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

BRIGE - 用于毒性评估和水质监测的原核实时基因表达谱分析

• 批准号: 0926284
• 负责人: April Gu
• 金额: $ 17.41万
• 依托单位: Northeastern University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0926284&HistoricalAwards=false
• 关键词: BRIGE; Prokaryotic; Real; Time; Gene

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.

PI: Gu， april提案号：0926284目标：与大量新出现的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。这个项目的目标是：A)探索原核生物实时基因表达谱在评估毒性作用和理解毒性机制方面的新应用的有效性；b)开发一种更复杂、信息更丰富、可行和可靠的毒性评估方法，用于评估和鉴定水样中的污染物；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。特别是妇女和代表性不足的群体，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转化研究和教育计划，专门应用基因组/分子生物技术进行水质监测。方法：一种新的原核实时基因表达谱分析方法，利用大肠杆菌中绿色荧光蛋白（GFP）与大约1900种不同基因启动子转录融合的综合文库（cellarray）。K12将用于水样的毒性评价和鉴定。活细胞中细胞水平集体状态基因表达活性的高时间分辨率和高通量测量将根据其毒性机制和作用方式，产生对特定化合物或化合物混合物响应的特征基因表达谱。建立各类新兴环境污染物的特征基因表达谱（生物标志物）数据库，揭示其毒性机制。多维层次聚类方法将应用于根据污染物在基因表达谱上的相似性，进而根据其潜在的毒性作用模式的相似性来确定和/或分类污染物。将真实环境样品中的基因表达谱结果与常规毒性评估方法的结果进行比较，以确认基因表达谱在毒性评估中的有效性。该教育计划是通过将一系列教学和教育推广活动与跨学科，体验式和多渠道的教学范式相结合而制定的，该教学范式将在课堂上学习现有知识，通过研究和发现发现新信息，并通过实践和应用应用和翻译知识。这些活动的亮点包括，通过与北大各个教育机构、地方组织和社区的联系，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，特别是妇女、少数民族和残疾人；发展一门新的跨学科课程，吸引其他学科的学生学习环境工程，并通过与北大和工研院的多层次合作，加强技术转让和信息传播。智力优势：本项目整合了生物技术、毒理学和环境工程等多种学科，为基于基因组的水质毒性评估研究开辟了新的领域。我们是第一个应用由大量生物发光gfp转化的大肠杆菌组成的原核细胞阵列。实时测量菌株对污染物的基因表达活性。与传统方法相比，该方法将产生更及时、更翔实的毒性评价结果。它比现有的基于微阵列的基因组分析方法具有更高的灵敏度和特异性，因为它为分析数据增加了一个时间维度，因此允许对污染物进行更系统和更高分辨率的毒性评估。此外，由于该方法具有更简单、更快速、更可靠的分析程序、更高的可重用性和定制细胞阵列文库所需的灵活性，因此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。广泛影响：本研究对确保水质，保护公众健康和提高生活质量具有重要意义。现有的水质监测毒性评价方法不足，严重阻碍了水质保护和改善工作的进展。该项目提出了一种新的跨学科方法，以满足对水中新出现的污染物的毒性效应进行有效和可靠的检测和评估的迫切需要。综合跨学科，体验式和多渠道的教学和教育模式将增加环境工程劳动力参与的多样性，并将使他们具备足够的知识和技能，以应对当今具有挑战性的环境问题。对首席研究员职业发展的支持将帮助她成为一名富有成效的学者，为改善水质和保护公众健康的技术和教育进步做出贡献，造福社会。