| Equine rhinopneumonitis was assigned as an animal epidemic of Class B by OIE, Equine rhinopneumonitis is an acute, feverish and contagious infectious disease, caused by equine Herpesvirus Type I(EHV-I)and equine Herpesvirus Type IV(EHV-IV)of alpha- herpesvirus which have the high homology. EHV-I and EHV-IV have significant differences in host ranges and pathogenicities. EHV-IV mainly infects the upper respiratory tract, while EHV-I shows a systemic distribution that can affect multiple organ systems. The symptoms not only are fever, loss of appetite and the respiratory symptom, but even can lead to abortions, the death of neonatal foal and other nervous system clinical symptoms.Although rhinopneumonitis’ death rate is not high, its sequelae may trigger many serious health problems.According to published gD gene sequence of the EHV-I on Genbank, designed a pair of primers to amplify, cloned the target fragments to the pMD19-T vector. Then cloned the genes of sequencing into the pVAX1 expression vector, which successfully built a eukaryotic expression vector pVAX1-g D containing equine herpes virus type I gD gene after testings of PCR, restriction enzyme cutting and sequencing. Then it was transfected to BHK-21 cells by mediating recombination plasmid. Western Blotting showed genes can be expressed in BHK-21 cells. Recombinant plasmid and gD protein single or mixed were injected in mice respectively. The levels of anti-protein IgG after twice immunities and the proliferative reaction of lymphocytes after the second immunity were detected, respectively. ELISA and MTT assays showed that combined immunodeficiency can effectively induce the production of specific serum antibodies, and significantly promote the proliferation of specific lymphocytes.This study successfully constructed the pVAX1-gD eukaryotic recombinant plasmid, and combined it with pET-30a-gD protein can induce humoral and cellular immunity. |
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