|Budget Amount *help
¥2,500,000 (Direct Cost : ¥2,500,000)
Fiscal Year 1997 : ¥600,000 (Direct Cost : ¥600,000)
Fiscal Year 1996 : ¥800,000 (Direct Cost : ¥800,000)
Fiscal Year 1995 : ¥1,100,000 (Direct Cost : ¥1,100,000)
In this reseasrch project for these three years, some soil-borne plant pathogens were successfully transformed with specific marker genes and their infection behavior in soil (before infection) and in planta (after infection into host plants) was monitored by detecting expression of the introduced marker genes. In order to evaluate feasibility or effectiveness of the present method, the author adopted different formae speciales of Fusarium oxysporum whic were indistinguishable one another by the conventional cultural method, and actually attempted to trace and distinguish each fungus. In this study, four formae speciales of F.oxysporum (F.oxy. f. sp. lycopersici, F.oxy. f. sp. fragarea, F.oxy. f. sp. melonis and F.oxy. f. sp. spinaciae) were separately transformed with four different marker genes, hygromycin-resistant gene, beta-glucuronidase gene, luciferase gene and green fluorescent protein gene. These genes were introduced into microconidia of these fungi by electroporation and the gene expression detected using an in situ hybridization. In the actual approach of the present system, host plants were transplanted in soil into which the gene marked pathogens had been introduced and the infected pathogens were detected from both soil and plant tissues as definate intervals after planting. The present method enabled us to simultaneously distinguish different formae speciales of the Fusarium pathogen by detecting different fluorescences emitted from the fluorochrome-labelled probes used for specific hybridization of each marker gene. Thus, the author succeeded in establishing the gene-marking method for monitoring filamentous soil-borne phytopathogenic fungi in the present project research.