Molecular transformation of Fusarium solani with an antibiotic resistance marker having no fungal DNA homology

Abstract

A vector was constructed for transformation of the plant pathogenic fungus Fusarium solani. The promoter 35Sp, from cauliflower mosaic virus, was fused to the bacterial gene APH(3′)II, which confers resistance to the aminoglycoside antibiotic G418. Two transformation procedures were developed: one using isolated fungal protoplasts, the other using germinated fungal spores. A transformation frequency of 3.3 G418-resistant colonies were obtained per microgram DNA. Of 14 colonies analyzed, 12 had vector sequences integrated into their high molecular weight DNA, and 2 were untransformed. Integration was sometimes accompanied by rearrangements of both the vector and flanking fungal DNAs. Primer-extension analysis of the mRNA from one transformant revealed two putative transcription initiation sites in the chimeric APH(3′)II gene. Both sites differed from the normal initiation site in plants. This vector will be useful in transformation systems in which integration by non-homologous recombination is desired.[/P ] © 1989 Springer-Verlag.