Chromatin immunoprecipitation detects Zfp423 binding in <i>Tulp3</i> intron 1.

<p>(<b><i>A</i></b>) UCSC browser window on <i>Arl4d</i> locus shows location of the ChIP-PCR assay (Arl4d_1) and a ChIP-Seq peak call from P19 cells (chr11-190), 7 kb 5’ to the annotated transcript (Arl4d). The peak notation indicates the 190^th best-supported peak on chromosome 11 in that experiment [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1006357#pgen.1006357.ref032" target="_blank">32</a>]. (<b><i>B</i></b>) Similar window on <i>Tulp3</i> locus includes 5 potential peaks from P19 ChIP-Seq data, three of which supported reliable quantitative PCR assays (Tulp3_1, _2 and _3) within 150 bp of the peak. (<b><i>C</i></b>) Graphs show fold enrichment relative to an IgG control for a control locus and the four candidate binding sites. Colored dots indicate measured values from independent biological replicates, bars indicate median values for each locus. A commercial anti-Zfp423 antibody developed in goat (green dots) showed modest but significant enrichment for Tulp3_3. A custom antibody developed in rabbit and affinity purified against the immunogen (blue dots) showed strong enrichment for Tulp3_3 and possible enrichment for Tulp3_2, 3 kb away. Non-parametric analysis of the combined data provides strong and conservative statistical support for binding at Tulp3_3 (p = 0.0087, Wilcoxon rank sum test with Bonferroni correction).