MOLECULAR ANALYSIS OF SULFONYLUREA HERBICIDE RESISTANT ALS GENES

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The enzyme acetolactate synthase (ALS) is the target of several classes of herbicides, including the sulfonylureas, the imidazolinones, and the triazolopyrimidines. ALS is required for the biosynthesis of isoleucine, leucine and valine. We have cloned ALS genes from a variety of monocotyledonous and dicotyledonous plants. The number of ALS genes in these species varies from one, as in Arabidopsis and sugar beets, to two or more, as in corn and soybeans. The deduced amino acid sequences of the mature plant ALS genes which have been sequenced are closely related. Three domains of the plant ALS proteins are also conserved relative to bacterial and yeast ALS proteins. ALS genes have been isolated and sequenced from sulfonylurea herbicide resistant plants and the resistance trait shown to result from single or double mutations in the conserved domains of the genes. To produce additional genes able to confer sulfonylurea herbicide resistance in transgenic crops, in vitro-generated mutations were introduced into the coding regions of cloned plant ALS genes. A number of single and double mutations were shown to be effective in conferring resistance, upon subsequent introduction of the altered genes into plants. The mutations conferred varying degrees of resistance, depending upon the site of the resulting amino acid substitution and upon the amino acid substituted at that site. Replacement of ALS regulatory sequences led to increases in the proportion of herbicide resistant enzyme such that up to 90% of the total ALS activity was herbicide resistant. The altered genes conferred sulfonylurea herbicide resistance in glasshouse trials of a wide variety of transgenic crop species. Additionally, transgenic tobacco, tomato and oilseed rape were shown to be herbicide resistant in field trials.