Biochemistry, Department of


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A THESIS Presented to the Faculty of The Graduate College at the University of Nebraska In Partial Fulfillment of Requirements For the Degree of Master of Science, Major: Biochemistry, Under the Supervision of Professor Kenneth W. Nickerson. Lincoln, Nebraska: December, 2010
Copyright 2010 Ryan Roberts


Photographs of an ecosystem are an important tool in macro-community ecology. The photograph is a permanent record of species phenotype. In microbiology, biochemical activity provides the most descriptive information of an organism’s phenotype. A method for fingerprinting all biochemical activities occurring within a microbial community is analogous to a photograph. Infrared spectroscopy in the region between wavelengths 2500 to 20,000 nm (mid-IR) is a well established instrumental method for fingerprinting the total biochemical profile of axenic cultures. Spectra are complex and sensitive as demonstrated by the ability to discriminate between strains of bacteria, fungi, and algae. This thesis develops the method to apply mid-IR spectroscopy in order to attain a biochemical fingerprint by attenuated total reflectance. Chapter 2 establishes methods for obtaining mid-IR spectra by prefiltering and concentrating lake water onto a 0.2 µm filter membrane. In order to optimize signal, a combined spectrum from PVDF and nylon membranes is used following a 20-25 µm prefilter. Chapter 3 verifies that the established method is capable of discriminating the biochemical fingerprints of six interdunal lakes from Western Nebraska with differing chemistries and community structures. Mid-IR spectra provide functional data, but do not mirror structure, based on 16S rRNA data, identically. Now, a record of biochemical activity within a microbial community can be captured for future comparison. This technique is culture-independent and provides functional, in situ information. In the future, more information can be deduced by comparing different functional communities to one another, assigning spectral information to particular biochemical activities in a community.

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