Four kinds of different hammerhead ribozymes (ribozymeA, ribozymeB,ribozymeC₁,ribozymeC₂) were designed and synthesized according to the secondary structure of HCV-RNA 5′-untranslated region and part of the neighbour C-region. Firstly, the cleavage of the four Rzs were tested in vitro, and only the ribozyme with GTA↓ motif at-11nt site of HCV-RNA showed cleavage activity. RzA-RNA and the combinated pCl-neo-luciferase in which a luciferase gene were ligated downstream the target sequence were then co-transfected into HepG₂ cell lines with lipofectine. The cleavage of RzA-RNA was tested by determined the expression of luciferase gene. Therefore, the gene of RzA was ligated into expression vector pCl-neo. This pCl-neo-RzA and the vector pCl-neo-luciferase were co-transfected into HepG₂ cell lines again with lipofectine. Since the pCl-neo-RzA was more stable than RzA-RNA in vivo and could produce RzA-RNA continously, it showed better cleavage activity.