Control Of Citrus Tristeza Virus By Cross-protection And RNAi Technologies

Citrus tristeza virus has seriously destroyed the citrus industry from 1930' in Brazil, and it is considered a major threat for the citrus industry. CTV produce decline and death of scion varieties which grafted on sour orange, especial pummelo and sweet orange. It is one of the most destructive and economically important diseases of commercial citrus worldwide. Most pummelo cultivars and sweet orange are susceptibled to CTV, especially the stem pitting citrus trisetza virus, which has emerged to be a threat to the development of citrus industry since 1980'when pommelo and sweet orange cultivation become increasing popular in China, some producing serious disease. Mild strain cross protection developed in western countries, has been proven to be one of the most effective ways in preventing sensitive citrus varieties from CTV damage where both severe CTV strains and brown citrus aphid co-exist. RNAi is an antiviral inherent mechanism in plant. Various studies revealed the expression of dsRNA in virus-resistant transgenic is efficiency and superiority of dsRNA-derived as a viable tool to achieve the silencing of plant virus. In this study, MSCP and RNAi are as the tool of studying the methods to control citrus tristeza virus.1. Pathogenicity of three CTV mild isolates(W6,W13,W17) have been identified by the biological methods, and combined with the biological symptoms and the gene p23 expressing analysis, the effect of cross protection(MSCP) has been estimated too. The results show that p23 gene group has consistency with the pathogenicity of CTV isolate in China, which can be applied to screening samples by PCR. Three mild isolate show differences in the cross protective, in which W17 isolate presented significant antagonistic affect. The homology of the gene p23 does not determine the protective effect by the resulets of gene analysis. Differences in the protective effect may be related to the multiple gene coordinate regulation to MSCP, the specialization of MSCP or other factors.2. According to the sequence of CTV p23 gene, we designed two pairs of specific primers with restriction site for the conservative region, and obtained two specific PCR fragments through PCR amplification. With the binary vector pBI 121, the RANi vector pCTV23- rnai is constructed.3. The RNAi vector pCTV23- rnai was transferred into'DA HONG'sweet orange via Agrobacterium mediated transformation.The npt II and the CTV-LS as primers were respectively successful confirmed in transformed citrus by PCR.