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Label-Free Fluorescent DNA Dendrimers for microRNA Detection Based On Nonlinear Hybridization Chain Reaction-Mediated Multiple G-Quadruplex with Low Background Signal
Bioconjugate Chemistry ( IF 4.0 ) Pub Date : 2018-03-16 00:00:00 , DOI: 10.1021/acs.bioconjchem.8b00098 Qingwang Xue 1 , Chunxue Liu 1 , Xia Li 1 , Li Dai 2 , Huaisheng Wang 1
Bioconjugate Chemistry ( IF 4.0 ) Pub Date : 2018-03-16 00:00:00 , DOI: 10.1021/acs.bioconjchem.8b00098 Qingwang Xue 1 , Chunxue Liu 1 , Xia Li 1 , Li Dai 2 , Huaisheng Wang 1
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Various fluorescent sensing systems for miRNA detection have been developed, but they mostly contain enzymatic amplification reactions and label procedures. The strict reaction conditions of tool enzymes and the high cost of labeling limit their potential applications, especially in complex biological matrices. Here, we have addressed the difficult problems and report a strategy for label-free fluorescent DNA dendrimers based on enzyme-free nonlinear hybridization chain reaction (HCR)-mediated multiple G-quadruplex for simple, sensitive, and selective detection of miRNAs with low-background signal. In the strategy, a split G-quadruplex (3:1) sequence is ingeniously designed at both ends of two double-stranded DNAs, which is exploited as building blocks for nonlinear HCR assembly, thereby acquiring a low background signal. A hairpin switch probe (HSP) was employed as recognition and transduction element. Upon sensing the target miRNA, the nonlinear HCR assembly of two blocks (blocks-A and blocks-B) was initiated with the help of two single-stranded DNA assistants, resulting in chain-branching growth of DNA dendrimers with multiple G-quadruplex incorporation. With the zinc(II)-protoporphyrin IX (ZnPPIX) selectively intercalated into the multiple G-quadruplexes, fluorescent DNA dendrimers were obtained, leading to an exponential fluorescence intensity increase. Benefiting from excellent performances of nonlinear HCR and low background signal, this strategy possesses the characteristics of a simplified reaction operation process, as well as high sensitivity. Moreover, the proposed fluorescent sensing strategy also shows preferable selectivity, and can be implemented without modified DNA blocks. Importantly, the strategy has also been tested for miRNA quantification with high confidence in breast cancer cells. Thus, this proposed strategy for label-free fluorescent DNA dendrimers based on a nonlinear HCR-mediated multiple G-quadruplex will be turned into an alternative approach for simple, sensitive, and selective miRNA quantification.
中文翻译：
基于低背景信号的非线性杂交链反应介导的多个G-四链体的无标记荧光DNA树状分子的microRNA检测。
已经开发了用于miRNA检测的各种荧光传感系统，但是它们大多包含酶促扩增反应和标记程序。工具酶的严格反应条件和标记的高昂费用限制了它们的潜在应用，尤其是在复杂的生物基质中。在这里，我们已经解决了难题，并报告了一种基于无酶非线性杂交链反应（HCR）介导的多个G-四链体的无标记荧光DNA树状大分子的策略，该方法可用于简单，灵敏和选择性地检测低RNA干扰的miRNA。背景信号。在该策略中，巧妙地在两个双链DNA的两端设计了一个分离的G-四链体（3：1）序列，该序列被用作非线性HCR组装的构建基块，从而获得了低背景信号。发夹开关探针（HSP）被用作识别和转导元件。在检测到靶标miRNA时，借助两个单链DNA助手启动了两个嵌段（嵌段-A和嵌段-B）的非线性HCR组装，导致DNA树枝状分子通过多个G-四链体结合而支链生长。 。将锌（II）-原卟啉IX（ZnPPIX）选择性地插入多个G-四链体中，可获得荧光DNA树状大分子，导致指数荧光强度增加。得益于非线性HCR的出色性能和低背景信号，该策略具有简化反应操作过程以及高灵敏度的特点。此外，建议的荧光传感策略还显示出较好的选择性，且无需修饰的DNA区块即可实现。重要的是，该策略也已在乳腺癌细胞中以高可信度进行了miRNA定量测试。因此，这种基于非线性HCR介导的多个G-四链体的无标记荧光DNA树状聚合物的拟议策略将变成一种用于简单，敏感和选择性miRNA定量的替代方法。
更新日期：2018-03-16
中文翻译：
基于低背景信号的非线性杂交链反应介导的多个G-四链体的无标记荧光DNA树状分子的microRNA检测。
已经开发了用于miRNA检测的各种荧光传感系统，但是它们大多包含酶促扩增反应和标记程序。工具酶的严格反应条件和标记的高昂费用限制了它们的潜在应用，尤其是在复杂的生物基质中。在这里，我们已经解决了难题，并报告了一种基于无酶非线性杂交链反应（HCR）介导的多个G-四链体的无标记荧光DNA树状大分子的策略，该方法可用于简单，灵敏和选择性地检测低RNA干扰的miRNA。背景信号。在该策略中，巧妙地在两个双链DNA的两端设计了一个分离的G-四链体（3：1）序列，该序列被用作非线性HCR组装的构建基块，从而获得了低背景信号。发夹开关探针（HSP）被用作识别和转导元件。在检测到靶标miRNA时，借助两个单链DNA助手启动了两个嵌段（嵌段-A和嵌段-B）的非线性HCR组装，导致DNA树枝状分子通过多个G-四链体结合而支链生长。 。将锌（II）-原卟啉IX（ZnPPIX）选择性地插入多个G-四链体中，可获得荧光DNA树状大分子，导致指数荧光强度增加。得益于非线性HCR的出色性能和低背景信号，该策略具有简化反应操作过程以及高灵敏度的特点。此外，建议的荧光传感策略还显示出较好的选择性，且无需修饰的DNA区块即可实现。重要的是，该策略也已在乳腺癌细胞中以高可信度进行了miRNA定量测试。因此，这种基于非线性HCR介导的多个G-四链体的无标记荧光DNA树状聚合物的拟议策略将变成一种用于简单，敏感和选择性miRNA定量的替代方法。
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