Wild oat (Avena fatua L.) (2n = 6x = 42) is an important grass weed that infests numerous crops worldwide. Herbicides inhibiting the plastidic acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) enzyme, have been widely used for wild oat control. However, recurrent use of ACCase-inhibiting herbicides has selected for resistance, resulting in the evolution of many herbicide-resistant wild oat populations. Herbicide resistance mechanisms in wild oat are known to include nonsynonymous point mutations in the ACCase target-site. To advance the study of ACCase target-site resistance in wild oat, a system was developed to test the ability of ACCase mutations to confer resistance using partial gene replacement in yeast (Saccharomyces cerevisiae).

The herbicide-sensitive domain of ACCase is found in the carboxyl transferase (CT) domain near the 3' end of the gene, and we sequenced full 3' regions of three ACCase gene copies, Acc1;1, Acc1;2 and Acc1;3, from a resistant wild oat biotype, UM1. Acc1;3 was of particular interest as its full 3' region was not previously available.

Sequence information from these three ACCase genes was used for partial gene replacement of ACCase in yeast, with the goal of confirming the ability of Cys₂₀₈₈ to Arg mutation identified in the ACCase of resistant wild oat, to confer herbicide resistance. A yeast integration plasmid was constructed to carry the susceptible wild oat sequence which was then manipulated by site-directed mutagenesis to a mutated sequence carrying Arg₂₀₈₈. Partial gene replacement was performed in wild-type yeast through homologous gene-targeting of the yeast ACCase with the mutated wild oat sequence. Yeast provides a uniform background for testing the mutation, and homologous gene-targeting eliminates the possibility of differences due to transgene position effects from random transformation of null-ACCase yeast.

Growth response of the yeast gene-replacement and wild-type (control) strains was studied in the presence of increasing concentrations of an ACCase-inhibiting phenylpyrazoline herbicide, pinoxaden. Significant differences in the growth response of yeast gene-replacement and the wild-type strains were observed at pinoxaden concentrations ranging from 4.8 × 10⁻³ g ai/ml to 19.2 × 10⁻³ g ai/ml, thus confirming the ability of Cys/Arg₂₀₈₈ mutation to confer resistance to an ACCase-inhibiting herbicide, pinoxaden.