CLONING AND ANALYSIS OF A 6.8-KB RDNA INTERGENIC SPACER REGION OF THEEUROPEAN ASH (FRAXINUS-EXCELSIOR L)

The 6.8-kb rDNA intergenic spacer region of F. excelsior was isolated from a CsCl/actinomycin-D gradient and cloned into pUC18 for further c haracterization. We observed the presence of subrepeats delimited by H aeIII enzyme sites. These subrepeats were subcloned and 11 clones were sequenced. These corresponded to subrepeated elements of either 32 bp or 41 bp that shared a 23-bp common sequence in the 5' end. Within ea ch family of subrepeats, the percentage of common nucleotides was 84.4 % for the 5 32-bp subrepeats and 67.4% for the 640-bp subrepeats. Non- repeated HaeIII fragments of 450 bp and 650 bp were also sub-cloned. T o compare homology at the IGS region between the rDNA spacers of F. ex celsior and the three related species (F. oxyphylla, F. americana, F. ornus), we conducted Southern hybridization analyses using each member of the 32-bp and 40-bp subrepeat families and the unique 450-bp and 6 50-bp fragments as probes. These analyses indicated that (1) the Ameri can ash is more genetically distant from the other three species that the latter are from each other and (2) F. oxyphylla and F. excelsior a re more closely related to each other than to F. ornus.