Construction of Mycobacterium tuberculosis protective antigens-ubiquitin system / 第二军医大学学报

ABSTRACT

Objective: To fuse Mycobacterium tuberculosis protective antigen gene with mice ubiquitin gene, constructing antigen ubiquitin system. Methods: Mice ubiquitin cDNA was amplified by RT PCR from mice testicle,and 4 antigen genes were obtained by PCR from cultured Mycobacterium tuberculosis . Ubiquitin and 4 antigen genes were linked by flexible adaptor respectively and the fusing genes were cloned into pVAX vector.The recombinant plasmids were digested with endonuclease and sequenced.Then the recombinant plasmids were transfected into COS7 cells and the expression was assayed by ELISA. Results:Ubiquitin and 4 antigen genes were 0.2,0.3,0.7,1.0,1.65 kb in length by agarose electrophoresis. Endonuclease digestion of the recombinant plasmids indicated that the fusion genes were correctly inserted into pVAX vector. Sequencing results of fusion nucleic acid vaccines were identical to those in GenBank.The recombinant plasmids expressed in COS7 cells. Conclusion: Four Chinese Mycobacterium tuberculosis protective antigens ubiquitin systems are successfully constructed and can be expressed in eukaryotic cells. This may provide a basis for diagnosis and therapy of tuberculosis.