Developing a tetO/TetR system in Neurospora crassa

Tinh-Suong Nguyen (1) , Eugene Gladyshev (1)

1 Epigénomique fongique - Fungal Epigenomics

The development of a tetO/TetR system in the fungus Neurospora crassa is described. The system includes (i) a synthetic gene encoding a TetR variant fused to GFP, and (ii) a standard tetO array integrated homologously, as a proof of principle, near the his-3 gene. The localization of TetR-GFP at the tetO array (observed by fluorescence microscopy) can be disrupted by the application of tetracycline. The full-length array is stable during vegetative growth, but it triggers strong repeat-induced point mutation (RIP) by the RID-dependent as well as the DIM-2-dependent pathways during the sexual phase. Thus, both RIP pathways must be inactivated to allow the faithful inheritance of the unmodified construct. In summary, this study introduces a new molecular tool into Neurospora research, and suggests that the standard tetO array can self-engage in recombination-independent homologous pairing.

Keywords: FROS; Fluorescent repressor operator system; Neurospora; RIP; Repeat-induced point mutation; TetR; tetO

Domains: Molecular biology

Tinh-Suong Nguyen, Eugene Gladyshev. Developing a tetO/TetR system in Neurospora crassa. Fungal Genetics and Biology, 2020, 136, pp.103316. ⟨10.1016/j.fgb.2019.103316⟩. ⟨pasteur-03261456⟩