TOOLS FOR DIFFERENTIAL METABOLOMICS

产品说明：该提案描述了一个由化学家，生物学家，工程师和信息科学家组成的团队的高度跨学科的努力，他们将合作开发一套新的工具，用于表征和识别模型生物代谢组的时间变化。将开发三套工具：增强代谢物组变化的量化的工具、通过大规模平行分析或差异分析增加通量的仪器以及改善代谢物采样和分辨率的分离材料和技术。使用大肠杆菌中的基因敲除突变体和酿酒酵母中的敲入突变体的集合，这些全球分析工具将用于识别代谢节点，控制网络，以及代谢组和基因组之间的联系。定量工作将侧重于稳定同位素编码试剂的制备和使用，以确定代谢物的相对浓度、绝对量和代谢通量。质谱和核磁共振仪器和工具将在这一提议中发挥关键作用。一个革命性的新质谱仪将建成，这是一个微型阵列的96个质量分析仪与每个分析仪监测一个小的质量范围连续。作为我们在体内监测代谢通量的努力的一部分，我们正在设计和建造用于NMR研究的流动池，该流动池将允许同时监测2-4个样品。 微型流动池沿着样品浓缩器将在代谢物的全球分析中至关重要。此外，我们正在开发工具，将提供第一个研究影响细胞利用矿物质离子的基因。最后，正在开发用于代谢物的液相分离的通用分离介质。

DESCRIPTION: This proposal describes a highly interdisciplinary effort by a team of chemists, biologists, engineers, and information scientists who will collaboratively develop a new set of tools for characterizing and recognizing temporal changes in the metabolome of model organisms. Three sets of tool will be developed; those that enhance the quantification of changes in the metabolome, instrumentation that augments throughput by either massively parallel analysis or differential analysis, and separation materials and techniques that improve sampling and resolution of metabolites. Using a collection of gene knockout mutants in Escherichia coli and a knockin mutant in Saccharomyces cerevisiae these global analysis tools will be used to identify metabolic nodes, control networks, and linkages between the metabolome and genome. Quantification efforts will focus on the preparation and use of stable isotope coded reagents for determining relative concentration, absolute amounts, and metabolic flux of metabolites. Mass spectral and nuclear magnetic resonance instruments and tools will play a critical role in this proposal. A revolutionary new mass spectrometer will be built that is a miniature array of 96 mass analyzers with each analyzer monitoring a small mass range continuously. As part of our effort to monitor metabolic flux in vivo we are designing and building miniaturize flow cells for NMR studies that will allow 2-4 samples to be monitored simultaneously. Miniaturized flow cells along with sample concentrators will be of critical importance in the global analyses of metabolites. In addition we are developing tools that will provide the first study of genes that impact utilization of mineral ions by cells. Finally universal separation media are being developed for liquid phase separations of metabolites.