弓形虫GJS株pcDNA3-MIC3核酸疫苗的研究  

Studies on pcDNA3-MIC3 nucleic acid vaccine of Toxoplasma gondii GJS strain

作　　者：张燕丽[1,2] 曹丽艳[1,2] 芦赟[1,2] 蔡志杰[1,2] 王艳华[2] 付宝权[2] 李文卉[2] 张德林[2] 

机构地区：[1]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室甘肃省动物寄生虫病重点实验室农业部兽医公共卫生重点开放实验室,甘肃兰州730046 [2]甘肃农业大学动物医学院,甘肃兰州730070

出　　处：《中国兽医学报》2009年第10期1299-1302,共4页Chinese Journal of Veterinary Science

基　　金：国家科技支撑计划项目(2007BAD40B05);公益性行业(农业)科研专项经费项目(200803017);甘肃省重大专项科技项目(2GS063-A43-013)

摘　　要：根据GenBank中MIC3基因序列设计1对引物,采用PCR技术从弓形虫GJS株基因组DNA中扩增微线体蛋白3(MIC3)基因片段,克隆到pMD18-T载体,经PCR、酶切及测序鉴定后,阳性重组质粒酶切并亚克隆到真核表达载体pcDNA3.1(+)后进行PCR、酶切及测序鉴定。重组质粒pcDNA3-MIC3肌肉注射免疫BALB/c小鼠,通过ELISA检测血清特异抗体;经腹腔攻击感染弓形虫GJS株速殖子,观察小鼠的生存时间。结果成功构建了pcD-NA3-MIC3质粒;免疫组小鼠血清检测到特异性抗体;攻击感染后免疫组小鼠平均存活时间较对照组明显延长。表明该核酸疫苗具有较好的免疫原性,能诱导小鼠产生良好的免疫保护作用。A pair of primers was designed according to the sequence of microneme protein 3 （MIC3） gene of T. gondii RH strain from GenBank. The gene fragment MIC3 was amplified by PCR from the genomic DNA of T. gondii GJS strain and cloned into the vector pMD18-T. After PCR, restrictive digestion and sequencing identification, the positive plasmid was digested and subcloned into an eukaryotic expression vector pcDNA3. 1 （＋）. The eukaryotic expression recombinant plasmid pcDNA3-MIC3 was identified by PCR, restriction enzyme digestion and sequencing analysis. After mice were injected with the recombinant plasmid pcDNA3-MIC3, the sera antibody were detected by indirect ELISA and survival time of mice after challenge with tachyzoites of T. gondii GJS strain were observed. The results showed that the recombinant plasmid pcDNA3-MIC3 was successfully constructed and the immunized mice showed higher specific antibody. The survival time of vaccined mice was longer than that of control groups. It indicated that the nucleic acid vaccine had a good immunity and could induce the immunoprotection in mice.

关 键 词：刚地弓形虫 微线体蛋白3(MIC3) 核酸疫苗 免疫保护 

分 类 号：S852.71[农业科学—基础兽医学]