GSM8477131: HCT116 cells, ID3, ChIP; Homo sapiens; ChIP-Seq
CRC cells were collected and treated with cell lysate. Ultrasonic disruption was performed, followed by centrifugation to remove sediment. ChIP dilution buffer and Protein A gel were added to the supernatant, and the mixture was incubated overnight at 4°C. Subsequently, ID3 (CST #9837) antibodies were added. The solution was inverted and mixed overnight, then centrifuged to remove the supernatant. The precipitated mixture was washed with a buffer, eluted with eluent, and treated with NaCl and proteinase K to dissolve the cross-linking. ChIP-Sequencing library preparation was performed according to Illumina's protocol Preparing Samples for ChIP Sequencing of DNA. 10 ng DNA of each sample was converted to phosphorylated blunt-ended; An 'A' base was added to the 3' end of the blunt phosphorylated DNA fragments; Illumina's genomic adapters were ligated to the A tailed DNA fragments; PCR amplification was performed to enrich ligated fragments. The enriched product of ~200-1500 bp was size selected using AMPure XP beads.