ChIP-seq of Danio rerio wild type and kdm4b mutant embryos to investigate kdm4b signaling
ChIP-seq of Danio rerio wild type and kdm4b mutant embryos to investigate kdm4b signaling
WT and kdm4b mutant embryos at 22 hpf were fixed in 1% formaldehyde and then lysed in SDS buffer on ice for 30 minutes. Chromatin was sonicated by Fisher Scientific's Sonic Dismembrator with 30% amplitude, pulse for 10 s on and 30 s off for a total sonication 'on'time of 3 min 30 s to generate 100~500 bp fragment DNA. Sonicated chromatin was precipitated by antibodies specific for H3K9me3 (ab8898, Abcam), H3K36me3 (ab9050, Abcam), FOXO3 (ab12162, Abcam), or Normal Rabbit IgG (Negative control, 2729, CST). 10 ng of DNA samples were prepared for Illumina sequencing as the standard steps. Illumina's genomic adapters were ligated to the DNA fragments. Enriched ligated fragments of ~200 - 500 bp was purified and quantified. The libraries were denatured, captured on Illumina flow cell and amplified in situ.