Construction Of Recombinant Poliovirus Expressing HIV-1 Gp120 Gene Antigen Determinant And Research On Expression

Background:Since Acquired Immune Deficiency Syndrom(AIDS) caused by human immunodeficiency virus(HIV)was identified,which is threatening the human health badly all along.Moreover,the price of drugs to resist HIV is expensive,and the drugs produce badness reactions easily,to use drugs excessively make the badness reactions increased, even bring death.In recent years,we pay more and more attention to the study on virus carrier vaccine.The gene gp120 of HIV is the emphases on the AIDS vaccine study.Poliovirus can induce cell-mediated,humoral and intestine mucus membrane immunity,poliovirus which lack P1 protein coding sequence can self-replicate,and it quality and safety detection means has been founded perfectly.So there are great applying views to construct recombinant poliovirus vaccine expressing HIV-1 gp120,at present,there were no reports about recombinant poliovirus vaccine expressing HIV-1 gene.Objective:To control AIDS prevalence effectively,the requirement of the HIV vaccine for preventing or treatment is exigent increasingly presently.This research mainly constructs recombinant poliovirus expressing HIV-1 gp120 gene antigen determinant and studies its expression. Methods:Chinese HIV-1B sub-type representative strains gp120 gene were chose,we used PCR to amplify gp120 gene fragments with plasmid pT7gp120 as template,gp120 amplification products and poliovirus expressing carders were catalyzed by EcoRI/SnaBI nd the target genes gp120 were inserted into the catalyzed carders correctly, recombinant polioviruses plasmids were constructed,recombinant plasmids were transferred into Hela cells using lipid transfer technique, transferred cells were chosen using G418 reagent,cell strains transferred recombinant poliovirus plasmids were gained,the expression of HIV-1 gp120 were detected with RT-PCR,ELISA;Recombinant vaccinia viruses provided P1 pre-protein for recombinant polioviruses,the purified recombinant poliovirus were gained.Large numbers of virus granule were observed by electron microscope;that recombinant poliovirus can infect 293 cells and express HIV-1 genes was detected by Western blot.Results:Construct recombinant poliovirus expressing HIV-1 gp120 gene antigen determinant,the experiment used RT-PCR,ELISA to indicate that the recombinant poliovirus can express HIV-1 gp120 in vivo, the recombinant poliovirus are regular icosahedrons with diameter 20-30nm on electron microscope,and the recombinant poliovirus can infect cells,and have heredity stability.Conclusion:Construct recombinant poliovirus expressing HIV-1 gp120 gene antigen determinant successfully,and it can express HIV-1 gp120 gene antigen determinant successfully,and it can express HIV-1 gp120 in vivo;the recombinant poliovirus are regular icosahedrons with diameter 20-30nm on electron microscope,and the recombinant poliovirus can infect cells,and have heredity stability;this experiment provide the foundation for further research on immunogenicity of recombinant poliovirus expressing HIV-1 gp120 gene antigen determinant,and humoral immunity and cell immunity induced by recombinant poliovirus,and provide experimental foundation for prevention and cure of HIV infection.