thrombin-aptamer and 6-carboxyfluorescein

A new target-responsive DNA strand displacement system via toehold-mediated click chemical ligation was designed and prepared for enzyme-free fluorescent-amplified aptasensors. The aptasensors significantly amplified fluorescent signals in response to targets based on target recycling processes.

Topics: Aptamers, Nucleotide; Click Chemistry; DNA; Fluoresceins; Fluorescent Dyes; Nucleic Acid Hybridization

We have developed a technique for the highly selective and sensitive detection of Pb(2+) and Hg(2+) using a thrombin-binding aptamer (TBA) probe labeled with the donor carboxyfluorescein (FAM) and the quencher 4-([4-(dimethylamino)phenyl]azo)benzoic acid (DABCYL) at its 5' and 3' termini, respectively. The TBA has a random coil structure that changes into a G-quartet structure and a hairpin-like structure upon binding Pb(2+) and Hg(2+) ions, respectively. As a result, the fluorescence decreases through fluorescence resonance energy transfer (FRET) between the fluorophore and quencher. These changes in fluorescence intensity allow the selective detection of Pb(2+) and Hg(2+) ions at concentrations as low as 300 pM and 5.0 nM using this TBA probe in the presence of phytic acid and a random DNA/NaCN mixture, respectively. The linear correlation existed between the fluorescence intensity and the concentration of Pb(2+) and Hg(2+) over the range of 0.5-30 nM (R(2) = 0.98) and 10-200 nM (R(2) = 0.98), respectively. To the best of our knowledge, this is the first example of a single DNA-based sensor that allows the detection of both Hg(2+) and Pb(2+) ions. This simple and cost-effective probe was also applied to separately determine Pb(2+) in soil samples and spiked Hg(2+) in pond samples.

Topics: Aptamers, Nucleotide; DNA; Fluoresceins; Fluorescence Resonance Energy Transfer; Ions; Lead; Mercury; Phytic Acid; Spectrometry, Fluorescence

Oligonucleotide derivatives, F-TBA-T and F-7TBA-T carrying thrombin-binding aptamer (TBA) sequence and FAM and TAMRA dyes at their each end were designed to achieve the fluorometric monitoring of potassium ion (K+) concentrations in a living organisms. These oligonucleotides could form a tetraplex structure upon addition of K+. Spectrophotometric experiments revealed that the fluorescence enhancement on the fluorescence resonance energy transfer (FRET) was larger in the case of F-7TBA-T having 7-nucleotides at 5'-end of TBA sequence than in the case of F-TBA-T due to the elimination of the quenching by dye-dye interaction.

Topics: Aptamers, Nucleotide; Cations; Fluoresceins; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Nucleic Acid Conformation; Potassium; Rhodamines