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Molecular Beacons are a type dual-labeled FRET probes incorporating a quencher and a fluorophore reporter molecule. They differ from traditional dual-labeled probes, due to the incorporation of a short (~5 b.p.) complementary stem sequence at the 3' and 5' ends.
These complementary sequences hybridize to form a “stem-loop” structure which holds the reporter and quencher close together in space. This alternate conformation makes Molecular Beacons more discriminatory, allowing them to resolve different target sequences differing by a single nucleotide. Molecular Beacons also differ from traditional dual-labeled probes because they do not rely on probe hydrolysis from the 5' exonuclease activity of Taq polymerase to generate its fluorescence, therefore, a post-PCR melt curve is possible.
Just like standard Dual-labeled BHQ® probes it is possible to multiplex assays using Molecular Beacons. Multiplexing allows for the detection of several targets simultaneously using multiple spectrally resolved fluorescent probes.