Version 1: A guide to Agrobacterium-mediated transformation of the chytrid fungus Spizellomyces punctatus

Chytrid fungi play key ecological roles in aquatic ecosystems, and some species cause a devastating skin disease in frogs and salamanders. Despite their importance, little is known about the basic cell biology of chytrids. Additionally, chytrids occupy a unique phylogenetic position– sister to the well-studied Dikarya (yeasts, sac fungi, and their multicellular relatives) and related to animals– making chytrids useful for studying important evolutionary questions. A major barrier to understanding chytrid biology has been a lack of genetic tools with which to test molecular hypotheses. Medina and colleagues recently developed a protocol for Agrobacterium-mediated transformation of Spizellomyces punctatus. In this manuscript, we describe the general procedure including planning and expected results. We also provide in-depth, step-by-step protocols and video guides for performing the entirety of this transformation procedure.