[Expression of the fusion protein CFP10-ESAT6 of Mycobacterium tuberculosis and the study of its immunogenicity].
OBJECTIVE
To express a recombinant fusion protein CFP10-ESAT6 of Mycobacterium tuberculosis, and obtain the polyclonal antibodies of this fusion protein by immune rabbit.


METHODS
The 630 bp cfpl0-esat6 fusion gene fragments were amplified from the genomic DNA of a Mycobacterium tuberculosis reference strain H37Rv and inserted into the expression plasmid pET32a (+) to generate the recombinant plasmid pET-cfp10-esat6. The recombinat expression plasmid was transformed into E. coli BL21 (DE3). The fused protein CFP10-ESAT6 with His-tag was expressed after inducing with IPTG and purified with affinity chromatography. This protein was used to immune the rabbit to obtained the polyclonal antibodies, and been analyzed with Western-blot and ELISA.


RESULTS
The recombinant plasmid pET-cfp10-esat6 was success fully constructed, the recombinant fusion protein CFP10-ESAT6 could be expressed at relatively high levels, and the polyclonal antibodies of fusion protein were obtained.


CONCLUSIONS
The successful construction and expression of the recombinant fusion protein CFP10-ESAT6 and the obtained polyclonal antibodies will be very helpful for the development of new anti-tuberculosis vaccine and the clinical serologic diagnosis.
Wang
Xiao-ying
XY
Research Unit of Infection and Immunity, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Bao
Lang
L
Zhao
Ming-cai
MC
Zhang
Hui-dong
HD
Long
Yang
Y
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000378:metabolism; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D032581:Tuberculosis Vaccines / Q000276:immunology
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C117940:CFP-10 protein, Mycobacterium tuberculosis; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D011993:Recombinant Fusion Proteins; D032581:Tuberculosis Vaccines