GSM8942383: Th2, Linoleic, DMSO, H3K27Ac ChIP, rep2; Mus musculus; ChIP-Seq
4×10^6 cultured Th2 cells were fixed with 1% (w/v) PFA at 37 °C for 10 min. Nuclei were extracted using lysis buffer and sonicated to reduce the DNA length to between 200 and 1,000 bp using a Bioruptor Pico Sonicator (Diagenode). The soluble fraction was incubated with antibody-conjugated protein-A coated beads overnight at 4 °C using a 1:1 mixture of anti-PPARγ antibody (E-8, from Santa Cruz Biotechnology, Dallas, TX, USA; and A3409A, from Cosmo Bio, Tokyo, Japan), anti-RXRα antibody (D6H10; Cell Signaling Technology), anti-H3K27Ac antibody (D5E4; Cell Signaling Technology), or negative control rabbit IgG antibody included in the kit. Enriched chromatin fragments were purified and de-crosslinked overnight at 65 °C, and immunoprecipitated DNA was extracted. The total input DNA (cellular DNA without immunoprecipitation) was extracted in parallel. DNA libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs, Ipswich, MA, USA) according to the manufacturer's protocol. DNA libraries were sequenced using an Illumina NovaSeq 6000 Sequencing System.