Ultrafast DNA detection based on turn-back loop primer-accelerated LAMP (TLAMP).

Area of Science:

• Molecular Biology
• Isothermal Amplification Techniques
• Pathogen Detection

Background:

• Rapid DNA detection is crucial for combating infectious diseases.
• Loop-mediated isothermal amplification (LAMP) is a widely used DNA detection method.
• Improving the speed and efficiency of LAMP is an ongoing research goal.

Purpose of the Study:

• To develop a novel LAMP variant with a superior amplification rate.
• To enhance DNA detection speed and efficiency for infectious disease diagnosis.
• To introduce turn-back loop primers (TLPs) for improved LAMP performance.

Main Methods:

• Development of a new LAMP variant named TLAMP.
• Design of bifunctional turn-back loop primers (TLPs) extending original loop primers.
• Utilizing TLPs to create multiple amplification pathways and structures.
• Testing TLAMP performance against conventional LAMP and other fast LAMP variants.
• Application of TLAMP for Monkeypox virus (MPXV) detection.

Main Results:

• TLAMP demonstrated a significantly reduced amplification-to-result time (within 20 min) compared to conventional LAMP (1 hour).
• TLAMP was approximately 10 minutes faster than existing fast LAMP variants, with comparable sensitivity and higher repeatability.
• The method successfully detected as few as 10 copies of MPXV pseudovirus within 20 min (real-time fluorescence) or 30 min (colorimetric assay).

Conclusions:

• TLAMP offers a sensitive, specific, reliable, and ultrafast DNA detection method.
• The novel TLAMP strategy significantly enhances amplification efficiency and speed.
• TLAMP presents a promising tool for rapid diagnosis of infectious diseases, addressing urgent public health challenges.