TaqMan probes based on oligonucleotide-hairpin minor groove ligand conjugates

Hybridization of TaqMan probes derived from oligonucleotides containing fluorophores (fluorescein, FAM, or tetramethylrhodamine, (Tamra)), fluorescence quenchers (BHQ1 or BHQ2), and a conjugated hairpin ligand (MGB) composed of two tripyrrolcarboxamide residues connected through an aminobutyric acid...

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Bibliographic Details
Published inBioorganicheskaia khimiia Vol. 33; no. 6; p. 661
Main Authors Kostina, E V, Riabinin, V A, Maksakova, G A, Siniakov, A N
Format Journal Article
LanguageRussian
Published Russia (Federation) 01.11.2007
Subjects
DNA - analysis
DNA - genetics
DNA Mutational Analysis - methods
DNA Probes - chemistry
Fluorescein - chemistry
Hepacivirus - genetics
Ligands
Nucleic Acid Conformation
Nucleic Acid Hybridization
Oligonucleotides - chemistry
Point Mutation
Polymerase Chain Reaction - methods
Rhodamines - chemistry
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Summary:Hybridization of TaqMan probes derived from oligonucleotides containing fluorophores (fluorescein, FAM, or tetramethylrhodamine, (Tamra)), fluorescence quenchers (BHQ1 or BHQ2), and a conjugated hairpin ligand (MGB) composed of two tripyrrolcarboxamide residues connected through an aminobutyric acid residue were proposed for discrimination of point mutations using the real time PCR technique. Identification of point A/C substitution was shown to be highly specific for hepatitis C virus subtypes 1a and 1b with two variants of the probe (5'-3'): ATTGAGCGGGTTTAp-BHQ2-MGB for subtype 1a and FAM-ATTGAGCGGGTTGAp-BHQ1-MGB for subtype 1b. Perfect duplexes (A.T and G.C pairs) increase fluorescence in the process of amplification, whereas imperfect duplexes (A.G and T.C pairs) induce no fluorescence changes. This phenomenon enables simultaneous genotyping of hepatitis C virus subtypes 1a and 1b.
ISSN:0132-3423