SGER: Development of a Non-invasive Method for Analysis, Mapping and Studying the Biological Interactions of the Root Secretone Using Raman Chemical Imaging Microscopy (RCIM)

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Although root secretions clearly represent a significant carbon cost to the plant, the mechanisms that regulate exudation are poorly understood. Most of the techniques known to date that analyze root secretions depend upon the destructive organic analysis of samples. In the past, these classical techniques that include colorimetric and chromatography have helped scientists to identify various exuded metabolites. However, none of these techniques are useful for monitoring secretions in real-time. Developing a technology to examine in situ-based non-invasive methods for analyzing real-time root secretions is a current goal of the scientific community. A non-invasive real time method to monitor root secretions will help to understand and elucidate various unknown novel chemical interactions between plant roots with other roots, microbes and nematodes. The purpose of this research is to explore Arabidopsis thaliana root secretions in situ using a novel technique known as Raman Chemical Imaging Microscopy (RCIM). RCIM allows rapid and non-destructive real-time analysis of nearly all secondary metabolites (no mass cut-off) based on their polarizability, without involving organic extraction and chromatographic separations. This project will investigate the following specific objectives: (1) to conduct a study of total root secretions in real-time from A. thaliana roots in response to abiotic and biotic stresses, (2) to utilize different secretion pump inhibitors to validate the results of Aim 1, and assort metabolites to the respective secretion pumps. The results obtained from this exploratory project will highlight RCIM as a powerful tool for answering basic questions in rhizosphere biology. Broader impacts. The unique application of RCIM in plant biology research is the observation and analysis in a standoff mode, which will facilitate the identification of secreted metabolites and provide data pertaining to the biological significance of these metabolites in the rhizospheric interactions. The outcome of this project will generate the most extensive collection of secretion profiling in A. thaliana. The methodology that will be developed through this project will be broadly applicable and will offer the ability to identify an unknown function for known secondary metabolites that are secreted and regulated in response to biotic and abiotic cues. The project will contribute to ongoing activities of the PI linking research and education. The career opportunities for a postdoctoral fellow and one undergraduate summer intern involved in the project will be enhanced in the areas of experimental design and technical expertise.