Identification And Analysis Of The Progeny Derived From Transgenic Brassica Napus With Disease-resistance And Insect-resistance

Rapeseed is one of the most important oil crops in our country. Its planting area and yield is at the leading position in the world. Rapeseed has always been subjected to the threat of biotic stresses in the whole life, such as disease, pests and weeds. Biotic stresses may lead to yield losses every year. In order to improve yield stability, the transgenic technology can be utilized to improve the resistance to disease or pests stresses in rapeseed.In this study, the insect-resistant gene Cry1C, Cry2A were first individually transferred into Brassica napus7-5or Westar by Agrobacterium-mediated transformation, and the plants with both Cry1C and Cry2A genes were later obtained by pyramiding. Moreover,the rice disease-resistant gene Ospgip was successfully transformed into B. napus materials7-5, Westar, P61-5and T45, respectively. The resistance or tolerance to Sclerotinia sclerotiorum of the transgenic plants has been improved. The main results of this study are as follows:1. By Agrobacterium-mediated transformation,169individuals with Ospgip gene,57individuals with Cry1C and39individuals with Cry2A were obtained. The positive rates of transgenic plants were90.4%,33.5%,48.8%respectively. PCR, RT-PCR and real-time PCR analysis of the transgenic progenies confirmed that Ospgip, Cry1C, Cry2A genes were stably inherited and expressed.2. The leaves from42transgenic seedlings of Ti generation which contained Ospgip gene were inoculated with Sclerotinia sclerotiorum in vitro. The results showed the plaque average sizes of33transgenic lines were less than the control. The plaque average sizes of33transgenic lines were less than the control inoculated at the adult stage.The correlation coefficient (r<0.2) of diseases resistance between the seedling stage and the adult stage of transgenic plants was no significant.3. Expression analysis of Ospgip gene in detached leaves after inoculation revealed enhanced expression in the transgenic plants. Coefficient (r=-0.72154) between the Ospgip expression and the plaque size was significant. 4. The Sclerotinia resistant test on the transgenic plants of three receptor materials (7-5, P61-5and T45) showed the disease-resistance have been evidently improved than the control.5. Insect bioassay test in To and T1generation transgenic plants showed that all plants with Cry1C exhibited remarkably resistace when feeding with Plutella xylostella larvae. The To transformants with Cry2A gene showed a mild resistance to the insect, while the T1transgenic plants had no different insect-resistance than the control.6. The insect-resistance of the transgenic plants pyramided Cry1C and Cry2A genes was stronger than the transgenic plants with single Cry2A gene, but no difference was observed between the pyramided and the Cry1C transformants.7. The transgenic plants with Cry1C/Bar and Cry2A/Bar were sprayed by Basta in field condition. The transgenic plants showed vigorously growth, but the non-transgenic plants became curly and yellow then withered in leaves, and finally died.