Monitoring the function of molecular beacon type probe at the single molecule level

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DNA site specifically modified with fluorescent molecule were synthesized. The lifetime of a fluorescent molecule in the triplet excited state was measured, which is reflected in the duration of the off time during the blinking of the fluorescence (tOFF). tOFF was determined by fluorescence correlation spectroscopy (FCS). It was demonstrated that the more a fluorescent molecule is exposed to a solvent, the faster its triplet excited state is quenched by molecular oxygen, resulting in a decrease of tOFF. DNA conformational change between a duplex and hairpin structure, function of the molecular beacon type probe, was successfully monitored by the blinking triggered by the changes in the solvent accessibility of the fluorescent molecule.