A novel mutated acetolactate synthase gene conferring specific resistance to pyrimidinyl carboxy herbicides in rice

Acetolactate synthase (ALS) is the first common enzyme in the biosynthetic pathway of branched-chain amino acids. Mutations of specific amino acids in ALS have been known to confer resistance to ALS-inhibiting herbicides such as sulfonylureas and pyrimidinyl carboxy (PC) herbicides. However, mutations conferring exclusive resistance to PC have not yet been reported to date. We selected PC resistant rice calli, which were derived from anther culture, using one of the PCs, bispyribac-sodium (BS), as a selection agent. Two lines of BS-resistant plants carrying a novel mutation, the 95th Glycine to Alanine (G95A), in ALS were obtained. In vitro ALS activity assay indicated that the recombinant protein of G95A-mutated ALS (ALS-G95A) conferred highly specific resistance to PC herbicides. In order to determine if the ALS-G95A gene could be used as a selection marker for rice transformation, the ALS-G95A gene was connected to ubiquitin promoter and introduced into rice. PC resistant plants containing integrated ALS-G95A gene were obtained after selection with BS as a selection agent. In conclusion, novel G95A mutated ALS gene confers highly specific resistant to PC-herbicides and can be used as a selection marker.