Aligner-mediated cleavage-triggered exponential amplification for sensitive detection of nucleic acids

Area of Science:

• Molecular Biology
• Biotechnology
• Analytical Chemistry

Background:

• Exponential amplification reaction (EXPAR) offers sensitive nucleic acid detection but faces challenges in trigger DNA generation.
• Current methods for trigger DNA require specific endonucleases and have limitations in universality and reaction condition optimization.

Purpose of the Study:

• To develop a simple, universal, and highly sensitive method for nucleic acid detection.
• To overcome the limitations of traditional EXPAR by improving trigger DNA generation.

Main Methods:

• Introduced aligner-mediated cleavage (AMC) using a single nicking endonuclease for programmable DNA cleavage.
• Established aligner-mediated cleavage-triggered exponential amplification (AMCEA) for nucleic acid detection.
• Validated AMCEA's performance in human serum samples.

Main Results:

• AMCEA demonstrated high sensitivity with a detection limit at the amol level.
• The method exhibited a broad dynamic range (5–6 orders of magnitude) and specificity, distinguishing single nucleotide mutations.
• AMCEA proved compatible with complex biological samples like human serum.

Conclusions:

• AMCEA offers a universal and simple platform for nucleic acid detection, overcoming EXPAR's limitations.
• The programmable cleavage via AMC enhances flexibility and broadens applicability.
• AMCEA shows significant potential for practical applications in biological sample analysis.