牛流行热病毒G1抗原表位基因在毕赤酵母中的表达和抗原性鉴定.pdfVIP

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牛流行热病毒G1抗原表位基因在毕赤酵母中的表达和抗原性鉴定.pdf

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VIROLOGICA SINICA, October 2007, 22 (5):347-352 CLC number: Q786 Document code: A Article ID: 1674-0769(2007)05-0347-06 Expression and Antigenic Characterization of the Epitope-G of the Bovine 1 * Ephemeral Fever Virus Glycoprotein in Pichia pastoris 1 1 1** 2 2 Fu-ying ZHENG , Guo-zhen LIN , Chang-qing QIU , Kui-zhang YUAN and Jun-ying SONG (1. Key Laboratory of Animal Virology of Ministry of Agriculture, State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China; 2. Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China) Abstract: The epitope-G1 gene of Bovine ephemeral fever virus (BEFV) glycoprotein was synthesised by PCR and cloned into expression vector pPIC9K to construct recombinant plasmid pPIC9K-G1. Then the pPIC9K-G1 was linearized and transformed into Pichia pastoris GS115. The recombinant P .pastoris strains were selected by a G418 transformation screen and confirmed by PCR. After being induced with methanol, an expressed protein with 26 kDa molecular weight was obtained, which was much bigger than the predicted size (15.54 kDa). Deglycosylation analysis indicated the recombinant G1 was glycosylated. Western blot and ELISA tests, as well as rabbit immunization and specificity experiments indicated that the target protein had both higher reaction activity and hi