Figure 4.

EMSA of SND1 Binding to the Promoter Sequence of MYB46 Gene.

The NAC domain of SND1 fused with MBP was expressed in E. coli, and the purified recombinant protein was incubated with biotin-labeled MYB46 promoter fragments. The samples were subjected to EMSA by polyacrylamide gel electrophoresis. The biotin-labeled DNA fragments were detected with the chemiluminiscent method.

(A) Diagram of the MYB46 promoter showing the DNA fragments used for EMSA. The positions of individual fragments relative to the start codon are shown above the line.

(B) EMSA of SND1 binding to a 553-bp fragment (located between −60 and −612 relative to the start codon) of the MYB46 promoter. MBP was used as a control protein. For competition analysis, unlabeled DNA fragments (competitors) in 30-fold (+) or 60-fold (++) molar excess relative to the labeled probes were included in the reactions.

(C) EMSA of SND1 binding to different regions of the MYB46 promoter. The locations of DNA fragments used for EMSA are shown in (A). The MYB46-P2 and MYB46-P6 fragments exhibited strong binding by SND1.