GSM8996698: SK-UT1 γH2AX DOX; Homo sapiens; ChIP-Seq
After culturing, 5 × 106 uLMS cells were collected using 0.25% trypsin-EDTA (FUJIFILM Wako Pure Chemical Corporation). The cell pellets were cross-linked with 1% formaldehyde solution (FUJIFILM Wako Pure Chemical Corporation) for 10 min at 37°C and treated with 0.125 M glycine (FUJIFILM Wako Pure Chemical Corporation) for 5 min at 25°C. After lysing the cells in lysis buffer (see Additional file 1: Table S3), DNA fragmentation was performed by sonication with a Covaris S220 (Covaris, Woburn, MA, USA) for 200 cycles for 10 min. The fragmented DNA underwent ChIP and was incubated overnight at 4 °C with Dynabeads Protein G (Thermo Fisher Scientific) with 2.5 µg of anti-gamma H2A.X (phospho S139) antibody (Abcam, Cat. #ab2893, Lot: GR3446449-2). Next, we washed the beads with low-salt, high-salt, and LiCl immune complex wash buffers (see Additional file 1: Table S3) at 4°C for 5 min, followed by washing with TE buffer (see Additional file 1: Table S3) for 5 min at 25°C. The fragments were extracted using an elution buffer (see Additional file 1: Table S3). Libraries were prepared using a ThruPLEX DNA-Seq Kit (TaKaRa Bio).