Study On The Resistant Mechanism Of Japanese Foxtail (Alopecurus Japonicus Steud.) To Haloxyfop-R-Methyl

The Japanese foxtail(Alopecurus japonicus) was a kind of worst grass weeds in oilseed rape and wheat fields in China. The occurrence of resistance to haloxyfop-R-methyl in A. japonicus had been reported, while its resistance mechanism remained unknown. In this paper, the resistant and sensitive populations of A. japonicus were used to identify the target enzyme resistance mechanism and to clone their acetyl coenzyme A carboxylase (ACCase) gene sequences on the basis of previous studies. The activity of cytochrome P450 (cytochrome P-450 monooxygenases, P450) reductase and glutathione-S-transferase (GST) were determined to enclose their role in resistance. Also the activeity of SOD (Superoxide dismutase), CAT (Catalase), POD (Peroxidase) and the contents of H2O2 (Hydrogen Peroxide), MDA(Malondialdehyde), chlorophyll were assayed to illuminate the mechanism of oxidative stress resistance Based on the above comprehensive of the final results, the resistance mechanisms of Alopecurus japonicus to haloxyfop-R-methyl was discovered initially.To illuminate the resistance mechanism on molecular level,three pairs of PCR primers were designed to specifically amplify fragments from the gene encoding CT region of chloroplastic ACCase in resistant and suspectible populations of A. japonicus. Other primers were designed to amplify other parts of ACCase.By comparing the sequence of resistant and sensitive plants, five point mutations were found (Ala-1686-Thr, Arg-1734-Gly, Met-1738-Leu, Ile-1826-Asn and Ile-2041-Asn). An amino acid mutation (Ile2041â†'Asn2041) found in resistant had been reported to confer resistance of A. myosuroides and Lolium rigidum to aryloxy-phenoxypropionate (AOPP),which was inferred to be one of the resistant mechanisms in A. japonicus to haloxyfop-R-methyl.The other mutant sites had not been reported before and their correlation with resistance is unknown.The cytochrome P450 monooxygenases(Cyt P450) reductase and glutathione- S-transferases (GST, EC2.5.1.18) activity of resistant and sensitive in Alopecurus japonicus were detected in order to clarify their relationship with metabolic resistance. The activity of Cyt P450 and GST in resistant plants were higher than that in sensitive ones without herbicide treatment. P450 reductase activity in both resistant and susceptible individuals showed increased firstly and then decreased.GST activity in the resistant biotype showed lower-higher, whereas sensitive ones for the lowe-higher-lower-increased trend.In sensitive populations, Cyt P450 and GST activity showed a trend of sharply changes, while the resistant population showed flat trend after herbicide spraying. The activity of Cyt P450 and GST in sensitive population were declined precede resistant one, and its activity was significantly lower than that of resistant population. Ten days later, the activity reached to the lowest level and remained below the initial. The results showed that haloxyfop-R-methyl may be detoxified by the catalyzing of Cyt P450, and then transformed into non-toxic or low toxic compounds under the GST's catalysis in resistance A. japonicus to prevent them from being harmed. This indicated that the increased metabolic activity may be one of the resistant mechanisms in A. japonicus to haloxyfop-R-methyl.The activity of SOD, CAT, POD and contents of H2O2, MDA, chlorophyll in resistant and suspectible populations of A. japonica were detected. Significant differences of all the indexes mentioned above were observed in resistant and sensitive populations after treated by haloxyfop-R-methyl. There was no significant change in resistant populations and the trend of resistant populations was slower than the sensitive one with the extension of herbicide stress time. After 14 days, all kinds of biochemical indicators of sensitive A. japonicus reached the extreme value. Therefore, resistance due to oxidative stress may be one of the mechanisms of resistance in A. japonicus.