ChIP-Seq Analysis of Ad4BP/SF-1 Binding

The DNA fragments prepared as described above were used to construct a ChIP-seq library using the NEBNext DNA sample prep master mix set 1 (NEB) and DNA Sample Prep kit, oligo only (Illumina). Adaptor-ligated DNA fragments (200–500 bp) were recovered and used for the following steps. After the quality of the library was validated on an Agilent Bioanalyzer 2100 (Agilent Technology), the ChIP-seq library was subjected to sequencing on the Illumina GAIIx platform. ChIP-seq reads were mapped to the reference mouse genome (mm10) using Bowtie^{41 (link)} (version 0.12.7), allowing up to three mismatches. Multiple-hit reads were excluded, and only reads uniquely mapped to the reference mouse genome were subjected to further analysis. To identify peaks and regions of Ad4BP/SF-1 enrichment, uniquely mapped reads were analyzed using MACS (Model-based Analysis for ChIP-Seq)^{42 (link)} program version 1.4.2 with the option “--nomodel”. The threshold P values for enriched regions of Ad4BP/SF-1 were set to 10⁻³.