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N Willetts

Publications and source records attributed to N Willetts.

At least 37 records · Page 2Linked to original sources
PubMed1980

Cloning the Tra1 region of RP1.

The Tra1 region of RP1 from a derivative with Tn7 inserted into the kanamycin resistance determinant was cloned, using EcoRI, into the multicopy vector plasmid pBR325. For one orientation of the cloned fragment the resultant chimeric plasmid was very frequently lost from the cell, but in the other orientation it was much more stable and also compatible with RP1. Complementation by the stable chimeric plasmid, pED800, of a series of RP1 tra mutants showed that the mutations of all those retaining sensitivity to the P-specific phages PRR1, Pf3, and PR4, or only to PR4, mapped in the Tra1 region, while only 2 out of 20 amber mutations leading to full P-specific phage-resistance did so.

Base Sequence↗
PubMed1979

Investigations of the F conjugation gene traI:traI mutants and lambdatraI transducing phages.

A series of traI point and deletion mutants of Flac, and a traM mutant, were characterised. Complementation tests with an amber Flac traI mutant confirmed their genotypes, and in addition all the traI mutants, but not the traM mutant, were complemented by pRS31 (PSC101 traDI) and EDlambda109 (lambdatraI). Judging from the efficiencies of plating of F-specific phages, none of the mutations affected pilus formation. The traI products of F and of the F-like plasmid R1 were interchangeable with each other but not with that of R100, while the traM product of F could not be replaced by those of R1 or of R100. Neither traI nor traM were needed for conjugal transfer of ColE1. Three lambda transducing phages carrying traI were isolated by in vivo or in vitro techniques, and characterised by genetic complementation tests, by analysis of the fragments produced by restriction endonucleases, and by measurement of heteroduplex molecules. The genetic structures together with the sizes and F coordinates, of the transfer regions carried by the phages were thereby determined. Comparison of the proteins synthesised in UV-irradiated cells by one of the lambdatraI phages with those made by a derivative carrying an amber traI mutation, allowed the traI product to be identified as a protein of molecular weight 174,000. In addition, the molecular weights of the traD (84,000), traS (18,000), and traT (25,000) products made by the lambdatraSTD1 phage EDlambda107 were measured. The possible roles of the traI and traM products in conjugation are discussed.

Coliphages↗
PubMed1979

The exploitation of lambda tra transducing phages in the study of bacterial conjugation.

A set of lambda tra transducing phages have been isolated from an abnormal lysogen in which the lambda prophage was inserted into the traB gene of Flac, and characterised both genetically and physicochemically. The phages were used in several ways to facilitate studies of bacterial conjugation: (a) they aided the identification and mapping of four new transfer genes, traU, traN, traV and traW: (b) they gave information about the location and orientation of the promoters for traM, traJ and traI: (c) a more precise physical map of the transfer region was developed from the lengths of the tra segments that they carried, and (d) the molecular weights of the traM and traJ proteins synthesised by a lambda ptraMJALEK phage were determined.

Bacteriophage lambda↗
PubMed1978

Plasmid cointegrates of Flac and lambda prophage.

Fifteen cointegrates of the plasmid Flac and prophage lambda that had suffered no detectable change in plasmid phenotype were isolated and characterized. The locations of the prophage insertions were determined by genetic analysis of deletion mutants obtained from each cointegrate as survivors of growth at 42 degrees C. In 11 cointegrates, the prophage was inserted between traI and lac, although probably in more than one location; in 3 others, it was on one side or the other of lac; and in 1 it was between lac and pif. Deletions covering all or part of the transfer region, as well as of lac and of pif, were obtained in the course of this analysis. Deletion mutants that had lost all known transfer genes were also oriT, but they retained the capacity to recircularize after transfer. Attempts were made to isolate lambda transducing phages for nearby plasmid genes from the cointegrates, and lambdaptraGD, lambdaptraD, lambdaptraI, and lambdadtraDI phages were obtained.

Coliphages↗
PubMed1978

Properties of lambda transducing bacteriophages carrying R100 plasmid DNA: mercury resistance genes.

Three lambdamer (resistance to Hg2+ and mercurials) transducing phages were prepared from three independent cointegrate isolates of bacteriophage lambda and plasmid R100. DNA heteroduplex and restriction nuclease analyses of the lambdamer DNA showed that all three phages had resulted from lambda insertion at kilobase coordinate 8.6 of plasmid R100, followed by loss of different lengths of lambda DNA and replacement with different lengths of R100 DNA. Two of the lambdamer phages were defective, containing deletions from lambdaatt through the lambdaN gene and into the lambdarex gene; the third, VAlambda14, was an N+ Spi- plaque-forming phage. With VAlambda14, N-dependent transcription of R100 mer from the lambdapL promoter suggested that transcription of mer proceeded in the direction from IS1b toward the sulfonamide resistance determinant (i.e., from a plasmid promoter in restriction nuclease fragment EcoRI-H toward fragment EcoRI-I). Phage-directed protein synthesis in a UV-irradiated lambdaind- lysogen showed the Hg2+-inducible synthesis of three major polypeptides of molecular weights 68,000, 11,500, and 8,500 and three minor ones of molecular weights 54,000, 33,000, and 13,500. The largest of the major polypeptides is identified as the subunit of the mercuric reductase enzyme. The functions of the smaller polypeptides are not known. Hg2+ reductase enzyme assays confirmed the regulation of mer synthesis during phage infection.

