Construction，expression and identification of the eukaryotic expression vector carrying Mycobacterium tuberculosis PPE68 gene

Objective:To establish the eukaryotic expression vector of recombinant PPE68 protein in Mycobacterium tuberculosis gene plasmid and lay the foundation for the later analysis of immunogenicity of PPE recombinant protein． Methods:The total DNA of Mycobacterium tuberculosis was extracted and PPE68 gene was amplified by PCR． The DNA fragment was inserted into pGEM-T，then the target gene was subcloned into pcDNA3． 1( + ) after the DNA fragment was cut from pGEM-T． New recombinant plasmid was transfected into hela cells，then expression product was confirmed by Western blot． Results:The size of the PCR product was 1 104 bp， the inserts of pGEM-T-PPE68 and pcDNA3． 1( + )-PPE68 digested by HindIII and EcoRI were as long as PCR product． The product that recombinant DNA be expressed in HeLa cells was about 40 000． Conclusions:The Results demonstate eukaryotic expression vector carring PPE68 gene has been set up and the expression products has been obtained and appraised．