GSM1173381: ChIP-seq H3K27ac in bud-stage embryo; Danio rerio; ChIP-Seq
For each ChIP experiment, ~2000 embryos were carefully staged. 500 embryos were then collected, transferred into PBS containing 20mM Na-butyrate, protease inhibitors (Sigma). Embryos were then transferred to 5ml syringe fitted with 21G needle. After leaving ~500ul buffer above the embryos, embryos were forced through the needle by pushing the pistons to dissociate the embryos. The resulting lysed extract was collected in 2ml eppendorf tubes, and crosslinked by adding formaldehyde solution to final concentration of 1.1%. Crosslinking was conducted for 15 minutes at room temperature while samples were mixed using nutator. After crosslinking, glycine was added to 0.125M to quench the formaldehyde, and incubated on ice for 5 minutes. Tubes were then centrifuged at 470g for 10 minutes at 4C to sediment the cells. The cells were then washed twice with 500ul ice-cold PBS/Na-butyrate/protease inhibitor solution, and flash-frozen and kept at -80C until the ChIP experiment. ChIP-seq libraries prepared using standard Illumina protocols