Powerful Amplification Cascades of FRET-Based Two-Layer Nonenzymatic Nucleic Acid Circuits.
Powerful Amplification Cascades of FRET-Based Two-Layer Nonenzymatic Nucleic Acid Circuits.
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DOI：
10.1021/acs.analchem.6b00609
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发表时间：
2016-05
影响因子：
7.4
通讯作者：
Ke Quan;Jin Huang;Xiaohai Yang;Yanjing Yang;Le Ying;He Wang;Nuli Xie;Min Ou;Kemin Wang
中科院分区：
文献类型：
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作者：
Ke Quan;Jin Huang;Xiaohai Yang;Yanjing Yang;Le Ying;He Wang;Nuli Xie;Min Ou;Kemin Wang
核酸电路在生物工程中发挥着重要的作用，越来越受到人们的关注。它们主要基于不同长度的核酸探针之间的核酸杂交和链置换反应。不依赖于蛋白酶的信号放大方案在分析应用中显示出巨大的潜力。虽然单个放大电路通常实现线性放大，但其可能无法满足以非常小的量检测目标的需要，因此非常有必要构建允许输入的较大放大的级联电路。在此，我们已经成功地设计了基于FRET的两层非酶核酸电路的强大扩增级联，其中催化发夹组装（CHA）的输出激活杂交链式反应（HCR）电路以诱导重复杂交，从而允许通过FRET信号实时监测自组装过程。在设计良好的高质量核酸电路放大器的帮助下，级联可以产生50000倍的信号放大。随后，通过结构转换适体的偶联，在缓冲液以及人血清中检测到低至200 pM的腺苷。据我们所知，我们首次实现了HCR对CHA电路的实时监测适配，实现了核酸和小分子的扩增检测，具有较高的灵敏度。
Nucleic acid circuits have played important roles in biological engineering and have increasingly attracted researchers' attention. They are primarily based on nucleic acid hybridizations and strand displacement reactions between nucleic acid probes of different lengths. Signal amplification schemes that do not rely on protein enzyme show great potential in analytical applications. While the single amplification circuit often achieves linear amplification that may not meet the need for detection of target in a very small amount, it is very necessary to construct cascade circuits that allow for larger amplification of inputs. Herein, we have successfully engineered powerful amplification cascades of FRET-based two-layer nonenzymatic nucleic acid circuits, in which the outputs of catalyzed hairpin assembly (CHA) activate hybridization chain reactions (HCR) circuits to induce repeated hybridization, allowing real-time monitoring of self-assembly process by FRET signal. The cascades can yield 50000-fold signal amplification with the help of the well-designed and high-quality nucleic acid circuit amplifiers. Subsequently, with coupling of structure-switching aptamer, as low as 200 pM adenosine is detected in buffer, as well as in human serum. To our knowledge, we have for the first time realized real-time monitoring adaptation of HCR to CHA circuits and achieved amplified detection of nucleic acids and small molecules with relatively high sensitivity.