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 ORIGINAL ARTICLE
Year : 2012  |  Volume : 30  |  Issue : 2  |  Page : 193-197

Construction of a recombinant plasmid harbouring the glyceraldenyde-3-phosphate dehydrogenase gene of periodic Brugia malayi and observation on DNA immunity


Department of Parasitology, School of Basic Medical Sciences, Nantong University, Nantong, Jiangsu - 226 001, China

Correspondence Address:
Z Fang
Department of Parasitology, School of Basic Medical Sciences, Nantong University, Nantong, Jiangsu - 226 001
China
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0255-0857.96691

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Purpose: Controlling and eliminating lymphatic filariasis will require further research of preventative measures and implementation. Parasite is dependent on glycolysis for ATP production. The glycolytic enzyme glyceraldenyde-3-phosphate dehydrogenase (GAPDH) plays an important role in glycolysis and therefore is either a potential target for anti-parasite drug development or a vaccine candidate. Therefore, we tried to investigate the DNA vaccine-elicited immune responses in BALB/c mice. Materials and Methods: We cloned a gene encoding the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) from periodic Brugia malayi into vector pcDNA3.1. Mice were injected at a dosage of 100 μg recombinant plasmid DNA with CpG intramuscular injection and immunized three times at 2-week intervals. pcDNA3.1 and normal saline were used as control. The tissue of muscles at the 4 weeks after the third injection was collected and target genes were detected using RT-PCR. The humoral responses elicited in mice by inoculation with the recombinant plasmid pcDNA3.1-BmGAPDH were detected using a standard ELISA. Two weeks after the third immunization, stimulation index (SI) was measured using the MTT method and the level of secreted IL-4 and INF-g were detected using ELISA. Results: Specific gene fragment coding GAPDH was amplified and the recombinant plasmid pcDNA3.1-BmGAPDH was constructed. Post-challenge sera from the mice immunized with the DNA vaccine had specific antibody titres of 1:1600 to 1:6400, and the highest titre was observed in the mice that were inoculated by pcDNA3.1-BmGAPDH/CpG at 6 weeks. At 4 weeks after immunization, the spleens of the mice were obviously enlarged. The proliferation of spleen T lymphocytes seen on the MTT assay was higher in the pcDNA3.1-BmGAPDH group than in the control group (P value <0.05). The levels of IL-4 and INF-g in serums from the immunized mice were significantly higher than that of the control (P value <0.05). Conclusions: We conclude that the recombinant eukaryotic plasmid pcDNA3.1-BmGAPDH could elicit humoral and cellular immune responses in mice.






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