GSM5736106: ChIP-Seq_CTCF_BRD2_degron_3h_input; Mus musculus; ChIP-Seq
The crosslinked cells were isolated into chromatin and digested using MNase (15 min at 37 °C), and then, sonicated in Bioruptor Sonication Device (high energy for 30s on and 60s off, work on 16 cycles).  Lysates were centrifuged twice, 40 μl supernatant were sampled as input, and the rest supernatant were transferred into 2 new DNase free tubes, 2 μg of BRD2 abs (CST, D89B4), 2 μg of BRD3 abs (Proteintech, 11859-1-AP), 2 μg of BRD4 abs (Abcam, ab128874), 1 μg of CTCF abs (ABclonal, A1133), 1 μg of Smc1a abs (Bethyl, A300-055A), 1 μg of Rpb1NTD abs (CST, D8L4Y) , 1 μg of Rpb1 PSer2 abs (CST, E1Z3G), 2.5 μg of TAF1 abs (CST, D6J8B), 2.5 μg of TAF3 abs (Proteintech, 18901-1-AP), 2.5 μg of TBP abs (Proteintech, 22006-1-AP), 4 μg of XRN2 abs (Proteintech, 11267-1-AP), and 1 μl GFP abs (Abcam, ab290) were added for each ChIP reaction, respectively, and equal isotype IgG control (Millipore, 12-370, 12-371) and incubated overnight at 4 °C with low speed rotation, the antibody enriched DNA was extracted by Tris saturated equilibrium phenol (25:24:1) (pH8.0). BRD2-degron mES, BRD2, BRD3, BRD4, GFP, CTCF, SMC1, Pol II-NTD, Pol II-S2P, TAF1, TAF3, TBP, and XRN2 ChIPed DNA were subjected for library construction using the NEBNext Ultra II DNA library prep kit, according to the manufacturer-supplied protocol, and then the 200~400 bps length amplified DNA were recovered and analyzed with Qubit assays and Fragment Analyzer, and finally subject to HiSeq Xten PE150 sequencing (Novogene, Beijing).