Purple non-sulfur photosynthetic bacteria (PNSB) such as R. capsulatus serve as a versatile platform for fundamental studies and various biotechnological applications. In this study, we sought to develop the class II RNA-guided CRISPR/Cas12a system from Francisella novicida for both genome editing and gene down-regulation in R. capsulatus. About 90% editing efficiency was achieved by using CRISPR/Cas12a driven by a strong promoter Ppuc when targeting ccoO or nifH gene. When both genes were simultaneously targeted, the multiplex gene editing efficiency reached >63%. In addition, CRISPRi using deactivated Cas12a was also evaluated using reporter genes gfp and lacZ, and the repression efficiency reached >80%. In summary, our work represents the first report to develop CRISPR/Cas12a mediated genome editing/transcriptional repression in R. capsulatus, which would greatly accelerate PNSB-related researches.
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