Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, 7-1-20 Minatojima-Minamimachi, Kobe 650-0047, Japan.
Graduate School of Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, 7-1-20 Minatojima-Minamimachi, Kobe 650-0047, Japan.
Anal Chem. 2020 Jun 2;92(11):7955-7963. doi: 10.1021/acs.analchem.0c01338. Epub 2020 May 15.
An RNA signaling aptamer is composed of two units: a sensing aptamer that binds the input target molecule and a working aptamer that binds the output target molecule to result in a detectable signal. A conformational change of the signaling aptamer that induces an allosteric interaction with the output target molecule in response to the input target molecule depends on a junction region, which connects the two aptamer units. Efficient and effective optimization of the junction region remains a technical challenge. In this study, we demonstrate a simple strategy for optimizing the junction region through functional RNA selection using RNA-capturing microsphere particles. From approximately 0.2 million sequence variants, a signaling aptamer that enabled intracellular detection of -adenosyl methionine with a high signal-to-noise ratio, which is approximately 2-fold higher relative fluorescence increment compared to the previously reported signaling aptamer, was obtained after single round of selection. The technology demonstrated here can be used to select RNA sequences that carry out specific functions in response to particular stimuli.
一种 RNA 信号适体由两个单元组成：一个感应适体，它结合输入目标分子，一个工作适体，它结合输出目标分子以产生可检测的信号。信号适体的构象变化诱导与输出目标分子的变构相互作用，以响应输入目标分子，这取决于连接两个适体单元的连接区。有效地优化连接区仍然是一个技术挑战。在这项研究中，我们展示了一种通过使用 RNA 捕获微球颗粒进行功能 RNA 选择来优化连接区的简单策略。从大约 200 万个序列变体中，我们获得了一种信号适体，它能够以高信噪比进行细胞内 - 腺苷甲硫氨酸检测，与之前报道的信号适体相比，相对荧光增量增加了约 2 倍。这里展示的技术可用于选择在响应特定刺激时执行特定功能的 RNA 序列。
