GSM6637480: E14_ESC_Esrrb_HiChIP; Mus musculus; OTHER
HiChIP was performed according to Arima HiC+ kit protocol (document part number: A160430 v00) with some modifications. Briefly, cells were crosslinked with 2% formaldehyde in PBS at room temperature for 10min, quenched for 5min and washed once with ice-cold PBS. After HiC reactions, the chromatins were sheared using Covaris S2 instrument with intensity setting 3 (equivalent to 105W), 5% duty cycle, and 200 cycles per burst for 7min. Following shearing, the chromatins were precleared using Dynabeads™ Protein A (Thermo Fisher Scientific, 10001D) for 1h at 4°C. Subsequently, 10µg of Esrrb antibody (PPH6705, Perseus Proteomics) was added to the precleared chromatins and incubated overnight at 4°C. Following the overnight incubation, BSA-blocked Dynabeads™ Protein A were added to the antibody-bound chromatins and incubated for 4h at 4°C, washed, eluted and decrosslinked according to Arima HiC+ kit protocol. Libraries were prepared according to Illumina's instructions with optimised rounds of PCR amplification, and finally analyzed the resulting fragments with paired-end sequencing.