title: GSM4799753: H3K27ac from UC Davis FAANG pig lung rep P350; Sus scrofa; ChIP-Seq
library_construction_protocol: ChIP-seq experiments were performed on frozen tissue using the iDeal ChIP-seq kit for Histones (Diagenode Cat.#C01010059, Denville, NJ) according to the manufacturer's protocol except for the following changes. 20-30 mg of frozen tissue was powdered using liquid nitrogen in pre-chilled mortar. Cross-linking was performed with 1% formaldehyde which was diluted from 16% methanol-free formaldehyde (Thermo Scientific, Cat.#28906, Waltham, MA) for 8 minutes and quenched with glycine for 10 minutes. Nuclei were harvested by centrifugation at 2000g for 5 minutes and resuspended in iS1 buffer for incubation on ice for 30 minutes. Chromatin was sheared using the Covaris E220 between 6-12 minutes depending on the tissue. For immunoprecipitation experiments, about 1000 ng of sheared chromatin (estimated from DNA extraction) was used as input after which the kit protocol was followed with 1 μg (histone modifications) or 1.5 μg (CTCF) of antibody. The following antibodies used were from Diagenode: H3K4me3 (in kit), H3K27me3 (#C15410069), H3K27ac (#C15410174), H3K4me1 (#C15410037), and CTCF (#15410210). An input (no antibody) was performed for each sample. NEBNext Ultra DNA library prep kit for Illumina libraries (New England Biolabs #E7645L, Ipswich, MA) was used for library construction, selecting for 150-200 bp (H3K4me3, H3K27ac, CTCF) or 200-400 bp (H3K27me3, H3K4me1) insert fragment sizes using Ampure beads (Beckman Coulter #A63881). Libraries were sequenced on Illumina's HiSeq 4000 with single-end 50 bp reads.
attributes.tag: GEO Accession
attributes.value: GSM4799753