Multicyclic reverse transcription-polymerase chain reaction assay system for quantification of GB virus-C/hepatitis G virus RNA in serum.
A new quantitative reverse transcription-polymerase chain reaction (RT-PCR) method is described for analyzing the amount of GB virus-C (GBV-C)/hepatitis G virus (HGV) RNA in serum. This multicyclic RT-PCR (MRT-PCR) method used oligonucleotide primers deduced from the 3' noncoding region (3'NCR) that is highly conserved among GBV-C/HGV isolates. Quantitation of GBV-C/HGV RNA using MRT-PCR ranged between 10(2) and 10(10) copies/ml when PCR cycle number was regulated at exponential amplification of the products. Competitive RT-PCR (CRT-PCR) was carried out with mutant RNA and sample that had been measured by MRT-PCR. Quantitation of GBV-C/HGV RNA using both methods agreed. MRT-PCR detected viral RNA in a single step PCR, and demonstrated a high degree of sensitivity that was equal to that of the RT-PCR procedure, which used nested primers deduced from the non-structural (NS) 3 region. The MRT-PCR method for quantitation of GBV-C/HGV RNA in serum may prove useful for diagnosis.
Saito
T
T
Basic Research Division, BioMedical Laboratories, Saitama, Japan. saitt@alk.co.jp
Matsumoto
S
S
Nojiri
O
O
Kageyama
T
T
Fukushi
S
S
Ishiyama
N
N
Kurihara
C
C
Katayama
K
K
D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D018067:Flaviviridae / Q000235:genetics / Q000302:isolation & purification*; D006525:Hepatitis, Viral, Human / Q000821:virology*; D006801:Humans; D015151:Immunoblotting; D012367:RNA, Viral / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014766:Viremia / Q000821:virology
D017931:DNA Primers; D012367:RNA, Viral