[Construction and expression of recombinant Mycobacterium bovis BCG with the ompA-like membrane protein gene Loa22 of Leptospira interrogans serovar].
OBJECTIVE
To study the immunity of Loa22 from Leptospira interrogans serovar Lai strain 56601 by expressing its protein in BCG.


METHODS
Amplified the mature peptide of Loa22 gene from the genome of of Leptospira interrogans serovar Lai strain 56601 and constructed recombinant plasmid rpMV36l-1oa22 with the E. coli-BCG integrating shuttle plasmid pMV361 and the Loa22 mature peptide gene. The rpMV36l-1oa22 plasmid was transformed into BCG by electroporation. The rBCG bearing rpMV36l-1oa22 was induced by high temperature of 45 degrees C and expressed protein was identified by SDS-PAGE and Western Blotting. Fifth 6-week-old BALB/c mice were randomly divided into five groups, which were inoculated intraperitoneally two times at 0-day and 21-day with BCG, rBCG-pMV361, rI3CG-1oa22, Loa22 and killed whole-leptospires respectively. All animals were dislocated from cervical vertebra on the 14Ih day after the last immunization. The proliferative reaction of splenic lymphocyte in tuitro were tested by XTT.


RESULTS
The rpMV36l-1oa22 plasmid was constructed successfully and transformed into BCG. The rBCG expressed a 19 X io  specifical protein identified by SDS-PAGE and Western Blotting. The splenic lymphocyte proliferate activity (SI) in rBCG-ioa22 group in intro was significantly higher than those in BCG group and rBCG-pMV361 group.


CONCLUSIONS
We explored the expressing feasibility of Loa22 in Mycobacterium bovis BCG. may therefore make further researches on the induction of protective immunity against human and animal leptospirosis.
Li
Dao-kun
DK
The Laboratory of Infection and Immunity, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Bao
Lang
L
Zhang
Ying
Y
Sun
Zhan
Z
D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D001428:Bacterial Vaccines / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D007114:Immunization; D007921:Leptospira interrogans / Q000235:genetics* / Q000276:immunology; D007922:Leptospirosis / Q000517:prevention & control*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009163:Mycobacterium bovis / Q000235:genetics / Q000378:metabolism*; D011897:Random Allocation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology
D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D011994:Recombinant Proteins; C078167:OMPA outer membrane proteins