title: GSM3386198: H3K27ac-ChIP-Seq; Mus musculus; ChIP-Seq
library_construction_protocol: Embryoid bodies from 20 10-cm dishes were harvested at day 6 and fixed immediately with 1% p-formaldehyde for 15 minutes at room temperature followed by quenching with 0.125M (final conc) Glycine. Lysates were sonicated with a microtip sonicator to shear the DNA. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 30 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4 C. Protein A agarose beads were added and incubation at 4 C was continued for another 3 hr. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65 C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation. Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on HiSeq 2500/NextSeq 500.
attributes.tag: GEO Accession
attributes.value: GSM3386198