Generation of small molecule-binding RNA arrays and their application to fluorogen-binding RNA aptamers.
Generation of small molecule-binding RNA arrays and their application to fluorogen-binding RNA aptamers.
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小分子结合 RNA 阵列的生成及其在荧光结合 RNA 适体中的应用。
DOI：
10.1016/j.ymeth.2019.04.021
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Henderson CA
中科院分区：
文献类型：
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作者：
Henderson CA
随着RNA功能的发现和工程改造，RNA靶向小分子的应用日益广泛。最近，已经开发了几种荧光结合RNA适体，其已经分别作为RNA标签或生物传感器应用于RNA和代谢物的活细胞成像。虽然这些基于荧光体结合RNA适体的装置的设计和应用在理论上是简单的，但在实践中，需要仔细优化。出于这个原因，能够定量荧光体-RNA适体相互作用的高通量体外筛选技术将是有益的。我们最近开发了一种产生功能RNA阵列的方法，并证明它们可用于检测荧光体-RNA适体相互作用。具体而言，我们能够可视化孔雀石绿色和孔雀石绿结合适体之间的相互作用。在这里，我们扩展这项研究，以证明功能RNA阵列可以用来量化荧光体-适体相互作用。作为概念验证，我们提供了用于生产孔雀石绿结合RNA适体和DFHBI结合菠菜RNA适体阵列的详细方案。此外，我们讨论了该技术的潜在效用的荧光结合RNA适体，包括作为一个分子生物传感器平台的应用。我们预计，功能性RNA阵列技术将有利于各种各样的生物学科。
The discovery and engineering of more and more functions of RNA has highlighted the utility of RNA-targeting small molecules. Recently, several fluorogen-binding RNA aptamers have been developed that have been applied to live cell imaging of RNA and metabolites as RNA tags or biosensors, respectively. Although the design and application of these fluorogen-binding RNA aptamer-based devices is straightforward in theory, in practice, careful optimisation is required. For this reason, high throughput in vitro screening techniques, capable of quantifying fluorogen-RNA aptamer interactions, would be beneficial. We recently developed a method for generating functional-RNA arrays and demonstrated that they could be used to detect fluorogen-RNA aptamer interactions. Specifically, we were able to visualise the interaction between malachite green and the malachite green-binding aptamer. Here we expand this study to demonstrate that functional-RNA arrays can be used to quantify fluorogen-aptamer interactions. As proof-of-concept, we provide detailed protocols for the production of malachite green-binding RNA aptamer and DFHBI-binding Spinach RNA aptamer arrays. Furthermore, we discuss the potential utility of the technology to fluorogen-binding RNA aptamers, including application as a molecular biosensor platform. We anticipate that functional-RNA array technology will be beneficial for a wide variety of biological disciplines.