Construction And Identification Of Transgenic Rice Plants With Resistance To Rice Black-streaked Dwarf Virus Disease

Rice black streaked dwarf virus disease is one of the rice major diseases in China, and the infected plants show dwarf, increased tiller, low seed setting rate and growth abnormalities phenotypes, which affect rice production seriously. Rice black streaked dwarf virus disease is caused by rice black streaked dwarf virus (RBSDV), and Laodelphax striatellus is the main transmitting medium of RBSDV. It can spread for a long time and widely without eggs. To control rice black streaked dwarf virus disease, it is needed to control both RBSDV and Laodelphax striatellus. In order to obtain the new rice varieties which resist to RBSDV, we constructed RBSDV S3 and Laodelphax striatellus trehalose gene (Memb) RNAi transgenic plants. Then screened positive transgenic rice and identified their resistance against RBSDV. The main results are as follows:1. We successfully constructed Memb gene RNAi vector based on the identification with colony PCR, enzyme digestion and sequencing. Then this vector was transformed into rice variety Wulingjing which is susceptible to RBSDV.2. Got stable genetic Memb RNAi and S3 RNAi transgenic rice plants. To identify positive transgenic plants, screened the plants with hygromycin, and amplified the inserted target fragment with PCR, then detected the transcriptional level of target fragment using real-time PCR. At present, We identified 9 positive lines of Memb RNAi T2 generation transgenic plants and 6 positive lines of S3 RNAi T3 generation transgenic plants.3. The disease resistance of the transgenic plants with high expression of target fragment was identified by artificial inoculation of RBSBV in greenhouse. It showed that both Memb RNAi T2 generation and S3 RNAi T3 generation plants are resistant to RBSDV, and the S3 RNAi plants are more resistant than Memb RNAi plants. According to the analysis of agronomic traits, there are no significant difference of agronomic traits between Memb/S3 RNAi transgenic plants and nontransgenic control plants in the field.The results in this study provide important genetic resources, reference information and new strategies for the molecular breeding of rice with resistance to RBSDV.