Date Presented

Spring 4-21-2006

Document Type

Thesis

Degree Name

Bachelor of Science

Department

Biology

First Advisor

Julius Sigler

Second Advisor

David O. Freier

Third Advisor

Nancy Cowden

Abstract

Identifying and confirming the presence of aquatic bacteria by molecular means presents many obstacles for a scientist. One bacterium that has been extensively studied because of its ability to adapt to and colonize a wide variety of ecological environments is Pseudomonas aeruginosa. The bacterium P. aeruginosa is a gram-negative bacterium that, in this case, was cultured from the water of College Lake on the campus of Lynchburg College in Lynchburg, Virginia. Samples of College Lake water were added to a biofilm reactor, and were run for three days. At that point slides were removed from the reactor, nutrient agar plates streaked and individual colonies isolated for strain and biochemical analysis. The purpose of this study was to confirm the identity of the bacterium as P. aeruginosa within the biofilms created, and begin preparation for determining the specific strain of P. aeruginosa present. First, chemical tests were carried out within a laboratory setting to confirm the bacterium’s identity and then bacterium samples were isolated and frozen to create a stock supply of the bacterium. In future research, the DNA of the bacterium stock supply will be isolated using Trizol - phenol/chloroform extraction. The quantity and purity of the DNA will then be determined spectrophotometrically and finally a test will be carried out to determine the presence of P. aeruginosa using Polymerase Chain Reaction (PCR) and a microplate reader. At that point in the research the DNA could be run on a sequencer to determine which specific strain of P. aeruginosa is present.

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