Hepatitis C virus (HCV) genotyping by annealing reverse transcription-PCR products with genotype-specific capture probes.
The genotype of the hepatitis C virus (HCV) strain infecting a given patient is an important predictive factor for the clinical outcome of chronic liver disease and its response to anti-viral therapeutic agents. We herein sought to develop a new easy, sensitive and accurate HCV genotyping method using annealing genotype-specific capture probes (AGSCP) in an automation-friendly 96-well plate format. The validation of our new AGSCP was performed using the Standard HCV Genotype Panel. We then used both our AGSCP and the commercially available INNO-LiPA assay to analyze the HCV genotypes from 111 Korean patients. Discordant results were analyzed by direct sequencing. AGSCP successfully genotyped the standard panel. The genotypes of 111 patient samples were also obtained successfully by AGSCP and INNO-LiPA. We observed a high concordance rate (93 matched samples, 83.8%) between the two assays. Sequencing analysis of the 18 discordant results revealed that the AGSCP had correctly identified 12 samples, whereas the INNO-LiPA had correctly identified only 6. These results collectively indicate that AGSCP assay is a convenient and sensitive method for large-scale genotyping, and it may be a promising tool for the determination of HCV and other genotypes in clinical settings.
Rho
Jungmin
J
Panbionet, Corp., POSTECH Biotech Center, Pohang 790-784, Republic of Korea.
Ryu
Jong Soon
JS
Hur
Wonhee
W
Kim
Chang Wook
CW
Jang
Jeong Won
JW
Bae
Si Hyun
SH
Choi
Jong Young
JY
Jang
Sung Key
SK
Yoon
Seung Kew
SK
D020121:5' Untranslated Regions / Q000235:genetics; D001331:Automation; D018076:DNA, Complementary / Q000235:genetics*; D004279:DNA, Viral / Q000235:genetics; D005838:Genotype; D016174:Hepacivirus / Q000235:genetics*; D006526:Hepatitis C / Q000821:virology; D006801:Humans; D009693:Nucleic Acid Hybridization / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
D020121:5' Untranslated Regions; D018076:DNA, Complementary; D004279:DNA, Viral