†The first three authors contributed equally to this work.
A genome-wide strategy for the identification of essential genes in Staphylococcus aureus
Article first published online: 25 APR 2002
Volume 43, Issue 6, pages 1387–1400, March 2002
How to Cite
Forsyth, R. A., Haselbeck, R. J., Ohlsen, K. L., Yamamoto, R. T., Xu, H., Trawick, J. D., Wall, D., Wang, L., Brown-Driver, V., Froelich, J. M., C., K. G., King, P., McCarthy, M., Malone, C., Misiner, B., Robbins, D., Tan, Z., Zhu, Z.-y., Carr, G., Mosca, D. A., Zamudio, C., Foulkes, J. G. and Zyskind, J. W. (2002), A genome-wide strategy for the identification of essential genes in Staphylococcus aureus. Molecular Microbiology, 43: 1387–1400. doi: 10.1046/j.1365-2958.2002.02832.x
- Issue published online: 25 APR 2002
- Article first published online: 25 APR 2002
To address the need for new approaches to antibiotic drug development, we have identified a large number of essential genes for the bacterial pathogen, Staphylococcus aureus, using a rapid shotgun antisense RNA method. Staphylococcus aureus chromosomal DNA fragments were cloned into a xylose-inducible expression plasmid and transformed into S. aureus. Homology comparisons between 658 S. aureus genes identified in this particular antisense screen and the Mycoplasma genitalium genome, which contains 517 genes in total, yielded 168 conserved genes, many of which appear to be essential in M. genitalium and other bacteria. Examples are presented in which expression of an antisense RNA specifically reduces its cognate mRNA. A cell-based, drug-screening assay is also described, wherein expression of an antisense RNA confers specific sensitivity to compounds targeting that gene product. This approach enables facile assay development for high throughput screening for any essential gene, independent of its biochemical function, thereby greatly facilitating the search for new antibiotics.