GSM9514280: Ishikawa cells, OE2-Input; Homo sapiens; ChIP-Seq
ChIP-seq libraries were prepared using the KAPA HTP Library Preparation Kit, complemented with NEXTflex DNA Barcodes from Bio Scientific. For both input and immunoprecipitation (IP) samples, 10 ng of DNA was used as starting material. Chromatin was first cross-linked using formaldehyde to capture protein-DNA interactions, followed by fragmentation into 200-1000 bp DNA fragments using sonication or enzymatic digestion. Immunoprecipitation was then performed with an antibody specific to the target protein to pull down protein-bound DNA fragments. The immunoprecipitated DNA was purified using Agencourt AMPure XP beads (Beckman Coulter) and washed to remove any contaminants. Library preparation involved end-repair, A-tailing, and adapter ligation using the KAPA HTP Library Preparation Kit, with NEXTflex DNA Barcodes used for multiplexing. The libraries were amplified by 13 cycles of PCR to ensure sufficient DNA for sequencing. Post amplification, libraries were size-selected at 250-450 bp using Agencourt AMPure XP beads to enrich for properly sized fragments. Finally, library quality was validated using the Agilent High Sensitivity DNA Kit to confirm the concentration and size distribution of the libraries. These libraries were then ready for high-throughput sequencing. Basecalls were performed using bcl2fastq v2.17 for Novaseq output.