Modular DNAzymes-Hydrogel Membrane Carriers for Highly Sensitive Isothermal Cross-Cascade Detection of Pathogenic...

The increasing prevalence of antimicrobial resistance has called for improved diagnostic testing of pathogenic bacteria. However, the development of rapid, cost-effective, and easy-to-use tests for bacterial infections remains a constant challenge. Here, we report a class of modular hydrogel membrane carriers incorporated with composite DNAzymes, which enable rapid and highly sensitive detection of pathogenic bacteria gene target analytes. We apply free radical polymerization to incorporate composite DNAzymes, consisting of an RNA substrate component and a DNAzyme component (e.g., 10-23 or 8-17 DNAzymes), into polyethylene glycol diacrylate polymer networks. Initiated by a nucleic acid target acting as an assembly facilitator, multicomponent DNAzymes are combined to cleave the RNA substrate component in the hydrogel carriers, which releases the DNAzyme component to cleave RNA reporter probes to generate fluorescence. We modulate the morphology, composition, and microporous structures of the DNAzyme carriers to achieve quantitative assay performance. We demonstrate a rapid and high-sensitivity detection of C. trachomatis gene target analytes as low as 50 fM in a short assay time of 25 min. The work represents a crucial step forward in the development of a generic, isothermal, and protein enzyme-free pathogenic bacteria testing platform technology.