Series Expression In Both Prokaryotic Cell And Eukaryotic Cells Of Cattle Interleukin18Gene And The Cattle Sergenti Theileria P23Surface Protein Gene

The cattle sergenti Theileriosis is the Tai Leke, Taylor of the genus Theileria sergenti (Theileria sergenti) parasitic in cattle red blood cells and lymphocytes, caused by a blood protozoosis. In recent years, with the growing cattle industry, cattle sergenti theileriosis occurrence of an upward trend, seriously affect the development of animal husbandry of the relevant countries. However, there is no clinical application of vaccines, seriously affect the healthy development of the cattle industry.Therefore, this study proceed from the bovine Theileria sergenti P23surface protein gene and bovine interleukin-18gene, according to the nucleotide sequence reported in GenBank, to design two pairs of specific primers SOE-PCR amplification of the T-p23,-IL-18fusion gene, was cloned into pMD18-T vector to construct pMD18-T-p23, of IL-18recombinant cloning vector by PCR and restriction enzyme digestion and sequenced, and then p23, IL-18fusion genecloned into pET-28a of the original nuclear expression plasmid and the true nuclear expression plasmid PVAX I, build the pET-28a-p23, of IL-18and PVAX-p23,-of IL-18reorganization expression plasmid after PCR and restriction analysis, prokaryotic plasmid of positive bacteriaexpression induced by IPTG, the expression product by SDS-PAGE and Western blotting analysis, positive bacteria, eukaryotic cells by RT-PCR reaction and indirect immunofluorescence assay, and to analyze the expression of the recombinant plasmid in eukaryotic cells.The results show that the successful cloning of p23,-IL-18fusion gene and successful expression in E. coli, expression of the protein size of approximately48ku can be Ngau Theileria sergenti-positive sera; fusion gene was expressed in BHK21cells, by RT-PCR and IFA experiments, indicating that the fusion gene was successfully transferred to the eukaryotic cells, BHK21, and that the gene expression in eukaryotic cells.The results laid the foundation for the cattle sergenti Theileriosis further immunization.