Fig. 6. An SCL-E12 complex binds specifically to the Gata2 VE-enhancer E box. No extract (lane 1) or nuclear extracts from 293T cells, which were mock transfected (lane 2), transfected with SCL alone (lane 3) or with SCL plus E12 expression plasmids (lanes 4-11), were incubated with radiolabeled E box oligonucleotide probe. To demonstrate binding specificity, unlabeled competitors (20- or 200-fold excess) containing wild-type (wt; lanes 5,6) or mutant (mut; lanes 7,8) E box, anti-SCL antibody (1 or 3 µl; lanes 9,10) or control mouse IgG (lane 11) were added to separate binding reactions. Formation of a lower-mobility complex was observed only when both SCL and E12 were present in the extract and was specifically disrupted by wild-type, but not mutant, competitors as well as by the addition of an anti-SCL antibody.