Title: Anther Specific Gene1(AEG1) promoter from cotton; its modification for tapetum specific expression
Abstract:
Tapetum-specific promoters are key to developing transgenics with barnase/barstar gene (for male sterility/restorer lines) for hybrid seed production. A gene expressing in the anther tissue named AEG1 and it’s 1.5kb upstream region (promoter) was isolated from cotton. The promoter was found to be active mainly in the tapetum tissue of cotton, in addition to weak activity in the roots. In order to modify the AEG1 promoter for its tapetum specificity, two promoters were developed (i) AEG1ΔB wherein the -230bp to -500bp of the promoter was deleted (the region carries several copies of the root-specific cis-elements, ROOTMOTIFAPOX1) and (ii) AEG1ΔBMut where the root motifs in the proximal region were mutated in the background of AEG1ΔB. The activity of the modified promoters was compared with that of wild-type AEG1 promoter in tobacco transgenics using β-glucuronidase as a reporter gene. It was observed that in tobacco the WT promoter drives its expression in the pollen and not in the tapetum and was also active in roots. In the case of the modified promoters, AEG1ΔB and AEG1ΔBMut, their activity in roots was reduced and abolished, respectively. Both modified promoters retained their activity in pollens. Further, transgenics in tobacco expressing the barnase gene under the wild-type AEG1 and AEG1ΔBMut promoters were developed. No proper transgenics could be developed with wild-type AEG1 promoter, whereas several male sterile lines were found with AEG1ΔBMut promoter. These lines did not set seed on self-pollination but did set seed following cross-pollination with untransformed tobacco lines. The possibility of using the modified promoter AEG1ΔBMut to develop male sterile plants in cotton will be discussed
