EAGER: Collaborative Research: Exploratory application of single-molecule real time (SMRT) DNA sequencing in microbial ecology research

EAGER：协作研究：单分子实时 (SMRT) DNA 测序在微生物生态学研究中的探索性应用

• 批准号: 1148118
• 负责人: K. Wommack
• 金额: $ 20万
• 依托单位: University of Delaware
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-15 至 2014-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1148118&HistoricalAwards=false
• 关键词: EAGER; Collaborative; Research; Exploratory; application

Microbial communities are the primary drivers of the global biogeochemical cycles that maintain the nutrient balance of ecosystems and ultimately shape overall ecosystem function. Today, we appreciate the critical role microbes play in the biochemical processing of nutrient elements; yet our understanding of how the structure of microbial communities influences the suite of biogeochemical processes within a given nutrient cycle is somewhat rudimentary. Limited understanding of structure-function relationships between microbial communities and biogeochemical cycles is due in large part to technological limitations in characterizing the ecology of microbial communities. Until recently, microbial ecologists simply had no means of unambiguously characterizing the richness and evenness of species within a microbial community and the prevailing biochemical capabilities of constituent microbial populations. Using high-throughput DNA sequencing and three methodological approaches,shotgun metagenomics, PCR amplicon sequencing, and genomics microbial ecologists are beginning to unveil the inner workings of microbial communities and connect genetic details with biogeochemical processes. While this work holds great promise for the advancement of microbial ecology, currently-available high-throughput sequencing technologies are not ideally suited to the high-sample throughput demands of ecosystem science, the small genome size of bacteria and viruses, and their genetic novelty. Moreover, in some cases the failure to adequately ground-truth application of next-generation DNA sequencers to environmental DNA samples has resulted in biased data and erroneous scientific conclusions. This "high-risk; high-reward" research seeks to explore and test the use of a new, next-generation DNA sequencer, the PacBio RS, which has several attributes that may make it better suited to the specific needs of microbial ecology research and has the potential to be highly transformative to this geoscience discipline. A series of controlled and carefully replicated experiments will be conducted that will test the use of PacBio sequencing for shotgun metagenomics, 16S PCR amplicon sequencing, and single cell genome sequencing. This project will leverage existing datasets from other high-throughput sequencing platforms (e.g., Illumina and 454) to directly compare the performance of PacBio in each of these application areas. Through a NSF Major Research Instrumentation award to the University of Delaware, the PIs will have access to one of the few PacBio RS instruments available at an academic institution. Ultimately, these investigations will constrain the experimental error within PacBio sequencing and serve as an initial demonstration of the utility of the instrument for microbial ecology research. The Broader Impacts of this proposal includes an effort to understand and constrain the sources of error and other biases within PacBio sequencing, and make technical recommendations that will shape the optimal use of the instrument within microbial science. In the course of this work, the PIs will mentor a Ph.D. graduate student and a post-doctoral researcher, and provide open access to all project data and findings.

微生物群落是全球生物地球化学周期的主要驱动因素，可维持生态系统的营养平衡并最终塑造整体生态系统功能。 今天，我们感谢微生物在营养元素的生化加工中所扮演的关键作用。然而，我们对微生物群落结构如何影响给定营养周期内生物地球化学过程的套件的理解是基本的。 对微生物群落与生物地球化学周期之间的结构 - 功能关系的理解有限，这在很大程度上是由于技术局限性在表征微生物群落的生态学方面的局限性。 直到最近，微生物生态学家根本没有明确表征微生物群落中物种的丰富性和均匀性，以及成分微生物种群的现行生化能力。 使用高通量DNA测序和三种方法学方法，shot弹枪宏基因组学，PCR扩增子测序和基因组学微生物生态学家开始公布微生物群落的内部运作，并将遗传细节与生物地球化学过程联系起来。 尽管这项工作对发展微生物生态学有很大的希望，但目前可用的高通量测序技术并不适合生态系统科学的高样本吞吐量需求，细菌和病毒的基因组大小和病毒及其遗传新颖性的较小基因组大小。 此外，在某些情况下，未能充分地将下一代DNA测序仪在环境DNA样品中实现，从而得出了偏见的数据和错误的科学结论。 这项“高风险”研究旨在探索和测试新的下一代DNA测序仪PACBIO RS的使用，该测序仪PACBIO RS具有多种属性，可能使其更适合微生物生态学研究的特定需求，并有可能对这项地理学纪律具有很高的变革性。 将进行一系列受控和仔细复制的实验，以测试PACBIO测序在shot弹枪元基因组学，16S PCR扩增子测序和单细胞基因组测序中的使用。 该项目将利用其他高通量测序平台（例如Illumina和454）的现有数据集直接比较这些应用程序领域的PACBIO的性能。 通过向特拉华大学的NSF重大研究仪器奖，PIS将可以访问学术机构可用的少数PACBIO RS仪器之一。 最终，这些研究将限制PACBIO测序中的实验误差，并作为微生物生态研究仪器效用的初步证明。该提案的更广泛的影响包括努力了解和限制PACBIO测序中错误和其他偏见的来源，并提出技术建议，以影响仪器在微生物科学中的最佳使用。 在这项工作的过程中，PIS将指导博士学位。研究生和博士后研究人员，并为所有项目数据和发现提供公开访问。

项目成果

CRISPR Spacers Indicate Preferential Matching of Specific Virioplankton Genes

• DOI: 10.1128/mbio.02651-18
• 发表时间: 2018-12
• 期刊: mBio
• 影响因子: 6.4
• 作者: Daniel J. Nasko;Barbra D. Ferrell;Ryan M. Moore;Jaysheel D. Bhavsar;Shawn W. Polson;K. E. Wommack

Family A DNA Polymerase Phylogeny Uncovers Diversity and Replication Gene Organization in the Virioplankton

• DOI: 10.3389/fmicb.2018.03053
• 发表时间: 2018-12-14
• 期刊: FRONTIERS IN MICROBIOLOGY
• 影响因子: 5.2
• 作者: Nasko，Daniel J.;Chopyk，Jessica;Wommack，K. Eric
• 通讯作者: Wommack，K. Eric