中国农业科学2012，Vol.45Issue(11)：2288-2294,7.DOI:10.3864/j.issn.0578-1752.2012.11.019
禽戊型肝炎病毒中国分离株ORF3蛋白的真核表达与抗原性分析
Eukaryotic Expression and Antigencity Analysis of Avian Hepatitis E Virus ORF3 Protein from China Isolate
摘要
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 DHlOBac 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.关键词
禽戊型肝炎病毒中国分离株/ORF3蛋白/真核表达/抗原性Key words
China avian hepatitis E virus/ ORF3/ eukaryotic expression/ antigenicity引用本文复制引用
胡守彬,赵钦,赵菲菲,肖一红,周恩民..禽戊型肝炎病毒中国分离株ORF3蛋白的真核表达与抗原性分析[J].中国农业科学,2012,45(11):2288-2294,7.基金项目
山东省“泰山学者”建设工程项目(040-72010) （040-72010）
