GSM7733657: K562 cells, DRB, Input for RIP; Homo sapiens; RIP-Seq
K562 cells (1.6x10^6) were crosslinked with 1% freshly made formaldehyde solution for 10 min at room temperature. Crosslinking was stopped by adding 1/10th volume of 2.66 M Glycine, kept for 5 min at room temperature and 10 min on ice. Cell pellets were washed twice with ice-cold PBS (freshly supplemented with 1 mM PMSF). Nuclei were extracted. DNase I treatment (250 U/ml) was performed for 30 min at 37°C, and EDTA (final concentration 20 mM) added to stop the reaction. Resuspended Nuclei were sonicated once for 20s (1 pulse every 3 seconds) at 30% amplitude (Branson Digital Sonifer, Danbury, CT). 5. 5 μg antibody or preimmune serum was used for IP - ORC1 (Abcam, ab85830), preimmune serum: IgG (ab171870) or no antibody, no serum (input). BGISEQ platform library preparation