Abstract:

Objective   To construct a HBV esiRNA library and to study its inhibitory effects on HBV replication in HepG2.215 cells. Methods   The GST-fusion protein of Escherichia coli endoribonucleaseⅢ (GST-RNaseⅢ) was prepared, purified，  and then used to cleave the double strand RNA(dsRNA) transcribed in vitro from a two-orientation-transcription template for 1-540 nucleotides of the HBV genome(T7-HBV-1). The HBV esiRNA library generated from the above digestion was then used at 50, 100 and 150nmol/L to transfect HepG2.215 cells carrying HBV. The inhibitory effects on HBsAg expression were examined with QRT and ELISA assays. Results   Active enzyme GST-RNaseⅢ was prepared and purified. The HBV esiRNA library was successfully generated from dsRNA by GSTRNaseⅢ digestion. HBsAg expression was significantly reduced in HepG2 cells after transfection of the esiRNA library. Conclusion   Biologically prepared esiRNA library could successfully inhibit the replication of HBV in HepG2.215 cells, providing a novel idea for the treatment of HBV infection.

Key words: RNA interference; Library; Hepatitis B virus; EndoribonucleaseⅢ