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Genotyping of Staphylococcus aureus by Ribosomal Spacer PCR (RS-PCR)
Published on: November 4, 2016
[Genetic analysis of the host range of the plasmid RSF1010]
1Department of Biology, Liaoning University, Shenyang.
Abstract:
32 host range mutants of pMO58, which contains a 8.0 kb fragment of RSF1010, were screened, using the transposon Tn5 insertion mutagenesis. To judge the genes which affect the host range of plasmid, the Tn5 insertion sites were located with restriction endonuclease mapping. The ability of the plasmid replication in different recipients was checked with DNA transformation. The results further confirmed the barrier to the host range of plasmid was their inability to replicate and maintain themselves in genetically different host. The repA repB and repC genes were necessary for the plasmid replication, but the repC gene may be a positive regulator and its function may be complemented by host gene products.
Insights
Plasmid host range is limited by replication and maintenance in different hosts. Genes repA, repB, and repC are essential for plasmid replication, with repC potentially aided by host factors.
Area of Science:
- Molecular Biology
- Microbial Genetics
- Plasmid Biology
Background:
- Plasmids are extrachromosomal DNA elements crucial for bacterial adaptation and gene transfer.
- Understanding plasmid host range is vital for genetic engineering and controlling microbial populations.
- RSF1010 is a well-characterized plasmid with a broad host range, making it a model system.
Purpose of the Study:
- To identify genes within the pMO58 plasmid fragment of RSF1010 that dictate its host range.
- To elucidate the mechanisms underlying replication barriers in genetically diverse bacterial hosts.
Main Methods:
- Generated host range mutants of pMO58 using Tn5 insertion mutagenesis.
- Located Tn5 insertion sites via restriction endonuclease mapping to identify affected genes.
- Assessed plasmid replication and maintenance in various recipient strains using DNA transformation.
Main Results:
- Confirmed that replication and maintenance deficiencies in genetically distinct hosts limit plasmid host range.
- Identified repA, repB, and repC genes as essential for pMO58 plasmid replication.
- Suggested that the repC gene may act as a positive regulator, potentially complemented by host factors.
Conclusions:
- Plasmid host range is primarily determined by the ability to replicate and persist in a given host.
- The repABC genes are critical determinants of pMO58 replication, highlighting their role in host compatibility.
- Host gene products may play a role in supporting the function of plasmid replication genes like repC.
