Figure 7.

CES binds to G-box motifs. (A, B) ChIP experiments with wild-type and 35S_p:CES-YFP plants using an anti-GFP antibody. G-box containing fragments of the promoters of CPD and CYP718 were quantified by real-time PCR amplification from immunoprecipitated samples, with the primer pairs listed in Supplementary Table S4. The primer pair 5S-F/5S-R (Li et al, 2010) was used for standardization. The standard deviation of at least three measurements is shown. (C) EMSAs analysing CES binding to a fragment of the CPD promoter. A radioactively labelled probe representing the same part of the CPD promoter as in (A) was incubated with GST–CES in the absence or presence of cold competitor oligonucleotides. The competitors C1–C5 contain different regions (indicated by a solid line; upper panel) while the G-box (CACGTG; light grey) was deleted in C6 or mutated to AAAAAA in C7. The competitors were used in 50 and 500-fold molar excess to the probe. P, probe; DPC, DNA–protein complexes.