title: GSM6764932: NPC1-/- mouse O4+ cells H3K27ac; Mus musculus; ChIP-Seq
library_construction_protocol: Cells were isolated from brains of 16-day-old mice by magnetic-activated cell sorting using microbeads targeting the antigen O4 and were immediately flash frozen. Cell pellets from 6 mice were pooled together and fixed with 1% formaldehyde for 15 minutes and quenched with glycine. After cell lysis, chromatin was sonicated and the DNA sheared to an average length of 300-500 bp. Genomic DNA was prepared by treating aliquots of chromatin with RNase, proteinase K, and heat, followed by ethanol precipitation. Soluble chromatin for H3K27me3 ChIP-seq was spiked-in with soluble Drosophila chromatin equivalent to 5-10% of mouse chromatin. The mix of suble chromatin was incubated with 4 ug of antibody against H3K27me3 or H3K27ac. Complexed were eluted with SDS buffer. ChIP-DNA was purified with phenol-chloroform extraction. *Drosophila spike-in was used for H3K27me3, but not H3K27ac libraries were prepared by consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. These steps were done with an automated system (Apollo 342, Wafergen Biosystems/Takara).