Construction and Identification of the Expression Plasmid Containing Ag85B Gene of Mycobacterium tuberculosis

To construct the plant expression plasmid containing Mycobacterium tuberculosis Ag85B genes,and transform the recombinant plasmid into Agrobacterium tumerfaciens LBA4404.We used pcDNA3-Ag85B as template to amplify Ag85B gene by polymerase chain reaction(PCR),and cloned Ag85B gene into the vector pCR2.1.The fusion fragment of promoter globulin-1 and the target gene Ag85B which get from double enzymes digestion of the recombinant plasmid pCRGAg85B was inserted into the plasmid pCAMBIA1300 which contains the bar gene for herbicide resistance,transformed recombinant plasmid pCAMG-Ag85B into Agrobacterium tumerfaciens LBA4404.Restriction enzyme digestion demonstrated that the inserted gene fragments were 0.8 bp,DNA sequence analyzing revealed that the Ag85B gene sequences were totally consistent with the GenBank reported.We have successfully constructed expression plasmid containing Ag85B gene and transformed it into LBA4404,and lays a foundation for further study a better vaccine against MTB.