Expression of hemagglutinin of avian influenza virus (AIV) and its application in diagnosis of AIV H9 subtype.
- Author:
Rui-Hua ZHANG
1
;
Mei-Lin JIN
;
Gui-Hua WANG
;
Zheng-Jun YU
;
Si-Ting ZHAO
;
Hong-Chao LI
;
Ya-Di TAN
;
Huan-Chun CHEN
Author Information
1. Lab of Animal virus, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China.
- Publication Type:Journal Article
- MeSH:
Cloning, Molecular;
Enzyme-Linked Immunosorbent Assay;
Escherichia coli;
genetics;
metabolism;
Hemagglutinin Glycoproteins, Influenza Virus;
biosynthesis;
genetics;
Humans;
Influenza A Virus, H9N2 Subtype;
genetics;
Influenza, Human;
diagnosis;
virology;
Recombinant Proteins;
biosynthesis;
genetics
- From:
Chinese Journal of Biotechnology
2005;21(2):315-319
- CountryChina
- Language:Chinese
-
Abstract:
In order to differently diagnose avian influenza virus (AIV) subtypes, the HA gene of AIV H9 subtype was cloned, expressed and utilized in an enzyme-linked immunoad sorbent assay (ELISA). HA gene (1683bp) of H9N2 AIV was amplified by RT-PCR from a strain of field isolated H9N2 AIV, and its identity was confirmed by sequencing. The HA gene was subcloned into prokaryotic expression vector pGEX-KG with its secretion signal sequence removed. The expressed HA-GST fusion protein in E. coli BL21 was characterized by SDS-PAGE and western blotting analysis as a 90kD protein with immunogenicity. The fusion protein was present primarily in inclusion bodies and was purified via denaturation and renenaturation. The HA-GST fusion protein was used to establish an indirect ELISA for the detection of antibodies to H9 subtypes of AIV. The assay has 91.57% specificity to H9 AIV, 92.31% sensitivity and excellent reduplication. It could be used to differently detect antibodies to H9 AIV.
