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Systematic insertional mutagenesis of a streptomycete genome: a link between osmoadaptation and antibiotic production

Bishop, A. and Fielding, S. and Dyson, P.J. and Herron, P.R. (2004) Systematic insertional mutagenesis of a streptomycete genome: a link between osmoadaptation and antibiotic production. Genome Research, 14. pp. 893-900. ISSN 1088-9051

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Abstract

The model organism Streptomyces coelicolor represents a genus that produces a vast range of bioactive secondary metabolites. We describe a versatile procedure for systematic and comprehensive mutagenesis of the S. coelicolor genome. The high-throughput process relies on in vitro transposon mutagenesis of an ordered cosmid library; mutagenized cosmids with fully characterized insertions are then transferred by intergeneric conjugation into Streptomyces, where gene replacement is selected. The procedure can yield insertions in upward of 90% of genes, and its application to the entire genome is underway. The methodology could be applied to many other organisms that can receive DNA via RK2/RP4-mediated intergeneric conjugation. The system permits introduction of mutations into different genetic backgrounds and qualitative measurement of the expression of disrupted genes as demonstrated in the analysis of a hybrid histidine kinase and response regulator gene pair, osaAB, involved in osmoadaptation in Streptomyces. The independently transcribed response regulator gene, osaB, is essential for osmoadaptation; when grown with supplementary osmolyte, an osaB mutant cannot erect aerial hyphae and produces up to fivefold greater antibiotic yields than the wild-type strain.

Item type: Article
ID code: 37861
Keywords: antibiotic production , insertional mutagenesis , streptomycete genome , osmoadaptation, Pharmacy and materia medica, Genetics, Genetics(clinical)
Subjects: Medicine > Pharmacy and materia medica
Department: Faculty of Science > Strathclyde Institute of Pharmacy and Biomedical Sciences
Related URLs:
    Depositing user: Pure Administrator
    Date Deposited: 24 Feb 2012 14:07
    Last modified: 04 Sep 2014 19:28
    URI: http://strathprints.strath.ac.uk/id/eprint/37861

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