AAT Bioquest

What are the properties of molecular beacons that affect nucleic acid detection?

Posted August 29, 2023


Molecular beacons have several properties that affect nucleic acid detection. MBs are ideal for the detection of SNP variations. Molecular beacons are single stranded oligonucleotide probes that are composed of a stem-and-loop structure. The loop possesses the probe sequence, which lies between two complementary arm sequences that form a hairpin stem. The hairpin structure induces the detection of SNPS with higher specificity than corresponding linear hybridization probes. MBs are designed so that over a wide range of temperatures only the perfectly complementary target hybrids are stable enough to open the stem hybrid. Thus, a wide range of temperatures exist where the perfectly complementary probe-target hybrids elicit a fluorogenic response, while the mismatched MB remain dark. The specificity of the MB also plays a role in the detection of nucleic acids as they are designed to only target specific certain sequences. Thus, it is crucial for the MB to be highly specific to the target nucleic acid of interest. The specificity of MBs has been shown to be more effective than probes that rely on the 5'-endonucleolytic cleavage activity of DNA polymerase. They allow for the detection of a small amount of mutant DNA in the presence of an abundant wild-type DNA. Additionally, MBs have a wide variety of differently colored fluorophores, and multiple targets can be distinguished in the same solution. SNPSs are identified by fluorescent colors generated in sealed amplification tubes. Amplification is done in the presence of two different molecular beacons and each allele is identified by the development of fluorescence of a specific color. On a side note, MBs detect SNP sites with significantly higher specificity than conventional oligonucleotide probes (due to their hairpin structure).  

Additional resources

Hybridization Efficiency of Molecular Beacons Bound to Gold Nanowires: Effect of Surface Coverage and Target Length

Real-Time PCR (qPCR)

Tide Quencher™ 1 acid [TQ1 acid]