Illumina HiSeq 2500 sequencing; Identify acetylation marks in adult mouse L4 vertebrae gene promoter regions from maternal high fat diet dams using H3K27ac ChIP-Seq
Identify acetylation marks in adult mouse L4 vertebrae gene promoter regions from maternal high fat diet dams using H3K27ac ChIP-Seq
After 17 weeks mice were sacrificed and L4 vertebrae was removed and cleaned. Tissue was then homogenized using tissue homogenizer and metallic beads. Briefly, DNA was crosslinked to protein using formaldehyde and immunoprecipitated using CHIP grade H3K27ac antibody. After immunoprecipitation, chromatin was purified from the mixture using Cell signaling procedure. Purified DNA was then eluted from chromatin using cell signaling protocol. DNA for each group were pooled. Purified immunoprecipitated and input DNA were utilized to prepare libraries for sequencing using NEBNext reagents. Briefly, 150 ng of DNA was used for end-filling, dA-tailing and ligation with Illumina's paired-end adapters using manufacturer's recommendations.  Adapter-ligated DNA was amplified using PCR and products around 200 – 350 bp were used for cluster generation. Quantification of the ChIP-seq libraries was done via Qubit dsDNA HS Assay kit.