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Related Experiment Video

Updated: Jun 8, 2026

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis

Published on: July 8, 2025

Templated chemistry for sequence-specific fluorogenic detection of duplex DNA.

Hao Li¹, Raphael M Franzini, Christopher Bruner

¹Department of Chemistry, Stanford University, Stanford, CA 94305-5080, USA.

Chembiochem : a European Journal of Chemical Biology

|September 23, 2010

Summary

This summary is machine-generated.

We developed a new fluorogenic chemistry method for detecting specific DNA sequences in solution. This technique uses specially designed probes to generate a strong fluorescence signal only when binding to the target DNA, enabling precise detection.

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Area of Science:

Biochemistry
Molecular Biology
Chemical Biology

Background:

Oligonucleotide probes are crucial for DNA detection.
Current methods may require harsh conditions or lack specificity.

Purpose of the Study:

To develop a novel templated fluorogenic chemistry for specific duplex DNA detection in solution.
To achieve sensitive and selective DNA sequence identification under native conditions.

Main Methods:

Design of two modified homopyrimidine oligodeoxynucleotide probes for triple-helix formation.
Utilizing a fluorescein-labeled probe with a quencher and a trigger probe with a triarylphosphine group.
Assessing fluorescence signal generation upon target DNA binding at different pH levels and with mismatched sequences.

Main Results:

Probes generated a strong fluorescence signal (approx. 60-fold increase) within minutes at pH 5.6 upon binding to complementary duplex DNA.
Signal generation was entirely dependent on the presence of the target DNA.
Using pseudoisocytosine enabled efficient reaction at pH 7, and single nucleotide mismatches significantly reduced the reaction rate, indicating high sequence selectivity.

Conclusions:

Templated fluorogenic chemistry offers a novel approach for detecting specific duplex DNA sequences.
This method allows for sensitive and selective DNA detection under native, nondenaturing conditions, particularly for homopurine-homopyrimidine targets.
The high sequence selectivity demonstrated by the probes opens possibilities for various molecular diagnostic applications.