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Plasmid. 2014 Nov;76:1-7. doi: 10.1016/j.plasmid.2014.08.001. Epub 2014 Sep 2.

Single-copy vectors for integration at the SaPI1 attachment site for Staphylococcus aureus.

Author information

1
Skirball Institute Program in Molecular Pathogenesis and Departments of Microbiology and Medicine, New York University Medical Center, New York, NY 10016, USA. Electronic address: john.chen@med.nyu.edu.
2
Department of Microbiology, New York University School of Medicine, New York, NY 10016, USA.
3
Skirball Institute Program in Molecular Pathogenesis and Departments of Microbiology and Medicine, New York University Medical Center, New York, NY 10016, USA.
4
Skirball Institute Program in Molecular Pathogenesis and Departments of Microbiology and Medicine, New York University Medical Center, New York, NY 10016, USA. Electronic address: richard.novick@med.nyu.edu.

Abstract

We have previously reported the construction of Staphylococcus aureus integration vectors based on the staphylococcal pathogenicity island 1 (SaPI1) site-specific recombination system. These are shuttle vectors that can be propagated in Escherichia coli, which allows for standard DNA manipulations. In S. aureus, these vectors are temperature-sensitive and can only be maintained at non-permissive (42 °C) temperatures by integrating into the chromosome. However, most S. aureus strains are sensitive to prolonged incubations at higher temperatures and will rapidly accumulate mutations, making the use of temperature-sensitive integration vectors impractical for single-copy applications. Here we describe improved versions of these vectors, which are maintained only in single-copy at the SaPI1 attachment site. In addition, we introduce several additional cassettes containing resistance markers, expanding the versatility of integrant selection, especially in strains that are resistant to multiple antibiotics.

KEYWORDS:

Complementation; Integration; SaPI; Single-copy

PMID:
25192956
PMCID:
PMC4346540
DOI:
10.1016/j.plasmid.2014.08.001
[Indexed for MEDLINE]
Free PMC Article

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