文章摘要
陈 霞,徐闻欢,徐 娟,赵 聃,王迎伟.结核杆菌ESAT-6抗原多表位DNA疫苗的构建与表达[J].南京医科大学学报,2008,28(4):412~415
结核杆菌ESAT-6抗原多表位DNA疫苗的构建与表达
Construction and expression of multiepitope DNA vaccine on ESAT-6 antigen of Mycobacterium tuberculosis in vitro
投稿时间:2007-09-18  
DOI:10.7655
中文关键词: 结核杆菌  ESAT-6  T细胞表位  DNA疫苗
英文关键词: Mycobacterium tuberculosis  ESAT-6  T cell epitope  DNA vaccine
基金项目:江苏省教育厅自然科学基金资助项目(05KJB310077);江苏省现代病原重点实验室开放课题(8853);南京医科大学校级重点项目(07NMUZ001)
作者单位
陈 霞 南京医科大学微生物与免疫学系江苏 南京 210029 
徐闻欢  
徐 娟  
赵 聃  
王迎伟  
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中文摘要:
      目的:构建含3个结核杆菌ESAT-6抗原T细胞表位及Flt3配体基因的重组质粒,并使其在大鼠肾小球系膜细胞(GMCs)中表达。方法:用计算机软件预测结核杆菌ESAT-6抗原的T细胞表位谱,选取3个T细胞表位,并以Ala-Ala-Tyr(AAY)序列作为接头,合成全基因序列,定向克隆入真核双表达载体pIRES及质粒pIRES-FL。在酶切分析与序列测定后,用PEI转染至GMCs细胞,并行Western blot 鉴定其体外表达。结果:核酸序列测定证实重组质粒构建正确,Western blot 证实该重组质粒能在体外GMCs细胞中表达融合蛋白。结论:成功构建了结核杆菌ESAT-6抗原多表位基因重组质粒。
英文摘要:
      Objective:To construct recombinant plasmids containing 3 T cell epitopes from ESAT-6 antigen of Mycobacterium tuberculosis and extra-cellular fragment of Flt3 ligand(FL) genes,and to express them in rat glomerular mesangial cells(GMC). Methods:The amino acid sequence of ESAT-6 antigen was analysed using predictive algorithms and 3 T cell epitopes were predicted. The oligonucleotide encoding and the linker Ala-Ala-Tyr(AAY) were synthesized and inserted into the bicistronic vector pIRES and pIRES-FL. The recombinant plasmids were transfected into GMC cells,the recombinant proteins expressing in GMC cells was examined by Western blot. Results:The recombinant plasmids were verified by sequencing and the recombinant fusion proteins were identified by Western blot. Conclusion:The multiepitope DNA vaccine from ESAT-6 antigen of Mycobacterium tuberculosis was constructed and expressed successfully.
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