Highlighted paper selected by editor-in-chief: Shuttle vectors derived from pN315 for study of essential genes in Staphylococcus aureus

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  • Matsuo Miki
    Laboratory of Microbiology, Graduate School of Pharmaceutical Sciences, The University of Tokyo
  • Kurokawa Kenji
    Laboratory of Microbiology, Graduate School of Pharmaceutical Sciences, The University of Tokyo National Research Laboratory of Defense Proteins, College of Pharmacy, Pusan National University
  • Lee Bok-Luel
    National Research Laboratory of Defense Proteins, College of Pharmacy, Pusan National University
  • Sekimizu Kazuhisa
    Laboratory of Microbiology, Graduate School of Pharmaceutical Sciences, The University of Tokyo

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  • Shuttle Vectors Derived from pN315 for Study of Essential Genes in Staphylococcus aureus

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Using the par to rep region of the 24653 bp plasmid pN315, which is present in Staphylococcus aureus strain N315, we constructed three vectors that can be shuttled between Escherichia coli and S. aureus and maintained stably in S. aureus. Due to plasmid incompatibility, the resident plasmid in S. aureus cells can be replaced via transformation with an entering plasmid, which carries a different drug resistance gene. To evaluate the applicability of this plasmid-based approach for identifying genes essential for S. aureus cell growth, the chromosomal mraY gene, which is involved in peptidoglycan biosynthesis, was deleted in cells harboring a resident plasmid with an intact mraY gene. The resultant disruptant was then transformed with an empty vector. Cells with a chromosomal mraY deletion but lacking the plasmid supplying mraY could not be recovered, suggesting that mraY is indispensable for staphylococcal cell growth or viability. In contrast, other two genes were shown to be dispensable by this system. Thus, the pN315-based plasmids appear to be useful for studying genes essential for S. aureus cell growth.

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