Graduate Thesis Or Dissertation

 

Purification and elementary analysis of Streptomyces griseus phage Public Deposited

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  • This investigation was undertaken to provide a chemical description of an actinophage for Streptomyces griseus, and to determine the base composition of the phage DNA moiety. It was believed of interest to learn whether the actinophage for the filamentous S. griseus host would be different in its chemical properties from the bacteriophages. No such chemical analysis of an actinophage has been reported. Complete chemical analyses are available only for a few of the Escherichia coli phages. A method for isolation and purification of the phage was developed, which consisted of isolation by several cycles of differential centrifugation and final purification by centrifugation in a cesium chloride density gradient. The purified phage was analyzed for phosphorus, DNA, nitrogen and protein by means of standard analytical methods and the resulting data reported, relative to the dry weight of the purified phage. The DNA was extracted from purified phage by the phenol extraction procedure. The purine and pyrimidine base composition of the DNA was determined by standard methods for the hydrolysis of DNA and analysis for the bases by paper chromatography, and also by calculation from the thermal denaturation temperature, or melting point, of the DNA. It was concluded that the S. griseus phage under study did not differ significantly in its chemical properties from most of the other bacteriophages that have been similarly analyzed. It was shown to consist of about 60% DNA and 40% protein. The DNA content of 60% was somewhat higher than the 50% value generally reported for phages. The analysis of the extracted DNA for purine and pyrimidine bases showed it to be composed of the four usual DNA bases. There was no evidence for the presence of an unusual base, The DNA base composition value of 53.4% guanine plus cytosine determined by chromatographic analysis agrees within experimental error with the 51.4% value calculated from the thermal denaturation temperature. The buoyant density in cesium chloride was found to be 1.55 g/cc.
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