title: GSM4314289: H3K27me3_ChIP_GHMT2m_Smad2; Mus musculus; ChIP-Seq
library_construction_protocol: Chromatin from D5 control MEFs or D7 GHMT2m-reprogrammed MEFs was isolated using the Magna ChIP A/G Chromatin Immunoprecipitation kit (Millipore 17-10085) as per the manufacturer's instructions. Briefly, 10 cm dishes of cells were crosslinked with 1% paraformaldehyde for 9 minutes at room temperature. Cells were then quenched with glycine (125 mM final concentration) for 5 minutes at room temperature. Chromatin was sheared to approximately 300 bp fragments using a Diagenode Bioruptor 300 ultrasonicator (D5 control MEFs: 15 cycles, 30 seconds on/90 seconds off, high intensity; D7 reprogrammed MEFs: 25 cycles, 30 seconds on/30 seconds off, high intensity). Chromatin was diluted 10 times and incubated overnight at 4 ˚C with 20 µL Protein A/G MagnaChIP beads (Millipore) and 8 µg anti-H3K27me3 (Millipore, 07-449) antibody in an end-over-end mixer. The next day, immunoprecipitated chromatin was washed, eluted, and reverse crosslinked with Proteinase K for 2 hours at 62 ˚C. Libraries were prepared from 1 ng ChIP DNA using the Ovation Ultralow Library System V2 1-16 (NuGEN).
attributes.tag: GEO Accession
attributes.value: GSM4314289