›› 2011, Vol. 31 ›› Issue (1): 118-.doi: 10.3969/j.issn.1674-8115.2011.01.028

• Technique and method • Previous Articles     Next Articles

Preparation of polyclonal antibodies of Escherichia coli type Ⅵ secretion system related proteins and detection of secreted proteins

SHI Xiao-lei1,2, CHEN Xiao-xi1,2, JIN Rui1,2, LI Hao1,2, XUE Pei1,2, YAO Yu-feng1, TAO Jing1   

  1. 1.Department of Medical Microbiology and Parasitology, Shanghai Jiaotong University School of Medicine, 2.Grade 2006, Department of The Eight-year Program of Clinical Medicine, Shanghai Jiaotong University, Shanghai 200025, China
  • Online:2011-01-28 Published:2011-02-01
  • Supported by:

    National Natural Science Foundation of China, 30870100

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Abstract:

Objective To prepare the polyclonal antibodies of AaiC, CRP and OmpA proteins for the research of function of Escherichia coli （E. coli） type Ⅵ secretion system, and detect the secreted protein AaiC. Methods Target genes were amplified by PCR and cloned into prokaryotic expression vector pET-22b or pQE80, and the recombinant plasmids were transformed into E. coli BL21 （DE3）. The fusion proteins were induced with IPTG, and were used to immunize New Zealand rabbits to generate polyclonal antibodies after purification. The specificity of polyclonal antibodies was evaluated by Western blotting. Secreted proteins of E. coli RS218 were detected by polyclonal antibodies of protein AaiC. Results The recombinant plasmids were successfully constructed, and the fusion proteins were highly expressed in E. coli. The specificity of polyclonal antibodies of AaiC, CRP and OmpA proteins was confirmed. AaiC antibody detection revealed that RS218 could secret protein AaiC. Conclusion The specific polyclonal antibodies of three recombinant fusion proteins AaiC, CRP and OmpA in E. coli have been successfully generated, and protein AaiC can be secreted by RS218, which can be applied in the research of function of Ⅵ secretion system of E. coli and pathogenesis of E. coli.

Key words: Escherichia coli, type Ⅵ secretion system, polyclonal antibody, secreted protein