IN-SITU HYBRIDIZATION OF RIBOSOMAL DNA TO ROSE CHROMOSOMES

The genus Rosa consists of approximately 200 species and 20,000 cultiv ars, most of complex hybrid origin. Few of the current cultivars, most of which are tetraploid with 2n = 4x = 28, can be traced unambiguousl y to their wild diploid progenitor species. Fluorescent in situ hybrid ization (FISH) offers the possibility to estimate the proportion of va rious donor genomes in complex advanced-generation hybrids. To establi sh the feasibility of FISH in rose and to explore differences, if any, in number and subchromosomal location of nucleolar organizer regions (NORs), mitotic chromosome spreads from five diploid species and a com mercial tetraploid cultivar were probed with part of the 18S-26S rDNA repeat from soybean. Shoot tips were the most convenient source of mit otic divisions for FISH. Cell suspensions from enzymatically digested shoot tips were spread with 3 ethanol:1 acetic acid on glass slides an d then squashed in 45% acetic acid under a coverslip. Prolonged digest ion in cellulase and pectolyase was required to obtain satisfactory ch romosome spreading and exposure for FISH. A pair of strong, subtermina l hybridization signals, corresponding to a single NOR per genome, was observed in the diploid species. The commercial tetraploid rose culti var had four strong, subterminal signals and thus one NOR per genome. However, two of the signals were consistently stronger than the other two. The results demonstrate the feasibility of detecting repetitive s equences in spread chromosomes of roses.