GENETIC-TRANSFORMATION OF THE ECTOMYCORRHIZAL FUNGUS PAXILLUS-INVOLUTUS BY PARTICLE BOMBARDMENT

Paxillus involvutus was transformed using gold particle mediated gene transfer. Transformation was determined using the hygromycin B phosphotransferase gene (hph) as the selectable marker and the beta-glucuronidase gene (GUS) as a reporter gene. Southern blot analysis confirmed that the vector DNA for both hph and GUS genes was integrated into the fungal genome. Variations in the number of multiple gene copies and insertions were found in the transformants. The hygromycin resistant transformants were mitotically stable maintaining both the hph and GUS genes in the fungal genome six months following transformation Western blot analysis determined that the GUS gene was capable of transcribing and translating its protein product in the transformed fungus. Enzyme assays of GUS extracts determined that beta-glucuronidase was active in the transformed fungi. Pure culture synthesis experiments showed that the ability of P. involutus to form ectomychorrhizae with Pinus resinosa was not altered by transformation. These results provide the first report of a successful transformation of an ectomycorrhizal fungus using particle bombardment.