Construction Of RNA Replicon Vaccine And DNA Vaccine Of HBV And Comparison Of Their Immune Efficiency In Mice

Hepatitis B Virus (HBV) may infect human by many ways, and itcan lead to the spread of hepatopathy among people. Since hepatis istransmited widely in China, it is essential to research the preventing andtreatment against hepatis. The vaccine currently used widely in ourcountry is producted by genetic engineering. There are still someproblems to be solved quickly in the vaccine application, such as immuneescape, not responsing or low response, cell immune failure, et al. DNAvaccine may create endogenous antigens, which may lead to specific CTLresponse. CTL is very important for preventing the infection of virus. Theaim of this research is to construct HBV RNA replicon vaccine andcompare its immune efficiency with DNA vaccine, and search for the bestpattern of RNA replicon vaccine and its best way of immunization.Gene design, PCR linking and clone: Two antigen genes weredesigned refer to bioinformatics and many references. They are S2S andMAG. There some CTL, HTL and B cell epitopes in MAG. S2S andMAG were obtained by PCR with many overlaping primers. Then S2Sand MAG were linked with pGEM-T. By screening and sequencing, thecorrect clones of pGEM-S2S and pGEM-MAG are obtained.Construction of RNA replicon vaccine and DNA vaccine: Eightvectors of vaccine have been constructed.pGEM-S2S and pGEM-MAG were digested by BamHⅠ. Meantime,pSFV and pcDNA3.1+ were digested by the same enzyme. Then thefragments were linked respectively, and 4 vectors were obtained:pSFV-S2S,pSFV-MAG,pcDNA-S2S,pcDNA-MAG。IL-2 was used as molecular adjuvant, so as to study its function inthe immunity of DNA vaccine and RNA replicon vaccine. Fragments ofIVS and IRES(named "II"in this research) were amplified frompIRES1neo. IL-2 was amplified from a plasmid witch composes IL-2.