A cascade amplification strategy for ultrasensitive Salmonella typhimurium detection based on DNA walker coupling with CRISPR-Cas12a.

Salmonella typhimurium (S. typhimurium) infection is one of leading causes of severe foodborne illness, which poses grievous threats to public health. Thus, the detection with ultra-sensitivity is highly demanded for timely prevention and diagnosis of S. typhimurium. In this study, we developed a novel detection machinery based on DNA walker and CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas12a technologies. Mechanistically, the S. typhimurium specific sequence triggers Nt.AlwI nicking endonuclease and produces particular signaling nucleotide, which further activates Cas12a for strong fluorescence signal output. This cascade amplification strategy exhibits excellent specificity and successfully decreases the limit of detection (LOD) of DNA walker by 2,000 folds to 5 CFU/mL. Collectively, this combinatorial approach offers great promises to effectively reduce foodborne diseases by ultrasensitive detection of S. typhimurium. As a proof of concept, this innovative design also shows prominent potential in detections of other biomolecules, cells and pathogens.