Systems Biology Strategies and Technologies for Understanding Microbes and Microbial Communities Genomic and Proteomic Strategies GTL Profiling Microbial Identity and Activity: Novel Applications of NanoSIMS and High Density Microarrays

Project Goals: The identification of microorganisms responsible for specific processes remains a major challenge in microbial ecology, one that requires the integration of multiple techniques. We propose to address this goal by developing a new methodology, “Chip-SIP”; combining the power of re-designed oligonucleotide microarrays with nano-scale secondary ion mass spectrometry (NanoSIMS) analyses, and linking the identity of microbes to their functional roles. Building upon the stable isotope probing approach (SIP), we will isotopically label microbial nucleic acids by growing organisms on 13C and 15N enriched substrates. Extracted RNA will be hybridized to a newly engineered high-density oligonucleotide microarray with a conductive surface and higher reproducibility relative to traditional microarrays. These advances in array surface chemistry will allow us to successfully analyze arrays by NanoSIMS, generating isotopic and elemental abundance images of the surface, and thereby indicating which organisms in complex consortia utilized the isotopically labeled substrate. Our first application of this new method will assign function to complex microbial community members dwelling in the hindgut of the wood-eating passalid beetle, Odontotaenius disjunctus. This microbial community represents a naturally-selected highly-efficient lignocellulose degrading consortium. Understanding the microbial processes by which woodingesting insects derive energy may aid large-scale conversion of lignocellulosic biomass into biofuels.