EXP-SA: Explosives Tracking: A Microsystem for Detection of Bacterial Endospores as Self-Replicating Nucleic Acid Taggants

本提案提出了一个综合的研究和教育计划，旨在生产、检测和鉴定用于爆炸跟踪的细菌孢子内标记物。虽然这项研究最直接的应用与阻止生物恐怖分子的活动有关，但生物工程和传感器科学与工程的预期基本进展将对其他应用产生重大的社会相关性，包括第一反应活动、医疗保健、食品安全以及避免和减轻污染。研究人员建议将嗜热脂肪芽孢杆菌内生孢子的生物工程与用于样品处理和标记物鉴定的微设备相结合。表面声波微滴混合/运输/培养系统将与分子挂锁探针技术相结合，用于生物工程内生孢子的灵敏鉴定。具体的研究任务是：(i)产生若干不同的芽孢杆菌孢子，每种芽孢杆菌都有一个或多个独特的DNA序列拼接到其基因组中；（ii）调查和确定芽孢萌发、营养细菌裂解、运输、混合和加热微液滴样品所需的最佳SAW装置设计；㈢设计DNA分离子系统；开发一种基于荧光的分子挂锁探针系统，用于DNA鉴定，该系统可与SAW制造微系统平台有效结合；制造和测试拟议的原型识别系统。更广泛的影响将通过以下项目和活动实现更广泛的影响：(i)通过缅因大学（UMaine）的两个主要正在进行的项目，即美国国家科学基金会教师研究经验（RET）和GK-12传感器，培训和互动高中观众；（ii）让缅因州和其他机构的本科生直接参与缅因州正在进行的美国国家科学基金会本科生研究经验（REU）计划下的研究项目；（iii）为本科生确定涉及跨学科研究和设计项目的适当顶点项目；（iv）加强现有的研究生水平课程(1)微型生物工程和(2)声波装置设计与制造，将研究成果纳入每门课程；㈤为涉及缅因州、杰克逊实验室和缅因州医学中心研究所的跨学科、多机构的国家科学基金会功能基因组学综合研究生教育和研究培训方案作出贡献；（六）为硕士生、博士生提供论文选题；在项目网站上以及通过会议和印刷文献传播研究和教育材料。

This proposal presents an integrated research and educational plan directed toward the production, detection, and identification of bacterial endospore taggants for explosive tracking. While the most immediate application of the research is related to stemming the activities of bioterrorists, the anticipated fundamental advances in bioengineering and sensor science and engineering will have significant societal relevance to other applications, including first-responder activities, healthcare, food safety, and pollution avoidance and mitigation. Intellectual Merit The investigators propose to combine bioengineering of Bacillus stearothermophilus endospores with microdevices for sample processing and taggant identification. A surface acoustic wave (SAW) microdroplet mixing/transport/incubator system will be coupled with molecular padlock probe technology for sensitive identification of bioengineered endospores. The specific research tasks are to: (i) Generate a number of different Bacillus spores, each with a unique DNA sequence or sequences spliced into its genome; (ii) Investigate and identify the optimal SAW device designs needed to germinate spores, lyse vegetative bacteria, transport, mix, and heat microdroplet samples; (iii) Design subsystems for DNA isolation; (iv) Develop a fluorescence-based molecular padlock probe system for DNA identification that can operate effectively in conjunction with the SAW fabrication microsystem platform; (v) Fabricate and test the proposed prototype identification system. Broader Impacts Broader impacts will be achieved through the following programs and activities to: (i) Train and interact with high school audiences through two major ongoing programs at University of Maine (UMaine), NSF Research Experiences for Teachers (RET) and the GK-12 Sensors; (ii) Involve undergraduates from Maine and other institutions directly into the research project under the umbrella of the ongoing NSF Research Experience for Undergraduates (REU) program at the UMaine; (iii) Identify appropriate Capstone projects for undergraduates involving cross-disciplinary research and design projects; (iv) Enhance existing graduate level courses (1) Microscale Bioengineering and (2) Design and Fabrication of Acoustic Wave Devices by incorporating research results into each course; (v) Contribute to the interdisciplinary multi-institutional NSF Integrative Graduate Education and Research Traineeship (IGERT) program in functional genomics, which involves UMaine, the Jackson Laboratory, and the Maine Medical Center Research Institute; (vi) Provide thesis topics for M.S. and Ph.D. students; (vii) Disseminate the research and educational material on a project website, and through conferences and printed literature.