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Inhibition of hepatitis B virus surface antigen expression by small hairpin RNA in vitro
Zheng-Gang Yang1, Zhi Chen, Qin Ni
1Institute of Infectious Diseases, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, Zhejiang Province, China.
Aim:
To explore the anti-hepatitis B virus effect of RNA interference (RNAi) using small hairpin RNA (shRNA) expression vector.
Methods:
Hepatitis B virus surface antigen green fluorescent protein (HBs-GFP) fusion vector and shRNA expression vectors were constructed and cotransfected transiently into HepG2 cells. mRNAs extracted from HepG2 cells were detected by real-time PCR. Fluorescence of HBs-GFP protein was detected by fluorescence-activated cell sorting (FACS). The effective shRNA expression vector was transfected into HepG2.2.15 cells. HBsAg and HBeAg in HepG2.2.15 cells were analyzed by radioimmunoassay (RIA) method.
Results:
FACS revealed that shRNA targeting at HBsAg reduced the GFP signal by 56% compared to the control. Real-time PCR showed that HBs-GFP mRNA extracted from HepG2 cells cotransfected with pAVU6+27 and HBs-GFP expression plasmids decreased by 90% compared to the empty vector control. The expressions of HBsAg and HBeAg were also inhibited by 43% and 64%, respectively.
Conclusion:
RNAi using shRNA expression vector can inhibit the expression of HBsAg, providing a fresh approach to screening the efficient small interfering RNAs (siRNAs).
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