Development of inducible promoter and CRISPRi plasmids functional in <i>Rickettsia rickettsii</i>

This work expands upon the genetic toolbox available for <i>R. rickettsii</i>. We describe both an inducible promoter and CRISPRi system compatible with <i>Rickettsia</i>, which may provide key instruments for the development of further tools. The development of an inducible promoter system allows for the overexpression of genes, which might be toxic when expressed constitutively. The CRISPRi system enables the ability to knock down genes with specificity, and critically, genes that may be essential and could not otherwise be knocked out. These developments may provide the foundation for unlocking genetic tools for other pathogens of the order Rickettsiales, such as the <i>Anaplasma</i>, <i>Orientia</i>, and <i>Ehrlichia</i> for which there are currently no inducible promoters or CRISPRi platforms.