A label-free, G-quadruplex DNAzyme-based fluorescent probe for signal-amplified DNA detection and turn-on assay of endonuclease

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• 作者：Zhixue Zhou^a ; ^b ; Yan Du^a ; ^b ; Libing Zhang^a ; ^b ; Shaojun Dong^a ; ^b ; ^{dongsj@ciac.jl.cn}
• 关键词：DNAzyme ; Molecular beacon ; Exonuclease III ; DNA detection ; Endonuclease
• 刊名：Biosensors and Bioelectronics
• 出版年：2012
• 期刊代码：12_09565663
• 类别：ch
• 出版时间：15 April, 2012
• 卷：34
• 期：1
• 页码：100-105
• 文件大小：844 K

摘要

A novel G-quadruplex DNAzyme molecular beacon (G-DNAzymeMB) strategy is developed for assays of target DNA and restriction endonuclease. The detection system consists of G-DNAzymeMB strand and a blocker DNA by using the fluorescence of 2鈥?7鈥?dichlorodihydrofluorescein diacetate (H₂DCFDA) catalyzed by G-DNAzymeMB as a signal reporter. G-DNAzymeMB exhibits peroxidase activity in its free hairpin structure, and forms a catalytically inactive hybrid when hybridized with blocker DNA. Upon displacement of blocker DNA by target DNA or cleavage by restriction endonuclease, G-DNAzymeMB is released and two lateral portions of G-DNAzymeMB form a G-quadruplex structure, resulting in the recovery of catalytic activity which acts as a cofactor to catalyze H₂O₂-mediated oxidation of H₂DCFDA. For DNA detection system, exonuclease III (Exo III)-catalyzed amplification strategy is introduced to improve the sensitivity and target DNA could be detected as low as 0.1 pM. With respect to restriction endonuclease detection system, 0.1 U/mL EcoRI endonuclease could be detected and this method could be easily transported to other restriction endonuclease analysis by simply changing the recognition sequence. These results demonstrate that the proposed G-DNAzymeMB strategy could be used as a label-free, simple, sensitive and cost-effective approach in analysis of target DNA and restriction endonuclease.