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Detection of hepatitis C virus genome in human serum by multi‐targeted polymerase chain reaction
Author(s) -
Okamoto M.,
Baba M.,
Kodama E.,
Sekine K.,
Takagi T.,
Kasukawa R.,
Shigeta S.
Publication year - 1993
Publication title -
journal of medical virology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.782
H-Index - 121
eISSN - 1096-9071
pISSN - 0146-6615
DOI - 10.1002/jmv.1890410103
Subject(s) - ns3 , primer (cosmetics) , hepatitis c virus , virology , reverse transcriptase , biology , polymerase chain reaction , microbiology and biotechnology , nested polymerase chain reaction , rna , virus , complementary dna , serial dilution , reverse transcription polymerase chain reaction , hepatitis b virus dna polymerase , polymerase , dna , gene , chemistry , messenger rna , genetics , medicine , organic chemistry , pathology , alternative medicine
A multi‐targeted “hemi‐nested” PCR (M‐PCR) assay in which the primer pairs derived from the 5′ non‐coding (5′NC) and the nonstructural protein 3 (NS3) regions of HCV genome were concurrently used for amplification in order to compare the sensitivity and specificity of polymerase chain reaction (PCR) with different primer pairs in detecting hepatitis C virus (HCV) genome. Sera from patients with virus‐associated liver diseases were examined for the presence of HCV RNA by the M‐PCR method following reverse transcription to cDNA. The amplified products derived from both the 5′NC and the NS3 regions were detected in 28 (70%) of the 40 HCV RNA‐positive samples. However, 12 samples (30%) were devoid of the signal of NS3‐derived product. Sensitivity tests using serial dilutions of HCV RNA revealed that the 5′NC‐derived band was still detectable in the 10 5 ‐fold diluted sample by the M‐PCR method, yet the NS3‐derived band could hardly be detected in the 10 4 ‐fold diluted sample. Thus, as previously demonstrated by a single‐targeted “nested” PCR assay, the present study using the M‐PCR assay has clearly shown that the 5′NC‐derived primers are more sensitive and specific than the NS3‐derived primers in detecting HCV RNA.