GSM8818986: Input_raw_MYC_ab_1; Homo sapiens; ChIP-Seq
Cells for ChIP-seq were crosslinked with 1% paraformaldehyde for 10 min and then quenched in 125 mM glycine for 5 min at room temperature with rotation. After washed with cold PBS, cells were lysed in lysis buffer (10 mM Tris_x0002_HCl, pH 7.5, 10 mM NaCl, and 0.5% NP-40) on ice for 10 min. The nuclei were spun down and then resuspended in 500 µl 1XChIP-seq buffer(50 mM Tris-HCl, pH 8.1, 10 mM EDTA, 100 mM NaCl, 1% Triton X-100, and 0.1% sodium deoxycholate). Sonication was per_x0002_formed for 5 cycles of 30 s on and 30 s off in Bioruptor. After centrifuged at 15,000 × g at 4 °C for 10 min, the supernatant was used for chromatin immunoprecipitation. Same amounts of Drosophila S2 cell chromatin were spiked in for normalization, 1% for input. 10 µg of Z22 antibodies was added into each sample and rocked at 4 °C overnight. Protein A beads were added and incubated for 3 h. After the beads were extensively washed, the bound DNA was eluted by elution buffer (10 mM Tris-HCl, pH 8.0, 10 mM EDTA, 150 mM NaCl, 5 mM DTT, and 1% SDS). Reverse_x0002_crosslinked at 65 °C overnight. The eluted DNA was treated with proteinase K and RNase A, and then purified with the Min-Elute PCR purification kit (Qiagen, Cat.#28006). Sample extracted was subjected to library preparation with VAHTS Universal DNA Library Prep Kit for Illumina V3 (Vazyme, Cat. #ND607-1) following the manufacturer's instructions.