Isolation of Circular Deoxyribonucleic Acid from
            <i>Salmonella typhosa</i>
            Hybrids Obtained from Matings with
            <i>Escherichia coli</i>
            Hfr Donors

Leavitt, Ronald W.; Wohlhieter, J. A.; Johnson, E. M.; Olson, Gary E.; Baron, L. S.

doi:10.1128/jb.108.3.1357-1365.1971

Cited by 13 publications

(7 citation statements)

References 21 publications

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“…As described previously (5), the plaques formed by OH on F-containing (male) E. coli K-12 strains are significantly smaller than those formed on non-F-containing (female) K-12 strains. Experiments to detect increases in phage titer were performed by adding phage to bacteria at a ratio of 1 to 1,000 in appropriate nutrient broth medium, incubating overnight, and assaying the supematants on sensitive indicator bacteria, as described previously (2,7). RESULTS Transfer of the scr elements to E. coli.…”

Section: Methodsmentioning

confidence: 99%

Characterization of transmissible genetic elements from sucrose-fermenting Salmonella strains

Wohlhieter¹,

Lazere²,

Snellings³

et al. 1975

J Bacteriol

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Two of seven sucrose-fermenting Salmonella strains obtained from clinical sources were found capable of conjugal transfer of the sucrose fermentation (Scr+) property to the Escherichia coli K-12 strain WR3026. The genetic elements conferring this Scr+ property, designated scr-53 and scr-94, were then conjugally transmissible from E. coli WR3026 Scr+ exconjugants to other strains of E. coli at frequencies of 5 x 10-I to 5 x 10-I for the scr-53 element and 10 6 to 10-s for the scr-94 element. In E. coli hosts, both of these elements were compatible with F-lac and with each of six previously characterized transmissible lac elements. No antibiotic resistance characteristics or colicin production were discovered to be associated with either scr-53 or scr-94. Neither scr element generated a male host response to the female-specific phage XII, but the scr-53 element rendered its E. coli host sensitive to the male-specific phage R-17. E. coli hosts containing scr-53 were susceptible to lysis by Plvir, and transduction of the scr-53 element was accomplished with this phage. The scr-53 element was isolated from E. coli WR3026, Scr+ transductants, and E. coli WR2036 Scr+ exconjugants as a covalently closed circular deoxyribonucleic acid molecule with a molecular weight (determined by electron microscopy) of approximately 52 x 106. Receipt of the scr-94 element rendered E. coli hosts of this element unsusceptible to lysis by Plvir, although adsorption of the phage by an E. coli WR3026 exconjugant containing scr-94 occurred as efficiently as it did on WR3026 itself. Repeated examination of E. coli strains harboring scr-94, as well as of the Salmonella strain which initially contained it, did not reveal the presence of circular deoxyribonucleic acid. The synthesis of the sucrose cleaving enzyme was inducible in E. coli exconjugants containing either scr-53 or scr-94.

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Section: Methodsmentioning

confidence: 99%

Characterization of transmissible genetic elements from sucrose-fermenting Salmonella strains

Wohlhieter¹,

Lazere²,

Snellings³

et al. 1975

J Bacteriol

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“…The entire lysate was centrifuged in an ethidium bromidecesium chloride (EtBr-CsCl) density gradient at 44,000 rev/min for 40 hr. Details of the method have been described by Leavitt et al (16). After centrifugation, the gradient was fractionated into 0.3-ml fractions, and 50-juliter samples were counted to determine which fractions contained the labeled DNA.…”

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confidence: 99%

“…Sucrose density gradient centrifugation. The method used for determination of the sedimentation coefficient of the supercoiled DNA fractions was described by Leavitt et al (16).…”

