摘要
[目的]构建猪繁殖与呼吸综合征和猪2型圆环病毒病的二联核酸疫苗并研究其免疫效果。[方法]将辽宁某猪场发病猪病料中猪繁殖与呼吸综合征病毒(PRRSV)的ORF5基因克隆至真核表达载体pIRES-neo的CMV启动子下游,然后用猪2型圆环病毒(PCV-2)的内蒙古分离株的ORF2基因替换pIRES的neo基因,构建真核表达质粒。通过W estern blot和IFA方法检测构建的重组质粒在BHK细胞中的表达情况。将重组质粒免疫Balb/c小鼠,检测免疫后的ELISA抗体水平和脾T淋巴细胞亚群的数量。[结果]成功构建了重组表达质粒pIRES-ORF2-ORF5,W estern blot和IFA试验证实该重组质粒在BHK细胞内成功表达了目的蛋白。与灭活苗相比,pIRES-ORF2-ORF5核酸疫苗免疫小鼠ELISA抗体水平差异不显著;其脾T淋巴细胞亚群数量显著高于灭活疫苗免疫组。[结论]构建的二联核酸疫苗能够诱导小鼠产生较好的体液免疫反应和细胞免疫反应,为更好地防制猪繁殖与呼吸综合征和猪2型圆环病毒病提供了条件。
[Objective] The aim of the study was to construct associated DNA vaccine of PRRS（Porcine reproductive and respiratory syndrome） and PCV-2（Porcine circovirus type 2） disease and its immunogenicity.[Methods] In this study,the ORF 5 gene of PRRSV isolated in Liaoning was cloned into pIRES-neo expression vector,and the neo gene of pIRES-neo expression vector was substituted by the ORF 2 gene of the Mongolia PCV-2 strain to construct the recombinant expression vector.The expression in BHK cells was detected through Western blot and IFA.Then the ELISA antibody level and the number of spleen T lymphocytes were detected after Balb/c mice were immunized with this DNA vaccine.[Results] The recombinant plasmid pIRES-ORF2-ORF5 was constructed successfully and could express the target proteins in BHK cells,as indicated by Western blot and IFA.There was no significantly difference in ELISA antibody between pIRES-ORF2-ORF5 immunized group and inactived vaccine immunized groups,while the number of spleen T lymphocytes induced by DNA vaccine was higher than that induced by inactived vaccine.[Conclusion] The recombinant plasmid pIRES-ORF2-ORF5 should induce good humoral immune response and cellular immune response in mice,providing the conditions for better prevention and control of PRRS and PCV-2 disease.
出处
《安徽农业科学》
CAS
北大核心
2009年第11期4991-4994,共4页
Journal of Anhui Agricultural Sciences
