Four asparagus cultivars were cytologically studied by conventional staining and in situ hybridization, in order to evaluate both the variability among cultivars and the distribution pattern of 18S-25S rRNA genes. lhe mitotic and meiotic analysis with conventional chromosome staining did not show heteromorphisms or differences between cultivars. In situ hybridization with 18S and 25S rONA probes showed 6 chromosomes with 18S-25S rONA sites in the 4 studied genotypes. lhe New Jersey, Waltam Washington and G4X 14 cultivars presented 4 chromosomes with larger subterminal sites and 2 with interstitial sites of smaller size. In the Oeco cultivar, heteromorphism was verified in the size of the rONA sites, which were larger and subterminal in 3 chromosomes, and small and interstitial in 3 other chromosomes. lhe results suggest that asparagus cultivars may present only small differences in the number and distribution of rONA loci