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Journal of ZheJiang University(Medical Science)  2013, Vol. 42 Issue (2): 164-170    DOI: 10.3785/j.issn.1008-9292.2013.02.006
Prokaryotic expression of Leptospira interrogans groEL gene and immunoprotection of its products in hamsters
LI Xiao-Yu1, WANG Yin-Huan1, YAN Jie2, CHENG Dong-Qing1
1.Department of Microbiology and Immunology,College of Life Science,Zhejiang Chinese Medical University,Hangzhou 310053,China; 2.Department of Microbiology and Parasitology,Zhejiang University School of Medicine,Hangzhou 310058,China
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Objective: To construct a prokaryotic expression system of groEL gene of Leptospira interrogans serogroup Icterohaemorrhagia serovar Lai strain Lai,and to determine the immunoprotective effect of recombinant GroEL protein(rGroEL) in LVG hamsters.
Methods: The groEL gene was amplified by high fidelity PCR and the amplification products were then sequenced.A prokaryotic expression system of groEL gene was constructed using routine genetic engineering technique.SDS-PAGE plus Bio-Rad Gel Image Analyzer was applied to examine the expression and dissolubility of rGroEL protein while Ni-NTA affinity chromatography was used to extract the expressed rGroEL.The immunoprotective rate in rGroEL-immunized LVG hamsters was determined after challenge with L.interrogans strain Lai.The cross agglutination titers of sera from immunized hamsters with different L.interrogans serogroups were detected using MAT.
Results: The nucleotide and amino acid sequences of the cloned groEL gene were the same as those reported in GenBank.The constructed prokaryotic expression system of groEL gene expressed soluble rGroEL.The immunoprotective rates of 100 and 200 μg rGroEL in LVG hamsters were 50.0% and 75.0%,respectively.The sera from the rGroEL-immunized LVG hamsters agglutinated all the L.interrogans serogroups tested with different levels.
Conclusion: The GroEL protein is a genus-specific immunoprotective antigen of L.interrogans and can be used to develop an universal genetically engineering vaccine of Leptospira.

Key wordsLeptospira interrogans      GroEL protein/genetics      Recombination,genetic      Gene expression      Sequence analysis,DNA      Immunization,passive      groEL gene      Prokaryotic expression      Immunoprotection     
Received: 28 December 2012      Published: 25 March 2013
CLC:  R 377.5  
Cite this article:

LI Xiao-Yu, WANG Yin-Huan, YAN Jie, CHENG Dong-Qing. Prokaryotic expression of Leptospira interrogans groEL gene and immunoprotection of its products in hamsters. Journal of ZheJiang University(Medical Science), 2013, 42(2): 164-170.

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结果:所克隆的groEL基因核苷酸和氨基酸序列与GenBank中相应基因完全相同。所构建的groEL基因原核表达系统能表达可溶性rGroEL。100和200 μg rGroEL对金地鼠的免疫保护率分别为50.0%和75.0%。rGroEL免疫金地鼠血清可不同程度地凝集各问号钩体血清群。

关键词: 问号钩端螺旋体,  GroEL蛋白质/遗传学,  重组,  遗传,  基因表达,  序列分析,  DNA,  免疫法,  被动,  groEL基因,  原核表达,  免疫保护 

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