Characterization of Bacteriophage gh-1 for Pseudomonas putida1

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Lee, Lucy F. (Michigan State University, East Lansing), and J. A. Boezi. Characterization of bacteriophage gh-1 for Pseudomonas putida. J. Bacteriol. 92:1821–1827. 1966.—Bacteriophage gh-1 of Pseudomonas putida A.3.12 was isolated and purified by differential centrifugation and diethylaminoethyl (DEAE) cellulose chromatography. An electron micrograph of the phage stained with uranyl acetate revealed a regular hexagonal outline about 50 mμ across with a short wedge-shaped tail attached at one corner of the head. The phage formed 10% as many plaques on P. putida C1S as on P. putida A.3.12, the organism used in the isolation procedure. No plaques were formed on P. fluorescens (ATCC 9712) or P. aeruginosa. The latent period of the infectious cycle was 21 min, and the average burst size was 103. The nucleic acid component of gh-1 is double-stranded deoxyribonucleic acid (DNA), with a base composition of 57.0% guanine plus cytosine (G + C) as determined by chemical analysis. The per cent G + C of P. putida A.3.12 DNA measured in a similar manner was 63.7%. The buoyant density of phage gh-1 measured by cesium chloride equilibrium centrifugation was 1.45 g/cm3, whereas that of gh-1 DNA, heat-denatured gh-1 DNA, and P. putida A.3.12 DNA was 1.716, 1.730, and 1.722 g/cm3, respectively. The per cent G + C of gh-1 DNA and P. putida A.3.12 DNA calculated from the buoyant densities was 57.1 and 63.3%, respectively. The sedimentation coefficients, S5020,w, of gh-1 and the phenol-extracted gh-1 DNA, measured by the boundary sedimentation velocity method, were 460 and 18.9, respectively. The molecular weight of phenol-extracted gh-1 DNA, calculated by use of the equation of Burgi and Hershey, is 6 × 106.

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