Department of Chemistry , University of Massachusetts , Amherst , Massachusetts 01003 , United States.
J Am Chem Soc. 2018 Jul 18;140(28):8739-8745. doi: 10.1021/jacs.8b03956. Epub 2018 Jul 9.
DNA and RNA nanotechnology has been used for the development of dynamic molecular devices. In particular, programmable enzyme-free nucleic acid circuits, such as catalytic hairpin assembly, have been demonstrated as useful tools for bioanalysis and to scale up system complexity to an extent beyond current cellular genetic circuits. However, the intracellular functions of most synthetic nucleic acid circuits have been hindered by challenges in the biological delivery and degradation. On the other hand, genetically encoded and transcribed RNA circuits emerge as alternative powerful tools for long-term embedded cellular analysis and regulation. Herein, we reported a genetically encoded RNA-based catalytic hairpin assembly circuit for sensitive RNA imaging inside living cells. The split version of Broccoli, a fluorogenic RNA aptamer, was used as the reporter. One target RNA can catalytically trigger the fluorescence from tens-to-hundreds of Broccoli. As a result, target RNAs can be sensitively detected. We have further engineered our circuit to allow easy programming to image various target RNA sequences. This design principle opens the arena for developing a large variety of genetically encoded RNA circuits for cellular applications.
DNA 和 RNA 纳米技术已被用于开发动态分子器件。特别是可编程无酶核酸电路，如催化发夹组装，已被证明是生物分析的有用工具，并将系统复杂性扩展到超出当前细胞遗传电路的程度。然而，大多数合成核酸电路的细胞内功能受到生物传递和降解方面的挑战的阻碍。另一方面，遗传编码和转录的 RNA 电路作为长期嵌入式细胞分析和调控的替代强大工具出现。在此，我们报道了一种基于遗传编码 RNA 的催化发夹组装电路，用于在活细胞内进行敏感的 RNA 成像。Broccoli 的分裂版本，一种荧光 RNA 适体，被用作报告子。一个靶 RNA 可以催化触发来自数十到数百个 Broccoli 的荧光。因此，可以灵敏地检测靶 RNA。我们进一步对我们的电路进行了工程设计，使其可以轻松编程以对各种靶 RNA 序列进行成像。该设计原理为开发用于细胞应用的各种遗传编码 RNA 电路开辟了领域。
