The construction of COFs functionalized CRISPR electrochemical sensor for ultrasensitive detection of bacteria by hyper-branched rolling circle amplification

Abstract

Rapid, sensitive, and low-cost foodborne pathogens detection is crucial for dealing with foodborne diseases caused by bacteria. In this work, we designed a new CRISPR/Cas9-mediated electrochemical biosensor for ultrasensitive detection of Escherichia coli (E. coli) based on multifunctional covalent organic framework-based nanocomposite (GOx-AuNPs-COF-H2) and hyperbranched rolling circle amplification (HB-RCA) reaction. When specific DNA targets were present in the system, the targets could be recognized and cleaved by CRISPR-Cas9 system, which triggered strand displacement reaction (SDR) and HB-RCA reaction. By introducing GOx-AuNPs-COF-H2 nanocomposite, H2 probe could hybridize with RCA products to bind to the electrode surface. After added glucose solution, GOx could catalyze it in the detection system and thus offered significant current signals for bacteria detection. This approach showed high analytical performance for assay of E. coli with the detection limit (LOD) of 10 CFU/mL. Furthermore, real samples were detected by this approach, demonstrating its promising potential for foodborne pathogens detection in food monitoring and clinical diagnosis.