C. The ChIPed DNA was purified using ChIP DNA clean & Concentrator (Zymo Research). Sequencing libraries were prepared  using the Nextera DNA Flex Library Prep Kit. 
GEO Accession
GSM5033141