Chipseq of Gallus Gallus HH22: FEZ
We performed ChIP-seq for H3K27ac, a known enhancer-associated histone modification. Briefly, frozen frontonasal tissue of chicken embryos (HH22) was pulverized with a mortar and pestle, resuspended in PBS, and cross-linked with 1% formaldehyde at room temperature for 10 min. Chromatin was sonicated to generate fragments averaging 200600 bp in size. Chromatin was incubated for 3 hours at 4C with 10g of Cell Signaling H3K27ac antibody (8173S, lot: 106590037-6). Protein A and G Dynabeads (Invitrogen) were then added to this chromatin/antibody mixture for 30 min at 4C. Immuno-complexes were sequentially washed, and protein/DNA complexes were eluted in an SDS buffer (1% SDS, 50 mM Tris pH 8.0, 10 mM EDTA) at 37C for one hour. Samples were treated with 1l Proteinase K (20mg/ml) at 65C and reverse-crosslinked overnight. Finally, the DNA was purified using Nucleic Acid Extraction Reagent (Y202-G30), and the quality was assessed on the Agilent Bioanalyzer. The ChIP-seq libraries were prepared using the AcceGen DNA Library Prep Kit from Illumina (AG1216). Libraries and input samples were pooled and sequenced with single-end 150bp reads on the Illumina NovaSeq 6000.