A New Interdisciplinary Frontier: Merging Molecular Biology with Isotopic Geochemistry

新的跨学科前沿：分子生物学与同位素地球化学的融合

• 批准号: 0311937
• 负责人: Ann Pearson
• 金额: $ 48.85万
• 依托单位: Harvard University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0311937&HistoricalAwards=false
• 关键词: New; Interdisciplinary; Frontier; Merging; Molecular

The recently appreciated scope of microbial diversity hints at vast hidden metabolic complexity in the environment. These processes govern the transfer of carbon and energy in natural systems. However, less than one percent of all known prokaryotes can be grown in pure culture. New, culture-independent methods are required to investigate the ecological roles and metabolic pathways of uncultivable prokaryotes. This project will demonstrate that carbon isotopic analysis of nucleic acids, specifically ribosomal RNA (rRNA), can be used to study microbial metabolism. The native rRNA in any sample is molecular biology's analogue to the "biomarker" concept of geochemistry. The key to realizing this important step is the recent development of a viable analytical method for RNA separation and isotopic analysis. Intact, species-specific rRNA is separated from mixtures of total RNA using magnetic bead capture methods. The pure material is transferred to a moving-wire interface attached to an isotope-ratio mass spectrometer. This represents a truly interdisciplinary achievement, as it permits the analysis of 13C/12C ratios of nanogram-sized samples of rRNA. It is possible to obtain both isotopic and phylogenetic information, simultaneously, from any natural sample. Scientific Goals This preliminary study will focus on topics of recent interest in geobiology. (i) What is the metabolic pathway of carbon assimilation in marine planktonic crenarchaeota? Is the hypothesis that this organism uses the 3-hydroxypropionate (3-HP) pathway correct? Is the magnitude of this isotopic fractionation the same in nature as it is in laboratory cultures? If not, what might be learned about the role of the 3-HP pathway in natural communities? (ii) Can we identify the major pathway to CH4 in sedimentary and geothermal environments? To what extent does anaerobic methane oxidation (AMO) contribute to total deep biosphere productivity? Is it possible to distinguish metabolic differences between different microbial taxa involved in AMO? The project fosters collaboration among research institutions and between biologists and geoscientists. In addition to the P.I., there are five individuals representing three research laboratories who also will participate in some aspect of the project. This work also supports the mission of NSF to foster educational development at the graduate and undergraduate levels. A graduate student Ph.D. thesis will result directly from this work; and some of the sampling will be performed by undergraduate students as the field component of a required academic course.

最近对微生物多样性范围的认识暗示了环境中隐藏的巨大代谢复杂性。 这些过程控制着自然系统中碳和能量的转移。 然而，只有不到百分之一的已知原核生物可以在纯培养物中生长。 需要新的、独立于培养的方法来研究不可培养的原核生物的生态作用和代谢途径。 该项目将证明核酸（特别是核糖体 RNA (rRNA)）的碳同位素分析可用于研究微生物代谢。 任何样品中的天然 rRNA 都是分子生物学中地球化学“生物标记”概念的类似物。 实现这一重要步骤的关键是最近开发出一种可行的 RNA 分离和同位素分析方法。 使用磁珠捕获方法从总 RNA 混合物中分离出完整的物种特异性 rRNA。纯材料被转移到连接到同位素比质谱仪的移动线接口。 这代表了真正的跨学科成就，因为它允许分析纳克大小的 rRNA 样品的 13C/12C 比率。 可以从任何自然样本中同时获得同位素和系统发育信息。 科学目标 这项初步研究将重点关注地球生物学最近感兴趣的主题。 (i) 海洋浮游缝隙菌碳同化的代谢途径是什么？ 该生物体使用 3-羟基丙酸 (3-HP) 途径的假设正确吗？ 这种同位素分馏的程度在自然界中与在实验室培养物中相同吗？ 如果不是，那么关于 3-HP 途径在自然群落中的作用，我们可以了解到什么？ (ii) 我们能否确定沉积和地热环境中 CH4 的主要途径？ 厌氧甲烷氧化（AMO）对深层生物圈总生产力的贡献有多大？ 是否有可能区分 AMO 涉及的不同微生物类群之间的代谢差异？该项目促进研究机构之间以及生物学家和地球科学家之间的合作。 除了 P.I. 之外，还有代表三个研究实验室的五名个人也将参与该项目的某些方面。 这项工作还支持 NSF 促进研究生和本科生教育发展的使命。 研究生博士。论文将直接来自这项工作；部分抽样将由本科生作为必修学术课程的现场组成部分进行。