Molecular beacons are single-stranded oligonucleotide detector probes that form a stem-and-loop structure. The loop portion of the molecule is a probe sequence that is complementary to a predetermined target sequence and the stem is formed by the annealing of complementary arm sequences that are present on either side of the probe sequence.
A fluorophore is covalently attached to the end of one arm and a nonfluorescent quencher is covalently attached to the end of the other arm. In the absence of target, the stem keeps the fluorophore and the quencher in close proximity to each other, preventing fluorescence (contact quenching). However, when molecular beacons bind to their target they undergo a conformational change that restores the fluorescence of the internally quenched fluorophore.
Molecular beacons have three key properties that enable the design of new and powerful diagnostic assays: 1) they only fluoresce when bound to their targets, 2) they can be labeled with a fluorophore of any desired color, and 3) they are so specific that they can easily discriminate single-nucleotide polymorphisms. As a result molecular beacons enable the development of cost-efficient multiplex diagnostic assays.
For more information on molecular beacons go to: www.molecular-beacons.org
A detailed desciption on the design and optimization of molecular beacon real-time PCR assays appeared in : Vet JA and Marras SAE (2005) Design and optimization of molecular beacon real-time polymerase chain reaction assays. Methods Mol Biol 288, 273-290. Click here for pdf