Abstract: DREB (dehydration-responsive element-binding protein) transcription factors play important roles in the stress response and regulation of plants growth and development. In traditional DNase I Foot printing, DNA probes is labeled with isotope and then performed polyacrylamide gel electrophoresis to separate digested labeled-DNA fragments, which takes various steps, with low differentiation rate and to detect fewer samples. To explore the mechanism oftranscriptional regulation of DREB3 in soybean, we used an improved DNase I Foot-printing method combining with EMSA (electrophoretic mobility shift assay) to identify binding region of proteins and to find out the core element in binding site. In this research, DNA was labeled using fluorescence instead of isotope and automated capillary electrophoresis polyacrylamide gel electrophoresis was replaced to detect digested DNA fragments. Finally, DNA binding site of GmMYB1 with GmDREB3 promoter was identified rapidly via the modified DNase I Foot printing. On the other hand, restriction enzyme was used to validate this result. To further confirm binding element in GmDREB3 promoter, we used a putative DNA binding element of GmMYB1 to complete EMSA, indicating that GmMYB1 can bind target DNA element in vitro. In short, compared with classic DNase I Foot printing, the modified method is more rapid, accurate and reliable, which will be advantageous as a high throughout method to largely identification of interaction between protein and target DNA sites in the future.

Key words: DNase I foot-printing, Electrophoretic mobility shift assay (EMSA), Capillary electrophoresis, Interaction between protein and DNA