Key Laboratory of Clinical Laboratory Diagnostics (Chinese Ministry of Education), College of Laboratory Medicine, Chongqing Medical Laboratory Microfluidics and SPRi Engineering Research Center, Chongqing Medical University, No. 1 Yi Xue Yuan Road, Chongqing, 400016, PR China.
Department of Laboratory, University-Town Hospital of Chongqing Medical University, Chongqing, 401331, PR China.
Biosens Bioelectron. 2022 Nov 15;216:114677. doi: 10.1016/j.bios.2022.114677. Epub 2022 Sep 5.
Nucleic acid nanotechnologies based on toehold-mediated strand displacement are ideally suited for single-nucleotide variations (SNVs) detection. But only a limited number of means could be used to construct selective hybridization probes via finely designed toehold and regulation of branching migration. Herein, we present a cooperative hybridization strategy relying on a dual-toehold and bulge-loop (DT&BL) probe, coupled with the strand displacement catalytic (SDC) cycle to identify SNVs. The dual-toehold can simultaneously hybridize the 5' and 3' ends of the target, so that it possessed the mutual correction function for improving the specificity in comparison with the single target-binding domain. Insertion of BLs into the dual-toehold probe allows tuning of Gibbs free energy change (ΔG) and control of the reaction rate during branching migration. Using the SDC cycle, the reactivity and selectivity of the DT&BL probe were increased drastically without elaborate competitive sequences. The feasibilities of this platform were demonstrated by the identification of three cancer-related genes. Moreover, the applicability of this biosensor to detect clinical samples showed satisfactory accuracy and reliability. We envision it would offer a new perspective for the construction of highly specific probes based on dynamic DNA nanotechnology, and serves as a promising tool for clinical diagnostics.
基于引发链位移的核酸纳米技术非常适合单核苷酸变异 (SNV) 的检测。但是，只能通过精心设计的引发点和分支迁移调控来构建选择性杂交探针。在这里，我们提出了一种依赖于双引发点和环突（DT&BL）探针的协同杂交策略，与链置换催化（SDC）循环相结合，用于识别 SNV。双引发点可以同时杂交目标物的 5'和 3'末端，因此与单个目标结合域相比，它具有相互校正功能，从而提高了特异性。将 BL 插入双引发点探针中，可以调整吉布斯自由能变化 (ΔG) 并控制分支迁移过程中的反应速率。通过 SDC 循环，DT&BL 探针的反应性和选择性大大提高，而无需精心设计竞争序列。通过鉴定三种与癌症相关的基因，证明了该平台的可行性。此外，该生物传感器在检测临床样本中的适用性表现出令人满意的准确性和可靠性。我们预计它将为基于动态 DNA 纳米技术构建高度特异性的探针提供新的视角，并成为临床诊断的有前途的工具。
