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High-frequency, site-specific recombination between lactococcal and pAM beta 1 plasmid DNAs
F Hayes1, C Daly, G F Fitzgerald
1Department of Food Microbiology, University College, Cork, Ireland.
Journal of Bacteriology
|June 1, 1990
Summary
A specific DNA fragment from the Lactococcus lactis plasmid pCI301 significantly increased recombination and mobilization with the enterococcal plasmid pAM beta 1. This site-specific recombination did not affect plasmid replication, and recombinant plasmids reverted to parental forms.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Plasmids are crucial mobile genetic elements in bacteria, facilitating gene transfer and adaptation.
- Understanding plasmid recombination is key to controlling genetic exchange and stability in microbial populations.
- Lactococcus lactis and Enterococcus species are important in food and clinical settings, respectively.
Purpose of the Study:
- To investigate in vivo recombination between Lactococcus lactis plasmid pCI301 and Enterococcus plasmid pAM beta 1.
- To identify and characterize the recombination site on pCI301 responsible for mediating interplasmid recombination.
- To analyze the specificity and consequences of this recombination event on plasmid replication and stability.
Main Methods:
- Cloning of a pCI301 fragment into a Lactococcus lactis-Escherichia coli shuttle vector.
- Mobilization and recombination assays to assess the frequency of interplasmid events.
- Site-directed mutagenesis and sequence analysis to determine recombination specificity.
- Analysis of recombinant plasmid replication and stability.
Main Results:
- A specific fragment from pCI301 greatly enhanced mobilization and recombination with pAM beta 1.
- Recombination was site- and orientation-specific on both pCI301 and pAM beta 1.
- Recombination occurred within the replication region of pAM beta 1 but did not impair its replication.
- Resolution of recombinant plasmids resulted in the regeneration of parental plasmid forms.
Conclusions:
- A distinct recombination site on pCI301 facilitates efficient, site-specific recombination with pAM beta 1.
- This recombination mechanism does not disrupt essential plasmid replication functions.
- The observed recombination and subsequent resolution suggest a potential pathway for plasmid maintenance or exchange in mixed bacterial populations.