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CLONING AND EXPRESSION OF MYCOBACTERIUM TUBERCULOSIS Ag85B IN E.COLI

Yue Zhao

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Abstract

Objective:To analyze the protective epitopes of Antigen 85B(Ag85B)of M.tuberculosis,and investigate the potential use of Ag85B as vaccine for the treatment and prevention of tuberculosis,expression recombinant of mature Ag85B was constructed.Methods:Ag85B gene was amplified with PCR,inserted into pUC18 and then sequenced.The cloned Ag85B was seperated and inserted in the downstream of PL promotor of expression vector pBV220.The SDS PAGE and Western Blot were used to identify the recombinant protein.Results:Both cloning and expression recombinants of Ag85B were obtained.DNA sequencing indicated that cloned Ag85B gene was similar to that reported previously.The mature Ag85B was expressed at high level(25%～30% of total bacterial protein)in E.coli.Conclusion:The cloning and high level expression of Ag85B in E.coli provide basis for future study on the preventive and therapeutic vaccine of tuberculosis.

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What this paper is about

Objective:To analyze the protective epitopes of Antigen 85B(Ag85B)of M.tuberculosis,and investigate the potential use of Ag85B as vaccine for the treatment and prevention of tuberculosis,expression recombinant of mature Ag85B was constructed.Methods:Ag85B gene was amplified with PCR,inserted into pUC18 and then sequenced.The cloned Ag85B was seperated and inserted in the downstream of PL promotor of expression vector pBV220.The SDS PAGE and Western Blot were used to identify the recombinant protein.Results:Both cloning and expression recombinants of Ag85B were obtained.DNA sequencing indicated that cloned Ag85B gene was similar to that reported previously.The mature Ag85B was expressed at high level(25%～30% of total bacterial protein)in E.coli.Conclusion:The cloning and high level expression of Ag85B in E.coli provide basis for future study on the preventive and therapeutic vaccine of tuberculosis.

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Available abstract

Objective:To analyze the protective epitopes of Antigen 85B(Ag85B)of M.tuberculosis,and investigate the potential use of Ag85B as vaccine for the treatment and prevention of tuberculosis,expression recombinant of mature Ag85B was constructed.Methods:Ag85B gene was amplified with PCR,inserted into pUC18 and then sequenced.The cloned Ag85B was seperated and inserted in the downstream of PL promotor of expression vector pBV220.The SDS PAGE and Western Blot were used to identify the recombinant protein.Results:Both cloning and expression recombinants of Ag85B were obtained.DNA sequencing indicated that cloned Ag85B gene was similar to that reported previously.The mature Ag85B was expressed at high level(25%～30% of total bacterial protein)in E.coli.Conclusion:The cloning and high level expression of Ag85B in E.coli provide basis for future study on the preventive and therapeutic vaccine of tuberculosis.

Key concepts: Cloning (programming), Mycobacterium tuberculosis, Recombinant DNA, Biology, Gene, Plasmid, Expression vector, Tuberculosis

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