Phoma glomerata (Corda) Wollenw. & Hochapfel is a pathogenic fungus causing spot diseases of plant leaves and fruits. This fungus is important in plant quarantine of seedlings and fruits in Korea. The aim of this study was to develop a sensitive and effective diagnostic method for P. glomerata detection in imported plants. The fungal species-specific PCR primers were designed based on the nucleotide sequences of the translation elongation factor 1 alpha gene and their specificity and sensitivity were tested. The designed primers named as PhoGlo-F and PhoGlo-R amplified specifically a 170 bp sized DNA band of the target gene from the genomic DNA of P. glomerata. No amplicon was produced from genomic DNAs of 16 other Phoma spp. and reference fungal species tested. Moreover, PhoGlo-F/PhoGlo-R primers successfully worked with real-time PCR technique. The detection limit of DNA content by conventional and real-time PCR were 10 pg and 1pg of the genomic DNA of P. glomerata, respectively. We believed that the developed makers would be very useful for P. glomerata detection.