Agrobacterium tumefaciens-Mediated Transformation Method for Fusarium oxysporum

Agrobacterium tumefaciens-mediated transformation (ATMT) is becoming a popular effective system as an insertional mutagenesis tool in filamentous fungi. An efficient Agrobacterium tumefaciens-mediated transformation approach was developed for the plant pathogenic fungus, F. oxysporum, the causal agent of Apple replant disease (ARD) in China. Four parameters were selected to optimize efficiencies of transformation. A. tumefaciens concentration, conidial concentration of F. oxysporum, and co-culture temperature and time have a significant influence on all parameters. Transformants emit green fluorescence under fluorescence microscopy. The integration of a mitotically stable hygromycin resistance gene (hph) in the genome is confirmed by PCR. The transformation efficiency can reach up to 300 transformants per 10⁶ conidia under optimal conditions. This ATMT method is an efficient tool for insertional mutagenesis of F. oxysporum.