[Cloning, prokaryotic expression of chicken interferon-alpha gene and study on antiviral effect of recombinant chicken interferon-alpha].
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.
Wei
Qin
Q
Lab of Animal Virus, State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan, China.
Peng
Gui-Qing
GQ
Jin
Mei-Lin
ML
Zhu
Yu-Dong
YD
Zhou
Hong-Bo
HB
Guo
Hong-Yan
HY
Chen
Huan-Chun
HC
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D015153:Blotting, Western; D002642:Chick Embryo; D003001:Cloning, Molecular; D053127:Influenza A Virus, H9N2 Subtype / Q000187:drug effects; D007370:Interferon Type I / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology*; D016898:Interferon-alpha / Q000235:genetics*; D009522:Newcastle disease virus / Q000187:drug effects; D010957:Plasmids; D011994:Recombinant Proteins
D000998:Antiviral Agents; D007370:Interferon Type I; D016898:Interferon-alpha; D011994:Recombinant Proteins