PMID- 20083192
OWN - NLM
STAT- MEDLINE
DCOM- 20110118
LR  - 20131121
IS  - 1096-1194 (Electronic)
IS  - 0890-8508 (Linking)
VI  - 24
IP  - 4
DP  - 2010 Aug
TI  - PCR with quenching probes enables the rapid detection and identification of 
      ganciclovir-resistance-causing U69 gene mutations in human herpesvirus 6.
PG  - 167-77
LID - 10.1016/j.mcp.2010.01.002 [doi]
AB  - A single-nucleotide polymorphism detection assay using PCR with quenching probes 
      (QP-PCR) was developed for the rapid detection of antiviral drug-resistance 
      mutations of human herpesvirus 6 (HHV-6). The mutations examined were in the 
      HHV-6 U69 gene, and were single-base mutations in sequences known to be 
      associated with ganciclovir (GCV) resistance in HCMV. We previously confirmed 
      that they conferred GCV resistance to recombinant baculoviruses (Nakano et al., 
      J. Virol. Methods 161:223-230, 2009). Six characterized mutations, including a 
      previously reported one that encodes a GCV-sensitive kinase-activity mutant 
      (Isegawa et al., J. Clin. Virol. 44:15-19, 2009), were used. The six mutations 
      were separated into three groups based on their location in the U69 protein, and 
      detected by the hybridization of three probes. We developed and validated a set 
      of assays for these mutations using PCR followed by differential melting of a 
      fluorescently labeled oligo probe, on a Roche Light Cycler platform. Nucleobase 
      quenching was used to detect the hybridized probe. The optimized assay could 
      distinguish the different mutants, and easily detected mutants representing 30% 
      of the DNA in a mixed sample. This QP-PCR assay permitted the rapid (1.5 h), 
      objective, and reproducible detection of drug-resistant mutations of HHV-6.
CI  - Copyright 2010 Elsevier Ltd. All rights reserved.
FAU - Isegawa, Yuji
AU  - Isegawa Y
AD  - Department of Infectious Disease Control, Graduate School of Medicine, Osaka 
      University, Suita, Osaka 565-0871, Japan. isegawa@bact.med.osaka-u.ac.jp
FAU - Matsumoto, Chisa
AU  - Matsumoto C
FAU - Nishinaka, Kazuko
AU  - Nishinaka K
FAU - Nakano, Kazushi
AU  - Nakano K
FAU - Tanaka, Tatsuya
AU  - Tanaka T
FAU - Sugimoto, Nakaba
AU  - Sugimoto N
FAU - Ohshima, Atsushi
AU  - Ohshima A
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20100118
PL  - England
TA  - Mol Cell Probes
JT  - Molecular and cellular probes
JID - 8709751
RN  - 0 (Antiviral Agents)
RN  - 0 (DNA Probes)
RN  - 0 (Mutant Proteins)
RN  - 0 (Nucleotides)
RN  - 0 (Viral Proteins)
RN  - P9G3CKZ4P5 (Ganciclovir)
SB  - IM
MH  - Antiviral Agents/pharmacology
MH  - Child, Preschool
MH  - DNA Probes/*metabolism
MH  - Drug Resistance, Viral/drug effects/*genetics
MH  - Ganciclovir/*pharmacology
MH  - Genes, Viral/*genetics
MH  - Herpesvirus 6, Human/*genetics
MH  - Humans
MH  - Male
MH  - Mutant Proteins/chemistry/genetics
MH  - Mutation/*genetics
MH  - Nucleic Acid Denaturation/drug effects
MH  - Nucleotides/genetics
MH  - Polymerase Chain Reaction/*methods
MH  - Sensitivity and Specificity
MH  - Transition Temperature/drug effects
MH  - Viral Proteins/chemistry/genetics
EDAT- 2010/01/20 06:00
MHDA- 2011/01/19 06:00
CRDT- 2010/01/20 06:00
PHST- 2009/09/30 00:00 [received]
PHST- 2010/01/06 00:00 [revised]
PHST- 2010/01/06 00:00 [accepted]
PHST- 2010/01/20 06:00 [entrez]
PHST- 2010/01/20 06:00 [pubmed]
PHST- 2011/01/19 06:00 [medline]
AID - S0890-8508(10)00003-4 [pii]
AID - 10.1016/j.mcp.2010.01.002 [doi]
PST - ppublish
SO  - Mol Cell Probes. 2010 Aug;24(4):167-77. doi: 10.1016/j.mcp.2010.01.002. Epub 2010 
      Jan 18.