TRANSFORMATION OF BIPOLARIS-SOROKINIANA WITH THE GUS GENE AND USE FOR STUDYING FUNGAL COLONIZATION OF BARLEY ROOTS

Bipolaris sorokiniana, a fungal pathogen of cereals, was transformed with the GUS (beta-glucuronidase) gene, using a plasmid (pGUS5) containing GUS A with the constitutive promotor GPD-1 and a hygromycin-resistance gene (hph) under the control of the same promotor. Hygromycin resistance was used as the selectable marker. Colonies growing on medium with hygromycin were obtained at a frequency of 6.5 X 10(-7) per viable fungal protoplast. Nineteen out of 20 tested hygromycin-resistant colonies were also GUS positive. Transformants that showed a stable expression of GUS activity and hygromycin resistance after several conidiation cycles and several months of growth on nonselective medium could be selected. However, some transformants lost their GUS activity and hygromycin resistance after two conidiation cycles on nonselective medium. The stable transformants did not differ from the wild-type strain in growth rate or pathogenicity. A significant, positive correlation was found between GUS activity and ergosterol content and between GUS activity and lesion size in barley roots infected with a transformed strain. GUS activity could be detected in the root tissue before disease symptoms appeared, and only negligible background GUS activity was found in roots infected with the wild-type Fungus, These results indicate that GUS-transformed strains of B. sorokiniana can be used for studying fungal colonization and growth in plant tissue.