Graphene oxide assisting the visual detection of Salmonella by CRISPR/ Cas12a

Rapid and accurate detection of Salmonella is extremely important to ensure food safety. Combining nucleic acid amplification with CRISPR/Cas12a can realize sensitive and specific detection of foodborne pathogens. However, present CRISPR/Cas-based methods mainly depend on fluorophore and quencher dual-labeled ssDNA as the reporter, which brings in high cost and steric hindrance. Herein, a new signal output format has been developed. The method takes advantage of graphene oxide adsorbing FAM-ssDNA and quenching the fluorescent emission while CRISPR/Cas12a recovering the fluorescent emission by trans-cleaving the FAM-ssDNA. Factors affecting the performance of the sensor have been optimized, and it is shown that the oligonucleotide of 12 nt with the fluorophore modified at the 3 ' end displays the highest signal to noise ratio. The signal output method has been combined with recombinase polymerase amplification for detection of Salmonella and presents consistent specificity and sensitivity (5 x 101 copies) as real-time PCR. By replacing the dual-labeled ssDNA reporter with FAM-ssDNA and GO, this signal output method saves similar to 65% cost. Notably, this method does not require a real-time thermal cycler or any other sophisticated instrument. Besides pathogens, this easy and convenient method can also be extended to detect other targets.