The acetohydroxy acid synthase S627N mutation confers herbicide tolerance in rice, and the rice variety containing this mutation produces good yields. This variety is commercially viable at Shanghai and Jiangsu regions in China. Weedy rice is a type of rice that produces lower yields and poorer quality grains than cultivated rice. It plagues commercial rice fields in many countries. One strategy to control its proliferation is to develop rice varieties that are tolerant to specific herbicides. Acetohydroxy acid synthase (AHAS) mutations have been found to confer herbicide tolerance to rice. Here, we identified a single mutation (S627N) in AHAS from an indica rice variety that conferred tolerance against imidazolinone herbicides, including imazethapyr and imazamox. A japonica rice variety (JD164) was developed to obtain herbicide tolerance by introducing the mutated indica ahas gene. Imidazolinone application was sufficient to efficiently control weedy rice in the JD164 field. Although the imazethapyr treatment caused dwarfing in the JD164 plants, it did not significantly reduce yields. To determine whether the decrease of the ahas mRNA expression caused the dwarfism of JD164 after imazethapyr application, we detected the ahas mRNA level in plants. The abundance of the ahas mRNA in JD164 increased after imidazolinone application, thus excluding the mRNA expression level as a possible cause of dwarfism. Activity assays showed that the mutated AHAS was tolerant to imidazolinone but the catalytic efficiency of the mutated AHAS decreased in its presence. Moreover, the activity of the mutated AHAS decreased more in the presence of imazethapyr than in the presence of imazamox. We observed no difference in the AHAS secondary structures, but homology modeling suggested that the S627N mutation enabled the substrate to access the active site channel in AHAS, resulting in imidazolinone tolerance. Our work combined herbicides with a rice variety to control weedy rice and showed the mechanism of herbicide tolerance in this rice variety.