表达PRRS病毒GP5、GP3和猪IL-18的核酸疫苗的构建及实验免疫研究  

Construction and immuninty of DNA vaccine expressing GP3/GP5 of PRRSV and IL-18 of swine

作　　者：沈国顺[1] 金宁一[1] 秦晓光[1] 尹荣兰[1] 庄天中[1] 马鸣潇[1] 郑敏[1] 

机构地区：[1]解放军军事医学科学院十一所全军基因工程重点实验室

出　　处：《免疫学杂志》2006年第6期629-634,共6页Immunological Journal

摘　　要：目的获得安全有效的防治PRRSV核酸疫苗。方法采用RT-PCR方法扩增了PRRSV长春分离株ORF5、ORF3基因,并将其插入真核表达载体pVAX1、pVIR-IL-18的CMV启动子下游,构建成真核表达质粒pVAX1-ORF5-ORF3和pVIR-IL18-ORF5-ORF3,通过Westernblot及IFA等检测目的基因表达产物。同时进行了Balb/c小鼠免疫试验,检测免疫后小鼠的ELISA抗体水平和脾T淋巴细胞亚群数量。结果所构建的2个重组DNA疫苗质粒在真核细胞能进行有效的转录,并表达了目的蛋白(GP5、GP3)。pVIR-IL18-ORF5-ORF3免疫组的小鼠的ELISA抗体水平高于pVAX1-ORF5-ORF3和PRRS灭活苗免疫组,但差异不显著。CD8+T淋巴细胞亚群数量显著高于pVAX1-ORF5-ORF3和PRRS灭活苗组。结论IL-18对猪的细胞免疫功能具有明显的促进作用。Objective To obtain a safe and effective DNA vaccine against porcine reproductive and respiratory syndrome virus （PRRSV）. Methods ORF5 and ORF3 genes of the CHANGCHUN strain of PRRSV was amplified using RT-PCR, and then insert into the down stream of CMV promoter from eucaryotic expressing vector pVAX1 and pVIR-IL-18. Expression products were detected by Western blotring and IFA. The two DNA vaccines were used to immune mouse Balb/c. Then the ELISA antibody and the numbers of the murine spleen T lymphocyte were detected. Results These two recombinant plasmid pVAXI-ORFS-ORF3 and pVIR-ILI8-ORFS-ORF3 were transcribed effectively and the objective proteins （GP5 and GP3） were expressed in eucaryotic cells, The ELISA antibody of pVIR-IL18-ORFS-OR_V3 exceeded to that of pVAX1-ORFS-ORF3 and inactivated vaccine, but there was not remarkable difference. The numbers of CD8^＋ T lymphocyte induced by pVIR-IL18-ORFS-ORF3 exceeded remarkably to those induced by pVAX1-ORFS-ORF3 and inactivated vaccine, Conclusion IL-18 could promote cell-mediated immunity of pig significantly.

关 键 词：PRRSV病毒 ORF5/ORY3基因 核酸疫苗 免疫 

分 类 号：S852.5[农业科学—基础兽医学]