表达HBsAg特异性siRNA的重组腺相关病毒载体的构建

doi:10.3969/j.issn.1672-5069.2010.03.001

Abstract

Abstract: Objective To construct the recombinant adeno-associated virus（rAAV）encoding HBsAg-shRNA and to observe its effect on HBsAg and HBeAg expression in HepG2215 cells in vitro. Methods pAAV-shHBs-hrGFP expressing plasmid was constructed by molecular biological techniques. The recombinants were cotransfected with p-RC and p-Helper into AAV-293 cells mediated by calcium acid phosphate. The rAAVs encoding HBsAg-shRNA（rAAV-shHBs-hrGFP）were harvested and infected HepG2215 cells. The silencing effect of this virus on HbsAg and HBeAg gene expression was assessed by ELISA. Results The plasmid of pAAV-shHBs-hrGFP was successfully constructed by identification of restrict enzyme and DNA sequencing. GFP expression was observed about 80% in AAV-293 cells. After cotransfection，the recombinant AAV-shHBs-hrGFP was harvested and the virus titer was tested. The HBsAg and HBeAg expression in HepG2.215 cells was down-regulated markedly after infection of rAAV-shHBs-hrGFP. Conclusion The recombinant AAV-shHBs-hrGFP vector is successfully constructed and can be used as a potential antivirus agent to inhibit HBV replication.

Key words: HepG2215 cells, AAV2, RNAi, HBsAg

HU Bin，YANG Yan，LIU Jia，et al.. Inhibition of HbsAg and HbeAg expression of recombinant adeno-associated virus encoding HBsAg-shRNA in HepG2.215 cells in vitro[J]. JOURNAL OF PRACTICAL HEPATOLOGY, 2010, 13(3): 161-165.

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Inhibition of HbsAg and HbeAg expression of recombinant adeno-associated virus encoding HBsAg-shRNA in HepG2.215 cells in vitro

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