The non-conventional yeast Yarrowia lipolytica is gaining interest in biotechnology as host for production of proteins, lipids and other biomolecules thanks to its capacity to achieve high biomasses, and to express high titers of proteins and lipids. Site-specific genome editing is however limited in this yeast. This was much improved by the recent development of a CRISPR-Cas9 genome editing protocol for Y. lipolytica based on a tRNA-sgRNA fusion. Nonetheless, in the latter protocol, Cas9 ...