Control of expression of a cloned yeast (Saccharomyces cerevisiae) gene (trp5) by a bacterial insertion element (IS2) [recombinant DNA]

A hybrid ColE1 plasmid [pYe(trp5)1], containing a yeast DNA segment that complements auxotrophic point mutations and deletions in the Escherichia coli tryptophan synthetase gene (trpAB), has been isolated. Expression of the yeast tryptophan synthetase activity from the cloned yeast gene (trp5) is re...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1978-12, Vol.75 (12), p.6172-6176
Main Authors: Walz, A, Ratzkin, B, Carbon, J
Format: Article
Language:English
Subjects:
Bacteriocin Plasmids
Bacteriophages
Clone Cells - metabolism
DNA
DNA, Recombinant
Enzymes
Escherichia coli - genetics
Gels
Genes
Genetic vectors
Hybridity
Operon
Plasmids
Saccharomyces cerevisiae - genetics
Translocation, Genetic
Transposons
Tryptophan - genetics
Yeasts
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Summary:A hybrid ColE1 plasmid [pYe(trp5)1], containing a yeast DNA segment that complements auxotrophic point mutations and deletions in the Escherichia coli tryptophan synthetase gene (trpAB), has been isolated. Expression of the yeast tryptophan synthetase activity from the cloned yeast gene (trp5) is relatively inefficient in E. coli, as measured by growth rates of trpAB/pYe(trp5)1 strains on minimal media lacking tryptophan and by enzyme assays. Faster growing variants occur spontaneously at a frequency of one in 104-105 cells plated and produce higher levels of the yeast enzyme. Plasmid DNA [pYe(trp5)2] from one of these variants was shown to contain a DNA insertion (1.3 kilobase pairs) in the cloned yeast DNA segment in relatively close proximity to the trp5 gene. This DNA insert was identified as a bacterial IS2 element, which carries a promoter for RNA transcription when inserted in the proper orientation. The spontaneous integration of a bacterial DNA insertion element into cloned eukaryotic DNA can result in more efficient expression of the foreign gene.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.75.12.6172