[Construction and identification of recombinant shuttle-plasmid with ESAT-6 from Mycobacterium tuberculosis].
OBJECTIVE
To construct a recombinant BCG secretively expressing ESAT-6 of Mycobacterium tuberculosis.


METHODS
alpha-antigen(alpha-Ag) signal sequence and esat-6 gene were amplified from the genome of Bacille Calmette-Guerin (BCG) and Mycobacterium tuberculosis by PCR respectively. esat-6 gene was cloned in E. coli-BCG shuttle-plasmid pMV261 to get pME. Then a new recombinant plasmid pSME was constructed by inserting BCG alpha-Ag signal sequence into pME.


RESULTS
The cloned genes alpha-Ag signal sequence and esat-6 were correctly inserted into the vector pMV261, which was confirmed by restriction endonuclease digestion and PCR amplification of pSME.


CONCLUSIONS
pSME was expected to secretively express ESAT-6 of Mycobacterium tuberculosis in BCG. This study provides the possibility of further researches on the development of new anti-tuberculosis vaccine.
Chen
Wei
W
Institute of Infection and Immunity, West China School of Preclinic and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Bao
Lang
L
Xie
Yongen
Y
Hu
Changhua
C
Zhang
Wanjiang
W
Li
Xuemin
X
Zhang
Huidong
H
D000818:Animals; D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D001500:BCG Vaccine / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000276:immunology*; D010957:Plasmids; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
D000942:Antigens, Bacterial; D001500:BCG Vaccine; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D011994:Recombinant Proteins