Stable transformation ofRhizoctonia solaniwith a modified hygromycin resistance gene

Wu, J. and O'Brien, P.A. (2009) Stable transformation ofRhizoctonia solaniwith a modified hygromycin resistance gene. Australasian Plant Pathology, 38 (1). pp. 79-84.

Abstract

A vector for Agrobacterium tumefaciens-mediated transformation of basidiomycetes was constructed by insertion of a modified hygromycin resistance gene into the plant binary vector pBin19. The hygromycin coding region is flanked by a basidiomycete promoter and terminator. Isolates from different anastomosis groupings (AG) of the phytopathogenic fungus Rhizoctonia solani (Kuhn) were transformed with this vector using A. tumefaciens. Hygromycin-resistant transformants were isolated from a single AG6 isolate, but not from AG3 or AG4 isolates. Of six transformants isolated, five showed enhanced growth on agar containing either 25 or 50 μg/mL hygromycin. However, as the hygromycin concentration increased, the difference between the transformants and the control reduced, until at 100 μg/mL there was no difference. The resistance phenotype was retained through repeated subcultures on non-selective media. The presence of the gene in the transformants was confirmed by PCR analysis and Southern hybridisation.