赖型钩端螺旋体外膜蛋白LipL32基因真核表达载体的构建及表达的初步研究  

Construction of the eukaryotic recombinant vector and expression of the outer membrane protein LipL32 gene from Leptospira serovar Lai

作　　者：黄毕[1] 鲍朗[1] 钟琪[1] 商正玲[1] 张会东[1] 张英[1] 

机构地区：[1]四川大学华西基础医学与法医学院感染免疫研究室,四川成都610041

出　　处：《细胞与分子免疫学杂志》2008年第2期109-112,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目(30471546);教育部博士点基金资助项目(20040610049)

摘　　要：目的:构建赖型钩端螺旋体外膜蛋白LipL32基因真核表达载体并在COS-7细胞中表达,为钩端螺旋体DNA疫苗的研究和开发奠定基础。方法:从赖型钩端螺旋体017株全基因组中PCR扩增出目的基因,双酶切构建重组质粒pcDNA3.1-LipL32。脂质体转染法将重组质粒转染COS-7细胞,通过RT-PCR、Westernblot检测目的基因的表达。结果:成功构建了LipL32基因的真核表达载体,并在COS-7细胞中获得瞬时和稳定表达。结论:赖型钩端螺旋体外膜蛋白LipL32基因真核表达载体能在哺乳动物细胞内表达,为钩端螺旋体DNA疫苗的应用提供了实验依据。AIM： To construct the eukaryotic experssion vector of LipL32 gene from Leptospira serovar Lai and express the recombinant plasmid in COS-7 cell. METHODS： The LipL32 gene was amplified from Leptospira strain 017 genomic DNA by PCR and cloned into pcDNA3.1, through restriction nuclease enzyme digestion. Then the recombinant plasmid was transformed into E. coli DH5α. After identified by nuclease digestion, PCR and sequencing analysis, the recombinant vector was transfected into COS-7 cell with lipsome. The expression of the target gene was detected by RT-PCR and Western blot. RESULTS： The eukaryotic experssion vector pcDNA3.1-UpL32 was successfully constructed and stably expressed in COS-7 cell. CONCLUSION： The eukaryotic recombinant vector of outer membrane protein LipL32 gene from Leptospira serovar Lai can be expressed in mammalian cell, which provides an experimental basis for the application of the Leptospira DNA vaccine.

关 键 词：钩端螺旋体 外膜蛋白LipL32 真核表达 

分 类 号：R377.5[医药卫生—病原生物学]