摘要
目的:构建含有肺炎链球菌(SPN)毒力因子CbpA抗原编码基因的重组质粒pcDNA3.1/CbpA,测定其诱导特异性免疫应答的能力并评价其保护效果.方法:PCR扩增SPNCbpA编码基因,将该编码基因克隆到pcDNA3.1(+)真核表达载体中,构建pcDNA3.1/CbpA重组质粒,经测序鉴定后转染COS-7细胞,Western Blot鉴定目的蛋白的表达,重组质粒免疫BALB/c小鼠,TIGR4型SPN攻击后,监测其生存时间.结果:CbpA能在COS-7细胞中表达;pcDNA3.1/CbpA重组质粒主动免疫BALB/C小鼠后,体内产生特异性IgG抗体,并诱导小鼠对SPN感染产生有效的保护.结论:成功构建了重组真核表达质粒pcDNA3.1/CbpA,可为SPN基因疫苗的研制提供依据.
Abstract
AIM: To construct a recombinant eukaryotic expression plasmid pcDNA3.1/CbpA and to detect its protective effect against invasive pneumocoecal infection,METHODS: CbpA gene was amplified from Streptococcus peumoniae TIGR4 by PCR and was then inserted into the eukaryotic expression vector pcD-NA3.1 ( + )to construct recombinant pcDNA3.1/CbpA. Western blot was used to detect the protein expression of the CbpA gene in transfected COS-7 cells. The recombinant pcDNA3. 1/CbpA was injected into BALB/C mice via an intramuscular route to examine its capacity of protecting mice against intraperitoneal challenged with TIGR4. RESULTS: pcDNA3. 1/CbpA recombinant plasmid was successfully constructed and cbpA protein was expressed in transfected COS-7 cells. Antigen-specific antibody was detected in mice immunized with pcDNA3. 1/CbpA. Protection assay demonstrated that the survival rate of the mice immunized with pcDNA3. 1/CbpA was higher than that of control group. CONCLUSION: pcDNA3. 1/CbpA induces protective immune response in mice and could be used as a potential vaccine to prevent pneumococcal infection.
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维普
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