Monitoring Genetic and Metabolic Potential for In-Site Bioremediation: Mass Spectrometry [report]

M.V. Buchanan
2000 unpublished
Transitions 21 Future Work 22 Appendix 25 Prof. Lidstrom's group developed molecular techniques for analyzing natural populations of methanotrophic bacteria. A DNA sequence database was developed for methanotrophs in Lake Washington sediment, and used to design assays specific for selected diagnostic genes in methanotrophs. These molecular tools were used to analyze the natural populations of methanotrophs in Lake Washington sediments by conventional hybridization techniques. A major surprise
more » ... A major surprise from this work was the finding that a significant segment of the natural population are sMMO-containing Methylomonas strains, which are of interest because they have the potential to carry out high rate degradation of trichloroethylene. New primers have been designed to detect this group of organisms. At ORNL, we have used these tailored products as a model system in developing a streamlined MALDI-MS protocol. We have characterized these products and examined the effects of interferences on our protocol. Aspects of this protocol include a rapid method for preparing PCR products for MALDI-MS analysis, parallel implementation of this purification, and automated MALDI-MS data acquisition. We have achieved advances in figures of merit for MALDI-MS analysis of PCR products, including mass range, resolution, and reproducibility. Relevance, Impact, and Technology Transfer: The issues addressed by this project involve technology to increase the applicability of in-situ bioremediation for DNAPL, or potentially any microbially-degraded pollutant. We have applied fundamental knowledge in molecular biology and analytical chemistry to the problem of rapidly characterizing microbial populations at potential cleanup sites. The project was a collaboration between the Organic and Biological Mass
doi:10.2172/885583 fatcat:7ejk6ltb5rbx7fa6dza3thp7cm