Biophysical and Biochemical Properties of Salmonella Potsdam Bacteriophage P3/2

<p>Addition of KH₂P0₄ salts to Fraser’s Medium produced higher titers of <em>Salmonella potsdam</em> bacteriophage P3/2, but it failed to maintain stable titers under normal cold storage conditions. The addition of ethylenediaminetetraacetate (EDTA) to the medium also produced high titers and helped to maintain phage viability indefinitely in storage. The presence of KH₂P0₄ with the EDTA did not increase the titers nor better the storage properties and was discontinued in favor of EDTA alone. This provided for the accumulation of the necessary amounts of P3/2 for DNA studies.</p> <p>The phage P3/2 adapted well to purification by the differential centrifugation technique. From 50-60 percent of the phage in a suspension can be concentrated by this method. In density gradient centrifugation, phage P3/2 maintained the most optimum viability in potassium tartrate. In potassium tartrate, P3/2 was calculated to have a density of 1.26 and a density of 1.42 in cesium chloride. The virulent mutant, P3/2^vir, had a density of 1.476 in cesium chloride.</p> <p>Phage P3/2^virwas employed in experiments on DNA characterization, DNA estimations by Dische method were reproducible over a period of time for a single phage preparation, but the titer increased upon initial storage in the cold. This made a DNA per pfu determination impractical. Calculations of protein per pfu, employing the Lowry technique, were likewise inconclusive due to their titer increase and the presence of extraneous protein material in the purest density gradient preparations.</p> <p>The average extinction to phosphorus value for two preparation of P3/2^virDNA was found to be 6065 which is comparable to the 6130 value for the highly polymerized calf thymus DNA.</p> <p>Formalin denaturation and heat denaturation test indicated that the P3/2^vir DNA was double-stranded.</p> <p>The Tm, the thermal denaturation temperature of P3/2^vir DNA, was determined to be 91.8°C. From the Tm value, the G-C content of the DNA was calculated to be 53.0 percent.</p> <p>By means of deoxyribonuclease and snake venom phosphodiesterase enzyme degradation and mononucleotide separation by ion-exchange chromatrography, the following molar ratios for the P3/2^virDNA were determined: G-C 54.7-55.1 percent; A-T 44.9-45.3 percent.</p>

Name thesis:degree_name

Master of Science (MS)

Level thesis:degree_level

Thesis

Discipline thesis:degree_discipline

Microbiology

Year

1968

Author dc:creator

• Kettering, James D.

Contributors dc:contributor

• Robert L. Nutter
• Leonard R. Bullas
• Richard E. Beltz

• Microbiology
• Salmonella; Bacteriophages

Statement dc:rights

• This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Language dc:language

English

Repository record dc:identifier

https://scholarsrepository.llu.edu/etd/1400

OAI identifier oai:identifier

oai:scholarsrepository.llu.edu:etd-2171