SGER: Assessing Genetic Mechanisms of DNA Repair in Ancient Ice Microbes through Analytical Flow Cytometry, High-Speed Cell Sorting, and Single Cell Genomics

SGER：通过分析流式细胞术、高速细胞分选和单细胞基因组学评估古代冰微生物 DNA 修复的遗传机制

• 批准号: 0907846
• 负责人: Kay Bidle
• 金额: $ 19.46万
• 依托单位: Rutgers University New Brunswick
• 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: 2009-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0907846&HistoricalAwards=false
• 关键词: SGER; Assessing; Genetic; Mechanisms; DNA

Technological advances in analytical flow cytometry and cell sorting are rapidly revolutionizing our understanding of microbial genetic diversity and its relationship to function in widely varied environmental systems. The in situ application of flow cytometry and high-speed sorting is now an indispensable component for interrogation of the physiological state and activity of subpopulations and assessment of the importance of specific genes to cell function. We propose to apply emerging techniques in analytical flow cytometry, high-speed cell sorting, and single-cell multiple displacement amplification (MDA) to Antarctic ice samples ranging 100 Kyr to 8 Myr in order to assess the relative proportion of viable vs. dead cells and to explore genetic strategies of DNA repair in ancient ice microbes. Recovery of viable microbes within ice exposed to cosmic radiation for ~8 Myr suggests that subpopulations of microbes within ancient ice possess particularly efficient DNA repair mechanisms. Given the strong influence of cosmic flux on DNA degradation in ancient ice samples and evidence of effective DNA repair, we hypothesize that the degree of DNA damage increases with time and that viable 8 Myr old bacteria possess particularly effective and novel DNA repair mechanisms for which we can obtain genetic signatures. The research goal is to obtain a library of genes used for DNA repair by viable bacteria from the oldest ice on earth. Using analytical techniques not previously applied to ancient ice microbes. Thus, the proposed research is an 'exploratory' endeavor with potentially high risk. It is likely that novel genetic capabilities will be discovered through our analyses, perhaps providing key molecular insight into the survival of life over geological time, and perhaps on other planets. Given the use of standard flow cytometry instrumentation and the increasing affordability of genome analyses, our research strategy is transferable to the scientific community and should offer key advantages to the study of ice microbes. This project will allow for continued professional development of a young PI and provide an opportunity whereby researchers with different educational backgrounds (undergraduate students, graduate students, technician, post-docs, and faculty) can interact and develop.

分析流式细胞术和细胞分选的技术进步正在迅速彻底改变我们对微生物遗传多样性及其与广泛不同环境系统中功能的关系的理解。流式细胞术和高速分选的原位应用现在已成为询问亚群的生理状态和活性以及评估特定基因对细胞功能的重要性的不可或缺的组成部分。我们建议将分析流式细胞术、高速细胞分选和单细胞多重位移扩增 (MDA) 等新兴技术应用于 100 Kyr 至 8 Myr 的南极冰样本，以评估活细胞与死亡细胞的相对比例，并探索古代冰微生物 DNA 修复的遗传策略。暴露于宇宙辐射约 8 Myr 的冰中活微生物的恢复表明，古代冰中的微生物亚群拥有特别有效的 DNA 修复机制。考虑到宇宙通量对古代冰样本中 DNA 降解的强烈影响以及有效 DNA 修复的证据，我们假设 DNA 损伤的程度随着时间的推移而增加，并且存活的 8 Myr 古老细菌拥有特别有效和新颖的 DNA 修复机制，我们可以通过这些机制获得遗传特征。研究目标是获得一个基因库，用于地球上最古老冰中的活细菌进行 DNA 修复。使用以前未应用于古代冰微生物的分析技术。 因此，拟议的研究是一项具有潜在高风险的“探索性”努力。通过我们的分析，很可能会发现新的遗传能力，或许可以为生命在地质时期甚至其他行星上的生存提供关键的分子洞察。鉴于标准流式细胞术仪器的使用以及基因组分析的日益可承受性，我们的研究策略可以转移到科学界，并且应该为冰微生物的研究提供关键优势。该项目将促进年轻PI的持续专业发展，并为不同教育背景的研究人员（本科生、研究生、技术人员、博士后和教师）提供互动和发展的机会。