DNA-Templated Click Ligation Chain Reaction Catalyzed by Heterogeneous Cu<sub>2</sub>O for Enzyme-Free Amplification and Ultrasensitive Detection of Nucleic Acids
Fan Wang, Chenglong Bao, Susu Cui, Jinlong Fan, Zijie Zhang, Weiwei Yang, Yongsheng Yu, Yingfu Li
Abstract
Nucleic acids play a pivotal role in the diagnosis of diseases. However, rapid, cost-efficient, and ultrasensitive identification of nucleic acid targets still represents a significant challenge. Herein, we describe an enzyme-free DNA amplification method capable of achieving accurate and ultrasensitive nucleic acid detection via D NA- t emplated c lick l igation c hain r eaction (DT-CLCR) catalyzed by a h eterogeneous n anocatalyst made of Cu 2 O (hnCu 2 O). This hnCu 2 O–DT-CLCR method is built on two cross-amplifying hnCu 2 O-catalyzed DNA-templated azide–alkyne cycloaddition-driven DNA ligation reactions that boast a fast reaction rate and a high DNA ligation yield in minutes, enabling rapid exponential amplification of specific DNA targets. This newly developed hnCu 2 O–DT-CLCR-enabled DNA amplification strategy is further integrated with two signal reporting mechanisms to achieve low-cost and easy-to-use biosensors: an electrochemical sensor through the conjugation of a methylene blue redox reporter to a DNA probe used in hnCu 2 O–DT-CLCR and a colorimetric sensor through the incorporation of the split-to-intact G-quadruplex DNAzyme encoded into hnCu 2 O–DT-CLCR. Both sensors are able to achieve specific detection of the intended DNA target with a limit of detection at aM ranges, even when challenged in complex biological matrices. The combined hnCu 2 O–DT-CLCR and sensing strategies offer attractive universal platforms for enzyme-free and yet efficient detection of specific nucleic acid targets.