title: Real-time PCR diagnostics failure caused by nucleotide variability within exon 4 of the human cytomegalovirus major immediate-early gene.
abstract: Here we report how variability in the human cytomegalovirus genome sequence may seriously affect the outcome of its molecular diagnosis. A real-time quantitative PCR assay targeting the major immediate-early gene failed to detect the viral load in some, but not all, clinical samples from hematooncological patients. By amplification and sequencing the DNA across the regions targeted by this assay we found a number of nucleotide substitutions which accounted for decreased primer/probe binding. This decreased the sensitivity of the assay up to 1,000-fold.
authors.affiliation: Center of Molecular Biology and Gene Therapy, Department of Internal Medicine-Hematooncology, University Hospital Brno, Brno, Czech Republic. mlengerova@fnbrno.cz
authors.forename: Martina
authors.initials: M
authors.lastname: Lengerova
authors.forename: Zdenek
authors.initials: Z
authors.lastname: Racil
authors.forename: Pavlina
authors.initials: P
authors.lastname: Volfova
authors.forename: Jana
authors.initials: J
authors.lastname: Lochmanova
authors.forename: Jitka
authors.initials: J
authors.lastname: Berkovcova
authors.forename: Dana
authors.initials: D
authors.lastname: Dvorakova
authors.forename: Jiri
authors.initials: J
authors.lastname: Vorlicek
authors.forename: Jiri
authors.initials: J
authors.lastname: Mayer
mesh_terms: D001483:Base Sequence; D003587:Cytomegalovirus / Q000235:genetics*; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis / Q000235:genetics; D005091:Exons / Q000235:genetics*; D005188:False Negative Reactions; D017781:Genes, Immediate-Early; D014644:Genetic Variation; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers; D004279:DNA, Viral