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. 2013 Nov 27;15(6):R113. doi: 10.1186/bcr3580

Figure 2.

Figure 2

Regulation of the MYB promoter by ZEB1. (A) (i) Outline of several CAT reporter constructs containing MYB promoter and intron 1 sequences with various numbers of E and Z boxes, which were used in the CAT ELISA assay. (ii) Western blot of HEK 293 cells transfected with various amounts of pcDNA3.1-ZEB1 expression construct, as surrogate validation of ZEB1 expression in the CAT ELISA assay. (iii) Results of a CAT ELISA assay, “+ZEB1” expressed as fold change from each individual CAT reporter alone, n = 1; error bars represent standard deviation. (B) Scheme of the MYB gene indicating (black bars) the position of E-boxes and Z-boxes, sites at which ZEB1 may bind [4]. Red bars indicate QRT-PCR primers used in the ChIP assay. (C) ChIP analysis of SCRsh-ET and ZEB1sh-ET cells using anti-ZEB1 (E20; Santa Cruz) and control anti-goat IgG. The graph depicts the enrichment of PCR-amplified immunoprecipitated DNA expressed as a percentage of total DNA immunoprecipitated with ZEB1 antibody or control anti-goat IgG relative to unprecipitated input sample. Data are representative of three independent assays (error bars represent SD). Additional data are shown in Additional file 9: Figure S9B.