In recent years, the CRISPR-Cas9 system has proven extremely useful for genome editing in many species, including the model yeast Saccharomyces cerevisiae and other yeast species such as Candida glabrata. Inducible CRISPR-Cas9 systems have the additional advantage of allowing to separate the transformation step of the organism by the CRISPR-Cas9 system, from the cutting and repair steps. This has indeed been developed in S. cerevisiae, where most inducible expression systems rely on the GAL promoters. Unfortunately, C. glabrata is gal- and lacks the GAL genes, like many other yeast species. We report here the use of a vector expressing cas9 under the control of the MET3 promoter, with the guide RNA cloned into the same plasmid. We show that it can be used efficiently in C. glabrata, for both described outcomes of CRISPR-Cas9-induced chromosome breaks; nonhomologous end joining in the absence of a homologous repair template; and homologous recombination in the presence of such a template. This system therefore allows easy editing of the genome of C. glabrata, and its inducibility may allow identification of essential genes in this asexual yeast, where spore lethality cannot be observed, as well as the study of double-strand break repair.

中文翻译：

---

一种新的诱导型CRISPR-Cas9系统，可用于光滑念珠菌的基因组编辑和双链断裂修复研究。

近年来，事实证明，CRISPR-Cas9系统对于许多物种的基因组编辑极为有用，包括酿酒酵母模型酵母和光滑念珠菌等其他酵母物种。诱导型CRISPR-Cas9系统的另一个优势是可以将CRISPR-Cas9系统的生物体转化步骤与切割和修复步骤分开。确实在酿酒酵母中已经开发出了这种酵母，其中大多数可诱导的表达系统依赖于GAL启动子。不幸的是，像许多其他酵母菌种一样，光滑念珠菌是gal-且缺少GAL基因。我们在这里报告在MET3启动子控制下表达cas9的载体的使用，其中指导RNA克隆到同一质粒中。我们证明它可以有效地用于C. glabrata，既描述了CRISPR-Cas9诱导的染色体断裂的结果; 在没有同源修复模板的情况下进行非同源末端连接；在这种模板存在下进行同源重组。因此，该系统可以轻松编辑光滑杯状梭菌的基因组，并且其可诱导性可以允许鉴定这种无性酵母中的必需基因，在这种无性酵母中无法观察到孢子致死性，并且可以研究双链断裂修复。