Citation: ZHONG Jin-dong, HUA Qun-yi, XIAO Rong-hai, XIA Xue-shan, YANG Yun-qing, ZHOU Xiao-li, DEN Zu-hong, . Cloning and Expression of VP1 Gene of Swine Vesicular Disease Virus .VIROLOGICA SINICA, 2006, 21(4) : 385-389.

Cloning and Expression of VP1 Gene of Swine Vesicular Disease Virus

  • Corresponding author: HUA Qun-yi, 
  • Available online: 20 July 2006
  • The VP1 gene of Swine vesicular disease virus (SVDV) was amplified by reverse transcription-polymerase chain reaction (RT-PCR) yielding a product of 849bp cDNA fragment. Using T-A cloning technique, the PCR product was cloned into pMD18-T vector. The purified VP1 gene was subcloned into the pBAD/Thio TOPO vector and the plasmid was identified by PCR. It was sequenced to confirm the authenticity of the sequence and orientations. SDS-PAGE and Western blotting revealed that the VP1 protein was expressed in Escherichia coli LGM194 at a high level and the recombinant fusion protein contained a N-terminal HP-Thioredoxin and a C-terminal polyhistidine tag. The optimal amount of the expressed fusion protein was 16% of total bacterial protein after being induced with L-arabinose at 0.002%concentration for 5 hours. It had a molecular mass of approximately 47.13 kDa and was immunologically reactive. The recombinant protein will be characterized and tested in an enzyme-linked immunosorbent assay (ELISA) format for potential application in the serodiagnosis of swine vesicular disease.

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    Cloning and Expression of VP1 Gene of Swine Vesicular Disease Virus

      Corresponding author: HUA Qun-yi,
    • 1. 1. Faculty of Bioengineering and Chemical Engineering, Kunming University of Science and Technology, Kunming 650224, China
    • 2. Technology Center of Yunnan Entry-Exit Inspection and Quarantine Bureau, Kunming 650228, China
    • 3. Xishuangbanna Veterinary Station, Jinghong 666100, China

    Abstract: The VP1 gene of Swine vesicular disease virus (SVDV) was amplified by reverse transcription-polymerase chain reaction (RT-PCR) yielding a product of 849bp cDNA fragment. Using T-A cloning technique, the PCR product was cloned into pMD18-T vector. The purified VP1 gene was subcloned into the pBAD/Thio TOPO vector and the plasmid was identified by PCR. It was sequenced to confirm the authenticity of the sequence and orientations. SDS-PAGE and Western blotting revealed that the VP1 protein was expressed in Escherichia coli LGM194 at a high level and the recombinant fusion protein contained a N-terminal HP-Thioredoxin and a C-terminal polyhistidine tag. The optimal amount of the expressed fusion protein was 16% of total bacterial protein after being induced with L-arabinose at 0.002%concentration for 5 hours. It had a molecular mass of approximately 47.13 kDa and was immunologically reactive. The recombinant protein will be characterized and tested in an enzyme-linked immunosorbent assay (ELISA) format for potential application in the serodiagnosis of swine vesicular disease.

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