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The hepatitis C virus (HCV) genotype is the most important factor in predicting the outcome of chronic hepatitis C treatment. Therefore, convenient and accurate HCV genotyping methods for routine laboratory testing are needed. In this study, to identify the HCV genotypes, an oligonucleotide DNA chip was designed using 15 probes from the 5'-untranslated region. Reverse transcription was combined with asymmetric polymerase chain reaction (PCR) to obtain an amplified product for hybridization without the need for a denaturation step. In addition, a biotin-labeled PCR product and streptavidin-Cy3 conjugate were cohybridized simultaneously. Clinical sera from Korean and French patients (n=112) were used to compare the chip results with those obtained by direct sequencing. The DNA chip showed 100% accuracy for the commercial panels and had a lower detection limit of 2.8x10(2) IU/ml. Agreement levels between the chip and sequencing results at the genotype and subgenotype levels were 100% and 94.6% (106/112), respectively. By the combined reverse transcription-asymmetric PCR and cohybridization methods, the DNA chip reduced assay time and was convenient to use. This DNA chip may be useful for identifying HCV genotypes in clinical laboratories., (2009 Elsevier B.V. All rights reserved.)