ChIP-seq in rat OPCs and iOLs, and mouse iOLs were performed as previously described7, with slight modifications. Briefly, for ChIP-seq, approximately 5 ×106 cells were cross-linked with 1% formaldehyde for 10 min at room temperature and then were quenched with 125 mM glycine for 5 min. Sonicated chromatin was used for immunoprecipitation by incubation with 4 μg appropriate antibodies (anti-OLIG2, Millipore, cat# ab9610; anti-H3K9me3, Active Motif, cat# 39161; anti-HA tag, Sant Cruz, cat# SC-7392) overnight at 4 °C. Immunoprecipitated complexes were collected using 40 μl protein A plus agarose beads (Millipore, cat#16-156). Subsequently, beads were washed sequentially with low-salt buffer, high-salt buffer, LiCl buffer and twice with TE and elution in 500 μl of elution buffer (1% SDS, 0.1 M NaHCO3). The elutes were heated at 42 °C for 2 h and treatment with proteinase K followed by 65 °C 10 h to reverse the cross-linking and were treated with RNaseA for 30 min before DNA was extracted and purified. The ChIP libraries were prepared using KAPA HyperPrep Kits (Roche, 07962347001), and then run on the Illumina sequencer Hiseq-Xten PE150.
