| In this research, IBV S, IBV S1 and ChIL-15 gene were modified and reconstructed firstly, then these genes were inserted into a eukaryotic expression vector, pcDNA3.1(+), to construct three new plasmids. Furthermore, a flexible linker which could express the (G4S)3 polypeptide was used to link the ChIL-15 gene with IBV S and S1 gene to construct two fusion gene plasmid of eukaryotic expression vetor-pcDNA3.1-IBV S1-linker-ChIL-15 and pcDNA3.1-IBV S-linker-ChIL-15. The results of the transient transfection of these two fusion plasmids in vitro showed that the fused gene could express together in Vero cells and could be located by specific anti-ChIL-15 monoclonal antibody and anti-IBV polyclonal antibody, therefore, it proved that the protein expressed by fusion gene could retain the bioactivities of two proteins, it also provided the theoretical foundation for the following immunization research in vivo.The experimental chicken were divided into 7 groups, the S group inoculated with the pcDNA3.1-IBV S, S1 group inoculated with the pcDNA3.1-IBV S1, the S+IL-15 group inoculated with the pcDNA3.1-IBV S and pcDNA3.1-Ch IL-15, the S1+IL-15 group inoculated with the pcDNA3.1-IBV S1 and pcDNA3.1- Ch IL-15, the S1-IL-15 group inoculated wiht the pcDNA3.1-IBV S1-linker-ChIL-15, the S-IL-15 group inoculate with the pcDNA3.1-IBV S-linker-ChIL-15 and the control group with empty pcDNA3.1 vetor. Thereafter, lymphocyte proliferative assay, enzyme linkered immunosorbent assay, Real Time RT-PCR and flow cytometric technique were used to detect the immune response of different groups. Finally chickens in different treatment groups were challenged with IBV Beaudette strain to compare the difference of the protection rate.The results showed that most of immune response of six inoculated groups are higher than the control group, especially, the S+IL-15, S1+IL-15, S-IL-15 and S1-IL-15 group which involved ChIL-15. The details were as follows: the T, B lymphocyte proliferative function; the number of CD4+, CD8+, TCRαβ+ and TCRγδ+T cell; the levels of special anti-IBV antibody in peripheral blood and IL-2 mRNA expression in thymus and spleen of immune groups were all higher than those of C group. And the S1-IL-15 group was the most significant among those groups. Meanwhile, when the S group was compared with the S1 group, the most obvious predominance of the S1 group was that S1 gene could induce cellular immunity and humoral immunity earlier than the S gene.The histopathological change of IBV target organs of each group were analyzed after challenge and the immunoprotection rates were compared among every group. The results proved that ChIL-15 could enhance the immunoprotection of the host to resist the virus infection. It also showd that the fusion genes could well display their bioactivities, and ChIL-15 could effectively reveal its immunologic enhancement with the help of the"linker".Above all, the results of this experiment showed that when the antigen genes of IBV were inoculated with ChIL-15, it could improve the host immune response and higher protection rate could be observed against the IBV chanllenge. Furthermore, IBV antigen genes and ChIL-15 fusion plasmid was first constructed in this experiment and the results observe here could provide solid scientific ground to the further study of ChIL-15 bioactivities, and the theoretical basis to the dicovery of new type immune-enhancing vaccine to virual diseases.
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