Highly Sensitive and Specific Detection of Mobilized Colistin Resistance Gene by CRISPR-Based Platform.

Mobilized colistin resistance () gene mediated by plasmid can cause the speediness dissemination of colistin-resistant strains, which have given rise to a great threat to the treatment of human infection. Hence, a rapid and accurate diagnosis technology for detecting is essential for the control of resistance gene. Here, a recombinase polymerase amplification (RPA) coupled with CRISPR/Cas12a platform was established for rapid, sensitive, and specific detection of gene. The analytical sensitivity of our assay is 420 fg per reaction in pure -positive isolates, and the threshold of this method in spiked clinical samples was down to 1.6 × 10 ~ 6.2 × 10 CFU/mL (1.6 ~ 6.2 CFU/reaction). Moreover, the RPA-CRISPR/Cas12a system perspicuously demonstrated no cross-reactivity with other resistant genes. The entire experimental process included rapid DNA extraction (15 min), RPA reaction (30 min), CRISPR/Cas12a cleavage (5 min), and fluorescence testing (<10 min), which could be completed within 60 min. In summary, the RPA-CRISPR/Cas12a assay designed here provides a rapid diagnostic way for monitoring in clinic and livestock farm. This study promises a rapid and accurate assay (RPA-CRISPR/Cas12a) for the surveillance of gene, which causes the efficacy loss of colistin in clinical treatments. In addition, the established method is fit for "on-site" surveillance especially.