Detection of hepatitis C virus by single-step hairpin primer RT-PCR.
We developed a novel single-step virus detection system using the fluorescent molecule with a hairpin primer on the reverse transcription (RT)-polymerase chain reaction (PCR). Primers that are specific to Hepatitis C virus (HCV) having a hairpin tag at the 5'-end were used for RT and PCR. The HCV-RNA template was reverse transcribed by reverse transcriptase with the hairpin primer (HP), and the resulting cDNA was amplified directly by HP-PCR (RT-HP-PCR). Using the RT-HP-PCR, we succeeded in demonstrating the detection of HCV-RNA in single-step. The RT-HP-PCR selectively detected HCV-RNA extracted from patients' sera containing contaminant materials. The method could be applicable to the quantitative detection of HCV.
The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan.
Fumie
F
Takei
Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Hideki
H
Tani
Research Institute for Microbial Diseases, Osaka University, 3-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Yoshiharu
Y
Matsuura
The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka 567-0047, Japan. Electronic address: nakatani@sanken.osaka-u.ac.jp.
Kazuhiko
K
Nakatani
D005453:Fluorescence; D016174:Hepacivirus / Q000235:genetics*; D006801:Humans; D012367:RNA, Viral / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
D012367:RNA, Viral
Detection method; Fluorescent molecule; HCV; Hairpin primer; PCR