[Expression of hemagglutinin of avian influenza virus (AIV) and its application in diagnosis of AIV H9 subtype].
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.
Zhang
Rui-Hua
RH
Lab of Animal virus, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China.
Jin
Mei-Lin
ML
Wang
Gui-Hua
GH
Yu
Zheng-Jun
ZJ
Zhao
Si-Ting
ST
Li
Hong-Chao
HC
Tan
Ya-Di
YD
Chen
Huan-Chun
HC
D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D053127:Influenza A Virus, H9N2 Subtype / Q000235:genetics*; D007251:Influenza, Human / Q000175:diagnosis* / Q000821:virology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D019267:Hemagglutinin Glycoproteins, Influenza Virus; D011994:Recombinant Proteins