A Paper-Based Synthetic Biology Platform for the On-Demand Testing of Water Quality

安全饮用水对公共健康至关重要，但日益受到人为活动和 老化的基础设施，用重金属和其他毒素污染它。这在超级基金附近尤其如此 以及已知或怀疑工业活动造成水污染的棕地地点 消息来源。最值得注意的是饮用水受到铅和砷的污染，这可能会导致铅 中毒和砷中毒，并可导致发育障碍、身体异常、 和癌症。一旦发现，这些污染物可以通过现有的净化技术来缓解。 然而，正如最近发生的事件，如密歇根州弗林特或新泽西州纽瓦克的危机所表明的那样，不恰当的组合 水管理和过滤失败正在导致重大的公共健康威胁。解决方案的一部分 安全水管理的挑战是频繁的水质检测。然而，可靠的测试仍然是有限的 到昂贵、耗时且需要大量实验室的分析化学技术 基础设施和技术专长。这使解决关键水问题所需的大规模测试变得复杂 管理问题，一直是用户对供水进行常规检测的一大障碍。这里 我们建议通过开发一种新的技术平台来解决这些问题，该平台将允许可靠、低成本的 对供水中的有害污染物进行成本、现场和按需监测。我们的技术已经建成 来自合成生物学的最新创新，允许重新调整天然变构转录因子的用途 它可以感知特定的有毒配体，如重金属，并通过激活基因表达来做出反应。这个 使用支持基因表达过程和可视基因表达的无细胞合成生物学反应 记者允许组装“无细胞生物传感器”，这是一种可以冷冻干燥的体外反应。 储存时间长，配送简单。这些传感器与水样的再水化然后激活 在有毒化合物存在的情况下，反应并产生可检测到的信号。此第一阶段建议书详细介绍了 一系列相辅相成的目标，以提高该生物传感平台的特异性和敏感性，在 检测铅和砷作为重大健康关切的示范目标污染物的背景，以及 将其整合到方便的纸质格式中，适合消费者使用。我们的方法包括 鉴定天然转录因子的生物信息学方法的发展和应用 具有改进的性能特征的同系物、高通量的无细胞合成生物学方法 快速表征它们的性能，以及嵌入和测试这些传感器的新制造技术 在纸基基板上。这一提议的成功结果将导致一种多路复用的纸质设备 这将解决可靠和负担得起的铅和铅污染物水质监测问题 砷。这项工作将建立这项技术的概念验证，使第二阶段目标能够实现 在真实世界的水样中实现用户指导的产品规格，并进行生产放大。

Safe drinking water is essential for public health, yet is increasingly threatened by anthropogenic activities and aging infrastructure that contaminate it with heavy metals and other toxins. This is especially true near Superfund and brownfield sites, where industrial activity is either known or suspected to have resulted in pollution of water sources. Most notable is the contamination of drinking water with lead and arsenic, which can result in lead poisoning and arsenicosis, respectively, and can contribute to developmental disorders, physical abnormalities, and cancer. Upon discovery, these contaminants can be mitigated by existing purification technologies. However, as recent events such as the crises in Flint, MI or Newark, NJ exemplify, the combination of improper water management and filtration failure is leading to major public health threats. Part of the solution to the challenge of safe water management is frequent water quality testing. However, reliable testing remains limited to analytical chemistry techniques that are costly, time consuming, and require substantial laboratory infrastructure and technical expertise. This complicates the large scale testing needed to address critical water management issues, and has been a large barrier for the routine testing of water supplies by consumers. Here we propose to address these issues by developing a new technology platform that will allow for the reliable, low- cost, on-site and on-demand monitoring of harmful contaminants within water supplies. Our technology is built from recent innovations in synthetic biology that allow the repurposing of natural allosteric transcription factors that can sense specific toxic ligands, such as heavy metals, and respond by activating gene expression. The use of cell-free synthetic biology reactions that support gene expression processes and visible gene expression reporters allows the assembly of “cell-free biosensors”, which are in vitro reactions that can be freeze-dried for long term storage and simple distribution. Rehydration of these sensors with a water sample then activates the reaction and produces a detectable signal in the presence of a toxic compound. This Phase I proposal details a series of complementary aims for achieving improved specificity and sensitivity of this biosensing platform, in the context of detecting lead and arsenic as model target contaminants of significant health concern, and incorporating it within a convenient paper-based format suitable for consumer use. Our approach includes the development and application of bioinformatic approaches to identify naturally-occurring transcription factor homologues with improved performance characteristics, high throughput cell-free synthetic biology approaches to rapidly characterize their performance, and new manufacturing techniques to embed and test these sensors on paper-based substrates. A successful outcome of this proposal will lead to a multiplexed, paper-based device that will address the problem of reliable and affordable water quality monitoring for the contaminants lead and arsenic. This work will establish a proof-of-concept of this technology that will enable the Phase II goals of achieving user-guided product specifications in real-world water samples and manufacturing scale-up.