GSM2576048: H3K27ac ChIP-matched input from dox- 24h ind SA-tTA-Ascl2 ESCs; Mus musculus; ChIP-Seq
ChIP-seq was performed on samples of 10M formaldehyde-fixed murine ES cells. Frozen samples were thawed and lysed to purify nuclei. Nuclei were then incubated in nuclear lysis/ChIP buffer and sonicated using a Diagenode standard sonicator for a total of 35 minutes. Samples were centrifuged to remove cellular debris, and antibodies added. Antibody-binding incubation was performed, rotating overnight at 4C. Sequential washes to remove nonspecific genomic template were performed using Life Technologies Protein G Dynabeads, and immunopurified samples were further purified. Samples were eluted from Dynabeads using a 70C benchtop shaker, and crosslink reversal performed overnight at 65C using heated shaker, prior to proteinase K and RNAse treatment.  Detailed description, including buffer components and formulae, is described in the text. Library preparation was performed according to manufacturer-supplied literature (NEB E6240L). Libraries were prepared using NEB ChIP purification reagents (E6240L) and Illumina indexing primers were used to facilitate demultiplexing.