AIM: To construct a live attenuated Salmonella ty- phimurium (S. typhimurium) strain harboring the H pylori neutrophil activating protein (HP-NAP) gene as an oral recombinant DNA vaccine, and to evaluate its immuno- genicity. METHODS: By genetic engineering methods, the ge- nomic DNA of H pylori was extracted as a template. The total length of the HP-NAP gene was amplified by poly- merase chain reaction (PCR) and cloned into pBT vector for sequencing and BLAST analysis, then subcloned into a eukaryotic expression vector pIRES followed by PCR identification and restriction enzyme digestion. The iden- tified recombinant plasmid pIRES-NAP was transfected into COS-7 cells for target fusion protein expression, and its antigenicity was detected by Western blotting. Then the recombinant plasmid was transformed into a live attenuated S. typhimurium strain SL7207 as an oral vac- cine strain, and its immunogenicity was evaluated with animal experiments. RESULTS: A 435 bp product was cloned using high homology with HP-NAP gene in GenBank (more than 98%). With identification by PCR and restriction enzyme digestion, a recombinant eukaryotic expression plasmid pIRES-NAP containing the HP-NAP gene of H pylori was successfully constructed. The expressed target protein had a specific reaction with H pylorii whole cell antibody and showed a single strip result detected by Western blotting. Oral immunization of mice with recombinant DNA vaccine strain SL7207 (pIRES-NAP) also induced a specific immune response. CONCLUSION: The successful construction of HP-NAP oral DNA vaccine with good immunogenicity may help to further investigate its immunoprotection effects and de- velop vaccine against H pylori infection.
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