Conditional Gene Deletion in Ustilago maydis Via CRE Recombinase In Planta

Site-specific recombinases have been extensively utilized in higher eukaryotes, particularly in animals, for various applications such as chromosomal translocations, large deletions, site-specific integration, and, notably, tissue-specific conditional knockouts. This technique has also been applied in simpler eukaryotes, including fungi and protozoa. However, in fungal research, especially involving phytopathogenic fungi, its use has often been limited to the recycling of marker genes during transformation experiments. In this study, detailed instructions are provided for generating conditional gene deletions in the corn phytopathogenic fungus Ustilago maydis, while infecting corn plants. This is accomplished through the transcriptional control of the recombinase by fungal promoters that are specifically activated during the biotrophic phase of fungal growth. This method enables gene deletions at defined developmental stages within plant tissues. These tools will prove valuable for investigating the roles of virulence-specific functions within the U. maydis-maize pathosystem under in planta conditions.