PCR amplification from a homolog of the bE mating-type gene as a sensitive assay for the presence of Ustilago scitaminea DNA

Ustilago scitaminea, the causal agent of sugarcane (Saccharum spp.) smut, has a bipolar mating system in that the haploid stage of the life cycle has sporidia of two mating types (designated plus and minus). Only haploid sporidia of complementary mating types can fuse to form the infectious, mycelial dikaryon. With the use of primers based on the U. maydis bE mating-type gene, DNA segments from sporidia were polymerase chain reaction (PCR)-amplified from both minus and plus mating types of U. scitaminea. These DNA fragments were sequenced and found to be approximately 70% identical in nucleotide sequence to the corresponding region of the bE gene in U. maydis and U. hordei. The mating behavior of U. scitaminea is similar to that of other bipolar Ustilago spp. This and the similarity in DNA sequence between the b genes suggests that mating-type genes of U. scitaminea are similar in structure and function to those of other Ustilago spp. previously studied. Use of one of the cloned fragments as a probe in Southern analysis of U. scitaminea revealed specific hybridization to single BamHI fragments of different sizes in the two mating types, confirming mating type-specific differences at this locus. Primers made to the cloned sequence have been used to amplify by PCR a DNA fragment of the predicted size from a 2,000:1 mixture of sugarcane and U. scitaminea dikaryon DNA.