Development of Analytical Protocols and Optimization of a Capillary Waveguide Biosensor for Marine Microbial Process Studies

P.I.Paul Kemp(纽约州立大学-SB)提议#：0352252国际和平研究所开发了一种毛细管波导生物传感器系统，用于研究水生系统中的微生物种群和过程，特别是沿海和河口水域和沉积物。生物传感器系统使用荧光检测来测量核酸与结合在毛细管内表面的探针分子的杂交。管壁充当光波导，而毛细管内部充当流动细胞和杂交室。通过适当地针对DNA、rRNA和mRNA序列，光纤生物传感器系统可以用于检测混合微生物群落中特定基因(DNA靶)的丰度、这些基因的表达(mRNA靶)或细胞的生理状态(rRNA靶)。这项建议的目标是：1)使用代表不同研究问题的模型生物系统，开发和测试应用仪器所需的分析方案；2)测试显示有望提高灵敏度的替代分析方案；以及3)进行理论分析，从而在仪器设计中进行实际改进。在研究过程中，将资助两名研究生，并将涉及其他高中、本科生和研究生。生物传感器将用与人类健康和海鲜行业直接相关的病原体进行测试。它不仅可用于检测海水中的病原体，还可用于检测加工海产品中的人类病原体、水产养殖设施中的鱼类病原体以及各种环境中的其他有害和滋扰物种。研究海洋环境中微生物丰度和种群动态、生物量生产和基因表达的能力对水质管理和海产品工业具有巨大的潜在效益，无论是在经济上还是在食品安全方面。除生态学研究外，其他潜在应用包括检测和量化用于河口管理的微生物污染物、贝类水产养殖和用于公共卫生目的的水质。生物传感器对国土安全问题也有影响，特别是对娱乐水域和水库以及沿海水域中潜在有害生物的存在进行监测。

P.I. Paul Kemp (SUNY-SB) Proposal #: 0352252 The PIs have developed a Capillary Waveguide Biosensor system to study microbial populations and processes in aquatic systems, particularly coastal and estuarine waters and sediments. The biosensor system employs fluorescence detection to measure the hybridization of nucleic acids to probe molecules bound to the interior surface of a capillary tube. The tube walls serve as an optical waveguide, while the capillary interior serves as a flow cell and hybridization chamber. By targeting DNA, rRNA and mRNA sequences appropriately, the fiber-optic biosensor system can be used to assay for the abundance of specific genes (DNA target), the expression of those genes (mRNA target), or the physiological status of cells (rRNA target) within mixed microbial communities. The goals of this proposal are to 1) develop and test the analytical protocols required to apply the instrument, using model biological systems representative of different research problems; 2) test alternative analytical protocols that show promise for increased sensitivity; and 3) conduct theoretical analyses leading to practical refinements in instrument design. During the course of this research, two graduate students will be supported and other high school, undergraduate and graduate students will be involved. The biosensor will be tested with pathogens of direct interest to human health and the seafood industry. It can be used not only for detection of pathogens in seawater, but human pathogens in processed seafood, fish pathogens in aquaculture facilities, and other harmful and nuisance species in a diverse range of environments. The capacity to examine microbial abundance and population dynamics, biomass production and gene expression in marine environments has enormous potential benefit for water quality management and the seafood industry, both economically and with respect to food safety. Other potential applications beyond ecological research include detection and quantification of microbial contaminants for estuary management, shellfish aquaculture, and water quality for public health purposes. The biosensor also has implications for homeland security issues, particularly monitoring for the presence of potentially harmful organisms in recreational waters and reservoirs, and in coastal waters.