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A R Wyman

Publications and source records attributed to A R Wyman.

7 recordsLinked to original sources
PubMed1986

Host/vector interactions which affect the viability of recombinant phage lambda clones.

A class of recombinant phage lambda clones are recovered from human genomic libraries on Escherichia coli recB21 recC22 sbcB15 cells, which fail to form plaques on wild-type cells. We report experiments which address the mechanism of this inhibition. The introduction of the recombination-stimulating sequence chi into one such clone allows growth of this phage on Rec+ cells. In addition, the insertion of lambda gam+ gene into a rec+-inhibited clone results in the ability of the phage to form plaques on wild-type cells. Since lambda Gam protein is an inhibitor of host RecBC enzyme, we tested a collection of such phage for growth on a variety of hosts altered in RecBC function. Host permissiveness correlated with the inactivation of the RecBC nucleolytic activities and not with the recombinational activities. These observations suggest that the inserted DNA sequences of these phage limit the production of packageable chromosomes. This conclusion is easily reconciled with our current knowledge of the interaction of the host recombination systems with lambda replication and encapsidation. Based on these experiments we have constructed strains, both recombination-proficient and recombination-deficient, which serve as improved hosts for the recovery of genomic sequences which are otherwise inhibitory to the growth of phage lambda.

Bacterial Proteins↗
PubMed1986

Factors which equalize the representation of genome segments in recombinant libraries.

Genomic segments which contain inverted repetitions longer than 300 bp are frequently lost from recombinant libraries grown on rec+ hosts. We have found that 9% of phage lambda clones that contain 15-20-kb insertions of human or Drosophila DNA are inhibited on rec+ hosts and as a result will become under-represented in amplified genomic libraries. We have therefore examined several factors of both host and vector origin which affect the fidelity of representation of genomic sequences in recombinant DNA libraries constructed in bacteriophage lambda vectors. This loss may be diminished if the vector carries either a chi element or a functional gam gene. The most successful approach, however, involves using a host with mutations in recB, recC, and sbcB, or in recD. We have shown that recombinant clones which require such mutant hosts for growth are somewhat more likely to contain DNA derived from loci in the genome which are polymorphic than are clones recovered on conventional hosts.

Animals↗
PubMed1985

Propagation of some human DNA sequences in bacteriophage lambda vectors requires mutant Escherichia coli hosts.

The growth of clones of human genomic DNA fragments in a bacteriophage lambda vector has been examined in a number of different Escherichia coli hosts. A large proportion (8.9%) of the phages carrying different fragments of the human genome fail to grow on standard rec+ hosts but will grow on hosts carrying mutations in the recB, recC, and sbcB genes. Heteroduplex analysis in the electron microscope of DNA from four of these phages revealed substantial secondary structure, including snap-back regions 200-500 base pairs in length. Such structures were not found in phages from the same DNA library that grow in rec+ hosts. These results are interpreted in the light of prior observations [Leach, D.R.F. & Stahl, F. (1983) Nature (London) 305, 448-451] showing that inverted repetitions cloned in phage lambda can be propagated in recB recC sbcB hosts but not in rec+ hosts.

Bacteriophage lambda↗
PubMed1979

Structural and image analysis of a crystalline layer from dipteran eggshell.

A crystalline layer has been identified as a constituent of the eggshell in the dipteran Drosophila melanogaster. This 400A thick intermediate chorionic layer (ICL) is composed of eight 50A thick sublayers and lies between the vitelline membrane and the endochorion. Whole mount views of isolated ICL after negative staining reveal P2 planar periodicity which, when analyzed further by optical diffraction and filtering, showed 1st (100A), 2nd, 3rd and 4th order reflections.

Animals↗