Dual Mode Analysis of Methicillin-Resistant <i>Staphylococcus aureus</i> by the CRISPR/Cas12a-Assisted Exonuclease-Mediated Signal Cycle

Methicillin-resistant <i>Staphylococcus aureus</i> (MRSA) is the main Gram-positive bacteria isolated from patients with ocular infections and remians a huge threat to public health. Therefore, sensitive and dual-mode analysis of MRSA is of great significance for evaluating MRSA infection. We depicted a clustered regularly interspaced short palindromic repeats (CRISPR)/Cas12a-based biosensor by integrating the exonuclease III (Exo III)-enhanced fluorescence and colorimetric signals for sensitive and dual-mode analysis of methicillin resistance in MRSA without needing nucleic acid amplification. In this method, Exo III-assisted signal recycling is triggered only when the target <i>mecA</i> gene activates the CRISPR/Cas12a complex, and the HP probes on the surface of magnetic nanoparticles are cleaved by the trans-cleavage activity of Cas12a. Taking the merits of the trans-cleavage activity of the CRISPR/Cas12a system and the Exo III-assisted signal recycling, this approach exhibits an exceptionally elevated detection threshold for the <i>mecA</i> gene. Besides MRSA detection, this accurate and sensitive sensor can be employed to assess additional biomarkers in disease diagnosis by simply changing the crRNA.