ChIP-Seq Protocol for Transcription Factor Binding

ChIP experiments were performed using ChIP-IT kit (Active Motif) as previously described^{71 (link)}. 5 µg of antibodies (Supplementary Table 5) were used. Immunoprecipitated-chromatin was eluted after several washes, reverse cross-linked and stored at −20 °C. ChIP-qPCRs were performed in the same way as RT-qPCR with 2 µL of ChIP or IgG samples instead of cDNA. Primer sequences are in Supplementary Table 5. Enriched DNA from EGR1-ChIP and input DNA fragments were used to generate libraries as previously described^{71 (link)}. Fifty-cycle single-end sequencings were performed using HiSeq 2000 (Illumina). Reads were aligned using human genome hg19 with BWA (v0.7.5a), peak calling assessed using MACS 2.0, and annotation done with HOMER (v4.7.2), with a p value of 0.01. Integrative Genomics Viewer (IGV 2.1) was used for representation.