Molecular Biology of the Staphylococci
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Cites methods from "Molecular Biology of the Staphyloco..."
...RN4220 carrying either an rplQ antisense clone (S1–760 A) or a lig antisense clone (S1–396 A) were grown to mid-log phase and diluted 1:10 into prewarmed Luria–Bertani (LB) broth....
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...The genomic library was amplified by passage through Escherichia coli to provide sufficient amounts of DNA, and DNA from the pooled library was then transformed into S. aureus strain RN4220....
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...Cell-based assay for target-specific inhibitors An overnight culture of RN4220 carrying plasmid S1- © 2002 Blackwell Science Ltd, Molecular Microbiology, 43, 1387–1400 1941 expressing antisense to the fab gene encoding bketoacyl-acyl carrier protein synthase was inoculated into fresh LB plus 34 mg ml–1 of chloramphenicol, and incubated with shaking at 37∞C. Exponential cultures were diluted to a final OD600 of 0.002 into two flasks of the same medium containing 0 and 12 mM xylose....
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...Staphylococcus aureus strains used include RN450 and RN4220 (Novick, 1990) and E. coli strains, DH5a and XL1Blue, which were obtained from Gibco-BRL and Stratagene respectively....
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...This plasmid library was electroporated into RN4220 to generate approximately 315 000 transformants that were recovered on LBG + 15 mg ml–1 chloramphenicol agar plates....
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