Bacterial Proteins↗
PubMed1976

Transfer gene expression during fertility inhibition of the Escherichia coli K12 sex factor F by the I-like plasmid R62.

Further understanding of how the FinQ fertility inhibition system of the I-like plasmid R62 inhibits transfer of the sex factor F has been gained by the use of a genetic assay for individual transfer gene products. The technique involved construction of a series of Flac plasmids carrying suppressible mutations in individual transfer genes together with a FinQ inhibitor-insensitive traQ mutation. The transfer of the Flac double mutants from a strain carrying wild-type Fhis and R62 then indicated the effect of R62-encoded transfer inhibition on the expression of individual F transfer genes. During such inhibition the products of genes traJ, traA, traE, traB and traC were present in quantities sufficient to permit efficient F transfer, whereas the levels of the traF, traH, traG and traD gene products were so reduced as to limit F transfer. These findings and a failure to obtain recombination between traC and traQ mutations suggest that the R62 fertility inhibition system terminates transcription of the transfer operon between traC and traF.

Coliphages↗
PubMed1976

Effect of tra mutations on F factor-specified immunity to lethal zygosis.

Hfr, F+, and F-prime cells are, unlike F- cells, insensitive to an excess of Hfr donor cells, indicating that there is an F factor mediated immunity to lethal zygosis (I1z). Results with Flac episomes carrying traJ, traS or various polar mutations in the tra region indicate that this immunity is independent of surface exclusion, of traJ control, and of all known genes within the tra operon. However, analysis of a series of strains with deletions in the F factor, extending from the right into the tra region, suggests that a gene for immunity to lethal zygosis is located within the tra region. We therefore conclude that I1z is genetically complex, and present a hypothesis to account for these results.

Cell Membrane↗
PubMed1976

Response of Cloudman S91 melanoma cells to melanocytestimulating hormone: enhancement by cytochalasin B.

Cloudmann S 91 mouse melanoma cells treated for 5 days with melanocyte-stimulating hormone (MSH), cytochalasin B (CB), or both, exhibited changes in cell volume, population, nucleation, and pigment production. Cells treated with CB or CB in combination with MSH were eight to nine times larger, rounder, multinucleated, and heavily pigmented. CB alone increased melanin and DNA per nucleus threefold. CB in combination with MSH increased melanin per nucleus 30-fold. Data on DNA per nucleus suggest that CB-treated cells remained in the G phase longer than did control cells. MSH alone caused a reduction in cell population and a fourfold increase in melanin per nucleus.

Animals↗
PubMed1974

Mapping loci for surface exclusion and incompatibility on the F factor of Escherichia coli K-12.

A series of Hfr deletion strains carrying deletions extending different distances into the integrated F factor have been used to map loci for surface exclusion (traS) and for incompatibility (inc) on the Escherichia coli K-12 sex factor F. traS mapped between traG and traD. It forms a part of the large operon, including all the known transfer genes except traJ, and is co-controlled with these. The product of traS is not required for formation of the F pilus. inc mapped between the phi(R) (11) locus and the origin of transfer; it is therefore one of the earliest loci transferred during conjugation.

Chromosome Mapping↗
PubMed1974

Plasmid specificity of the origin of transfer of sex factor F.

The ability of F-like plasmids to promote transfer from the F origin of transfer was determined. Chromosome transfer was measured from plasmid derivatives of RecA(-) Hfr deletion strains which had lost all the F transfer genes but which in some cases retained, and in others had also lost, the origin sequence. ColV2 and ColVBtrp could initiate transfer from the F origin, but R100-1, R1-19, and R538-1 drd could not. These results can be correlated with the plasmid specificity of the traI components of the different plasmid transfer systems, supporting the hypothesis that the origin of transfer is the site of action of the traI product. Most F-like plasmids, including R1-19 and R538-1 drd, could transfer ColE1, consistent with previous findings that the (plasmid-specific) traI product is not necessary for ColE1 transfer by Flac; ColE1 transfer may be initiated by a ColE1-or host-determined product. R100-1 and R136fin(-) could not transfer ColE1 efficiently, apparently because of differences residing in their pilus-forming genes.

Chromosomes, Bacterial↗