GSM5593407: KOinput-1; Homo sapiens; ChIP-Seq
Briefly, 30X106 K562 cells were fixed with 1% formaldehyde, sonicated and immunoprecipitated with antibodies against H3K27ac, HA and BRG1 . Chromatin was pulled down using Dynabeads Protein G, washed twice with RIPA-LS, RIPA-HS, LiCL washing buffer, followed by one wash with 10mM TrisHCl pH8.0. Precipitated DNA was reverse cross link and eluted for ChIP or was tagmentated (ChIPmentation)on magnetic beads with 0.2ul of tagmentation enzyme (Nextera XT DNA Library Preparation Kit, Illumina # FC-131-1024) for 10 minutes at 37 degree while shaking. The beads with tagementated DNA were washed twice with RIPA-LS and TE buffers. The DNA was eluted and de-crosslinked in ChIP elution buffer (20 mM Tris, pH 7.5, 5 mM EDTA, pH 8.0, 50 mM NaCl, 1% SDS, and 50 μg/mL proteinase K) for 1 hour at 55 degree and incubated at degree  overnight. ChIP-mentation Libraries were prepared using Nextera XT DNA Library Preparation Kit, Illumina # FC-131-1024. Eluted DNA was purified using SRI beads (1.8x) and PCR amplified with Nextera primers. Libraries were purified (1.8x) and size selected (0.65x/ 0.15x) with SRI beads. ChIP Libaries were processing followed manufacturer's protocol (Takara Bio, R400675)