Preparation, Purification, and Properties of E. Coli Virus T2

Abstract

Many reports (1-9) on the purification of bacterial viruses have appeared since Northrop's paper on staphylococcus phage (24). In general they all agree that these viruses are nucleoproteins and of those analyzed virtually all have contained principally desoxyribose type nucleic acid (DNA). Taylor (11) who made the most complete analysis of purified T~ virus of E. coli B reported the presence of more than 1 per cent pentosenucleic acid (PNA) and lipid in addition to the protein and DNA.in the present study the method of preparation and the homogeneity of the purified product have been improved. Lysates of T2 in 6 to 10 liter quantities have been prepared which titered 2 to 5 X 1011 infectious units per ml. Purification was effected by (a) decanting the virus solution from the settled cell debris, (b) precipitation of the virus in the cold at pH 4.0, (c) enzymatic digestion of extraneous DNA with desoxyribonuclease (DNAse), and finally differential centrifugation. These steps resulted in a remarkably stable product having a high specific infectivity yet containing virtually no lipid or PNA. A variety of fractionafion experiments designed to furnish evidence on the homogeneity of the purified preparations failed to demonstrate the presence of impurities. Table I contains data on eight different preparations of synthetic medium T2 lysates in 6 to 10 liter quantities. Titers of the lysates varied from 2 to 5 X 1011 plaque-forming units per ml. In test tube quantities lysate titers have run as high as 8 X 10n/ml, Other workers (25,5,6) have also reported high titer lysates. Trial runs using the coli viruses T2, and T4 also yielded lysates titerlng 2 X 10n/ml. In some preliminary experiments T~ lysates titering 2 X 10n/ml. were prepared in nutrient broth of four times the usual concentration. Fig. 1 is an electron microscope picture of an unfractionated high titer T2 * Aided by a grant from The National Foundation for Infantile Paralysis. EXPERIMENTAL RESULTS Preparation of Lysa~s.--

Amino acid and sugar transport in Escherichia coli (ColIb) during abortive infection by bacteriophage T5

“…Media and growth of bacteria. Basic growth medium is defined as M9 phosphate-buffered balanced salt solution (21), supplemented with glucose (0.5%), yeast extract (0.05%), CaCl2 (5 x 10-4 M), and thymine (50 jug/ml). Other additives were present as indicated.…”

Sampling Submicron T1 Bacteriophage Aerosols

“…900 HARSTAD on July 3, 2020 by guest http://aem.asm.org/ Downloaded from VOL. 13, 1965 SAMPLING SUBMICRON Ti BACTERIOPHAGE AEROSOLS nucleic acid and ribonucleic acid decreased the viscosity of the suspension and enhanced the separation of the phage component by centrifugation (Herriott and Barlow, 1952). The suspension was clarified by low-speed centrifugation and then subjected to six complete cycles of differential centrifugation, i.e., alternate high-speed (60,000 X g) and low-speed (10,000 X g) centrifugation.…”

Electron Microscopy of in Vitro Endocytosis of T2 Phage by Cells From Rabbit Peritoneal Exudate

“…1"2 Baderiophage.--The bacteriophage was grown on Escherichia coli B in M-9 medium (9) and purified by differential centrifngation (10). Stocks of 10 TM plaque-forming units/ml were maintained in sterile isotonic saline containing 0.001 M MgSO4 at pH 7.0.…”