Biosensors are key components in engineered biological systems, providing a means of measuring and acting upon the large biochemical space in living cells. However, generating small molecule sensing elements and integrating them into in vivo biosensors have been challenging. Using aptamer-coupled ribozyme libraries and a novel ribozyme regeneration method, we developed de novo rapid in vitro evolution of RNA biosensors (DRIVER) that enables multiplexed discovery of biosensors. With DRIVER and high-throughput characterization (CleaveSeq) fully automated on liquid-handling systems, we identified and validated biosensors against six small molecules, including five for which no aptamers were previously found. DRIVERevolved biosensors were applied directly to regulate gene expression in yeast, displaying activation ratios up to 33-fold. DRIVER biosensors were also applied in detecting metabolite production from a multi-enzyme biosynthetic pathway. This work demonstrates DRIVER as a scalable pipeline for engineering de novo biosensors with wide-ranging applications in biomanufacturing, diagnostics, therapeutics, and synthetic biology.

中文翻译：

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多重，自动化的进化流程可实现生物传感器的可扩展发现和表征

生物传感器是工程生物系统中的关键组件，提供了一种测量并作用于活细胞中巨大生化空间的手段。然而，产生小分子感测元件并将其整合到体内生物传感器中一直是具有挑战性的。使用适体偶联的核酶文库和新颖的核酶再生方法，我们开发了RNA生物传感器（DRIVER）的从头快速体外进化，该生物传感器能够进行生物传感器的多重发现。通过在液体处理系统上实现全自动的DRIVER和高通量表征（CleaveSeq），我们鉴定并验证了针对六个小分子的生物传感器，其中包括五个以前未发现适体的生物传感器。DRIVERevolved生物传感器直接应用于调节酵母中的基因表达，显示出高达33倍的激活率。DRIVER生物传感器还用于检测多酶生物合成途径的代谢产物。这项工作证明了DRIVER是用于从头设计生物传感器的可扩展管道，在生物制造，诊断，治疗和合成生物学中具有广泛的应用。