ENHANCEMENT OF SHORT-TERM AND MEDIUM-TERM EXPRESSION OF TRANSGENES IN EMBRYOGENIC SUSPENSIONS OF PICEA-ABIES (L) KARST

Biolistic transfer and subsequent reporter gene expression was enhanced 3-20-fold in embryogenic suspension cultures of Picea abies by treatment with osmotica before and after bombardment. Myoinositol was more effective than mannitol+sorbitol and about as effective as sucrose in enhancing expression id after bombardment. For both myoinositol and sucrose the optimal concentration was 0.25 M. Abscisic acid interacted with myoinositol to give a further enhancement. Maintaining the cells on 0.25 M myoinositol after bombardment enhanced the expression measured 10 d later. These results provide evidence for a metabolic component of the mechanism of osmotic enhancement in addition to an effect depending on partial plasmolysis at the time of bombardment. Expression driven by the polyubiquitin promoter 10 d after bombardment was more than twice that driven by an enhanced 35S, and 20 times that driven by a simple 355 promoter. Experiments using plasmids with and without scaffold attachment regions indicated that a greater proportion of the cells that expressed the reporter gene at 6 d compared with 1 d after bombardment had been integratively transformed.