草原兔尾鼠卵透明带3DNA疫苗pVAX1-sig-LTB-lZP3-C3d3的构建表达及其免疫不育的研究  

A study on the construction,expression and immunosterility of Lagurus laguru zona pellucida 3 DNA Vaccine pVAX1-sig-LTB-lZP3-C3d3

作　　者：李晨晨[1] 于继云[2] 姜敏[1] 涂亦娴[1] 马晓林[1] 张富春[1] 

机构地区：[1]新疆大学生命科学与技术学院新疆生物资源基因重点实验室,新疆乌鲁木齐830046 [2]军事医学科学院基础医学研究所,北京100850

出　　处：《细胞与分子免疫学杂志》2011年第9期941-944,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目(30760136);新疆动物学重点学科资助项目(20100702)

摘　　要：目的:构建草原兔尾鼠卵透明带3 DNA疫苗pVAX1-sig-LTB-lZP3-C3d3进行小鼠的黏膜免疫,增强该疫苗的免疫不育效果。方法:将两种佐剂分子大肠杆菌不耐热肠毒素B亚基和C3d基因通过酶切鉴定,构建重组质粒pVAX1-sig-LTB-lZP3-C3d3,采用RT-PCR和W estern b lot技术,检测其在mRNA和蛋白水平的表达,并通过肌肉注射、滴鼻和灌服3种途径免疫雌性C57BL/6小鼠,通过ELISA检测抗体水平及分型。结果:酶切鉴定,RT-PCR和W estern b lot结果表明重组质粒构建正确并可在mRNA和蛋白水平的表达,ELISA结果表明以重组质粒pVAX1-sig-LTB-lZP3-C3d3免疫诱导的特异性IgG、IgA的水平明显高于对照组(P<0.01)。抗生育实验表明该疫苗免疫的小鼠平均生仔数与对照组相比差异极显著(P<0.01)。结论:构建的草原兔尾鼠卵透明带3DNA疫苗重组质粒pVAX1-sig-LTB-lZP3-C3d3可高效地激发小鼠特异性免疫应答,提高抗生育的效果。AIM： To enhance the immunocontraceptive effect of Lagurus lagurus zona pellucida 3 DNA vaccine,and to achieve the prospect of application through the pVAX1-sig-LTB-lZP3-C3d3 different immunity pathway.METHODS： Two adjuvant molecules were constructed into the recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 as DNA vaccine which contains Escherichia coli heat-labile enterotoxin B subunit and the molecular adjuvant 3 copies of C3d.The results of RT-PCR and western blot showed that the DNA vaccine was expressed in mRNA and protein level.The female C57BL/6 mice were immunized by three ways： intramuscular injection,intranasal or oral route.Antibody levels and types were detected by ELISA.RESULTS： ELISA results showed that recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 immunization induced specific IgG,IgA levels were significantly different comparing with control（P0.01）.Antifertility experiment showed that the experimental group reduced the average fertility significantly different compared with the control group（P0.01）.CONCLUSION： Restriction analysis,RT-PCR and Western blot showed that the recombinant plasmid constructed correctly and can be the expression of mRNA and protein levels.It resulted that the recombinant plasmid pVAX1-sig-LTB-lZP3-C3d3 can induce the specific immune response efficiently and enhance the immunocontraceptive effects.

关 键 词：草原兔尾鼠 卵透明带3 DNA疫苗 佐剂分子 免疫不育 小鼠 

分 类 号：Q78[生物学—分子生物学]