Screening of fungi for the presence of the trichodiene synthase encoding sequence by hybridization to the Tri5 gene cloned from Fusarium poae

Abstract

A trichodiene synthase gene (Tri5) was amplified from F. poae by polymerase chain reaction using synthetic primers constructed on the basis of the coding portion of the same gene from F. sporotrichioides. Sequence analysis showed a high degree of similarity with other trichodiene synthase genes. A 378 bp HindIII fragment of the gene that contains the genetic information for the putative active site of the trichodiene synthase enzyme was radiolabelled and used for dot blot analysis. This probe could detect Tri5 hybridization in 1-10 ng DNA of fusaria that have the genetic potentiality to synthesize toxic trichothecene compounds, but gave no reaction with trichothecene nonproducing members of the genus. When other fungi reported to produce trichothecenes (Myrothecium, Stachybotrys, Trichoderma, Trichothecium spp.) were tested, only strains of Myrothecium and Stachybotrys gave strong positive reaction. Faint but consistent hybridization signals were obtained in four species (F. semitectum, F. tricinctum, Trichoderma viride and Trichothecium roseum) indicating the presence of nonhomologous evolutionary variants or inactive remnants of the Tri5 gene in these fungi.