REPRESSION OF ACETOLACTATE SYNTHASE ACTIVITY THROUGH ANTISENSE INHIBITION - MOLECULAR-ANALYSIS AND BIOCHEMICAL-ANALYSIS OF TRANSGENIC POTATO (SOLANUM-TUBEROSUM L CV DESIREE) PLANTS

Acetolactate synthase (ALS), the first enzyme in the biosynthetic path way of leucine, valine, and isoleucine, is the biochemical target of d ifferent herbicides. To investigate the effects of repression of ALS a ctivity through antisense gene expression we cloned an ALS gene from p otato (Solanum tuberosum L. cv Desiree), constructed a chimeric antise nse gene under control of the cauliflower mosaic virus 35S promoter, a nd created transgenic potato plants through Agrobacterium tumefaciens- mediated gene transfer. Two regenerants revealed severe growth retarda tion and strong phenotypical effects resembling those caused by ALS-in hibiting herbicides. Antisense gene expression decreased the steady-st ate level of ALS mRNA in these plants and induced a corresponding decr ease in ALS activity of up to 85%. This reduction was sufficient to ge nerate plants almost inviable without amino acid supplementation. In b oth ALS antisense and herbicide-treated plants, we could exclude accum ulation of 2-oxobutyrate and/or 2-aminobutyrate as the reason for the observed deleterious effects, but we detected elevated levels of free amino acids and imbalances in their relative proportions, Thus, antise nse inhibition of ALS generated an in vivo model of herbicide action. Furthermore, expression of antisense RNA to the enzyme of interest pro vides a general method for validation of potential herbicide targets.