title: GSM3752830: 16-14T-Old-H3K9ac; Homo sapiens; ChIP-Seq
library_construction_protocol: Nuclei were isolated from 200 mg frozen brain tissue by dounce homogenization in nuclei isolation buffer followed by ultracentrifugation on a 1.8 M sucrose cushion. Nuclei pellets were resuspended and crosslinked in 1% formaldehyde for 10 min at RT, quenched with 125 mM glycine for 5 min and washed twice in cold PBS. 2 x 106 nuclei were lysed in nuclei lysis buffer and chromatin was sheared using a Covaris S220 sonicator to a final ~250 bp. Equal aliquots of sonicated chromatin were used per immunoprecipitation (IP) reaction with antibody (either H3K27ac, or H3K9ac or H3K122ac or H3K4em1) preconjugated to Protein G Dynabeads. 10% of each IP was used for the Inputs. ChIP reactions were incubated overnight at 4°C followed by ChIP washes and DNA was eluted in elution buffer (1% SDS, 50 mM Tris-HCl pH 8, 10 mM EDTA) at 65°C. Eluted DNA from ChIP and Input samples was reverse crosslinked and purified using PCR columns (Qiagen). Sequencing libraries for ChIP experiments were prepared using NEBNext Ultra II reagents (New England Biolabs). All ChIP samples and Input were single-end sequenced on an Illumina NextSeq 500.
attributes.tag: GEO Accession
attributes.value: GSM3752830