FIG. 2.

Design and validation of Y1H system. (A) Specific DSX-DNA recognition regulates the expression of lacZ via fusion protein containing an N-terminal Gal4 AD and a C-terminal DSX domain. A 48-bp Drosophila promoter fragment derived from fbe was inserted upstream of lacZ to provide DSX-binding sites dsx^A and dsx^B. CTD, C-terminal dimerization domain of DSX. (B) Wild-type DNA-binding sites and inactive variants fbe^AB, containing two wild-type sites and control elements (respectively designated fbe^AX, fbe^XB, and fbe^XX) in which one, the other, or both sites are inactivated by twin A→G transitions at center of target sites (bold). (C) Colonies on an X-gal indicator plate in the presence of wild-type or variant enhancer elements. (D) Histogram describing β-galactosidase activity in the presence of wild-type or variant DNA sites (columns 1 to 4), empty reporter vector (column 5) or reporter (column 6) plasmid lacking fbe sites.