GSM5018128: H3K27F_NT_R2; Mus musculus; ChIP-Seq
For ChIP-Seq, cells were collected and either cross-linked first in 2mM disuccinimidyl glutarate (DSG) then in 1% formaldehyde (for Hdac3 CST-85057 ChIP-seq) or only in 1% formaldehyde (for H3K27ac and Hdac3 ab7030 ChIP-seq).  After quenching the excess formaldehyde with 125 mM glycine, the fixed cells were washed, nuclei were isolated and then sheared in a Covaris E229 sonicator for 8 minutes to generate DNA fragments between ~ 200-1000 bp.  After clarification of insoluble material by centrifugation, the chromatin was immunoprecipitated overnight at 4C with antibodies against Hdac3 or H3K27ac then bound to Protein A Dynabeads (Invitrogen). Antibody bound DNA were washed and treated with Proteinase K and RNase A and the purified ChIP DNA was used for library generation for subsequent sequencing. ChIP-Seq libraries were generated using the NuGen Ovation Ultralow Library System V2 following the manufacturer's instructions.