Use of CRISPR interference for efficient and rapid gene inactivation in < em > Fusobacterium nucleatum < /em >

Appl Environ Microbiol. 2024 Jan 8:e0166523. doi: 10.1128/aem.01665-23. Online ahead of print.ABSTRACTHow can we effectively investigate the gene functions in Fusobacterium nucleatum, given the dual challenges of gene inactivation and the inherent genetic resistance of many strains? Traditional methods have been cumbersome and often inadequate. Addressing this, our work introduces a novel inducible CRISPR interference (CRISPRi) system in which dCas9 expression is controlled at the translation level by a theophylline-responsive riboswitch unit, and single-guide RNA expression is driven by the robust, constitutive rpsJ promoter. This approach simplifies gene inactivation in the model organism (ATCC 23726) and extends its application to previously considered genetically intractable strains like CTI-2 and Fusobacterium periodonticum. With CRISPRi's potential, it is a pivotal tool for in-depth genetic studies into fusobacterial pathogenesis, potentially unlocking targeted therapeutic strategies.PMID:38185820 | DOI:10.1128/aem.01665-23
Source: Applied and Environmental Microbiology - Category: Microbiology Authors: Source Type: research