T-DNA TAGGING OF A SEED COAT-SPECIFIC CRYPTIC PROMOTER IN TOBACCO

T-DNA tagging with a promoterless beta-glucuronidase (GUS) gene genera ted a transgenic Nicotiana tabacum plant that expressed GUS activity o nly in developing seed coats. Cloning and deletion analysis of the GUS fusion revealed that the promoter responsible for seed coat specifici ty was located in the plant DNA proximal to the GUS gene. A 3.3 kb fra gment corresponding to the insertion site was isolated from untransfor med plants. No long open reading frames were detected in this region. Northern blots and RNase protection assays failed to detect transcript s from this region in untransformed plants. Furthermore, the insertion site was situated within the N. tomentosiformis genome of the allotet raploid species N. tabacum, in a region which is not conserved within the genus Nicotiana. It is concluded that seed coat-specific GUS expre ssion in this transgenic plant resulted from T-DNA insertion next to a cryptic promoter. These results suggest that at least some of the fus ions generated to marker genes in promoter trapping studies are not as sociated with conventional gene promoters. The possibility that simila r insertion events play a role in gene evolution is discussed.