Construction and expression of fused gene vaccine esat6-cfp10 of Mycobacterium tuberculosis

AIM: To construct and express fused gene vaccine esat6-cfp10 of Mycobacterium tuberculosis. METHODS: cfp10 gene was amplified by PCR from H37Rv virulent strain of MTB and was then inserted into pGEM-7zf(+) cloning vector containing esat6 gene. After sequenced, the esat6-cfp10 fused gene was cloned into eukaryotic vector pcDNA3.1(+). This recombinant plasmid was transfected into Chinese Hamster Ovary (CHO) cells by cation liposome. The expression of mRNA and the fused protein were detected by RT-PCR and indirect immunofluorescence technique. RESULTS: The length of PCR product was 350 bp, identical with that reported by GenBank. The 630 bp fragment of the fused esat6-cfp10 gene was obtained by restriction enzyme digestion. RT-PCR suggested that the mRNA was expressed in CHO cells and the positive cells were stained positively by indirect immunofluorescence technique. CONCLUSION: cfp10 gene was successfully cloned and the eukaryotic recombinant plasmid fused esat6-cfp10 was constructed successfully and expressed in CHO cells.