Terminal-Optimized Aptamer-Guide-DNAzyme Triplex Boosts Clostridium butyricum Argonaute-Driven Multiplex Foodborne...

科学分野:

• Biotechnology; Food Science; Molecular Biology

背景:

• Multiplexed and sensitive detection of foodborne pathogens is crucial for disease control.; Mesophilic Clostridium butyricum Argonaute (CbAgo) has endonuclease activity for precise DNA-directed cleavage, useful for pathogen detection.; CbAgo's activity constraints can be overcome by engineered guide DNA (gDNA) and synergistic enzymatic mechanisms.

研究 の 目的:

• To engineer CbAgo's gDNA with a DNAzyme-functionalized fragment (cDNA) to enhance cleavage activity.; To investigate the synergistic integration of DNAzyme and CbAgo activities for improved signal amplification.; To develop a multiplexed biosensor for simultaneous detection of foodborne pathogenic bacteria.

主な方法:

• Engineered gDNA by incorporating a DNAzyme-functionalized fragment (cDNA) for substrate cleavage.; Investigated terminal modification group preferences for optimal DNAzyme-CbAgo interaction.; Developed a DNAzyme-CbAgo synergistic fluorescence aptamer biosensor.

主要な成果:

• Engineered CbAgo with cDNA demonstrated enhanced cooperative cleavage activity.; Synergistic integration of dual enzymatic activities significantly improved signal amplification.; The developed biosensor achieved multiplexed detection of three pathogenic bacteria with a low detection limit (69 CFU mL⁻¹).

結論:

• The DNAzyme-CbAgo synergistic system offers enhanced efficiency for pathogen detection.; The developed aptamer biosensor enables simultaneous detection of multiple foodborne pathogens.; This platform provides a potential method for foodborne pathogen detection without DNA extraction and amplification.