Genetic Transformation Of Brassica Napus With Insect-Resistant Gene Cry2A~* And Cry1C~* Mediated By Agrobacterium Tumefaciens

Rapeseed is one of major oil crops in China, but the yield and quality of which are highly susceptible to pests and weeds during the large scaled production, finally bringing us huge loss to rape production. In order to improve the genetic characterstics of resistance to the larva of Lepidoptera and herbicide, in this research, using two novel synthetic Bt genes Cry2A* and Cry1C* with the modification and optimization in the codon regions of the DNA sequences were transferred into one B.napus cultivar Westar via Agrobacterium-mediated transformation, we got a batch of transgenic plants. The main results of this study are as follows:1. The optimal concentration for screening Westar (B. napus) tolerant to Basta at the callus and differentiation stage was identified as 5-7mg/L by Basta sensitivity test.2. Using Agrobacterium-mediated transformation, we had obtained 18 and 42 transgenic plants with the vector of UBI:Cry2A* and 42 UBI:Cry1C*, transformation efficiency was 2.02% and 3.34%, respectively; RT-PCR analysis confirmed that the Cry2A* and Cry1C* genes are expressed stably at the transcriptional level in transgenic plants; Southern blotting analysis proved that the exogenous genes insert into genome of transgenic plants with 1-3 copies; the T1 generation plants were analyzed based on PCR, which proved that the exogenous gene in the transgenic progeny can be stably inherited.3. Insect bioassay test in To and T1 generation transgenic plants showed that Cry1C* transgenic plants can inhibit larval eating apparently, resulting in growth retardation and even death; on the contrary, Cry2A* transgenic plants didn’t exhibit significant resistance to larvae, the larve developed normally.4. The leaves of transgenic plants at T1 generation were spreaded with herbicide Basta on different concentrations, which suggested that diluted 2500 times Basta is the critical value of herbicide resistance in transgenic plants; after spraying whole plant with diluted 2500 times Basta, transgenic plants with Cry2A* and Cry1C* showed an increasing resistance to Basta compared to non-transgenic plants.