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ISSN 1672-9145 Acta Biochim Biophys Sin 2005, 37(3): 199–204 CN 31-1940/Q

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Co-expression and Immunity of Legionella pneumophila mip Gene and Immunoadjuvant ctxB Gene

Tao WANG, Jian-Ping CHEN*, Hong LI, Ke-Qian ZHI¹, Lei ZHANG, Chun-Lei YANG², and Da-Chang TAO²

School of Preclinical and Forensic Medicine of West China, Sichuan University, Chengdu 610041, China;

¹School of Stomatology, Xi’an Jiaotong University, Xi’an 710004, China;

²School of Life Science, Sichuan University, Chengdu 610041, China

Abstract The mip gene of Legionella pneumophila and the ctxB gene of Vibrio cholerae were amplified by PCR respectively. The amplified cDNA was ligated to the pcDNA3.1(+) vector. The recombinant plasmids pcDNA3.1-mip and pcDNA3.1-ctxB were identified by restriction analysis and PCR, and further confirmed by sequencing analysis. NIH3T3 cells were transfected with pcDNA3.1-mip and pcDNA3.1-ctxB according to the Lipofection method. Transient and stable products of the co-expression of the mip gene and ctxB gene were detected by immunofluorescence and Western blotting. The results showed that NIH3T3 cells were successfully transfected, and that the transiently and stably co-expressed products can be detected in the transfected cells. To detect the humoral and cellular immune response in immunized mice induced by the co-mmunization of the mip and ctxB genes, female BALB/c mice were immunized intramuscularly with pcDNA3.1-mip and pcDNA3.1-ctxB. The results showed that the specific antibody titer and the cytotoxic T-lymphocyte response for pcDNA3.1-mip immunization and co-immunization were increased compared with that of pcDNA3.1(+) immunization. Furthermore, the specific antibody titer and cytotoxic T-lymphocyte response for co-immunization were increased compared with that of pcDNA3.1-mip immunization. Statistical analysis using one-way analysis of variance (ANOVA) showed that there was a significant difference between the groups (P<0.01). The results indicated that the ctxB gene enhanced the humoral and cellular immune response to the mip gene immunization. These findings provide experimental evidence to support the development of the L. pneumophila DNA vaccine.

Key words Legionella pneumophila; co-expression; humoral immunity; cellular immunity

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Received: November 23, 2004 Accepted: February 1, 2005

This work was supported by a grant from the National Natural Science Foundation of China (No. 30300302)

*Corresponding author: Tel, 86-28-85501248; E-mail, [email protected]