Hydrogel Digital LAMP with Suppressed Nonspecific Amplification for Rapid Diagnostics of Fungal Disease in Fresh Fruits

Abstract

Fungal disease, mainly caused by Alternaria alternata infection, can generate severe economic losses and health hazards. However, rapid nucleic acid test without nonspecific reaction still remains challenging. Here, we reported the hydrogel digital loop-mediated isothermal amplification (HdLAMP) with suppressed nonspecific amplification for rapid diagnosis of fungi in fresh fruits. The introduction of hydrogel offered a simple platform to achieve absolute quantification. By breaking the 3′end G-C anchor, the nonspecific amplification of primers could be suppressed, while the specific positive reaction in HdLAMP was not affected. This method could be applied for A. alternata detection in 9 min with excellent performances in speed, specificity, reproducibility, sensitivity, and detection limit down to a single copy. Finally, the real diseased jujubes during postharvest storage were successfully diagnosed as an A. alternata infection. HdLAMP promotes the molecular diagnosis of fungal diseases and broadens the application of hydrogels in the agricultural and food industry.

Nucleic Acid Plate Culture: Label-Free and Naked-Eye-Based Digital Loop-Mediated Isothermal Amplification in Hydrogel with Machine Learning

“…, blood cells, pulps, or dietary fiber in the samples) might strongly interfere with the target partition and droplet generation, resulting in an uneven fluorescence background. Furthermore, biological species, such as chlorophyll, porphyrins, proteins, and alkaloids, may also yield self-fluorescence, giving a high background yet a weak positive signal (low S/N). , In the case of the hydrogel-based culture assay, the cross-linking network of hydrogel ensures the perfect isolation of each single bacteria for digital counting . In addition, the visible light scattering signals from MPPs are free of self-fluorescence background interference.…”

“…Because of its simple, facile, visible, and accurate characteristics, the plate culture method has been extensively used by numerous laboratories and individuals. In addition, the hydrogel platform has also been developed for digital nucleic acid analysis based on fluorescence counting since the movement of nucleic acid is limited in hydrogel. , …”

Focusing on New Discoveries in Food Technology and Creating a New Future of Nutrition and Health: An Introduction to the 4th International Symposium of Food Science, Nutrition and Health in Dalian, China

“…This work revealed that the constructed aptasensor provided a rapid, accurate, and sensitive method for the analysis of OTA in food samples. Dr. Xingyu Lin discussed hydrogel digital loop-mediated isothermal amplification (HdLAMP), specifically designed for the rapid diagnosis of fungi in fresh fruits . This method showcased a reduced nonspecific amplification, enhancing accuracy and efficiency.…”