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Isolation and characterization of an R-prime plasmid from Rhizobium meliloti
Abstract:
Using a simple enrichment procedure, we isolated an R-prime derivative of plasmid R68.45 carrying a 17.8-megadalton segment of the Rhizobium meliloti 41 chromosome. The chromosomal segment carried on this plasmid (pGY1) includes the markers cys-24+, cys-46+, and att16-3. Plasmid pGY1 mobilized the chromosome in a polarized way starting from the region of homology, but cannot promote chromosome transfer from other sites. The att16-3 site on pGY1 allowed the integration of phage 16-3 into pGY1, and a composite plasmid of 91.8 megadaltons was formed. This vector (pGY2) is suitable for the introduction of Rhizobium bacteriophage 16-3 into other gram-negative bacteria.
Insights
Researchers created a novel R-prime plasmid, pGY1, containing a Rhizobium meliloti chromosomal segment. This plasmid facilitates polarized chromosome mobilization and, as pGY2, enables Rhizobium bacteriophage 16-3 introduction into gram-negative bacteria.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Plasmid R68.45 is a well-characterized conjugative plasmid.
- Rhizobium meliloti is a key nitrogen-fixing bacterium.
Purpose of the Study:
- To construct a novel R-prime plasmid carrying specific Rhizobium meliloti chromosomal markers.
- To evaluate the chromosome mobilization capabilities of the constructed plasmid.
- To develop a vector for introducing Rhizobium bacteriophage into other bacteria.
Main Methods:
- Enrichment procedure for R-prime isolation.
- Plasmid construction and characterization.
- Conjugation experiments for chromosome mobilization.
- Bacteriophage integration and vector formation.
Main Results:
- Isolation of R-prime plasmid pGY1 with a 17.8-megadalton Rhizobium meliloti chromosomal segment (cys-24+, cys-46+, att16-3).
- pGY1 demonstrated polarized chromosome mobilization from the region of homology.
- Integration of phage 16-3 into pGY1 formed a composite plasmid, pGY2 (91.8 megadaltons).
Conclusions:
- The R-prime plasmid pGY1 is a functional tool for Rhizobium meliloti genetic studies.
- The composite plasmid pGY2 serves as a viable vector for transferring Rhizobium bacteriophage 16-3 to other gram-negative bacteria.