Development Of Transgenic Rice Tolerant To Two Herbicides

The total loss resulted from weeds is valued approximately at100billion US$worldwide. Herbicides are widely used as a perfered method for weed control. Nicosulfuron and Mesotrione are two major herbicides that have been used widely for weed control. Nicosulfuron is a highly effective broad-spectrum sulfonylurea herbicide. It is mainly used in corn field and is very useful in controlling most annual grasses, some broad-leaf weeds and some sedge. Nicosulfuron can kill weeds by inhibiting the action of acetolactate synthase (ALS), thus causing the lack of valine, leucine and isoleucine. Nicosulfuron is widely welcomed for its good herbicidal property, miscibility, and lower dosage. Mesotrione is an effective broad-spectrum cyclohexanedione-type herbicide. It is useful to control most of the broad-leaf weeds and some of the grass weeds. Mesotrione is an effective inhibitor of4-hydroxyphenylpyruvate dioxygenase (HPPD), which is a key enzyme to regulate the biological synthesis of plastoquinone and tocopherol in plant. Since plastoquinone is a critical assistant factor of phytoene desaturase, the application of mesotrione makes weeds albino and dead by causing the failure of plastoquinone. Mesotrione can efficiently control the weeds resistant to triazine and inhibitor ALS herbicide.We scanned the various weeds by spraying nicosulfron and observed the Zoysia sinica is highly resistance to nicosulfuron. We postulated that this plant may have P450genes degrading nicosulfron. Primers were designed according to conservative nucleotide regions of rice CYP81A6(DQ341412.1), a barley P450(AK363181.1) and a ryegrass P450(AF321855.1). A1445bp fragment was amplified from Zoysia sinica genome. The full-length DNA of this P450gene was then obtained by tail-PCR method, and was named KC3-1.Agrobarcterium T-DNA vector was constructed for plant expression of KC3-1, and was used to transformed rice. During the rooting stage, the transgenic rice plants were transferred on medium containing nicosulfuron (0.1mg/L) for analyzing the resistance to nicosulfron. The results indicated that KC3-1transgenic rice seedling had resistance to nicosulfuron. Total of30independent events were obtained. In the hydroponic culture stage in green house, all transgenic rice plants were sprayed with80mg/L nicosulfuron. We found that20of the30events showed significant nicosulfron resistance. Further testing suggested that event KC3-1-7is one the best in tolerance to nicosulfuron, with tolerance up to240mg/L～320mg/L without visible damage.During4-5leaves stage of seedling, we assayed the resistance of K.C3-1-7transgenic rice to herbicides of different modes of actions, mesotrione, duel, paraquate and metolachlor. The results indicated that KC3-1-7transgenic rice is resistant to mesotrione. The tolerance of KC3-1-7to mesotrione was up to300mg/L～400mg/L.Management of development of tolerance by weeds to herbicides is a major challenge. One way to prevent or delay the resistance development is to apply combined herbicides with different modes of actions. This study may open a technology for utilizing nicosulfron and mesotrione together in weed control in transgenic crops. Furthermore, the transgenic plants could be used as a remedy to soil and water that polluted by nicosulfron and mesotrione.