Fig. 2. Binding of in vitro synthesized Su(Hw) to two putative Su(Hw) binding sites in the 125 bp DNA fragment. (A) The sequence of the 125 bp DNA fragment is shown. Putative Su(Hw) binding sites are boxed. The primers used to obtain the DNA fragments are shown as arrows. The mutated residues are indicated below the sequence. The consensus for the Su(Hw) binding site was taken from Scott and Geyer (Scott and Geyer, 1999). (B) Electrophoretic mobility shift assays. The radioactively labeled gypsy, 125 bp fragment, Su(Hw)#1, Su(Hw)#2, Su(Hw)#1^*, Su(Hw)#1^**, 454 bp and 454^* bp fragments were used as probes, incubated with in vitro-synthesized Su(Hw) protein and run on a 1.5% agarose gel (Materials and Methods). One shifted band (indicated by arrows) presumably corresponds to a protein-DNA complex formed by Su(Hw) with only one Su(Hw) binding site. The binding of Su(Hw) to the 125 bp DNA fragment was examined in the presence of competitors. The binding is competed by excess unlabeled Su(Hw)#1^* fragment but not by the Su(Hw)#1^** fragment with the mutated Su(Hw) binding site.