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Copy number mutants of the broad-host-range Streptomyces plasmid pMG200
Abstract:
pMG200, isolated from the bacteriocin-releasing strain Streptomyces chrysomallus, was further physically mapped. Variants of S. chrysomallus were isolated which inhibited the parental strain. Two types of plasmids, pMG210 and pMG220, were isolated from these variants, with copy numbers of 10-30 and 300, respectively, compared with 1-3 for pMG200. pMG210 is apparently physically identical to pMG200 but presumably differs at a level not detected by simple restriction mapping; pMG220 is deleted for 1.6 kb. Genes for thiostrepton and viomycin resistance were subcloned from pIJ364 on to pMG200 and a fragment containing the gene for nourseothricin resistance was subcloned on to pMG220. In this way nonessential sites were identified.
Insights
Researchers physically mapped plasmid pMG200 from Streptomyces chrysomallus. New plasmid variants, pMG210 and pMG220, were identified with altered copy numbers and deletions, aiding in nonessential site identification.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Plasmid pMG200 was isolated from the bacteriocin-releasing bacterium Streptomyces chrysomallus.
- Streptomyces chrysomallus variants were identified that inhibited the parental strain, suggesting genetic modifications.
- Characterization of these variants is crucial for understanding plasmid biology and strain improvement.
Purpose of the Study:
- To physically map plasmid pMG200.
- To characterize newly isolated plasmid variants (pMG210, pMG220) from Streptomyces chrysomallus.
- To identify nonessential sites on these plasmids through gene subcloning.
Main Methods:
- Physical mapping of plasmid pMG200 using restriction analysis.
- Isolation and characterization of novel plasmids (pMG210, pMG220) from S. chrysomallus variants.
- Subcloning of antibiotic resistance genes (thiostrepton, viomycin, nourseothricin) onto pMG200 and pMG220.
Main Results:
- Plasmid pMG200 was physically mapped.
- Two new plasmids, pMG210 (physically similar to pMG200) and pMG220 (1.6 kb deletion), were isolated with significantly higher copy numbers.
- Antibiotic resistance genes were successfully subcloned, enabling the identification of nonessential plasmid regions.
Conclusions:
- The study successfully mapped pMG200 and characterized novel variants with altered properties.
- Subcloning antibiotic resistance genes proved effective for identifying nonessential sites on the plasmids.
- These findings contribute to the understanding of plasmid diversity and genetic manipulation in Streptomyces.