H3K27ac ChIP-seq, sample matched input, for specimen F24, sample GW024, isolated Microglia
Microglia were crosslinked with paraformaldehyde, sorted and snap-frozen in liquid nitrogen. Microglia were thawed, lysed with IGEPAL, then sonicated and cleared by centrifugation. Supernatants were immunoprecipitated with antibodies bound to Protein A Dynabeads, washed 3 times on ice and eluted. Crosslinking was reversed by incubation in a hot air oven, digested with RNase A and proteinase K, then purified with Sera-Mag SpeedBeads. Libraries were then subjected to blunting, A-tailing and adapter ligation using barcoded adapters and digested with uracil DNA glycosylase to generate strand-specific libraries. Libraries were PCR-amplified for 12-15 cycles and size-selected for 200-350 bp fragments by gel extraction, then sequenced.