Label-Free Sequence-Specific Visualization of LAMP Amplified Salmonella via DNA Machine Produces G-Quadruplex DNAzyme

Salmonella is one of four key global causes of diarrhea, and in humans, it is generally contracted through the consumption of contaminated food. It is necessary to develop an accurate, simple, and rapid method to monitor Salmonella in the early phase. Herein, we developed a sequence-specific visualization method based on loop-mediated isothermal amplification (LAMP) for the detection of Salmonella in milk. With restriction endonuclease and nicking endonuclease, amplicons were produced into single-stranded triggers, which further promoted the generation of a G-quadruplex by a DNA machine. The G-quadruplex DNAzyme possesses peroxidase-like activity and catalyzes the color development of 2,2'-azino-di-(3-ethylbenzthiazoline sulfonic acid) (ABTS) as the readouts. The feasibility for real samples analysis was also confirmed with Salmonella spiked milk, and the sensitivity was 800 CFU/mL when observed with the naked eye. Using this method, the detection of Salmonella in milk can be completed within 1.5 h. Without the involvement of any sophisticated instrument, this specific colorimetric method can be a useful tool in resource-limited areas.

Automated summary

A sequence-specific visualization method based on loop-mediated isothermal amplification (LAMP) was developed for the detection of Salmonella in milk. The method used restriction endonuclease and nicking endonuclease to produce amplicons into single-stranded triggers, which further promoted the generation of a G-quadruplex by a DNA machine. The G-quadruplex DNAzyme possessed peroxidase-like activity and catalyzed the color development of ABTS as the readouts. The method demonstrated a sensitivity of 800 CFU/mL when observed with the naked eye and could detect Salmonella in milk within 1.5 h without the need for any sophisticated instrument.