Multianalyte, dipstick-type, nanoparticle-based DNA biosensor for visual genotyping of single-nucleotide...

Area of Science:

• Biotechnology
• Molecular Diagnostics
• Genetics

Background:

• DNA biosensors offer sensitive detection of nucleic acids through molecular recognition and signal transduction.
• Recent advancements include disposable, visual DNA biosensors that eliminate the need for complex instrumentation.
• Single-nucleotide polymorphisms (SNPs) are key genetic markers for disease association studies.

Purpose of the Study:

• To develop and validate a multianalyte DNA biosensor for the visual genotyping of two single-nucleotide polymorphisms (SNPs).
• To enable simultaneous detection of four alleles for two distinct SNPs using a single assay.
• To demonstrate the utility of the biosensor for clinical sample analysis.

Main Methods:

• Utilized a primer extension (PEXT) reaction with four allele-specific primers on PCR-amplified DNA flanking two SNP sites.
• Incorporated biotin-dUTP during primer extension for subsequent capture and detection.
• Employed a disposable, dipstick-type biosensor with immobilized oligonucleotides and antibiotin-functionalized gold nanoparticles for visual readout.

Main Results:

• The DNA biosensor successfully detected the presence or absence of specific alleles through visual color changes (red zones).
• A control zone confirmed proper system performance by capturing excess nanoparticles.
• The assay accurately genotyped two common SNPs in twenty clinical samples from the MBL2 gene.

Conclusions:

• The developed multianalyte DNA biosensor provides a simple, visual, and instrument-free method for SNP genotyping.
• This approach facilitates rapid and simultaneous detection of multiple genetic variations.
• The biosensor shows promise for point-of-care genetic testing and diagnostics.