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泛素基因介导下的猪繁殖与呼吸综合征病毒M基因对小鼠的免疫试验
引用本文:张维军,童铁钢,白宇,王群,徐树兰,孙庆歌,田野,杨涛,童光志,吴东来.泛素基因介导下的猪繁殖与呼吸综合征病毒M基因对小鼠的免疫试验[J].微生物学报,2009,49(6):799-806.
作者姓名:张维军  童铁钢  白宇  王群  徐树兰  孙庆歌  田野  杨涛  童光志  吴东来
作者单位:1. 中国农业科学院哈尔滨兽医研究所,兽医生物技术国家重点实验室,农业部兽医公共卫生重点开放实验室,哈尔滨,150001
2. 中国农业科学院上海兽医研究所,上海,200241
基金项目:国家高科技发展(863)计划”(2006AA10A203);黑龙江省自然科学基金重点项目(ZJN-0602-01)
摘    要:摘要:【目的】获得猪繁殖与呼吸综合征病毒(Porcine Reproductive and Respiratory Syndrome virus, PRRSV)M蛋白基因及其与泛素(Ubiquitin, Ub)基因融合的真核表达质粒,并进一步研究Ub对M基因免疫效果的影响。【方法】利用RT-PCR技术以PRRSV CH-1a株和BALB/c小鼠脾脏组织总RNA为模板,分别扩增出PRRSV M蛋白基因与小鼠的Ub基因,利用SOE技术将PRRSV M基因与小鼠Ub基因进行融合,最终构建真核表达质粒pVAX1-M与pVAX1-U-M。以两种真核表达质粒DNA肌肉免疫BALB/c小鼠后,分别检测体液免疫反应与细胞免疫反应,比较M单基因与Ub-M融合基因DNA免疫所诱生的免疫应答强度【结果】IFA试验表明,pVAX1-M与pVAX1-U-M均能BHK-21细胞内成功表达目的基因;两种重组质粒免疫小鼠后均可诱生PRRSV ELISA抗体与细胞免疫反应,但是pVAX1-U-M诱导的细胞免疫反应明显高于pVAX1-M,差异显著(P<0.05);而其诱导的抗体水平明显低于pVAX1-M,二者差异也显著(P<0.05)。【结论】泛素在一定程度上可以促进PRRSV M基因诱导细胞免疫反应,但对体液免疫未见到同样作用。

关 键 词:关键词:猪繁殖与呼吸综合征病毒  M基因  泛素  体液免疫  细胞免疫  DNA疫苗
收稿时间:2008/12/9 0:00:00
修稿时间:3/2/2009 12:00:00 AM

Effects of fusion gene of ubiquitin and Porcine Reproductive and Respiratory Syndrome virus M gene on the immune response in inoculated mice
Weijun Zhang,Tiegang Tong,Yu Bai,Qun Wang,Shulan Xu,Qingge Sun,Ye Tian,Tao Yang,Guangzhi Tong and Donglai Wu.Effects of fusion gene of ubiquitin and Porcine Reproductive and Respiratory Syndrome virus M gene on the immune response in inoculated mice[J].Acta Microbiologica Sinica,2009,49(6):799-806.
Authors:Weijun Zhang  Tiegang Tong  Yu Bai  Qun Wang  Shulan Xu  Qingge Sun  Ye Tian  Tao Yang  Guangzhi Tong and Donglai Wu
Affiliation:State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China;Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China;State Key Laboratory of Veterinary Biotechnology, The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China
Abstract:Abstract: Objective] To evaluate the effects of the fusion gene of ubiquitin (Ub) and Porcine Reproductive and Respiratory Syndrome virus (PRRSV) M gene on the immune response in inoculated mice. Methods] Mouse Ub gene and PRRSV M gene were amplified by RT-PCR from BALB/c mice spleen cells and PRRSV Ch-1a strain, respectively, and the M and Ub gene (U-M) was fused by SOE PCR. Therefore, pVAX1-U-M and pVAX1-M recombinant plasmid were constructed for eukaryotic expression. Results] The fusion U-M and M protein expressions were verified in transfected BHK-21 cells by indirect fluorescence assay. Furthermore, both pVAX1-M and pVAX1-U-M induced specific humoral and cellar immune responses against PRRSV in the recombinant plasmid injected mice. However, pVAX1-U-M was able to induce higher level of T cell response then that of pVAX1-M (P< 0.05), but lower level of antibody (P< 0.05). Conclusion] Expression of U-M fusion gene had ability to enhance specific T cell response against PRRSV, but no effect on stimulation of humoral response in inoculated mice.
Keywords:Keywords: PRRSV  M gene  ubiquitin  humoral immunity  cellular immunity  DNA vaccine
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