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

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. au...

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Bibliographic Details
Published in:Biological & Pharmaceutical Bulletin 2010/02/01, Vol.33(2), pp.198-203
Main Authors: Matsuo, Miki, Kurokawa, Kenji, Lee, Bok-Luel, Sekimizu, Kazuhisa
Format: Article
Language:English
Subjects:
Bacterial Proteins - genetics
Escherichia coli
Genes, Bacterial
Genes, Essential
Genes, Lethal
Genetic Vectors - chemical synthesis
Genetic Vectors - genetics
incompatibility
mraY
plasmid shuffling
Plasmids - chemical synthesis
Plasmids - genetics
pN315
Staphylococcus aureus
Staphylococcus aureus - genetics
Staphylococcus aureus - growth & development
Transferases - genetics
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Summary: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.
ISSN:0918-6158
1347-5215
DOI:10.1248/bpb.33.198