THE NIA GENE OF FUSARIUM-OXYSPORUM - ISOLATION, SEQUENCE AND DEVELOPMENT OF A HOMOLOGOUS TRANSFORMATION SYSTEM

The Fusarium oxysporum gene nia, encoding nitrate reductase (NR), was isolated from a cosmid library by direct complementation of an F. oxys porum nia- mutant. The gene specifies a protein of 905 amino acids and contains a 57-bp intron. Comparison of the deduced aa sequence with N R of other fungi revealed a high degree of similarity and conservation in the intron position. The cloned nia made it possible to develop th e first homologous transformation system for this fungus. Transformati on frequencies of up to 600 transformants per mug of DNA were achieved . Gene replacements, single-copy homologous integrations and integrati ons at non-homologous sites were observed. Direct comparison between p lasmids and cosmids carrying the same gene showed a higher frequency o f targeted transformation using cosmid vectors. Gene replacement event s were observed in about 50% of the transformants analysed with each t ype of vector used. This high frequency of substitution offers new app lications for the transformation system in F. oxysporum.