Study On The Cleavage Activity Of The Optimized M1GS Ribozymes That Target The Core Gene Of Hepatitis C Virus

More than 170 million individuals worldwide are currently infected with the hepatitis C virus (HCV). Chronic HCV infection frequently results in serious liver disease, including steatosis, cirrhosis and hepato-cellular carcinoma. At present, there is still no specific, effective treatments. In this case it is significant to exploring a new anti-HCV treatment strategy.In this artical,the core gene (C) of HCV genome was chose as a target gene to be cleaved because it is relatively conservative and plays a key role in the process of viral replication. Basing on the ribyzyme M1GS-HCV/C₁₆₇ which was constructed before, two corresponding sequence-specific M1GS ribozymes that target the potential sites in the core gene of HCV RNA (site 52th, 141st) had been constructed. The cleavage activity of the two M1GS ribozymes was tested by the extracellular cleavage .Results showed that both the two M1GS ribyzyme could produce targeting cleavage and their cleavage activity is better than M1GS-HCV/C₁₆₇ .In addition, to determine whether the M1GS ribozymes have introcellular inhibition to the expression of target gene or not, two recombinant plasmids pc-M1GS52,pc-M1GS141 which expressing in eukaryotic cell have been construced. And then pc-HCV/core and pc-M1GS eukaryotic expressive plasmids were co-transfected into Huh-7. It was confirmed by semi-quantative RT-PCR and Western Blot that M1GS ribozyme M1GS-HCV/C52 could decrease the level of the target mRNA and reduce the expression of HCV core protein significantly.In conclusion, a M1GS ribozyme which possessing extracelluar and introcellular cleavage activities to the target mRNA of HCV c gene was successfully optimized. It provided direct experimental materials for the future research as well as the foundation for reaserch of anti-HCV treatment using the new ribozyme technology—M1GS.