Cloning of a Streptomyces clavuligerus DNA fragment encoding the cephalosporin 7 alpha-hydroxylase and its expression in Streptomyces lividans
| Autor: | P. J. Barker, Gilberto A Hintermann, A. L. Demain, Xinfa Xiao, F. Foor, Jacqueline Piret, A. Häusler |
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| Rok vydání: | 1993 |
| Předmět: |
DNA
Bacterial Molecular Sequence Data Oligonucleotides Gene Expression Streptomyces clavuligerus Molecular cloning Polymerase Chain Reaction Streptomyces Mixed Function Oxygenases Nucleic acid thermodynamics Restriction map Bacteriophages Pharmacology (medical) Amino Acid Sequence Cloning Molecular Chromatography High Pressure Liquid Southern blot Recombination Genetic Pharmacology biology Hybridization probe Nucleic Acid Hybridization biology.organism_classification Molecular biology Blotting Southern Infectious Diseases Primer (molecular biology) DNA Probes Research Article |
| Zdroj: | Antimicrobial Agents and Chemotherapy. 37:84-88 |
| ISSN: | 1098-6596 0066-4804 |
| Popis: | A 26-mer DNA probe was designed from N-terminal sequence data for the cephalosporin 7 alpha-hydroxylase (CH) of Streptomyces clavuligerus NRRL 3585 and used to screen a DNA library from this organism. The library was constructed in the lambda GEM-11 phage system. After plaque purification and reprobing, positive recombinant phages were chosen for further analysis. Characterization of the cloned DNA by restriction mapping and Southern hybridization showed that a 1.5-kb SalI fragment hybridized to the probe. Polymerase chain reaction assays using this fragment as a template and the probe as a primer indicated that the fragment carries the entire putative CH gene (cmcI). This was confirmed through the expression of CH enzymatic activity when the fragment was introduced into Streptomyces lividans. A putative beta-lactamase activity was detected in S. lividans. |
| Databáze: | OpenAIRE |
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