We here designed an in vitro selection scheme for obtaining an aptamer with which to rationally construct an artificial riboswitch as its component part. In fact, a nanosized DNA-binding aptamer obtained through this scheme allowed us to easily and successfully create eukaryotic riboswitches that upregulate internal ribosome entry site-mediated translation in response to the ligand (nanosized DNA) in wheat germ extract, a eukaryotic cell-free expression system. The induction ratio of the best riboswitch ligand-dose-dependently increased to 21 at 300 μM ligand. This switching efficiency is much higher than that of the same type of riboswitch with a widely used theophylline-binding aptamer, which was in vitro selected without considering its utility for constructing riboswitches. The selection scheme described here would facilitate obtaining various ligand/aptamer pairs suitable for constructing artificial riboswitches, which could serve as elements of synthetic gene circuits in synthetic biology.

中文翻译：

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与纳米级 DNA 结合的 RNA 适体的体外选择用于构建人工核糖开关

我们在这里设计了一种体外选择方案，用于获得适体，以合理地构建人工核糖开关作为其组成部分。事实上，通过该方案获得的纳米级 DNA 结合适体使我们能够轻松并成功地创建真核核糖开关，以响应小麦胚芽提取物中的配体（纳米级 DNA）上调内部核糖体进入位点介导的翻译，这是一种真核生物无细胞表达系统。在 300 μM 配体时，最佳核糖开关配体的诱导率以剂量依赖性方式增加至 21。这种转换效率远高于具有广泛使用的茶碱结合适体的同类型核糖开关的转换效率，这是在体外选择时不考虑其用于构建核糖开关的效用。这里描述的选择方案将有助于获得适合构建人工核糖开关的各种配体/适体对, 可作为合成生物学中合成基因回路的元素。