Construction Of HCMV DNA Vaccine And Immune Response In Mice Induced By The Vaccine

Human cytomegalovirus (HCMV) infection is usually asymptomatic in immunocompetent people. However, it may cause severe and, sometimes, fatal disease in immunologically immature or immunocompromised individuals, such as transplant recipients, human immunodeficiency virus (HIV)-infected patients, and developing fetuses . The clinical significance of HCMV has increased due to the increased number of organ allograft transplant recipients and HIV-infected individuals. Furthermore, HCMV is the leading infectious cause for birth defects in newborns. Development of a safe and effective HCMV vaccine is thus required for those people at risk for HCMV infection and disease and has been placed in the top priority by the Institute of Medicine for the clinical and economic benefit that a vaccine would produce .pp65 gene was obtained from HCMV containing the pp65 DNA fragment using PCR and inserted into vector pMD18-T, to form pMD18-T/pp65. The recombinant plasmid pMD18-T/pp65 was digested with KpnI and NheI . The DNA fragment recovered by electrophoresis was ligated into the vector pVAX1 digested by KpnI /NheI, the resulting plasmid, pVAX1/pp65 was gotten.After analysis of restriction and sequencing,plasmids extracted large quantities by alkali lysis and purified by sepharose 4FF chromatography technique.Injected the plasmids intramuscularly directly into BALB/c mice,contrast to the pVAX1 negative control groups simultaneously.ELISA detected the induced humoral immunal response,and cellular immunal response was evaluated by spleen lymphocyte cells transformation experiment/MTT method.Successfully amplified the pp65 gene of human cytomegalovirus AD169 strain and Constructed the eukaryotic expression vector of pVAX1/pp65 The recombinated plasmids could induce the BALB/c mice to produce antibodies and the titre achieve to 1:80 at the fourth,sixth,fifth week respectively, furthermore the time last long. The mean value of ConA stimulated indexes were 1.748±0.051 in the DNA vaccines immunized mice spleen cells seprately, while in the pVAX1 control groups, the mean values were 1.095±0.060.We have demonstrated that HCMV DNA vaccine was active as an immunogen. The DNA vaccines of pVAX1/pp65 could induce the BALB/c mice to produce specific immune response depending on the route of administration and immune status of the animal. In conclusion, more and more attentions have been paid to the study on vaccine using DNA in the recent years, and many great achievements have been attained in this field. However, many problems still need to be solved. There is a long way to go before practical use of DNA-derived vaccine.