Primers were designed according to the published sequence of the apple genome and a Mdgal gene promoter of Malus× domestica Borkh ‘Gala’ were cloned. Online function prediction by PLACE and Plant CARE showed that Mdgal promoter contained a large number of phytohormone responsive elements, such as gibberellinresponsive elements (GAREmotif, Pbox), auxinresponsive element (TGAelement), MeJA responsive elements (CGTCAmotif, TGACGmotif) and some stress responsive elements. Meanwhile, to identify the upstream region of the Mdgal gene required for promoter activity, we constructed five Mdgal promoter deletion derivatives. Each deletion construct was analyzed by Agrobacteriummediated transient transformation in apple fruits and tobacco leaves. Then the transgenic apple and tobacco with GUS histochemical staining and quantitative detection of GUS enzyme activity were analyzed. The results indicated that all the five promoters had the promoter activity, and both activation and repression regulatory elements coexisted in the Mdgal promoter sequence. Positive regulatory elements may be located in a region from -1 206 to -754 bp and negative regulatory elements are present at position from -754 to -597 bp. Finally, hormone treatment assay suggested that the cisregulatory regions in the Mdgal promoter was responsive to different exogenous hormones in tobacco, and the highest level of GUS activity was observed in MeJA treatment. Taking together, our results demonstrate that Mdgal promoter with the basic structural characteristics of eukaryotes gene promoters has the up and downregulation function and hormonal treatment responsiveness, which might be involved in apple fruit development and ripening.