The Coexpression Of NDV F Gene And IBDV VP0 Gene In RFPV And Detection Of Immunogenicity

The Newcastle disease and Inferious burasal disease virus are very dangerous disease to avian. The sequence, the fusion(F) protein gene of NDV, was determined, which is 1792-base-pair sequence, encodes a F protein which has essential features previously established for the F protein of virulent NDV strains. The construct protein (VPO) of IBDV was clone and preserved which is 3125bp sequence encode prominent protecting antigen VP2 of IBDV .F protein is the main protective antigen of NDV and the primary structure of the virulence of the virus. Hence, the F gene and VPO gene is important for constriction of recombine genetic engeering vaccine .To express IBDV VPO gene and NDV F gene in recombinant FPV 282E4 strain and prepare for producting recombinant FPV vaccine which can protect chickren from NDV and IBDV infecting, Transfering vector pATI2- F-VPO was constructed by inserted NDV F gene and IBDV VPO gene into TK gene of FPV DNA downstream of two synthetic promoter p7.5-ATI which were conjugated at opposite direction. The pATI2-F-VPO plasmid was obtained, The result of restriction map show that exogenous fragment were inserted in correct direction. Then plasmad of pATI2-F-VPO was transfected with DQTAP Liposomal into CEF cultures infected with FPV to select stable recombinant, the homologous recombinants were passaged in chicken embryo giboblast culture treated with BrdU and then screened for the expression of NDV F protein by indirect immunofluorescene assay(IFA). The expressed products were examinated by Western blot. 2 recombinants were isolated which can express F and VPO protein at same time. Western-blot demonstrate that the expressed products can react with specific PcAb to IBDV and NDV. The result show that Lymphocyte were induced.Cellular and humoral immune responses were examined in spleen cell of chickren by Lymphocyte converting experiment. Spleen lymphocyte of the immuned chicken were cultured, then were stimulated by ConA and PHA in culture medium with 3H-TdR, detect the radioactivity of cell. The resault show that immuned group have higer reaction than control group which were not46immuned. This prove that recombine virus can stimulate Lymphocyte effectualy. The recombined virus has satisfactory immunogenicity.