An RP4-prime containing a 285 kb fragment of rhizobium meliloti pSym megaplasmid: Structural characterization and utilization for genetic studies of symbiotic functions controlled by pSym
| Autor: | Anne Marie Garnerone, Ilona Dusha, Marie Hélène Renalier, P. Boistard, J. S. Julliot, Betty Terzaghi |
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| Rok vydání: | 1984 |
| Předmět: |
Genetics
Transposable element Cointegrate food and beverages Mutagenesis (molecular biology technique) biochemical phenomena metabolism and nutrition Biology biology.organism_classification medicine.disease_cause chemistry.chemical_compound Plasmid chemistry medicine Rhizobium Homologous recombination Molecular Biology Escherichia coli Prophage |
| Zdroj: | Molecular and General Genetics MGG. 193:17-26 |
| ISSN: | 1432-1874 0026-8925 |
| DOI: | 10.1007/bf00327408 |
| Popis: | We integrated the RP4 plasmid into a selected region of the pSym megaplasmid of Rhizobium meliloti 2011 by homologous recombination between pSym and a cloned fragment of pSym present in the RP4. This cointegrate was used to mobilize into Escherichia coli a Tn5 transposon located on pSym in the vicinity of the site of integration of the RP4. By this technique we obtained a series of RP4-primes that contained large fragments of the pSym megaplasmid and that were most probably generated by IS8 promoted deletions in the RP4-pSym cointegrate. One of them, pGMI42, which carries nitrogenase genes nifD and H as well as nodulation genes, was used for mutagenesis of the corresponding region of pSym after insertion of the Mu prophage into the tet gene. When various (pGMI-42:: Mu)::Tn7 were introduced into R. meliloti 2011 by conjugation, homologous recombination allowed insertion of Tn7 into pSym whereas the pGMI42::Mu was lost due to the suicide effect of Mu. In this way we obtained several symbiotic mutants deficient in either nodulation (Nod-) or nitrogen fixation (Fix-) in association with the host plant Medicago sativa. |
| Databáze: | OpenAIRE |
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