Genetically Encoded Fluorogenic DNA Aptamers for Imaging Metabolite in Living Cells.

Area of Science:

• Biotechnology
• Molecular Biology
• Cellular Imaging

Background:

• Genetically encoded fluorescent proteins and RNA sensors are vital for cellular biomolecule imaging.
• Existing sensors face limitations in generalizability and stability.
• Expanding the toolkit for molecular imaging is crucial for biological research.

Purpose of the Study:

• To develop a novel genetically encoded fluorogenic DNA aptamer (GEFDA) sensor.
• To improve the stability and generalizability of genetically encoded sensors.
• To enable real-time monitoring of small molecules, specifically adenosine triphosphate (ATP), in live cells.

Main Methods:

• A fluorogenic DNA aptamer for dimethylindole red was linked to an ATP aptamer.
• The combined aptamer construct was integrated into a plasmid for genetic encoding.
• The GEFDA sensor was tested for ATP detection in live bacterial and mammalian cells.

Main Results:

• The GEFDA sensor demonstrated a 4-fold enhancement in red fluorescence at 650 nm in the presence of ATP.
• Dimerization of the sensor improved the signal-to-noise ratio by 2-3 fold.
• Successful implementation of the GEFDA sensor for ATP detection in both bacterial and mammalian cells was achieved.

Conclusions:

• Genetically encoded fluorogenic DNA aptamer sensors offer enhanced stability compared to protein and RNA-based sensors.
• GEFDA sensors represent a novel tool for real-time monitoring of small molecular metabolites.
• This approach expands the capabilities of genetically encoded biosensors for diverse biological applications.