title: liver chromatin immunoprecipitation sequencing (ChIP-seq)
design: Liver tissues from four healthy adult Yorkshire pigs at Iowa State University were grounded under liquid nitrogen and cross-linked with 1% formaldehyde at room temperature with rotation for 15 min. Cross-linking was quenched, then was washed and flash frozen. Nuclei were extracted, lysed, and sonicated to produce sheared liver chromatin with an average length of 200 to 500 bp using a Bioruptor (Diagenode, Inc., Seraing, Belgium) for a total of processing time of 30 min (15s on, 45s off), with power setting at H. Fifty micrograms of final soluble chromatin was used for each ChIP reaction and combined with Protein G Dynabeads (Diagenode) prebound with 1g of antibodies to H3K4me3 (Diagenode Cat. No. C01010051) and 1g of antibodies to H3K36me3 (Diagenode Cat. No. C15410192-10). Immunoprecipitated chromatin was washed five times with 1 mL of wash buffer. Immunoprecipiated chromatin was eluted, cross-links were reversed, and DNA was purified.The sequencing libraries were prepared from the ChIP and Input DNA samples using the Illumina (San Diego, CA) TruSeq DNA Sample Preparation kit following the manufacturer's instructions, and DNA was subjected to ultra-high throughput sequencing on a Illumina HiSeq 2000 DNA sequencers.