摘要
AIM: To study the effect of siRNA expressed from DNA vector on HBV replication.METHODS: Human U6 promoter was amplified from genomic DNA and cloned into plasmid pUC18 to construct a mammalian siRNA expression vector pUC18U6. Then oligonucleotides coding for a short hairpin RNA against HBV were cloned into pUC18U6 to form pUC18U6HBVsir which was introduced into 2.2.15 cells by using liposome-mediated transfection.2.2.15 cells transfected by pUC18U6 and pUC18U6GFPsir which expressed siRNA against green fluorescent protein and mock-transfected 2.2.15 cells were used as controls.Concentration of HBsAg in the supernatant of the transfected cells was measured by using solid-phase radioimmunoassay.RESULTS: A mammalian siRNA expression vector pUC18U6 was 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 inhibited by siRNA expressed from the DNA vector.
AIM:To study the effect of siRNA expressed from DNA vector on HBV replication. METHODS:Human U6 promoter was amplified from genomic DNA and doned into plasmid pUC18 to construct a mammalian siRNA expression vector pUCt8U6.Then oligonucleotides coding for a short hairpin RNA against HBV were cloned into pUC18U6 to form pUCi8U6HBVsir which was introduced into 2.2.15 cells by using liposome-mediated transfection. 2.2.15 cells transfected by pUC18U6 and pUC18U6GFPsir which expressed siRNA against green fluorescent protein and mock-transfected 2.2.15 cells were used as controls. Concentration of HBsAg in the supernatant of the transfected cells was measured by using solid-phase radioimmunoassay. RESULTS:A mammalian siRNA expression vector pUC18U6 was 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 inhibited by siRNA expressed from the DNA vector.
基金
Supported by National Natural Science Foundation of China,No.30100157
Medical Research Fund of Chinese PLA,No.01MA184
and Innovation Project of FMMU,No.CX99005
