Generi Biotech - Compounds and Services for Molecular Biology
> Real-Time PCR Probes > CATALOGUE AND PRICELIST > Dual Labelled Probes > About Dual Labelled Probes
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About Dual Labelled ProbesTechnology of dual labelled probes (also called hydrolyzation probes or TaqMan® probes) utilizes 5´-3´ exonuclease activity of Taq DNA polymerase to cleave a probe labelled with a pair of fluorophores at its ends, where one fluorophore is fluorescent (reporter) and the other is not (quencher). The probe is designed to becomplementary to a certain target sequence inside the amplified region and has 3` end modified to avoid elongation. As long as the probe is intact, the proximity of the quencher greatly reduces the fluorescence emitted by the reporter dye. Upon hybridization with the target sequence, the probe is cleaved by the 5´-3´exonuclease activity of Taq DNA polymerase during the elongation phase of PCR. This cleavage leads to separation of the fluorophores and enhancement of fluorescent activity of the reporter.
Hydrolyzation probes are usually labelled with the reporter at the end 5´, with the quencher placed at the end 3´. Choice of the reporter depends on excitation and emission channels of the used real-time PCR cycler (almost each real-time PCR cycler provides a channel for fluorescein - FAM). Today, BHQ (Black Hole Quencher) type quenchers are usually used as quenchers; they quench a broad spectrum of fluorophores and show no autofluorescence. GENERI BIOTECH supplies hydrolyzation probes labelled with FAM and HEX reporters in combination with the quencher BHQ1 and Cy3, Cy5 and PULSAR in combination with BHQ2. |
