Cloning, prokaryotic expression of chicken interferon-alpha gene and study on antiviral effect of recombinant chicken interferon-alpha.
- Author:
Qin WEI
1
;
Gui-Qing PENG
;
Mei-Lin JIN
;
Yu-Dong ZHU
;
Hong-Bo ZHOU
;
Hong-Yan GUO
;
Huan-Chun CHEN
Author Information
1. Lab of Animal Virus, State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China.
- Publication Type:Journal Article
- MeSH:
Animals;
Antiviral Agents;
pharmacology;
Blotting, Western;
Chick Embryo;
Cloning, Molecular;
Influenza A Virus, H9N2 Subtype;
drug effects;
Interferon Type I;
biosynthesis;
isolation & purification;
pharmacology;
Interferon-alpha;
genetics;
Newcastle disease virus;
drug effects;
Plasmids;
Recombinant Proteins
- From:
Chinese Journal of Biotechnology
2006;22(5):737-743
- CountryChina
- Language:Chinese
-
Abstract:
The full length of chicken interferon alpha (ChIFN-alpha) gene was amplified by the polymerase chain reaction (PCR) from total liver genome of Sanhuang meat-chicken and sequenced. The amplified gene was about 582bp. The coding region for mature protein (489bp) was subcloned into pET-28a(+). The recombinant plasmid pET-28a(+)-IFNalpha was identified by enzyme digestion and DNA sequencing. Data of SDS-PAGE and Western-blot indicated that a 22kD fusion protein was expressed in the form of inclusion bodies with good immunity. The purity of inclusion bodies was above 70% and that of protein purified by nickel affinity chromatography was 95%. The recombinant protein could inhibit H9N2 avian influenza virus (H9N2 AIV) replication on chick embryo fibroblast. 2 microg of recombinant IFN-alpha could completely protect Chick embryo from H9N2 AIV infection. The recombinant IFN-alpha can also delay Newcastle disease virus (NDV) replication on chick embryo for 12 approximately 48h. Chicken administered recombinant IFN-alpha can resist the H9N2 AIV infection. The bioactivities of recombinant IFN-alpha purified by affinity chromatograph were 20 times higher than that of inclusion bodies.
