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G-四链体二聚体/核酸外切酶Ⅰ辅助信号放大纸芯片检测黄曲霉毒素B1OA北大核心CSTPCD

G-Quadruplex Dimer/ExonucleaseⅠAssisted Signal Amplification Strategy for Rapid Determination of Aflatoxin B1 Using a Paper Chip

中文摘要英文摘要

本研究利用DNA四面体(TDN)精准调控识别配体的取向和密度,结合核酸外切酶Ⅰ(ExoⅠ)对单链DNA的特异性切割活性,借助G-四链体(G4)二聚体对硫黄素T(ThT)的荧光增强作用,构建了旋转型纸芯片,用于快速高灵敏检测黄曲霉毒素B1(AFB1).以含有G4二聚体与AFB1识别序列的单链DNA(G4二聚体探针)为识别探针,其中AFB1识别序列与TDN的延伸手臂链通过碱基互补配对形式结合.当AFB1不存在时,G4二聚体探针上的G4二聚体与ThT结合,体系呈现出较强的荧光信号;当AFB1存在时,AFB1与G4二聚体探针结合,使其脱离TDN.此时,ExoⅠ切割脱离的G4二聚体探针,释放出AFB1,AFB1能够再次与TDN上的G4二聚体探针结合,如此循环,使得TDN上的G4二聚体探针减少,导致体系剩余的G4二聚体减少,荧光强度降低.此旋转型纸芯片在AFB1浓度为0.0001~500 ng/mL范围内具有良好的线性关系,检出限为0.1 pg/mL.将本方法用于花生和红酒样品中AFB1的检测,回收率为85.2%~118.2%.本研究开发的的TDN/G4二聚体/ExoⅠ检测策略显著增强了体系的特异性和灵敏度.

In this work,a tetrahedral DNA nanostructure(TDN)functionalized rotational paper-based analytical device(RPAD)was constructed for rapid and highly sensitive detection of aflatoxin B1(AFB1)using exonucleaseⅠ(ExoⅠ)and G-quadruplex(G4)dimer.Herein,a single-stranded DNA,containing both of the G4 dimer sequence and AFB1 recognition sequence,was used as the recognition probe(G4 dimer probe).TDN was used to precisely regulate the orientation and distribution density of G4 dimer probe to improve the recognition efficiency of the system.ExoⅠas a single stranded DNA specific nuclease was introduced for effective amplification of the detection signal.G4 dimer was employed to enhance the fluorescence signal of thioflavin T(ThT).In the absence of AFB1,the G4 dimer structure of G4 dimer probe could specifically bind with ThT to generate dramatic fluorescence enhancement.However,in the presence of AFB1,AFB1 could specifically bind with G4 dimer probe,resulting in the dissociation of G4 dimer probe from TDN and further be digested by ExoⅠ.At the same time,the released AFB1 could bind to G4 dimer probe on the TDN again by this way to generate signal amplification.After this cycle,the amount of aptamer on the TDN was decreased,accompanied by the reduction of G4 dimer on TDN.In this case,the fluorescence intensity of the system was reduced.The designed RPAD showed a good linear response in AFB1 concentration range of 0.0001-500 ng/mL and the limit of detection was 0.1 pg/mL.Moreover,the proposed strategy was successfully applied to detection of AFB1 in peanut and wine.The developed TDN/G4 dimer/ExoⅠstrategy improved the specificity and sensitivity of the system significantly.

贺璇;齐骥;余子辉;陈燕;付秀丽

烟台大学化学化工学院,烟台 264005中国科学院烟台海岸带研究所,海岸带环境工程技术研究与发展中心,中国科学院海岸带环境过程与生态修复重点实验室,烟台 264003烟台大学物理与电子信息学院,烟台 264005山东农业工程学院资源与环境工程学院,济南 250100

旋转型纸芯片核酸外切酶ⅠG-四链体二聚体DNA四面体黄曲霉毒素B1

Rotational paper-based analytical deviceExonucleaseⅠG-quadruplex dimerTetrahedral DNA nanostructureAflatoxin B1

《分析化学》 2024 (008)

1094-1102,中插1-中插5 / 14

山东省高等学校青创科技支持计划项目(No.2019KJF029)和国家自然科学基金项目(Nos.61801274,22106179)资助. Supported by the Qingchuang Science and Technology Program of Shandong Province,China(No.2019KJF029)and the National Natural Science Foundation of China(Nos.61801274,22106179).

10.19756/j.issn.0253-3820.231149

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