Cloning of DNA fragments specific to the pathotype and race of Verticillium dahliae

摘要

Japanese isolates of Verticillium dahliae, a causal agent of wilt disease in many plants, are classifiable into pathotypes based on their pathogenicity. Because these pathotypes are morphologically indistinguishable, establishinga rapid identification method is very important for the control of this pathogen in Japan. For cloning DNA fragmentsthat are useful for identification and specific detection of V. dahliae pathotypes, we performed random amplified polymorphic DNA (RAPD) analyses using various isolates. Onepolymerase chain reaction (PCR) product, E10-U48, was specific to isolates pathogenic to sweet pepper. The otherproduct, B68-TV, was specific to race 1 of isolates pathogenic to tomato. The specificity of these sequences wasconfirmed by genomic Southern hybridization. Further analyses revealed that the region peripheral to B68-TV obtained from the genomic DNA library includes the sequencespecific to all isolates pathogenic to tomato (races 1 and 2). Moreover, sequence tagged site (STS) primers designedfrom B68-TV and its peripheral region showed race-specific and pathotype-specific amplification in a PCR assay. Theprobes and primers obtained in this study are likely to be useful tools for the identification and specific detection ofpathotypes and races of V. dahliae.