A new class of label-free molecular beacon (MB) system based on DNA strands that contain abasic (AP) sites (AP-DNA) and adopt stem-loop structures, in combination with fluorescent ligands that bind these AP sites, has been developed. Unlike a conventional MB, which requires covalent labeling of the MB with a fluorophore and a quencher, the developed system (APMB) does not require covalent attachment of signal transduction units. Detailed sensing functions of a series of APMB systems were examined with the aid of the fluorescent ligand named ATMND to provide insight into the design strategy for APMB systems. The effects of the stem length and the position of the AP site in the stem moiety on the fluorescence response of the APMB system were examined. Genotyping of a G/C SNP of PCR amplification products was successfully demonstrated with the APMB system and blue-fluorescent ATMND as a ligand. The APMB system was further extended to a system that utilized green-fluorescent lumiflavin.
- abasic sites
- DNA recognition
- fluorescent probes
- molecular beacons
- single-nucleotide polymorphism