传染性造血组织坏死病毒糖蛋白基因的原核表达及抗原性分析

Prokaryotic expression and antigenicity analysis of the glycoprotein gene of infectious hematopoietic necrosis virus

应用RT-PCR方法扩增了传染性造血组织坏死病毒IHNV-ZYX株编码的糖蛋白(G)基因,将G基因克隆至原核表达载体pET30b,并在大肠杆菌Rosetta(DE3)中得到了表达。SDS-PAGE分析表明,重组菌诱导后得到了预期大小的G蛋白。提取G蛋白的包涵体,并制备抗血清。间接ELISA和Western-blotting结果显示,重组表达的G蛋白与天然的IHNV G蛋白一样具有抗原性。试验为进一步研究IHNV G基因的免疫保护作用,为灵敏高效的检测传染性造血组织坏死病的方法的建立和基因工程疫苗的研制奠定了基础。

The gene encoding the viral glycoprotein （G） was amplified by RT- PCR from IHNV-ZYX strain and cloned into pET30b vector. The expression of recombinant plasmid pET30b-G in E. coil Rosetta （DE3） was induced and detected by SDS-PAGE analysis. The molecular weight of expressed recombinant G protein was approximately 52 ku as predicted. The inclusion body containing fusion protein was extracted and immunized in rabbits to obtain the antisera against G protein. Antigenicity of G protein was analyzed by indirect ELISA and Western-blotting. The results showed that the immunogenical and antigenical characters of the expressed G protein was in common with native IHNV G protein. This study laid an important material foundation for further studying on the G gene function of IHNV-ZYX in immune protection, establishing sensitive and efficient methods in detecting infectious hematopoietic necrosis, and manufacturing genetic engineering vaccine.