Expression Pattern of Entire Cytochrome P450 Genes and Response of Defenses in a Metabolic-Herbicide-Resistant Biotype of .

Enhanced herbicide metabolism mediated by cytochrome P450s has been proposed as one of the major mechanisms of resistance to fenoxaprop--ethyl in a metabolic-herbicide-resistant biotype of Asia minor bluegrass ( Nees ex Steud.). Upon pre-treatment with the P450 inhibitor piperonyl butoxide, a remarkable reduction in metabolic rates of the phytotoxic fenoxaprop- has been observed in the resistant plants, implying that constitutive and/or fenoxaprop--ethyl-induced up-regulation of specific P450 isoforms are involved in the fenoxaprop--ethyl resistance. However, which P450 gene(s) were responsible for the metabolic resistance is still unknown. In this present study, based on the abundant gene resources of established previously, a total of 48 putative P450 genes were isolated from the metabolic-herbicide-resistant plants and used for gene expression analysis. The most suitable reference genes for accurate normalization of real-time quantitative PCR data were first identified in and recognized as actin (), 18S rRNA (), and ribulose-1,5-bisphosphate carboxylase oxygenase () under fenoxaprop--ethyl stress, glyceraldehyde-3-phosphate dehydrogenase () and elongation factor 1α () under mesosulfuron-methyl stress, and , , eukaryotic initiation factor 4a (), and 25S rRNA () at different growth stages. Expression analysis of the putative P450 genes revealed that six genes, respectively, annotated as , , , , , and were up-regulated more than 10-fold in the resistant plants by fenoxaprop--ethyl treatment, and all of them exhibited constitutively and/or herbicide-induced higher transcript levels in the fenoxaprop--ethyl-resistant than in the susceptible plants. Three genes, respectively, annotated as , , and constantly up-regulated in the resistant than in the susceptible plants after fenoxaprop--ethyl treatment. Up-regulated expressions of these specific P450 genes were consistent with the higher P450 contents determined in the resistant plants. These results will help to elucidate the mechanisms for P450-mediated metabolic-herbicide resistance in as well as other grass weed species.