ChIP-Seq Analysis of Transcription Factor Binding

For ChIP-Seq, 100 μg human cell line chromatin was immunoprecipitated with 5 μg mouse anti-GR antibody. ChIP-Seq libraries were sequenced on an Illumina High-Seq 2000. Sequence reads for each sample were mapped to the hg19 or mm9 genome assemblies as relevant with Bowtie (38 (link)). Duplicate reads were removed, and the remaining unique reads were normalized to 10 million reads. Peak calling was performed by HOMER (39 (link)) using an FDR cutoff of 0.001, a cumulative Poisson p-value of <0.0001, and required a 4-fold enrichment of normalized sequenced reads in the treatment sample over the control/input sample. Normalized sequence reads around each peak were counted in 25 bp bins. Motif discovery was conducted with HOMER package v4.2. In ChIP-Seq data we used the following settings: -size 150 –S 10 –bits (39 (link)). We limited the motif analysis to a 150 bp DNA region around each peak summit. Distance to gene and gene annotations for ChIP-Seq peaks were obtained using GREAT v1.82.