Objective: Recently, HCV subgenomic RNA that replicates in vitro in a certain cell line have been elucidated. Since the 5) end of the genome of positive strand RNA viruses is often modified with a cap structure or a covalently linked protein, we have assessed structural feature of the HCV genome obtained from Huh7 cells in which HCV subgenomic RNA has been shown to efficiently self-replicate. Methods: HCV subgenomic RNA was obtained from the Huh7 and was analyzed for its 5) end. Results: Phosphorylation of the genomic RNA by polynucleotide kinase was observed only after treatment with phosphatase. The labeling efficiency of the genome with polynucleotide kinase was not enhanced by treatment with pyrophosphatase. Conclusion: It is suggested that the 5) end of HCV genomic RNA obtained from HCV replicon cells is not modified except phosphorylation. Furthermore, analysis of the 5) end of the HCV RNA obtained from the HCV subgenome self-replicating cells revealed the presence of two types of subgenomic RNA that contained either guanylate or adenylate at the 5) end. This result indicates that the 5) end of the subgenome in Huh7 cells is redundant and there is no significant evolutionary advantage between the two genomes. Copyright (C) 2005 S. Karger AG, Basel.