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Toehold switches are biosensors useful for the detection of endogenous and environmental RNAs. They have been engineered to detect virus RNAs in cell-free gene expression reactions. Their inherent sequence programmability makes engineering a fast and predictable process. Despite improvements in the design, toehold switches suffer from leaky translation in the OFF state, which compromises the fold change and sensitivity of the biosensor. To address this, we constructed and tested signal amplification circuits for three toehold switches triggered by Dengue and Sars-CoV-2 RNAs and an artificial RNA. The serine integrase circuit efficientl contained leakage, boosted the expression fold-change from OFF to ON, and decreased the detection limit of the switches by three to four orders of magnitude. Ultimately, the integrase circuit converted the analog switches’ signals into digital-like output. The circuit is broadly useful for biosensors and eliminates the hard work of designing and testing multiple switches to find the best possible performer.
中文翻译：
Toehold 开关是可用于检测内源性和环境 RNA 的生物传感器。它们被设计用于检测无细胞基因表达反应中的病毒 RNA。它们固有的序列可编程性使工程成为一个快速且可预测的过程。尽管设计有所改进，但脚趾开关在 OFF 状态下会发生泄漏翻译，这会影响生物传感器的倍数变化和灵敏度。为了解决这个问题，我们构建并测试了由登革热和 Sars-CoV-2 RNA 以及人工 RNA 触发的三个立足点开关的信号放大电路。丝氨酸整合酶电路有效地包含了泄漏，提高了从关闭到打开的表达倍数变化，并将开关的检测极限降低了三到四个数量级。最终，集成酶电路将模拟开关的信号转换为类似数字的输出。该电路广泛用于生物传感器，消除了设计和测试多个开关以找到最佳性能的艰苦工作。 |
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