Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (11): 2288-2294.doi: 10.3864/j.issn.0578-1752.2012.11.019

• VETERINARY SCIENCE • Previous Articles     Next Articles

Eukaryotic Expression and Antigencity Analysis of Avian Hepatitis E Virus ORF3 Protein from China Isolate

 HU  Shou-Bin, ZHAO  Qin, ZHAO  Fei-Fei, XIAO  Yi-Hong, ZHOU  恩Min   

  1. 1.山东农业大学动物科技学院预防兽医系免疫生物学实验室，山东泰安 271018
    2.西北农林科技大学兽医免疫学研究所/西北农林科技大学动物医学院，陕西杨凌 712100
  • Received:2011-09-20 Online:2012-06-01 Published:2012-03-23

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Abstract: 【Objective】The objective of the study is to obtain recombinant protein of avian hepatitis E virus (HEV) ORF3 of China isolate (CaHEV) and analyze its antigencity. 【Method】Total RNA was extracted from CaHEV and ORF3 gene was amplified by RT-PCR and cloned into pFastBac-HT. The recombinant plasmid was transformed into DH10Bac and transfected into sf9 insect cells to express the recombinant ORF3 protein. The recombinant ORF3 protein was identified by SDS-PAGE, Western bolt and IFA methods. The purified ORF3 protein was used to immunize Balb/c mice to produce polyclonal antibodies and then to analyze the epitopes in ORF3 using 3 truncated ORF3 proteins 【Result】The results showed that ORF3 protein was expressed with the highest expression level at 4 days post inoculation. The titer of anti-ORF3 polyclonal antibodies was 104 determined with the ELISA method. The dominant epitopes of ORF3 were located between amino acids 74 and 88 in C-terminal region.【Conclusion】 Recombinant ORF3 protein from avian HEV China isolate was expressed successfully and the dominant epitopes were located between amino acids 74 and 88 in C-terminal region. These results paved a way for future study of the structure and function of ORF3 protein.

Key words: China avian hepatitis E virus, ORF3, eukaryotic expression, antigenicity

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