FUNCTIONAL-ANALYSIS OF 2 BEAN NITRATE REDUCTASE PROMOTERS IN TRANSGENIC TOBACCO

Transcription of nitrate reductase (NR) genes is regulated by nitrate as well as a variety of other environmental and developmental factors. Two NR genes have previously been characterized in bean (Phaseolus vu lgaris). Both genes are nitrate- and light-inducible and their mRNA ex pression level displays a circadian rhythm. The two genes, NR1 and NR2 , show differences in developmental and tissue-specific expression. In order to deliniate elements in the NR1 and NR2 promoters that are inv olved in these expression patterns, eight 5' deleted Versions of the N R1 promoter and one fragment of the NR2 promoter were cloned in front of the GUS reporter gene and transformed into tobacco. Approximately 5 0% of the regenerated transformants expressed detectable levels of GUS activity. For the eight NR1 promoter deletion constructs, differences in promoter strength were observed indicating both positive and negat ive elements in the promoter. For both the NR1 and the NR2 promoter-GU S constructs no significant nitrate regulation could be detected in th e transgenic plants. A possible explanation for this is that elements needed for proper nitrate regulation of the NR1 and NR2 genes are loca ted outside the analysed 3 500 bp and 1 400 bp promoter regions, respe ctively. A diurnal rhythm regulation similar to that seen for the endo genous nitrate reductase in bean plants, could be found for the GUS mR NA expression in several of the transformants containing the NR1 and N R2 promoter-GUS constructs.