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AIM:To study the effect of siRNA expressed from DNA vectoron HBV replication.METHODS:Human U6 promoter was amplified from genomicDNA and doned into plasmid pUC18 to construct a mammaliansiRNA expression vector pUCt8U6.Then oligonucleotidescoding for a short hairpin RNA against HBV were clonedinto pUC18U6 to form pUCi8U6HBVsir which was introducedinto 2.2.15 cells by using liposome-mediated transfection.2.2.15 cells transfected by pUC18U6 and pUC18U6GFPsirwhich expressed siRNA against green fluorescent proteinand mock-transfected 2.2.15 cells were used as controls.Concentration of HBsAg in the supernatant of the transfectedcells was measured by using solid-phase radioimmunoassay.RESULTS:A mammalian siRNA expression vector pUC18U6was constructed successfully.Compared with controls,pUC18U6HBVsir which expressed siRNA against HBV decreasedconcentration of HBsAg significantly by 44%(P<0.05).CONCLUSION:HBV replication in 2.2.15 cells is inhibitedby siRNA expressed from the DNA vector.
AIM: To study the effect of siRNA expressed from DNA vectoron HBV replication. METHODS: Human U6 promoter was amplified from genomic DNA and doned into plasmid pUC18 to construct a mammalians RNAi vector pUCt8U6.Then oligonucleotides encoding for a short hairpin RNA against HBV were clonedinto pUC18U to form pUCi8U6HBVsir which was introduced in 2.2.15 cells by using liposome-mediated transfection.2.2.15 cells transfected by pUC18U6 and pUC18U6GFPsirwhich expressed siRNA against green fluorescent protein and mock-transfected 2.2.15 cells were used as controls. Confluence of HBsAg in the supernatant of the transfected cells were measured by using solid-phase radioimmunoassay.RESULTS: A mammalian siRNA expression vector pUC18U6was constructed successfully.Compared with controls, pUC18U6HBVsir which expressed siRNA against HBV decreased concentration of HBsAg significantly by 44% (P <0.05) .CONCLUSION: HBV replication in 2.2.15 cells is inhibitedby siRNA expressed from the DNA vector.