Molecular beacons (MBs) have been used as viable fluorescent probes in nucleic acid analysis. Many researchers around the world continue to modify the MBs to suit their needs. As a result, a number of nucleic acid probing systems with close resemblance to the MBs are being reported from time to time. Quencher-free molecular beacons (QF-MBs) are a significant modification of the conventional MB; in QF-MBs the quencher part has been eliminated. Despite the absence of the quencher, the QF-MBs can identify specific target DNA. They can also be used in SNP typing and in real-time PCR analysis for quantification of DNAs. The design, factors behind functioning and applications of different types of QF-MBs and closely related quencher-free nucleic acid probing systems (QF-NAPs) have been described in this tutorial review.
We inserted a fluorene-labeled deoxyuridine derivative, synthesized using Sonogashira coupling, efficiently into the loop region of a DNA hairpin using phosphoramidite chemistry. This molecular beacon, which features no additional fluorescence quencher, discriminates between perfect and one-base-mismatched base pairing by changes in its fluorescence intensity. The discrimination factor is 14.7 for the recognition of a single (A/C) base mismatch.
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