GSM8150688: TGFb_H2Az-Ac_Rep2; Mus musculus; ChIP-Seq
Cells were lysed in lysis buffer (10 mM NaCl, 0,2% NP-40 + Prot Inhibitors) and Chromatin was extracted by Sonication (Bioruptor Nano) in 0.5% SDS, 5mM EDTA + Prot Inhibitors. Then, chromatin fragments were extracted using 4 volumes of 25 mM HEPES, 185 mM NaCl, 1.25% Tx-100 + Protease Inhibitors. Next, samples were incubated for 10-12 hours with the H2Az-Ac antibody, then captured using 25 uL Magna ChIP Prot A+G and incubating for 3h at 4 ºC. Next, ChIPed DNA was released and reversed crosslinking by 30 min incubation with 2 uL of Proteinase K in ChIP Elution buffer followed by 1-hour incubation at 68 ºC. Finally, the ChIPed DNA was purified with a 2.2X SPRI Cleanup and quantified by Qbit. ChIP-seq libraries were prepared from 5 ng of DNA with NEB Next Ultra II kit, QCed for concentration (Qbit) and size (Tape-Station) and sequenced to 50 million reads per sample.