Switch-conversional ratiometric fluorescence biosensor for miRNA detection.

Area of Science:

• Biochemistry
• Molecular Biology
• Biotechnology

Background:

• MicroRNAs (miRNAs) are critical regulators of gene expression and cellular processes.
• Accurate detection of miRNAs is essential for clinical diagnostics and therapeutic monitoring.
• Existing detection methods face challenges in sensitivity and specificity.

Purpose of the Study:

• To design and develop a novel switch-conversional ratiometric fluorescence biosensor (SCRF biosensor) for highly sensitive microRNA detection.
• To utilize amplicon fragments for structure conversion of a DNA switch, enabling ratiometric fluorescence.
• To establish a quantitative and rapid method for miRNA detection with broad application prospects.

Main Methods:

• A structure-convertible DNA switch modified with two fluorophores (Cy3 and Cy5) and a quencher was designed.
• Amplicon fragments (c*) were generated via exponential amplification.
• Hybridization of amplicon fragments to the DNA switch induced structural conversion and fluorescence resonance energy transfer (FRET) between Cy5 and Cy3.

Main Results:

• The SCRF biosensor demonstrated a wide detection range for target miRNAs, from 100 fM to 100 nM.
• An excellent limit of detection (LOD) of 70.9 fM was achieved.
• The ratiometric fluorescence signal allowed for quantitative and rapid miRNA detection.

Conclusions:

• The developed SCRF biosensor offers a highly sensitive and quantitative platform for miRNA detection.
• This biosensor holds significant potential for applications in clinical diagnosis and molecular therapy.
• The switch-conversional mechanism provides a robust approach for fluorescence-based biosensing.