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Fusarium oxysporumf. sp.cubense, particularly the tropical race 4 (TR4), is the most destructive fungal pathogen of banana and can cause significant economic losses. However, the molecular mechanisms underlying the pathogenicity and virulence of the fungus are poorly understood. In this study, we characterized the mutant W2987, which was derived through T-DNA insertional mutagenesis of a wild-type TR4 isolate. W2987 showed slower growth, produced fewer conidia and was less virulent than the wild-type strain. Thermal asymmetric interlaced-PCR (TAIL-PCR) analysis showed that the T-DNA was inserted in the 3′ untranslated region (UTR) of an open reading frame (ORF) in W2987. BLAST search results indicated that the ORF encodes a hypothetical protein with unknown function. Phylogenetic analysis of the gene sequences from variousFusariumisolates showed that theF. oxysporumf. sp.cubenseTR4 isolates were distinct from otherF. oxysporumisolates. One pair of primers based on the 3′ UTR sequence of the putative virulence gene amplified a specific PCR product from theF. oxysporumf. sp.cubenseTR4 isolates but not from other races andFusariumspecies. This result indicates that this genetic locus possibly associated with virulence is unique and reliable for diagnosis and detection ofF. oxysporumf. sp.cubenseTR4. [ABSTRACT FROM PUBLISHER]