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Updated: Jun 10, 2026

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
Real-time nucleic acid sequence-based amplification (NASBA) using an adenine-induced quenching probe and an
Y Kouguchi1, M Teramoto, M Kuramoto
1Fukuyama City Public Health Center, Fukuyama, Hiroshima, Japan. shiken-kensa@city.fukuyama.hiroshima.jp
Aims:
We found that an adenine base caused fluorescence quenching of a fluorescein (FL)-labelled probe in DNA:RNA hybrid sequences, and applied this finding to a nucleic acid sequence-based amplification (NASBA) method.
Methods And Results:
The present NASBA method employed a probe containing an FL-modified thymine at its 3' end and ethidium bromide (EtBr) on the basis of a combination of adenine-induced quenching and fluorescence resonance energy transfer (FRET) between the FL donor and EtBr acceptor. This NASBA was used to detect Shiga toxin (STX) stx-specific mRNA in STX-producing Escherichia coli, demonstrating rapid quantification of the target gene with high sensitivity.
Conclusion:
Although the inherent quenching effect of adenine was inferior to that of guanine, FRET between the FL and EtBr moieties enhanced the adenine-induced quenching, allowing rapid and sensitive real-time NASBA detection.
Significance And Impact Of The Study:
This study gives a novel real-time diagnostic system based on NASBA for a sensitive mRNA (or viral RNA) detection.
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