基于DNA模板点击化学和催化发夹型DNA自组装反应的新型均相电化学生物传感器检测miRNA-21
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摘要
构建了一种利用DNA模板点击化学和催化发夹型DNA自组装反应(Catalyzed hairpin assembly,CHA)指示并放大信号,检测miRNA-21的新型均相电化学生物传感器。根据目标物miRNA-2的序列设计了两种发夹结构的探针,分别修饰5-氨基-2,3-二氰基-1,4-萘醌(ADNQ)和3(2-呋喃)丙酸(FPA),利用目标物miRNA-2引发的两种探针间的催化发夹组装使ADNQ和FPA的分子间距缩小,发生Diels-Alder反应,在实现目标物循环信号放大的同时,破坏ADNQ的化学结构,导致电化学响应信号降低,据此检测miRNA-21的浓度。采用循环伏安法(CV)、差分脉冲伏安法(DPV)和凝胶电泳等考察了此传感器的分析性能。结果表明,在0.1～1.0×10~4 pmol/L浓度范围内,此传感器响应电流变化值与miRNA-21浓度的对数呈线性关系,检出限为0.037 pmol/L(S/N=3)。将此传感器用于小鼠血清和全血裂解液样品的检测,结果表明,本传感器适用于miRNA-21的检测。
A homogeneous electrochemical biosensor for miRNA-21 detection based on DNA-templated click chemistry and catalytic hairpin assembly was developed. 5-Amino-2,3-dicyano-1,4-naphthoquinone(ADNQ) and 2-furanpropanoic acid(FPA) were linked to two suitable probes of hairpin structure, respectively. In the presence of miRNA-21, the catalytic hairpin assembly between the two probes could be triggered. Therefore, ADNQ and FPA linked to the probes got very close to each other and reacted by a Diels-Alder reaction to change the quinone structure of ADNQ intoto ketone, making it no longer electro-active. Thus, the current difference(ΔI) could be used to indicate the concentration of miRNA-21. The feasibility and detecting performance of the proposed biosensor were also examined by cyclic voltammetry(CV), differential pulse voltammetry(DPV) and polyacrylamide gel electrophoresis(PAGE). The biosensor was successfully applied to sensitively detect miRNA-21 in a linear concentration range from 0.1-1.0×10~4 pmol/L with a detection limit of 0.037 pmol/L(S/N=3). The biosensor was suitable for detection of miRNA-21 in blood samples.
A homogeneous electrochemical biosensor for miRNA-21 detection based on DNA-templated click chemistry and catalytic hairpin assembly was developed. 5-Amino-2,3-dicyano-1,4-naphthoquinone(ADNQ) and 2-furanpropanoic acid(FPA) were linked to two suitable probes of hairpin structure, respectively. In the presence of miRNA-21, the catalytic hairpin assembly between the two probes could be triggered. Therefore, ADNQ and FPA linked to the probes got very close to each other and reacted by a Diels-Alder reaction to change the quinone structure of ADNQ intoto ketone, making it no longer electro-active. Thus, the current difference(ΔI) could be used to indicate the concentration of miRNA-21. The feasibility and detecting performance of the proposed biosensor were also examined by cyclic voltammetry(CV), differential pulse voltammetry(DPV) and polyacrylamide gel electrophoresis(PAGE). The biosensor was successfully applied to sensitively detect miRNA-21 in a linear concentration range from 0.1-1.0×10~4 pmol/L with a detection limit of 0.037 pmol/L(S/N=3). The biosensor was suitable for detection of miRNA-21 in blood samples.
引文
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2 Farazi T A,Spitzer J I,Morozov P.J.Pathol.,2015,223(2):102-115
3 Gasparri M L,Besharat Z M,Besharat A R.Current Knowledge of miRNAs as Biomarkers in Breast Cancer,Recent Trends in Cancer Biology:Spotlight on Signaling Cascades and microRNAs,Springer,Cham.,2018:221-231
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8 Fu S W,Chen L,Man Y G.J.Cancer,2011,2(1):116-122
9 Wang J.Biosens.Bioelectron.,2006,21(10):1887-1892
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