Ipure beads v2

ChIP-seq assays were performed as described previously (Marie et al; 2018) , using iDeal ChIPseq kit for Transcription Factors (Diagenode, C01010055). Briefly, O4 + MACSorted cells were fixed in 1% formaldehyde (EMS, 15714) for 10 min at room temperature and the reaction was quenched with 125 mM glycine for 5 min at room temperature. Lysates were sonicated with a Bioruptor Pico sonicator (Diagenode, total time 8 min) and 4μg of antibodies were added to sheared chromatin (from 4 million cells for Olig2 and from 1 million cells for histone marks) and incubated at 4°C overnight on 10 rpm rotation. Antibodies used were: mouse anti-Olig2 antibody (Millipore, MABN50), rabbit anti-H3K4me3 antibody (Active motif, 39060), rabbit anti-H3K27Ac antibody (Active motif, 39034), rabbit anti-H3K4me1 antibody (Ozyme, 5326T), mouse anti-H3K27me3 antibody (Abcam, ab6002). Mock (Rabbit IgG) was used as negative control. Chromatin-protein complexes were immunoprecipitated with protein A/G magnetic beads and washed sequentially according to the manufacturer (Diagenode, C01010055). DNA fragments were then purified using IPure beads v2 (Diagenode, C01010055). Input (nonimmunoprecipitated chromatin) was used as control in each individual experiment.
The ChIP-seq libraries were prepared using ILLUMINA Truseq ChIP preparation kit and sequenced with ILLUMINA Nextseq 500 platform.