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技术的进步正在迅速改变我们对微生物遗传多样性及其与各种环境系统功能关系的理解。流式细胞术和高速分选的原位应用现在是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。我们建议将新兴技术应用于分析流式细胞术，高速细胞分选，单细胞多重置换扩增（MDA）到南极冰样本范围为100 Kyr至8 Myr，以评估活细胞与死细胞的相对比例，并探索古冰微生物DNA修复的遗传策略。在暴露于宇宙辐射的冰中恢复存活的微生物约800万年，表明古冰中的微生物亚群具有特别有效的DNA修复机制。考虑到宇宙通量对古冰样品中DNA降解的强烈影响和有效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800万年前的细菌具有特别有效和新颖的DNA修复机制，我们可以获得遗传特征。研究目标是从地球上最古老的冰中获得用于活细菌DNA修复的基因库。使用了以前没有用于古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”奋进。通过我们的分析，很可能会发现新的遗传能力，也许会为地质时期的生命生存提供关键的分子见解，也许在其他星球上。考虑到标准流式细胞仪的使用和基因组分析的可负担性越来越高，我们的研究策略可以转移到科学界，并应为冰微生物的研究提供关键优势。该项目将允许年轻PI的持续专业发展，并提供一个机会，使具有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）可以互动和发展。