草莓镶脉病毒侵染性克隆的构建及其ORFVI基因的原核表达.pdfVIP

Abstract Strawberry vein banding virus (SVBV) is a latent virus inducing very serious damage in strawberry plants all over the world. Recently, SVBV is detected in the Henan, Hebei and Jilin provinces in China and causes severe losses. SVBV is classified as a member of Caulimoviridae. SVBV has a double-stranded and circle DNA genome of approximately 7.8 kb and contains 7 ORFs which encode different functional proteins. SVBV infectious clone was constructed to study the replication, movement of virus and the relationship between virus and protein factors of host plant. In addition, pathogenicity of SVBV has close relationship with P6 protein encoded by gene ORF VI of SVBV. The gene ORF VI was cloned and prokaryotic expression and antiserum preparation were carried out. The work establish the basis to research the concrete functions of P6 protein and has the significant effect on illuminating the pathogenic mechanism of plant virus. 1. Construction of an infectious clone of SVBV The pSVBV-DE3 is a clone plasmid of complete sequence of SVBV about 7.8 kb, containing a single BamH I and a single EcoR I. An infectious clone of SVBV was constructed by the two restriction enzymes. 0.5SVBV about 4 kb was digested from pSVBV-E3 via BamH I and EcoR I and was inserted into binary vector pBinPLUS via the same enzymes, forming pBin-0.5SVBV. Then pSVBV-E3 was digested by EcoR I to obtain the complete SVBV. Finally, the recombination vector pBin-1.5SVBV was obtained by inserting complete sequence of SVBV into pBin-0.5SVBV. 2. Transformation, inoculation and detection of the infectious clone of SVBV The recombination plasmid pBin-1.5SVBV was transformed into Agrobacterium tumefaciens GV3101 and the positive clone was