BRIGE - Prokaryotic Real Time Gene Expression Profiling For Toxicity Assessment And Water Quality Monitoring

PI：GU，4月建议编号：0926284目标：与过多的新污染物相关的健康风险需要在水质监测的毒性评估技术方面取得突破。该项目的目标是：a)探索原核实时基因表达谱在评估毒性效应和了解毒性机制方面的新应用的有效性，b)开发一种更复杂、更有信息量、但可行和可靠的毒性评估方法，以评估和识别水样中的污染物，以及c)建立一个创造性的综合教育计划，以吸引和教育来自不同学科和身份的人员，特别是妇女和代表不足的群体，成为环境工程领域的贡献者。长期目标是建立一个独特的跨学科和翻译研究和教育计划，专门应用基因组/分子生物技术进行水质监测。方法：利用绿色荧光蛋白(GFP)与约1900个不同基因启动子在大肠杆菌中的转录融合的综合文库(细胞阵列)，建立一种新的原核实时基因表达谱方法。K12，将用于水样的毒性评估和鉴定。对活细胞中细胞水平集体状态基因表达活动的高时间分辨率和高通量测量将产生响应特定化合物或化合物混合物的特征基因表达谱，这取决于它们的毒性机制和作用模式。将建立各类新兴环境污染物的标志性基因表达谱(生物标记物)数据库，并揭示其毒性机制。将采用多维等级聚类法，根据污染物在基因表达谱上的相似性，进而根据其潜在毒性作用模式的相似性，对污染物进行识别和/或分类。将实际环境样品的基因表达谱结果与传统毒性评价方法得到的结果进行比较，以证实基因表达谱用于毒性评价的有效性。教育计划是通过将一系列教学和教育推广活动与跨学科、体验式和多渠道教学模式相结合而制定的，这种教学模式将包括在课堂上学习现有知识，通过研究和发现发现新信息，并通过实践和应用来应用和转化知识。这些活动的亮点包括通过与南大不同的教育身份、当地组织和社区建立联系，扩大开创性的BEST(生物技术促进环境-展示和培训)计划，以增加环境工程领域参与者的多样性，特别是针对妇女、少数民族和残疾人；开发一门新的跨学科课程，以吸引和利用其他学科的学生学习环境工程，并通过多层次的合作、NU的合作计划和通过ITRI(工业转化研究计划)加强技术转让和信息传播。我们是第一个应用由大量生物发光的绿色荧光蛋白转化的大肠杆菌组成的原核细胞阵列。菌株来测量对污染物的实时基因表达活性。这种方法将导致比传统方法更及时和更有信息量的毒性评价结果。它比现有的基于微阵列的基因组图谱方法具有更高的灵敏度和特异性，因为它增加了图谱数据的时间维度，从而允许更系统和更高分辨率的污染物毒性评估。此外，该方法大大提高了基因表达谱用于毒性评估的可行性和成本效益，因为它具有更简单、更快速和可靠的分析程序，更高的重用性和良好的细胞阵列库定制灵活性。现有的水质监测毒性评价方法还不够完善，严重阻碍了水质保护和改善的进程。该项目提出了一种新的跨学科方法，以满足对有效和可靠的方法来检测和评估水中新出现的污染物的毒性影响的迫切需要。综合的跨学科、体验式和多渠道的教学和教育模式将增加参与环境工程劳动力的多样性，并将使他们掌握足够的知识和技能，以应对当今具有挑战性的环境问题。对她事业发展的支持将帮助她发展成为一名多产的学者，通过为改善水质和保护公共卫生的技术和教育进步做出贡献，造福社会。

PI: Gu, AprilProposal Number: 0926284Goals: The health risk associated with the plethora of emerging contaminants necessitates a breakthrough in toxicity assessment technology for water quality monitoring. The goals of this project are: a) to explore thevalidity of a novel application of prokaryotic real time gene expression profiling for assessing toxic effects and understanding toxicity mechanisms, b) to develop a more sophisticated and informative, yet feasible andreliable toxicity assessment methodology for evaluation and identification of contaminants in water samples and c) to establish a creative and integrated education program to attract and educate personnel from various disciplines and identities, especially women and underrepresented groups, to become contributors in the environmental engineering field. The long-term goal is to build a unique interdisciplinary and translational research and educational program that specializes in applying genomic/molecular biotechnology for waterquality monitoring.Methods: A new prokaryotic real-time gene expression profiling method, using a comprehensive library (cellarray) of transcriptional fusions of Green Fluorescence Protein (GFP) to each of about 1900 different gene promoters in E. coli. k12, will be employed for toxicity evaluation and identification in water samples. The high-temporal-resolution and high-throughput measurements of cellular-level collective state geneexpression activities in the living cell will yield characteristic gene expression profiling in response to a specific compound or a mixture of compounds, depending on their toxic mechanisms and modes of action. A database of signature gene expression profiles (biomarkers) for various categories of emerging environmental pollutants will be established and the toxic mechanisms will be revealed. Multidimensional hierarchical clustering methods will be applied to identify and/or classify pollutants based on their similarity in gene expression profiles and, by extension, similarity in their underlying modes of action of toxicity. Geneexpression profiling results in real environmental samples will be compared with those obtained by conventional toxicity evaluation methods in order to confirm the validity of gene expression profiling fortoxicity assessment. The education plan is developed by integrating a series of teaching and educationoutreach activities with an interdisciplinary, experiential and multi-channel teaching paradigm that will incorporate the learning of existing knowledge in classrooms, discovering new information through researchand discovery, and applying and translating knowledge through practice and application. Highlights of theseactivities include expanding the pioneering BEST (Biotechnology for the Environment-Showcase and Training) program, through connections with various NU education identities, local organizations andcommunities, to increase the diversity of participants in the environmental engineering field, especially forwomen, minorities and people with disabilities; developing a new interdisciplinary course to attract andexpose students from other disciplines to environmental engineering and enhancing technology transfer andinformation dissemination through multi-level collaborations, Co-op program at NU and via ITRI (IndustrialTranslational Research Initiative).Intellectual Merits: This project integrates diverse disciplines of biotechnology, toxicology and environmental engineering and opens new ground for research in genomic-based toxicity assessment for water quality monitoring. We are the first to apply a prokaryotic cell-array consisting of a large number of bioluminescent GFP-transformed E. coli. strains to measure the real-time gene expression activities in response to contaminants. This approach will lead to timely and more informative toxicity evaluation results than conventional methods. It has higher sensitivity and specificity than the existing microarray-based genomic profiling approach because it adds a temporal dimension to the profiling data and therefore allows for more systematic and higher-resolution toxicity evaluation of pollutants. Moreover, the proposed method greatly improves the feasibility and cost-effectiveness of gene expression profiling for toxicity assessment due to its simpler, faster and reliable assay procedures, higher reusability and desirable flexibility for customization of the cell-array library.Broader Impacts: This research has significant impact on ensuring water quality for public health protectionand life quality improvement. Current available toxicity assessment methods for water quality monitoring are not sufficient, and they severely hamper our progress in water quality protection and improvement. This project presents a new interdisciplinary approach to meet the extremely urgent need for effective and reliable methods to detect and evaluate toxicity effects of emerging contaminants in water. The integrated interdisciplinary, experiential and multi-channel teaching and education paradigm will increase the diversityof participation in environmental engineering workforce and it will equip them with adequate knowledge and skills to tackle today's challenging environmental issues. The support for the PI's career development will help her develop as a productive scholar who benefits society by contributing to the technological and educational advancement of water quality improvement and public health protection.