INDUCTION OF A WHEAT EM PROMOTER BY ABA AND OPTICALLY PURE ABA ANALOGS IN WHITE SPRUCE (PICEA-GLAUCA) PROTOPLASTS

In white spruce (Picea glauca) protoplasts, abscisic acid (ABA) and op tically pure ABA analogs induced expression of a reporter gene under r egulation of a wheat ABA-responsive promoter. A fusion of a 650 bp pro moter fragment from the wheat Em gene promoter and the Escherichia col i uidA sequence encoding beta-glucuronidase (GUS) was linked in the pl asmid pBM 113Kp. Expression of the Em-uidA fusion varied among 6 white spruce genotypes. Protoplasts from 4-day-old embryogenic suspension c ultures gave the highest GUS activity relative to other stages in the 7-day growth cycle of suspension cultures. Racemic ABA [R,S-(+/-)-ABA] induced a significant increase of protoplast GUS activity over backgr ound at a concentration of 1 x 10(-5) M, but maximum GUS activity was found at 1 x 10(-3) M. ABA stereochemistry had a significant effect on gene expression. The natural isomer of ABA [S-(+)-ABA] was an effecti ve inducer at a concentration as low as 1 x 10(-7) M, but a concentrat ion of greater than 1 x 10(-4) M was required for induction by [R-(-)- ABA]. Moreover, analogs with the same configuration at C-1' as that of natural ABA were more effective for induction of expression from the Em-uidA insert at 1 x 10(-4) M than were their enantiomers. Plasmid pB I511, carrying the chloramphenicol acetyl transferase (CAT) gene drive n by the constitutively expressed, tandemly duplicated cauliflower mos aic virus 35S promoter, was co-electroporated with pBM113Kp for monito ring the influence of addition of exogenous ABA or ABA analogs on hete rologous gene expression in protoplasts. CAT activity was not signific antly affected by the presence or absence of ABA or the analogs used.