A TGACGT motif in the 5'-upstream region of alpha-amylase gene from Vigna mungo is a cis-element for expression in cotyledons of germinated seeds

Alpha-amylase is expressed at high levels in cotyledons of germinated seeds of Vigna mungo. The mRNA for alpha-amylase appeared in cotyledons of the seeds at 1 d after imbibition started (DAI). Two TGACGT motifs at -445 and at -125 in the promoter region of the gene interacted with nuclear proteins from cotyledons of dry seeds and the activities were detected until 3 DAI. A transient assay with particle bombardment showed that the downstream region from -135 in the promoter was required for high level expression in the cotyledons and the activity was reduced by mutation of the TGACGT motif at -125. The activities to bind the TGACGT motifs were detected in the axes of the seeds at 1 DAI but disappeared at 4 DAI, although the mRNA for alpha-amylase in the axes appeared at 4 DAI and increased in level by 6 DAI. A transient assay experiment showed that a positive regulatory element for the expression in the axes was located in the region from -630 to -453. These results indicated that the TGACGT motif at -125 was required for high level expression of the gene in the cotyledons of the germinated seeds.