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Biomedical subjects

W J Brammar

Publications and source records attributed to W J Brammar.

9 recordsLinked to original sources
PubMed1979

In vitro insertion of the lambda attachment site into the plasmid RP4.

The region of the phage lambda chromosome containing the attachment site (P.P') and the genes int and xis, excised by the action of endonuclease R.EcoRI, has been inserted into the unique site for that enzyme on the promiscuous conjugative plasmid, RP4, generating the recombinant plasmid RP4att lambda. Transformants containing the hybrid plasmid were recognised by their ability to allow efficient lysogenization by phage lambda b2 (Weil and Signer, 1968; Echols et al., 1968) containing the mutant attachment site delta. P'. The construction and properties of the hybrid plasmid RP4att lambda are described.

Attachment Sites, Microbiological↗
PubMed1979

Aberrant immunity behaviour of hybrid lambda imm21 phages containing the DNA of ColE1-type plasmids.

Hybrid lambda and lambda imm21 bacteriophages carrying various ColE1-type plasmids have been constructed in vitro. The lambda imm21/plasmid recombinants display aberrant immunity behaviour, giving clear plaques under conditions where the parental phages give turbid ones and being able to grow on homoimmune lysogens. lambda imm lambda/plasmid recombinants show no such unusual behaviour. Studies with hybrids of a lambda imm21 cITS phage carrying pMB9 DNA showed the operation of the plasmid's replication system to be the basic cause of the aberrant immunity behaviour. The plasmid replication system could act as a complete alternative to the phage system during vegetative phage growth. The probable reason that lambda imm21 phages show such altered phenotypes when carrying a functional plasmid replication origin, whereas lambda imm lambda and lambda imm434 (Mukai et al., 1978) phages do not, is the relative ease of titration of the phage 21 repressor to allow transcription from pR21. Various uses are considered for the altered phenotypic behaviour of lambda imm21/ColE1-type plasmid hybrids.

Bacteriocin Plasmids↗
PubMed1977

Lambdoid phages that simplify the recovery of in vitro recombinants.

Derivatives of phage lambda are described for use as vectors for fragments of DNA generated with the HindIII and EcoRI restriction enzymes. With some vectors, hybrid molecules are recognised by a change from a turbid to a clear plaque morphology resulting from the insertion of a fragment of DNA into the lambda gene coding for the phage regressor. Other vectors contain a central, replaceable fragment of DNA which imparts a readily recognisable phenotype. This central fragment may include either a gene for a mutant transfer RNA (suppressor) or a part of the lacZ gene of E. coli able to complement a lacZ host. The appropriate lacZ host and indicator plates permit the ready distinction between recombinant and vector phages by the colour of the plaques.

Coliphages↗
PubMed1976

The use of specialised transducing phages in the amplification of enzyme production.

Two types of lambdatrp phages have been used as model systems to investigate ways of optimising the expression of bacterial genes from transducing phage genomes. Excellent yields of trp enzymes were achieved by infecting a trpR- host with Q- or Q-S- derivatives of lambdatrpAM1, which expresses its trp genese exclusively from the trp promoter. The five trp geneproducts constituted more than 50% of the total soluble protein of infected cells under these conditions, and an even higher proportion of the protein synthesized after infection. In a trpR+ host, phage DNA replication was easily able to override tryptophan-mediated repression by titration of the trp promoter were equally productive, while having the advantage of being much simpler to construct and propagate. lambdatrp phages lacking the trp promoter were used to investigate ways of optimising gene expression initiated at the phage promoter, PL. Though very powerful, the latter promoter is more difficult to harness then the trp promoter. Derepression of transcription from PL by the use of cro- mutations is accompanied by poor replication of transducing phage DNA. Attempts to circumvent this difficulty using virulent of cro,cII double mutants have not been successful. Nevertheless, cells infected with a lambdatrp phage expressing its trp genes exclusively from PL made up to 16 per cent of their protein as trp gene-products.

Anthranilate Synthase↗
PubMed1976

Control of cI gene expression in bacteriophage lambda imm434, studied in an immunity/trp fusion made in vitro.

The trp genes of a lambda imm434 trp-transducing phage have been fused to the immunity region by deletion, in vitro, of the DNA between two targets for the restriction enzyme R.EcoRI. The resulting phage has been used to study the control of expression of the cI gene in vivo. The constitutive rate of expression of the cI gene is between 2 and 5% of the maximally stimulated rate. The products of the cII and cIII genes enhance expression of cI on infection of a sensitive host. The requirement for the cII product is more stringent than that for the cIII protein. The phage 434 repressor present in a 434-immune cell stimulates the rate of cI expression from a superinfecting homoimmune phage about fifteen-fold. This result strongly suggests that repressor stimulates its own synthesis by a direct effect on transcription of the cI gene.

Chromosome Mapping↗
PubMed1976

The construction in vitro of transducing derivatives of phage lambda.

Methods are described for the construction of plaque-forming, transducing derivatives of phage lambda, using appropirate receptor genomes and fragments of DNA generated by the restriction enzymes endo R.EcoRI and endo R.HindIII. The general properties of the transducing derivatives are described and discussed. Plaque-forming phages carrying the E. coli trp, his, cysB, thyA, supD, supE, sup F, hsd, tna and lig genes have been isolated.

Chromosomes, Bacterial↗