Examining Complex Microbial Communities Using the Terminal Restriction Fragment Length Polymorphism Method and Dedicated TRFLP Analysis Software
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Date
2010-08-10
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Abstract
The purpose of this research was to develop a robust methodology that could be used to determine the composition of complex microbial communities found in a variety of environments. In an effort to profile these communities, a 16S rDNA directed, sequencing independent method, Terminal Restriction Fragment Length Polymorphism (TRFLP) was used to generate fragment profiles from genomic DNA isolated from each sample. A custom designed software package, In Silico©, was then used to match these terminal restriction fragments in the sample to patterns of 16S rDNA fragments in a custom database of reference patterns. Identifying these patterns allowed for inferences to be made about the structure of the microbial populations in the environments sampled. TRFLP was chosen as the method of identification since it is a high throughput, cost effective method for community profiling. A combination of this method and the In Silico© software was used to examine microbial communities associated with open air swine waste lagoon systems, the large intestine of the Trichechus manatus latirostrus, Florida manatee, and the microbial populations found in rumen fed fermentors. The community analysis methodology we developed and employed was able to not only detect large and diverse microbial populations within each sample, but was also able to show differences between the detected microbial populations under the various experimental regimes used in the studies presented.
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Keywords
Microbial Community Analysis, 16S rDNA, Swine Lagoons, TRFLP
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Degree
PhD
Discipline
Microbiology
