C/EBPϵ can directly bind to the Mad1 promoter in vivo. C/EBPϵ-ER MPROs were treated with 200 nM 4-OH-tamoxifen (Tam). ChIP was performed using an anti-C/EBPϵ antibody and the binding of C/EBPϵ to the Mad1 promoter assessed by quantitative real-time PCR. (A) Schematic representation of the murine Mad1 gene, with consensus C/EBP-binding sites and PCR amplicons as indicated. (B) Representative quantitative PCR plot showing amplification of site D. NTC indicates no template control; input, total amount of input chromatin; no Ab, no antibody control immunoprecipitation. The arbitrary amplification threshold is depicted as the horizontal bar running across the graph. (C) Quantitation of C/EBPϵ binding to the Mad1 promoter 2 and 4 hours after the addition of Tam for 4 consensus C/EBP-binding sites. Data expressed as fold enrichment relative to 0 hour time point from a representative experiment (n = 3 experiments).