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Abstract: Objective: To determine the effective nucleotide sites of ribozymes against HCV, and obtain a highly effective, nontoxic and inexpensive antisense ribozyme specific for HCV. Methods: Two effective ribozymes, targeted to HCV 5'NC region and C region, were designed and synthesized.Eukaryotic expression vectors, pSV2-gpt. CD-SR, containing either HCRZNC or HCRZC were constructed and transfected into MT-2 cells, which had been infected by HCV. Quantitative PCR and hybridization were used to determine the effect of inhibition of HCV by ribozymes. Results: HCRZNC and HCRZC suppressed the replication of HCV by 54.7% and 2.1%, respectively. Furthermore, when both ribozymes were cotransfected into cells, they suppressed replication by 78.8%. Conclusion: Two specific antisense ribozymes have strong inhibitory effects on the replication of HCV in cultured cells, and have better effect when used together.

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