Accurate MRSA identification through dual-functional aptamer and CRISPR-Cas12a assisted rolling circle amplification

Infectious diseases have become one of the most threatening global challenge with high morbidity and mortality, bringing great difficulties to clinical diagnosis and treatment. New strategy for high-specific and sensitive bacteria detection are urgently needed in facing the crisis of worldwide antibiotic resistance. Herein, a novel method through the integration of dual aptamer technology and CRISPR-Cas12a assisted rolling circle amplification (RCA) was present to obtain both accurate identification and high-sensitive detection of Methicillin-Resistant Staphylococcus Aureus (MRSA). The specificity inherited from the dual functionalized aptamers initiated bioconjugation to specifically recognize the protein targets on the surface of bacteria. Besides the target activity, the functionalized aptamer could also convert the protein recognition to nucleic acids signals. Through the integration of attached RCA and CRISPR-Cas12a assisted trans-cleavage, dual amplification of the nucleic acid signal was obtained. Based on this, we have extended the application of CRISPR-Cas12a from the nucleic acid detection to bacteria detection. As a result, the proposed method was demonstrated to be with significantly improved sensitivity towards MRSA detection. We believe that the novel integrated strategy would diversify the existing pool of bacterial detection and inspire the development of promising drug candidates in the future.