Figure 2

CSE2 (YY1 binding sites) does not function as an enhancer blocker. In order to determine the minimal regions for the previously reported insulator activity in human PEG3-DMR [9], the hYY1-1 fragment was further divided into 4 smaller fragments with 200-bp size (hYY1-a through -d), and these fragments were subcloned into the pNI-CD vector to test insulator activity. These inserts were localized between the erythroid-specific enhancer (E) and Neomycin resistance gene (Neo) that is driven by the γ-globulin promoter. The values on the X axis of the graph indicate the fold difference of the survived colony number in the G418 selection relative to the value of the negative control lacking any insulator, pNI-CD(AscI). The last fragment, hYY1-d, did not show any insulator activity, and thus was omitted in this figure. The two YY1 binding sites (oval) within the first fragment, hYY1-a, were mutated and tested again, which is indicated by hYY1-a*. The mutated YY1 binding sites are indicated by black ovals.