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confidence: 99%

Isolation and Characterization of the Fertility Factor P of Vibrio cholerae

et al. 1973

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Vibrio cholerae strains with the transmissible fertility factor P contained a supercoiled circular deoxyribonucleic acid (DNA) component amounting to between 2 and 6% of the total DNA obtained from the cells. Such a component was not observed in V. cholerae strains lacking the fertility factor. This supercoiled circular DNA was isolated from P+ cells, and the molecular weight was determined by sedimentation velocity experiments and electron microscopy to be approximately 80 million daltons. These supercoiled circular DNA molecules, which have a guanine plus cytosine (G + C) composition of 42%, were concluded to be the extrachromosomal P factor. It was calculated that there is approximately one copy of the P factor per chromosome. A small amount of,supercoiled circular DNA was occasionally isolated from the Pstrains of V. cholerae. The function of this component, which has a molecular weight of 40 million daltons, is not known. The molecules found in the Pstrains were readily distinguished from the P+ circular molecules by their smaller molecular weight and different G + C composition. Recombination of chromosomal genes inVibrio cholerae depends on the presence of a fertility factor designated P (3, 4). V. cholerae strains containing this factor are referred to as P+ and behave as donor strains. V. cholerae cultures lacking the P factor, which can act as recipients, are termed P-. Although the recombination frequency for chromosomal characters in matings between P+ and Pstrains is low, the P factor itself is transmitted at high frequency from P+ to P-cells. Apparently, the P factor only occasionally causes transfer of chromosomal genes. High-frequency donors, similar to the Hfr types of Escherichia coli in which the fertility factor is chromosomally fixed, have not been found in the V. cholerae mating system.A number of recent studies have established that fertility agents as well as other plasmids exist as autonomously replicating deoxyribonucleic acid (DNA) molecules, functionally and physically distinct from the bacterial chromosome. In this extrachromosomal state, such elements can be isolated as supercoiled circular DNA molecules (13). In the present . 90024. report, we describe the isolation of the P factor from the P+ V. cholerae strains and its characterization as a supercoiled circular DNA molecule.MATERIALS AND METHODS Media. Meat extract agar (MEA) and Difco brain heart infusion (BHI) broth were used for routine cultivation of the V. cholerae strains. The nutrient gelatin agar (NGA) used for the lacunae assay was described by Enomoto (10). Tritium-labeled DNA was isolated from cells grown in the minimal medium for V. cholerae described by Bhaskaran and Rowley (5) to which 1% casein hydrolysate and 20 gCi of 3Hthymidine/ml (specific activity 6.7 Ci/mmole) was added.Bacterial strains. All bacterial strains used are listed in Table 1. The P+ strains were lyophilized to ensure active donor cultures, and a fresh vial was opened for each experiment. Infection of nondonor strains with the P factor. Amounts ...

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“…In a recent attempt (10) to gain some knowledge of the manner in which supernumerary genes are conserved in partially diploid cells, we examined unstable Salmonella typhosa hybrids derived from matings with Escherichia coli K-12 Hfr strains. Some of these S. typhosa hybrids were found to contain supercoiled circular deoxyribonucleic acid (DNA) molecules, whose presence was correlated with the presence of the added E. coli genetic material.…”

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Conservation of Salmonella typhimurium Deoxyribonucleic Acid in Partially Diploid Hybrids of Escherichia coli

et al. 1973

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Partially diploid Escherichia coli K-12 hybrids recovered from mating with a Salmonella typhimurium Hfr strain were found to differ with respect to the manner in which they conserved the added Salmonella deoxyribonucleic acid (DNA). Five of the diploid hybrids examined appeared to maintain the Salmonella DNA as part of a functional F-merogenote; these hybrids were sensitive to the male-specific phage, R-17, responded as males to the female-specific phage, φII, and transferred their inherited Salmonella genetic markers at high frequency in conjugation experiments. Six diploid hybrids were observed which were not sensitive to R-17, and from which the added Salmonella DNA was not transmissible in conjugation tests; nevertheless, these hybrids responded as males to φII, and the Salmonella chromosomal fragments were conserved in them as parts of supercoiled, circular DNA elements. It was concluded that these circular DNA elements were defective F-merogenotes, unable to direct the synthesis of F-pili. Three diploid hybrids were found which were not sensitive to R-17, and which responded as females to φII; no circular DNA was found in them, and it was concluded that their conservation of the Salmonella genetic fragments was accomplished in some manner which did not involve association with F or assumption of the supercoiled circular configuration. Other partially diploid hybrids were observed which appeared similar to these latter three hybrids with regard to their conservation of the Salmonella DNA, but which also contained an infecting F-factor; in these hybrids, both genetic and molecular experiments indicated that the unstably conserved Salmonella DNA was not associated physically with the F-factor.

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Isolation of Circular Deoxyribonucleic Acid from Salmonella typhosa Hybrids Obtained from Matings with Escherichia coli Hfr Donors

Cited by 13 publications

References 21 publications

Characterization of transmissible genetic elements from sucrose-fermenting Salmonella strains

Characterization of transmissible genetic elements from sucrose-fermenting Salmonella strains

Isolation and Characterization of the Fertility Factor P of Vibrio cholerae

Conservation of Salmonella typhimurium Deoxyribonucleic Acid in Partially Diploid Hybrids of Escherichia coli

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