GSM3970382: KI167 JUN BJ5ta; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
TFs and others
Cell type Class
Attributes by original data submitter
anti-Jun (Santa Cruz H-79 sc-1694)
Sequenced DNA Library
Cells in culture dishes were crosslinked in 1% formaldehyde for 15 min and the reaction was quenched by 125 mM glycine. Cross-linked cells were washed with LB1 (50 mM HEPES-KOH pH7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP40, 0.25% Triton X-100) and then with LB2 (200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, and 10 mM Tris-HCl pH 8.0). Chromatin was extracted by sonication in LB3-Triton (1 mM EDTA, 0.5 mM EGTA, 10 mM Tris-HCl pH 8, 100 mM NaCl, 0.1% Na-Deoxycholate, 0.5% N-lauroyl sarcosine, 1% Triton). LB1, LB2, and LB3-Triton used during extraction were supplmented with 1x protease inhibitor cocktail (Calbiochem 539131) and 100 nM phosphatase inhibitor Nodularin (Enzo ALX-350-061). One million cells were used in a single 200-µL ChIP reaction. Antibodies used in ChIP are: rabbit monoclonal anti-phospho-Ser22-LMNA antibody D2B2E (Cell Signaling 13448S, Lot # 1; 5 µL per IP); mouse monoclonal anti-unphospho-Ser22-LMNA antibody E1 (Santa Cruz Biotechnology sc-376248, Lot # H2812; 10 µL per IP); mouse monoclonal anti-acetyl-Lys27 histone H3 antibody (Wako MABI0309, Lot # 14007; 2 µL per IP); mouse monoclonal anti-trimethyl-Lys4 histone H3 antibody (Wako MABI14004, Lot # 14004; 2 µL per IP); mouse monoclonal anti-Ty1 antibody (Diagenode C15200054, Lot # 005; 1 µL per IP). Immunoprecipitated DNA was reverse-crosslinked and used to construct high-throughput sequencing libraries using NEBNext Ultra DNA Library Prep Kit (New England Biolabs, E7370).