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BACKGROUND: DNA immunization provides a promis- ing approach to elicit protective humoral and cellular im- mune responses against HBV. This study was to construct an eukaryotic expression plasmid containing helper T lym- phocyte epitope, which will enhance the immunogenicity of a novel hepatitis B virus ( HBV) fusion protein DNA vaccine. METHODS: The target gene containing pan-DR helper T cell epitopes (PADRE) and HBsAg was amplified by poly- merase chain reaction (PCR). The PCR products were linked with PMD-18T vector. Plasmid DNA was purified from transformed E. coli competent cell JM109 and digested with Hind and EcoR . Then, the target gene was cloned in pcDNA3.1(+) digested by Hind and EcoR . Finally, the identity of DNA was verified by digestion and DNA sequencing. RESULTS: The recombinant expression vectors of pcDNA 3. l(+)-PADRE/HBs were identified by restriction enzyme digestion and DNA sequencing. The insert DNA fragment was consistent with the expected sequence. CONCLUSIONS: The constructed eukaryotic expression plasmid of pcDNA3.1(+)-PADRE/HBs is convenient for further study of eukaryotic transfection and response for cellular and humoral immunity against HBV.
BACKGROUND: DNA immunization provides a promis- ing approach to elicit protective humoral and cellular im-mune responses against HBV. This study was to construct an eukaryotic expression plasmid containing helper T lym- phocyte epitope, which will enhance the immunogenicity of a novel hepatitis B METHODS: The target gene containing pan-DR helper T cell epitopes (PADRE) and HBsAg was amplified by poly-merase chain reaction (PCR). The PCR products were linked with PMD-18T vector. Plasmid DNA was purified from transformed E. coli competent cells JM109 and digested with Hind and EcoR. Then, the target gene was cloned in pcDNA3.1 (+) digested by Hind and EcoR. Finally, the identity of DNA was verified by digestion and DNA sequencing. RESULTS: The recombinant expression vectors of pcDNA 3. l (+) - PADRE / HBs were identified by restriction enzyme digestion and DNA sequencing. The insert DNA fragment was consistent with the expected sequence. The constructed eukaryotic expression plasmid of pcDNA3.1 (+) - PADRE / HBs is convenient for further study of eukaryotic transfection and response for cellular and humoral immunity against HBV.