Flavin Binding Allosteric Aptamer with Noncovalent Labeling for miR Sensing.

Area of Science:

• Biotechnology
• Molecular Biology
• Analytical Chemistry

Background:

• Label-free detection methods are crucial for simplifying biological assays.
• Modular allosteric aptamers offer a versatile platform for molecular recognition and signal transduction.
• Existing methods often require complex target labeling, limiting their efficiency.

Purpose of the Study:

• To design and characterize a novel aptamer scaffold for label-free detection of microRNA Let-7i.
• To develop a system where target binding triggers a measurable signal via conformational change.
• To evaluate the sensitivity, selectivity, and linear range of the developed aptasensor.

Main Methods:

• Design of a modular allosteric aptamer with distinct recognition and signaling domains.
• Incorporation of a hairpin loop structure and a signaling trigger (FAD).
• Assay development for microRNA Let-7i detection based on FAD release upon target hybridization.

Main Results:

• The aptamer scaffold successfully formed a hairpin loop in the presence of FAD.
• Selective hybridization with Let-7i induced a conformational shift, releasing FAD and generating a signal.
• Achieved sensitive detection of Let-7i with a wide linear range (0.1 pM to 1 μM) and a low detection limit (150 fM).
• Demonstrated high selectivity, discriminating between sequences with 1- and 2-nucleotide differences.

Conclusions:

• The developed modular allosteric aptamer provides an efficient and label-free method for microRNA detection.
• This strategy offers a sensitive, selective, and robust platform for biological sensing applications.
• The aptasensor's ability to discriminate closely related sequences highlights its potential for complex biological samples.