Interruption-deficient mutants of bacteriophage T5 I. Isolation and general properties

Rogers, Stephen L.; Godwin, E A; Shinosky, E S; Rhoades, Marc

doi:10.1128/jvi.29.2.716-725.1979

Cited by 24 publications

(25 citation statements)

References 27 publications

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“…We used DNA extracted from TSHA23 to avoid the effects of nonrandom breaks in DNA during our manipulations. This DNA is free of genetically predetermined breaks (26), but otherwise appears to be indistinguishable from wild-type T5 DNA (unpublished data).…”

Section: Resultsmentioning

confidence: 89%

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Physical locus of the DNA polymerase gene and genetic maps of bacteriophage T5 mutants

Fujimura

Tavtigian

Choy

et al. 1985

J Virol

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Segments of DNA that contained the DNA polymerase gene of bacteriophage T5 were isolated. The physical locus of the gene was identified by transforming Escherichia coli with purified DNA fragments generated by restriction enzyme digestions, and the transformed cells were used to rescue amber mutants of T5 with mutations in the gene for DNA polymerase. The method is applicable to any other gene that has mutations with low reversion frequencies. We studied the following mutations of the T5 DNA polymerase gene, reading from left to right by the standard convention (D. J. McCorquodale, Crit. Rev. Microbiol. 4:101-159, 1975): D7, D8, aml, ts5E-ts53, am6, and D9. These loci were found to reside within three pieces of DNA with a total length of 3,600 base pairs. Because the structural gene for T5 DNA polymerase is estimated to be 2,600 base pairs long, the whole structural gene may reside in these segments. These are located 58.3 to 61.3% of the distance from the left end of the DNA. The left-end piece of the DNA (1,100 base pairs) containing the polymerase gene has loci D7 and D8, and the right-end piece (1,600 base pairs) has locus D9, according to the results of the transformation assay. These results are consistent with the genetic map.Plaskon CTFE2300 powder and Adogen 464 were the generous gifts of the late G. D. Novelli. A column (90 by 0.6 cm) was equilibrated with 1 M NaCl in TE buffer (10 mM Tris hydrochloride [pH 7.6], 1 mM EDTA) and then with 0.5 M NaCl in TE buffer. A sample (240 ,ug) was adjusted to 0.5 M NaCl and loaded through a pump. The column was washed with 400 ml of TE buffer containing an increasing NaCl concentration (0.55 to 0.6 M) at a pressure of 300 to 400 lb/in2. The DNA was eluted with a linear gradient made with 495 on July 6, 2020 by guest

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Section: Resultsmentioning

confidence: 89%

“…Mutant strain T5HA23 has none of the single-* Corresponding author. strand breaks that are present in the wild type (26). The Escherichia coli Su+ strain was that used by Hendrickson and McCorquodale (13); E. coli F was used as the Su-host and has been described previously (11).…”

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confidence: 99%

Physical locus of the DNA polymerase gene and genetic maps of bacteriophage T5 mutants

Fujimura

Tavtigian

Choy

et al. 1985

J Virol

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“…The GCGC sequences in the nick regions are nested in 9-nucleotide (nt) sequences that differ slightly from each other but can be found only in the regions of nicks (Table S3). Single-stranded nicks of conserved 5= end sequence in one DNA strand of virion DNA are also a characteristic feature of T5 phage (31,32), its close relative BF23 (31), KMV-like viruses (33), and Pseudomonas putida phage PpG1, an evolutionarily divergent member of the LUZ24-like phage group (34). The deduced consensus sequence of nick sites in PR1 DNA [5=-(R/G)CGCRNDR] resembles that of major nick sites in T5 DNA [5=-(R/G)CGCRGG].…”

Section: Importancementioning

confidence: 99%

“…Alternatively, the 3= ends of nicked strands may serve to initiate recombinational replication or repair. The lack of obvious difference between the development of T5 nick-free mutants and wild-type phages in a laboratory (24,32) suggests that there may be an alternative pathway for the process in which the nicks are involved or that the nicks are important under conditions that have not yet been studied.…”

Section: Importancementioning

confidence: 99%

A Lytic Providencia rettgeri Virus of Potential Therapeutic Value Is a Deep-Branching Member of the T5virus Genus

Oliveira

Pinto

Hendrix

et al. 2017

Appl Environ Microbiol

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is emerging as a new opportunistic pathogen with high antibiotic resistance. The need to find alternative methods to control antibiotic-resistant bacteria and the recent advances in phage therapy motivate the search for new phages able to infect spp. This study describes the isolation and characterization of an obligatory lytic phage, vB_PreS_PR1 (PR1), with therapeutic potential against drug-resistant PR1 is a siphovirus. Its virion DNA size (118,537 bp), transcriptional organization, terminal repeats (10,461 bp), and nicks in the 3'-to-5' strand are similar to those of phage T5. However, sequence similarities of PR1 to phages of the genus at the DNA and protein levels are limited, suggesting that it belongs to a new species within the family. PR1 exhibits the ability to kill antibiotic-resistant strains, is highly specific to the species, and did not present known genomic markers indicating a temperate lifestyle. The lack of homologies between its proteins and proteins of the only other sequenced prophage, Redjac, suggests that these two phages evolved separately and may target different host proteins. The alarming increase in the number of bacteria resistant to antibiotics has been observed worldwide. This is particularly true for Gram-negative bacteria. For certain of their strains, no effective antibiotics are available. sp. has been a neglected pathogen but is emerging as a multidrug-resistant bacterium. This has revived interest in bacteriophages as alternative therapeutic agents against this bacterium. We describe the morphological, physiological, and genomic characterization of a novel lytic virus, PR1, which is able to kill drug-resistant clinical isolates. Genomic and phylogenetic analyses indicate that PR1 is a distant relative of genus representatives. The lack of known virulence- or temperate lifestyle-associated genes in the genome of PR1 makes this phage a potential candidate for therapeutic use. Analysis of its genome also improves our knowledge of the ecology and diversity of T5-like siphoviruses, providing a new link for evolutionary studies of this phage group.

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“…Denaturation of T5 DNA results in the formation of over 40 discrete single-stranded fragments that can be resolved by agarose gel electrophoresis (5). As recently demonstrated by Rogers et al (10), the DNA content of single T5 plaques can be analyzed in a similar manner. T5 deletion mutants can therefore be recognized, as was the case for interruption-deficient mutants, by alterations in the pattern of single-chain fragments.…”

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confidence: 93%

New deletion mutant of bacteriophage T5

Rhoades¹,

Schwartz²,

Wahl³

1980

J Virol

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Interruption-deficient mutants of bacteriophage T5 I. Isolation and general properties

Cited by 24 publications

References 27 publications

Physical locus of the DNA polymerase gene and genetic maps of bacteriophage T5 mutants

Physical locus of the DNA polymerase gene and genetic maps of bacteriophage T5 mutants

A Lytic Providencia rettgeri Virus of Potential Therapeutic Value Is a Deep-Branching Member of the T5virus Genus

New deletion mutant of bacteriophage T5

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