Key Laboratory of Luminescence Analysis and Molecular Sensing, Ministry of Education, College of Chemistry and Chemical Engineering, Southwest University, Chongqing, 400715, PR China.
Guangxi Key Laboratory of Natural Polymer Chemistry and Physics, College of Chemistry and Materials Science, Nanning Normal University, Nanning, 530001, PR China.
Biosens Bioelectron. 2022 Feb 1;197:113809. doi: 10.1016/j.bios.2021.113809. Epub 2021 Nov 16.
The sensing of intracellular microRNAs (miRNAs) is of significance for early-stage disease diagnosis and therapeutic monitoring. DNA is an interesting building material that can be programed into assemblies with rigid and branched structures, especially suitable for imaging intracellular biomolecules or therapeutic drug delivery. Here, by introducing the palindromic sequences into the programmable DNA hairpins, we describe an endogenous target-responsive three-way branched and palindrome-assisted catalytic hairpin assembly (3W-pCHA) approach for imaging miRNA-155 of living tumor cells with high sensitivity. The miRNA-155 triggers autonomous assembly of the fluorescently quenched signal hairpin and two hairpin dimers formed via hybridization of their respective palindromic sequences to yield branched DNA junctions, which carry the unopened hairpins and thus provide addressable substrates for continuous assembly formation of DNA nanostructures. During the formation of the DNA nanostructures, the miRNA-155 is cyclically reused and many signal probes are unfolded to show highly intensified fluorescence for detecting miRNA-155 down to 6.9 pM in vitro with high selectivity. More importantly, these probes can be transfected into live cancer cells to initiate the assembly process triggered by intracellular miRNA-155, which provides a new way for imaging highly under-expressed miRNAs in cells. Besides, this approach can also be employed to differentiate miRNA-155 expression variations in different cells, indicating its promising potentials for early-stage disease diagnosis and biological studies in cells.
细胞内 microRNAs (miRNAs) 的感应对于早期疾病诊断和治疗监测具有重要意义。DNA 是一种有趣的建筑材料，可以被编程成具有刚性和分支结构的组装体，特别适合于成像细胞内生物分子或治疗药物的递运。在这里，通过将回文序列引入可编程 DNA 发夹中，我们描述了一种内源性靶标响应的三向分支和回文辅助催化发夹组装 (3W-pCHA) 方法，用于高灵敏度地对活肿瘤细胞中的 miRNA-155 进行成像。miRNA-155 触发荧光猝灭信号发夹的自主组装以及通过其各自回文序列杂交形成的两个发夹二聚体，从而产生分支 DNA 连接，这些连接携带未打开的发夹，从而为 DNA 纳米结构的连续组装提供了可寻址的底物。在 DNA 纳米结构的形成过程中，miRNA-155 被循环重复利用，并且许多信号探针被展开，以显示高度增强的荧光，用于体外检测低至 6.9 pM 的 miRNA-155，具有高选择性。更重要的是，这些探针可以转染到活癌细胞中，以启动由细胞内 miRNA-155 触发的组装过程，这为在细胞中对高度低表达的 miRNAs 进行成像提供了一种新方法。此外，该方法还可用于区分不同细胞中 miRNA-155 表达的变化，表明其在早期疾病诊断和细胞生物学研究中具有广阔的应用前景。
