Antisense RNA technology has been used effectively to downregulate gene expression in a variety of bacterial systems. Regulated antisense RNA strategy provides an important approach to identify and characterize essential genes critical to bacterial growth in vitro and in vivo. This strategy allows selective genes to be turned on or off and to be expressed at certain levels. The availability of the Staphylococcus aureus (S. aureus) genome sequence makes it feasible to generate a gene-specific antisense RNA library. The combination of regulated antisense RNA technology and the gene-specific antisense RNA library allows for genome-wide analyses of functions of staphylococcal gene products for growth in culture and survival, during infection.