Abstract

Efficient point-of-care (POC) molecular diagnostic techniques can be designed using highly selective hybridization of G-quadruplex-based DNA-nanomachine (G-4DNM) with specific analytes that produce readable outputs. We propose the design of G-4DNM that can unwind an analyte's secondary structures, solving the problem of nucleic acid analytes' recognition at ambient temperatures. S.aureus is a pathogen responsible for a number of nosocomial infections. For detection, we chose a specific gene that is constitutively expressed. S.aureus was analyzed by G-4DNM equipped with 3 long analyte binding arms to tightly bind and unwind single-stranded analytes. Only when all arms bind the analyte a G-quadruplex (G-4) structure is formed. This structure can form a complex with hemin, which exhibits peroxidase activity. In the presence of the G-4/hemin complex, the chemiluminescence (CL) of luminol molecules, activated by hydrogen peroxide, is enhanced. We evaluated the CL kinetics for several minutes, comparing them with the background signal using a highly sensitive photon counting head. The results indicated that S.aureus was recognized with high selectivity and sensitivity up to attomolar concentrations in a quartz cuvette. Our research provides the basis for rapid and affordable POC diagnostics. In the future, this system may become a full-fledged lab-on-a-chip for the detection of marker-sequenced nucleic acids.

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