Ratiometric fluorescence sensor based on carbon dots as internal reference signal and T7 exonuclease-assisted signal amplification strategy for microRNA-21 detection
Ratiometric fluorescence sensor based on carbon dots as internal reference signal and T7 exonuclease-assisted signal amplification strategy for microRNA-21 detection
复制标题
基于碳点作为内参信号和T7核酸外切酶辅助信号放大策略的比率荧光传感器用于microRNA-21检测
DOI：
10.1016/j.aca.2019.12.068
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发表时间：
2020-03-22
影响因子：
6.2
通讯作者：
Weng, Shaohuang
中科院分区：
文献类型：
--
作者：
Wang, Zhenzhen;Xue, Zhiqiang;Weng, Shaohuang
miRNA-21的表达水平与肿瘤的发生、发展密切相关，尤其是在胃肠道肿瘤中。监测miRNA-21在胃肠道肿瘤的诊断和评估中具有临床应用价值。以碳量子点（CD）和FAM标记的ssDNA为信号源，建立了一种基于T7核酸外切酶介导的循环酶促扩增法的miRNA-21开启比率荧光生物检测方法。CD展示了在该策略中内置的内部荧光、探针载体和猝灭剂的三重功能。在不存在miRNA-21的情况下，FAM标记的ssDNA将被CD吸附并淬灭。miRNA-21的加入诱导了T7核酸外切酶从50端开始的环状水解，然后释放出短切的FAM标记的寡核苷酸。然后，通过比率测定法检测增加的FAM信号（F-FAM）和稳定的CD信号（F-CD），以定量miRNA-21。F-FAM/F-CD值与miRNA-21浓度在0.05-10 nM范围内呈良好的线性关系，检测限为1 pM，具有良好的选择性和重现性。此外，该传感器成功评估了健康个体和胃肠道肿瘤患者临床血液样本中miRNA-21的表达水平，结果与qRT-PCR高度一致，提示在与miRNA-21表达水平相关的癌症诊断中具有较大的临床应用价值。(c)2020 Elsevier B. V.保留所有权利。
The expression level of miRNA-21 is closely related to the occurrence and development of cancer, especially in gastrointestinal cancer. Monitoring miRNA-21 has clinical application in the diagnosis and evaluation of gastrointestinal cancer. A turn-on ratiometric fluorescence bioassay based on the T7 exonuclease-mediated cyclic enzymatic amplification method was developed for miRNA-21 determination by using carbon dots (CDs) and FAM-labeled ssDNA as the signal source. CDs demonstrated the triple functions of built-in internal fluorescence, probe carrier, and quencher in this strategy. In the absence of miRNA-21, FAM-labeled ssDNA would be adsorbed and quenched by CDs. The addition of miRNA-21 induced cycle hydrolysis from the 50 end by the T7 exonuclease and then released the short-cleaved FAM-labeled oligonucleotides. Then, the increased FAM signal (F-FAM) and the stable CD signal (F-CDs) would be tested through a ratiometric routine for the quantification of miRNA-21. The F-FAM/F-CDs value showed a good linear relationship with the concentration of miRNA-21 in the range of 0.05-10 nM, and the detection limit for miRNA-21 was 1 pM with excellent selectivity and reproducibility. Furthermore, this sensor successfully evaluated the expression level of miRNA-21 in clinical blood samples from healthy individuals and gastrointestinal cancer patients, and the results were highly consistent with those of qRT-PCR, suggesting the great clinical application value in the diagnosis of cancer associated with miRNA-21 expression levels. (c) 2020 Elsevier B.V. All rights reserved.