Integrated NMR/SIP technologies for the functional and structural characterization of microbial communities
EMSL Project ID
25690
Abstract
We propose to combine stable isotope probing (SIP) of DNA (SIP-DNA) with real-time on-line SIP-nuclear magnetic resonance (SIP-NMR) analysis of microbial community metabolism. SIP-NMR provides access to metabolic information that is not obtainable with other technologies. Specific aims are: 1) Development of in-vivo NMR spectroscopy platforms that measures microbial community function at scales of the microenvironment (?l) and bench-scale (ml), 2) Application of in-vivo NMR spectroscopy platforms for analysis of metabolic fluxes within a complex microbial community using isotopically labeled (13C) substrates (SIP-NMR) and 3) Coupling of metabolic fluxes to identification of active microbial populations within the microbial community with SIP-DNA. This innovative approach can sharply focus community genomics and other "omics" efforts by identifying relevant metabolic activities and interactions within communities, and restricting omics analyses to newly-synthesized 13C macromolecules. We will examine a complex anaerobic microbial community that degrades plant cell walls while operated in controlled bioreactor environment. The project entails technology development of in situ NMR spectroscopic analyses at bench and microscopic scales. These tools will be coupled with isotopically labeled (13C) substrates to interrogate the functional (metabolic) pathways in the complex microbial community. In order to couple kinetic metabolic analyses with functional properties of community members, stable isotope probing will be applied by sequence analysis of 16S rRNA genes into which specific 13C substrates were incorporated during metabolic analysis. The latter will be a quantitative approach that will allow for answering the question: "how many species are active" and "how active are they" (as defined by the sensitivity of the approach) during the experiments.
Research is geared to quantitatively analyze the metabolic functionality of a complex microbial community by making use of stable isotope approaches that can initially be coupled to analysis of functional genes but be extended in the near future to community genomics. In doing so, we develop NMR technologies that can be applied to metabolic analysis of a broad array of microbial communities and habitats. The application of stable isotopes in this project will give us the confidence, as a proof-of-principle, that quantitative metabolic analysis can be coupled to stable isotope probing. We are confident that future technological breakthroughs will make it relatively easy to couple such metabolic analyses to functionally expressed genes or gene products with the help of transcriptomics and proteomics. The fundamental technology developed in this project can be applied especially to DOE mission areas in Bioenergy, Bioremediation and Carbon Sequestration.
Project Details
Project type
Large-Scale EMSL Research
Start Date
2007-09-01
End Date
2008-09-07
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members