A novel mutant acetolactate synthase gene from rice cells, which confers resistance to ALS-inhibiting herbicides

Abstract

A two-point mutated gene of acetolactate synthase (ALS) was isolated from rice cells, which was cultured together with an ALS-inhibiting herbicide, bispyribac-sodium (BS). The mutations involved residues of tryptophan at position 548 to leucine (W548L) and serine at position 627 to isoleucine (S627I). The ALS expressed in Escherichia coli from this gene showed resistance to multiple herbicides including pyrimidinylcarboxylate (PC), sulfonylurea and imidazolinone herbicides, and showed stronger resistance to PC herbicides than to other herbicides. BS, a PC herbicide, had almost no effect on the enzyme even at 100 mM, which is an approximately 10,000-fold higher concentration than the concentration required for 50% inhibition of the wild-type. The resistance level of W548L/S627I mutating ALS to BS was stronger than the additive effect predicted from the degree of resistance of each single amino acid mutated ALS. Transformed rice cells carrying this gene and a regenerated rice plant expressed resistance to BS, suggesting that this gene is useful as a selectable marker for introducing foreign traits into rice when used with PC herbicides.

Functional analysis of transgenic rice plants expressing a novel mutated ALS gene of rice

“…In our preceding paper, 3) we generated T 0 transgenic rice plants harboring the W548L/S627I mutated ALS gene through hygromycin selection, and confirmed that heterozygous sequences of endogenous and mutated ALS were detected in genomic DNAs of transgenic plants. One of these plants, W38, showed resistance to BS at 1 kg a.i./ha application.…”

“…2) We have isolated a novel mutated ALS gene from rice cells, which confers resistance to PC herbicides. 3,4) Before isolation of this mutated ALS gene, no paper had reported a mutated ALS gene conferring resistance specific to PC herbicides. The mutated gene had two point mutations resulting in W548L/S627I mutations in the ALS protein.…”

“…The mutated ALS protein showed similar specific activity to that of the wild type. A transgenic rice plant harboring this gene, which was obtained through hygromycin selection by Agrobacterium-mediated genetic transformation, showed high resistance to BS, 3) suggesting that this novel mutated ALS gene could be used as a selectable marker in combination with BS. Indeed, it has been shown that rice plants transformed with this gene driven with rice ALS promoter are efficiently selected by BS using the Agrobacterium-mediated transformation method.…”

Transgenic Tall Fescue and Maize with Resistance to ALS-Inhibiting Herbicides

“…Double mutations have been found in the rice ALS gene through cell culture using bispyribac-sodium (BS), a PC herbicide (Kawai et al, 2007b). Before isolation of the mutated ALS gene, no paper had reported a mutated ALS gene as conferring resistance to PC herbicides.…”

“…Table 1. Mutations in ALS conferring resistance to ALS-inhibiting herbicides (Adapted from Kawai et al, 2007b).…”