Cloning, Expression Of Canine Interferon Alpha And Its Biological Activity

Interferon (IFN) is a multigene family of inducible cytokines. IFNs which are commonly grouped into two types, generated by specific cells induced by inducer, are a kind of glycoprotein that has anti-viral activity, anti-tumor activity and immunoregulation function on the homogenecity animal cells. IFN type I is known as viral IFN and includes IFN-α(leukocyte), IFN-β(fibroblast) and IFN-ω. IFN Type II is known as immune IFN (such as IFN-γ). At present, IFN-αreseach is hot, which is synthesized mainly by Leukocytes and has a common receptor with beta, omega of IFNs. IFN–αcan inhibit virus replication, viral protein synthesis and promote MHC class I molecules presenting antigen.CaIFN-α1 gene was amplified by RT-PCR from the peripheral blood lymphocytes of beagle dogs. The PCR product was linked to pGEM-T vector and transformed into E. coli DH5α. The positive clones were screened by restriction enzyme digestion and sequenced. The results showed that CaIFN-α1 gene was 100% homology with that published in GenBank (No.M28624). Then CaIFN-α1 gene was cloned into the eukaryotic expression vector pFastBacI through a series of restriction enzyme digestion and ligation reactions. The positive recombinant plasmids were screened and identified by PCR. At the last, CaIFN-α1 was expressed in Sf9 insect cells. Supernatant of the cells infected with the recombinant virus could inhibit the cyto-pathic effect (CPE) in CEF infected with vesicular stoma-titis new jersey virus (VSV) . This result indicated that the CaIFN-α1 expressed in Sf9 cells has interfer activity to virus repilcation. However it could not inhibit CPE in CEF infected with CVI988 or NDV(F48E8). Also same result was found in MDBK cells infected with VSV or CPV- F81.