Mini-Mulac transposons with broad-host-range origins of conjugal transfer and replication designed for gene regulation studies in Rhizobiaceae.
Novel mini-Mu derivatives were constructed, carrying a truncated lacZYA operon fused to the terminal 117 bp of the Mu S-end, for the isolation of translational lac fusions by mini-Mu-mediated insertion mutagenesis. Different selectable markers (chloramphenicol resistance; gentamycin resistance) were introduced to allow selection for mini-Mu insertions in different replicons and bacterial strains. A mini-Mulac derivative carrying the site for conjugal transfer of plasmid RP4 (oriT) and the origin of replication of the Agrobacterium rhizogenes Ri plasmid (oriRiHRI) was constructed to enable one-step lac-fusion mutagenesis of cloned (plasmid-borne) regions in Escherichia coli and efficient conjugal transfer of gene fusions to to a variety of Gram-negative bacteria. The conjugation frequency, stability and copy number of replicons carrying mini-Mulac derivatives with oriT and oriRiHRI in members of the Rhizobiaceae such as Rhizobium meliloti, Azorhizobium caulinodans ORS571 and Agrobacterium tumefaciens C58 was examined.
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Abteilung Genetische Grundlagen der Pflanzenzüchtung, Max-Planck-Institut für Zuchtungsforschung, Köln F.R.G.
Schell
J
J
de Bruijn
F J
FJ
D010583:Bacteriophage mu / Q000235:genetics*; D003227:Conjugation, Genetic; D004251:DNA Transposable Elements; D005798:Genes, Bacterial; D005822:Genetic Vectors; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012093:Replicon; D012230:Rhizobiaceae / Q000235:genetics*
D004251:DNA Transposable Elements; D011993:Recombinant Fusion Proteins