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目的:构建以人细胞毒性T淋巴细胞抗原4(Cytotoxic T lymphocyte associated antigen 4,CTLA4)为引导的抗牙龈卟啉单胞菌菌毛蛋白FimA的靶向牙周炎DNA疫苗,并检测其在体外真核细胞中的表达。方法:采用基因重组技术构建携带CTLA4-Ig基因和牙龈卟啉单胞菌fimA基因的重组质粒pCTLA4-FimA,转染COS7细胞,蛋白免疫印记和间接免疫荧光法检测重组质粒的蛋白表达。结果:重组质粒pCTLA4-FimA携带CTLA4-Ig和fimA基因片断。pCTLA4-FimA表达的抗原可与特异性抗FimA抗体发生免疫反应。pCTLA4-FimA转染的COS7细胞中可检测到目的蛋白的表达。结论:成功构建了靶向牙周炎DNA疫苗pCTLA4-FimA,并可在真核细胞中正确表达。
OBJECTIVE: To construct a DNA vaccine against periodontitis of Pseudomonas gingivalis Fimbriae induced by cytotoxic T lymphocyte associated antigen 4 (CTLA4) Expression in eukaryotic cells. Methods: Recombinant plasmid pCTLA4-FimA carrying CTLA4-Ig gene and finkA gene of Porphyromonas gingivalis was constructed by gene recombination technique. COS7 cells were transfected into COS7 cells. The protein expression of recombinant plasmid was detected by Western blot and indirect immunofluorescence. Results: The recombinant plasmid pCTLA4-FimA carries CTLA4-Ig and fimA gene fragments. Antigens expressed by pCTLA4-FimA are immunoreactive with specific anti-FimA antibodies. The expression of the target protein can be detected in pCTLA4-FimA transfected COS7 cells. Conclusion: DNA vaccine targeting periodontitis pCTLA4-FimA was successfully constructed and expressed correctly in eukaryotic cells.