SRA

The study of molecular mechanisms and interactions hinges upon robust model organisms. One of the most important steps in setting up such model systems is establishing a genetic system. Several model organisms are in place for the study of archaeal cell biology, such as the euryarchaeal Haloferax volcanii. However, the established euryarchaeal model organisms tend to have few associated viruses, making the detailed study of virus-host interactions difficult. The environmental strain Hfx. gibbonsii LR2-5 has been shown to be infected by multiple diverse viruses and is closely related to Hfx. volcanii. In this study, this organism was adapted as a new model organism. The most commonly used basis for genetic systems among archaea is a pyrE gene deletion, resulting in uracil auxotrophy. This auxotrophy is then used as a selection marker. Here, gene deletion was performed in Hfx. gibbonsii LR2-5. Furthermore, the genetic toolbox of Hfx. volcanii was tested in Hfx. gibbonsii. The pyrE gene encoded on expression and deletion plasmids of Hfx. volcanii was able to rescue the auxotrophic mutant. Next, protein expression using an established promoter was shown to be fully functional by expressing a fluorescent fusion protein, which showed distinct cellular localization. Finally, gene deletions were generated using pyrE as a selectable marker. Genes could be deleted and a deletion phenotype rescued by expressing the deleted gene from a plasmid. Thus, the versatile genetic toolbox of Hfx. volcanii can now be applied to the study of virus-host interactions in Hfx. gibbonsii and its associated viruses.