PAPM EAGER: Microfluidic Root Exudate Sampler with High Spatio-Temporal Sampling Resolution

PAPM EAGER：具有高时空采样分辨率的微流控根分泌物采样器

• 批准号: 1650182
• 负责人: Liang Dong
• 金额: $ 30万
• 依托单位: Iowa State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-15 至 2021-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1650182&HistoricalAwards=false
• 关键词: PAPM; EAGER; Microfluidic; Root; Exudate; PAPM; EAGER; Microfluidic; Root; Exudate

Roots interact with and respond to the biotic and abiotic environment in which they live (the rhizosphere) by qualitatively and quantitatively modulating material exuded by roots. These exudates use the language of chemistry to communicate between and among the biotic and abiotic components. Powerful DNA sequencing platforms and analytical tools for identifying chemical components are now available for profiling these interactions among the rhizosphere and the root components. The integrated application of these analytical strategies is limited by the ability to access and isolate exudates from roots and the rhizosphere. Existing exudate sampling tools are bulky, require large amounts of soil, and significantly alter the soil structure. This difficulty of sampling exudates has slowed the process of linking plant genetic determinants to rhizosphere microbiome genomic and metabolic features. This project addresses key design, fabrication, integration, and operation problems faced in developing next-generation root exudate sampling tools. The research will develop greatly-needed tools for probing the chemical exchange between plants and the micro- and macro-organisms in the rhizosphere. The root exudates are critical drivers of microbiome assembly and plant-pest/pathogen outcomes. The dynamic and environmentally responsive nature of root exudates illustrates the importance of developing sampling tools that are functional in a real-world situation, rather than the current tools that are limited to use in primarily artificial hydroponic and polymer-embedded systems. The samplers that will be developed will significantly impact the pace of research on rhizosphere microbiome by enabling continuous, spatially-resolved sampling of the microbes and exudates on roots grown in real-world conditions. This enhanced capability will meet societal needs to increase agricultural productivity for an increasing global population in the face of the uncertainties associated with climate-change, and thus develop new strategies to impact gains in agricultural productivity. This research will enhance interdisciplinary STEM workforce development by hosting at least two under-represented students in an undergraduate Howard Hughes Medical Institute summer internship program, providing research opportunities to four undergraduate senior students, and providing hands-on workshops to a high school Science Bound program to engage students in tech-transfer endeavors, while highlighting plant-microbe contributions to agriculture and global food security. This project will elaborate advanced technology for gathering high spatiotemporal resolution data of metabolites and microbes in the rhizosphere. This objective will be met by developing a modular toolkit for the localized sampling of rhizosphere exudates from roots grown in soil matrices. This toolkit will consist of (i) a single site exudate sampler, which will serve as a building block of more complex modular sampling systems; (ii) distributed exudate samplers able to extract exudates from key locations with high spatial resolution; (iii) spine-like flexible exudate samplers, providing conformational fitting at the root-soil interface, which will maximize sampling at this crucial interface; and (iv) parallel gradient samplers positioned radially outward from a root, providing access to radial gradients of exudates. These samplers will be uniquely coupled with microfluidic sorters to enable automated separation and isolation of microbes from the collected exudates for simultaneous analysis of both the microbes and the soluble exudates. Furthermore, these samplers will integrate miniature tensiometers, which will allow monitoring of local soil potential condition at the sampling sites, and trigger the automatic start of sampling. Rendering such a "smart" device will improve temporal resolution of sampling. Validating the utility of these integrated devices will involve installing them, collecting and sorting samples, and analyzing the interactions between the rhizosphere and genetically specified maize roots, grown under gnotobiotic conditions with and without microbes. The multidisciplinary research has drawn expertise ranging from microsystems design and construction, microbiome and metabolomics, to address the proposed goal and deliver on the specific aims.

根部通过定性和定量调节根部散发的材料来与其居住的生物和非生物环境相互作用并响应。这些渗出物使用化学语言在生物和非生物成分之间进行交流。现在，用于识别化学成分的强大DNA测序平台和分析工具现在可以在根际和根部成分之间分析这些相互作用。这些分析策略的综合应用受到访问和隔离根和根际渗出液的能力的限制。现有的渗出液采样工具是笨重的，需要大量的土壤，并显着改变土壤结构。采样渗出物的这种困难使将植物遗传决定因素与根际微生物组基因组和代谢特征联系起来的过程减慢了。该项目解决了开发下一代根渗出液采样工具时面临的关键设计，制造，集成和操作问题。该研究将开发出急需的工具，用于探测根际中植物与微生物之间的化学交换。根部渗出物是微生物组组装和植物 - 基因/病原体结果的关键驱动因素。根部渗出的动态和环境响应性质说明了开发在现实情况下起作用的采样工具的重要性，而不是当前用于主要用于人工水培和聚合物包含系统的工具。将要开发的采样器将通过在现实情况下对根的连续，空间分辨的采样来显着影响根际微生物组的研究速度。这种增强的能力将满足社会需求，以提高农业生产力，以面对与气候变化相关的不确定性，从而增加全球人口，从而制定新的策略来影响农业生产力的增长。 This research will enhance interdisciplinary STEM workforce development by hosting at least two under-represented students in an undergraduate Howard Hughes Medical Institute summer internship program, providing research opportunities to four undergraduate senior students, and providing hands-on workshops to a high school Science Bound program to engage students in tech-transfer endeavors, while highlighting plant-microbe contributions to agriculture and global food security. 该项目将详细阐述高级技术，以收集根际代谢物和微生物的高时空分辨率数据。通过开发一个模块化工具包来实现这一目标，用于从土壤基质中生长的根源的根际渗出液的局部采样。该工具包将由（i）单个站点渗出液采样器组成，该采样器将作为更复杂的模块化采样系统的构建块； （ii）分布式渗出液采样器能够从空间分辨率高的关键位置提取渗出液； （iii）类似脊柱的柔性渗出液采样器，在根部土壤界面提供构象拟合，这将在此关键界面处最大化采样； （iv）并行梯度采样器从根部向外放置径向向外放置，从而可以访问渗出液的径向梯度。这些采样器将与微流体分落者唯一耦合，以实现从收集的渗出液中自动分离和分离微生物，以同时分析微生物和可溶性渗出液。此外，这些采样器将集成微型张力仪，这将允许监测采样位点的局部土壤潜在条件，并触发采样的自动启动。渲染这样的“智能”设备将改善抽样的时间分辨率。验证这些集成设备的效用将涉及安装它们，收集和分类样品，并分析根际和遗传指定的玉米根之间的相互作用，这些玉米根在具有和没有微生物的gnotobiotic条件下生长。多学科研究吸引了从微系统设计和建筑，微生物组和代谢组学等方面的专业知识，以解决拟议的目标并以具体目的实现。