草鱼呼肠孤病毒内衣壳蛋白VP6和增强绿色荧光蛋白在昆虫细胞中的构建和共表达.pdf

VIROLOGICA SINICA, October 2007, 22 (5):397-404 CLC number: Q786 Document code: A Article ID: 1674-0769(2007)05-0397-08 Construction and Co-expression of Grass Carp Reovirus VP6 Protein and * Enhanced Green Fluorescence Protein in the Insect Cells 1** 2 1# 1,3 Qin FANG , Eng Khuan Seng , Wen DAI and Lan-lan ZHANG (1. State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China; 2. School of Chemical Life Sciences, Nanyang Polytechnic, 180 Ang Mo Kio Ave 8, 569830, Singapore; 3.Graduate school of the Chinese Academy of Sciences,Beijing 100039,China) Abstract: Grass carp reovirus (GCRV), a disaster agent to aquatic animals, belongs to Genus Aquareovirus of family Reoviridea. Sequence analysis revealed GCRV genome segment 8 (s8) was 1 296 bp nucleotides in length encoding an inner capsid protein VP6 of about 43kDa. To obtain in vitro non-fusion expression of a GCRV VP6 protein containing a molecular of fluorescence reporter, the recombinant baculovirus, which contained the GCRVs8 and eGFP (enhanced green fluorescence protein) genes, was constructed by using the Bac-to-Bac insect expression system. In this study, the whole GCRVs8 and eGFP genes, amplified by PCR, were constructed into a pFastBacDual vector under polyhedron (PH) and p10 promoters, respectively. The constructed dual recombinant plasmid (pFbDGCRVs8/eGFP) was transformed into DH10Bac cells to obtain recombinant