Identification of species in tribe Brassiceae by dot-blot hybridization using species-specific ITS1 probes

Simple, reliable methods for identification of species are required for management of many species and lines in a plant gene bank. Species-specific probes were designed from published sequences of the ITS1 region in rDNA of 16 species in Brassica and its related genera, and used as probes for dot-blot hybridization with plant genomic DNA. All the probes detected species-specific signals at dot-blots of genomic DNAs of the 16 species in Brassica, Diplotaxis, Eruca, and Raphanus. Signals of the Brassica digenomic species in the U's triangle, i.e., B. napus, B. juncea, and B. carinata, were detected by the probes of their parental monogenomic species, i.e., B. rapa, B. nigra, and B. oleracea. The probe for B. oleracea showed signals of B. balearica, B. cretica, B. incana, B. insularis, and B. macrocarpa, which have the C genome as B. oleracea. Eruca vesicaria DNA was detected by the probe for E. sativa, which has been classified as a subspecies of E. vescaria. DNA of leaf tissue extracted by an alkaline solution and seed DNA prepared by the NaI method can be used directly for dot-blotting. Misidentification of species was revealed in 20 accessions in the Tohoku University Brassica Seed Bank. These results indicate dot-blot hybridization to be a simple and efficient technique for identification of plant species in a gene bank.