钩端螺旋体赖型017株外膜脂蛋白LipL41基因的克隆及原核表达  

Molecular Cloning and Expression in E.coli of the Surface exposed Lipoprotein LipL41 Gene of Leptospira lai

作　　者：李学敏[1] 鲍朗[1] 胡昌华[1] 谢勇恩[1] 晏菊芳[1] 张会东[1] 

机构地区：[1]华西医科大学基础医学院感染免疫研究室,成都610041

出　　处：《华西医科大学学报》2001年第3期341-343,448,共4页Journal of West China University of Medical Sciences

摘　　要：目的　构建钩端螺旋体外膜脂蛋白 L ip L41基因的重组原核表达质粒 ,为进一步制备以 L ip L41为目的基因的亚单位疫苗提供实验依据。方法　根据钩端螺旋体 L.kirscneri RM5 2株外膜脂蛋白 L ip L41基因序列设计引物 ,以赖型钩端螺旋体 0 17株基因组 DNA为模板 ,进行 PCR扩增并 DNA测序 ,以质粒 p GEX1λT为载体 ,插入 L ip L41基因片段构建重组原核表达质粒 ,并检测 L ip L41的表达。结果　测序结果示所得片段为 L ip L41的编码序列 ,酶切及 PCR分析证实重组质粒构建成功 ,SDS- PAGE和 Western blotting分析重组质粒可高效表达蛋白质 L ip L41。结论　 L ip L41基因的重组原核表达质粒构建成功 。Objective To construct a recombinant expressive plasmid using LipL41gene of leptospira lai for further research of subunit vaccine of leptospira. Methods A pair of oligonucleotide primers were designed based on the sequence of LipL41 of leptospira kirschneri RM 52 in Genbank. With genomic DNA of leptospira lai 017 as template, a fragment was amplified by PCR and DNA sequencing analysis showed this fragment to be the gene that encodes LipL41. A recombinant vector was constructed using plasmid pGEX1λT and the expression of LipL41 gene was tested. Results The production of PCR was LipL41 gene. The recombinant plasmid was constructed and testified by nuclease digestion and PCR, and LipL41 gene could express in E.coli. Conclusion The recombinant plasmid has been constructed successfully and LipL41 gene can express stably at high level.

关 键 词：钩端螺旋体赖型017株 LipL41基因 基因克隆 基因表达 膜脂蛋白 

分 类 号：R377.5[医药卫生—病原生物学]