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Streptococcal plasmid pIP501 has a functional oriT site

A Wang1, F L Macrina

  • 1Department of Microbiology and Immunology, Virginia Commonwealth University, Richmond 23298-0678, USA.

Journal of Bacteriology
|August 1, 1995
PubMed
Summary

The study identified a crucial origin of transfer (oriT) site in the gram-positive plasmid pIP501, essential for its conjugal transfer. This site facilitates plasmid mobilization, similar to mechanisms in gram-negative bacteria.

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetics

Background:

  • Gram-positive conjugative plasmids utilize specific DNA sequences for transfer.
  • The plasmid pIP501, found in gram-positive bacteria, was investigated for its transfer mechanisms.

Purpose of the Study:

  • To determine if a putative origin of transfer (oriT) site in plasmid pIP501 is essential for conjugal transfer.
  • To elucidate the role of the oriT site in plasmid mobilization and DNA processing.

Main Methods:

  • Plasmid mobilization assays were performed in Enterococcus faecalis using recombinant plasmids containing pIP501 oriT fragments.
  • Alkaline gel electrophoresis and DNA sequencing were used to analyze DNA cleavage at the oriT site.
  • Conjugal transfer frequencies were quantified to assess the oriT site's function.

Main Results:

  • The oriT region of pIP501 significantly increased the mobilization frequency of a nonconjugative plasmid.
  • A specific single-stranded DNA break was detected at the predicted oriT site.
  • The oriT site was located upstream of genes involved in conjugal transfer.

Conclusions:

  • The identified oriT site in pIP501 is critical for conjugal plasmid mobilization.
  • Specific nicking at the oriT site is a key step in the mobilization process.
  • The mechanism of plasmid mobilization by pIP501 resembles that observed in gram-negative bacteria.

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