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Co-Expression of Porcine Interferon-Alpha and Interferon-Gamma Using a Baculovirus and Its Inhibitant Activity on PRRSV

WANG Yan-bin, SUN Xiang-li, WEI Zhan-yong, CHEN Hong-ying, YANG Ming-fan, CUI Bao-an   

  1. College of Animal Science and Veterinary Medicine,Henan Agricultural University/Key Laboratory of Animal-Derived Food Safety of Henan Province, Zhengzhou 450002
  • Received:2010-04-20 Online:2011-05-05 Published:2011-02-21

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Abstract: 【Objective】Compound recombinant porcine interferon α and interferon γ were prepared for clinical antiviral investigation 【Method】 Nucleotide sequences encoded mature porcine interferon-alpha and interferon-gamma were cloned and inserted into pFastBacDual under the control of PH promoter and P10 promoter, respectively. The authentic signal sequences of porcine interferon-alpha and interferon-gamma were substituted with the honeybee melittin signal sequence, and fused with C-terminal 6×Histidine tag for easy purification. The constructed plasmid was transformed into DH10 competent cells for homologous recombination, the recombinant shuttle plasmid Bacmid then transfected to Sf9 insect cells to obtain recombinant baculovirus. After infection with recombinant baculovirus, the recombinant protein activity was detected.【Result】The recombinant proteins were successfully detected in Sf9 cells by immunofluorescence assay and Western blot analysis. The expression products were mainly detected in the culture medium. The compound recombinant interferon-α & γ were verified to be of high antiviral activity by inhibiting the cytopathic effect of vesicular stomatitis virus in PK-15 cells, which is about 3.24×107 U·mL-1 in supernatant. The results also suggest that in vitro rpIFN-α & γ protected Marc-145 from cytopathic effect by PRRSV of 100 TCID50. 【Conclusion】 The baculovirus expression system to achieve porcine interferon-α and γ were successfully secreted in insect cells, the recombinant protein in cells inhibited the replication of PRRSV.

Key words: porcine , interferon , baculovirus , expression , PRRSV

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