Department of Chemical and Biological Engineering, Northwestern University, Evanston, IL, USA.
Center for Synthetic Biology, Northwestern University, Evanston, IL, USA.
Nat Biotechnol. 2020 Dec;38(12):1451-1459. doi: 10.1038/s41587-020-0571-7. Epub 2020 Jul 6.
Lack of access to safe drinking water is a global problem, and methods to reliably and easily detect contaminants could be transformative. We report the development of a cell-free in vitro transcription system that uses RNA Output Sensors Activated by Ligand Induction (ROSALIND) to detect contaminants in water. A combination of highly processive RNA polymerases, allosteric protein transcription factors and synthetic DNA transcription templates regulates the synthesis of a fluorescence-activating RNA aptamer. The presence of a target contaminant induces the transcription of the aptamer, and a fluorescent signal is produced. We apply ROSALIND to detect a range of water contaminants, including antibiotics, small molecules and metals. We also show that adding RNA circuitry can invert responses, reduce crosstalk and improve sensitivity without protein engineering. The ROSALIND system can be freeze-dried for easy storage and distribution, and we apply it in the field to test municipal water supplies, demonstrating its potential use for monitoring water quality.
缺乏安全饮用水是一个全球性问题，而能够可靠且便捷地检测污染物的方法可能会带来变革。我们报告了一种无细胞体外转录系统的开发，该系统使用配体诱导激活的 RNA 输出传感器（ROSALIND）来检测水中的污染物。高度持续的 RNA 聚合酶、变构蛋白转录因子和合成 DNA 转录模板的组合调节荧光激活 RNA 适体的合成。目标污染物的存在会诱导适体的转录，并产生荧光信号。我们将 ROSALIND 应用于检测一系列水污染物，包括抗生素、小分子和金属。我们还表明，添加 RNA 电路可以在不进行蛋白质工程的情况下反转响应、减少串扰并提高灵敏度。ROSALIND 系统可冻干保存，便于运输和分发，我们将其应用于现场测试市政供水，展示了其在水质监测方面的潜在用途。
