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循环的主要驱动力，维持生态系统的营养平衡，并最终塑造整个生态系统的功能。 今天，我们认识到微生物在营养元素的生物化学过程中所起的关键作用;然而，我们对微生物群落结构如何影响给定营养循环中的生物地球化学过程的理解有些初步。 对微生物群落和生物地球化学循环之间的结构-功能关系的有限理解在很大程度上是由于表征微生物群落生态学的技术限制。 直到最近，微生物生态学家还无法明确地描述微生物群落中物种的丰富性和均匀性，以及组成微生物种群的主要生化能力。 使用高通量DNA测序和三种方法学方法，鸟枪宏基因组学，PCR扩增子测序和基因组学微生物生态学家开始揭示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。 虽然这项工作为微生物生态学的进步带来了巨大的希望，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品通量需求，细菌和病毒的小基因组大小及其遗传新奇。 此外，在某些情况下，未能将下一代DNA测序仪充分地应用于环境DNA样本，导致了有偏见的数据和错误的科学结论。 这种“高风险;高回报”研究旨在探索和测试新的下一代DNA测序仪PacBio RS的使用，该测序仪具有多个属性，可能使其更适合微生物生态学研究的特定需求，并有可能对这一地球科学学科产生重大变革。 将进行一系列受控和仔细重复的实验，这些实验将测试PacBio测序在鸟枪宏基因组学、16 S 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）直接比较PacBio在这些应用领域的性能。 通过授予特拉华州大学的NSF重大研究仪器奖，PI将获得学术机构中为数不多的PacBio RS仪器之一。 最终，这些研究将限制PacBio测序中的实验误差，并作为该仪器用于微生物生态学研究的初步证明。该提案的更广泛影响包括努力了解和限制PacBio测序中的错误和其他偏差来源，并提出技术建议，以塑造微生物科学中仪器的最佳使用。 在这项工作的过程中，PI将指导博士。该项目由一名研究生和一名博士后研究员组成，并开放获取所有项目数据和研究结果。

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