CYTOLOGICAL STUDIES OF THE NUCLEOLUS ORGANIZING REGIONS IN THE MEDICAGO COMPLEX - SATIVA-COERULEA-FALCATA

A cytological examination of the nucleolus organizing regions (NORs) o f three species from the Medicago sativa complex was conducted to eval uate the structural and functional evolution of the ribosomal RNA (rRN A) loci that encode the 18S, 5.8S, and 26S rRNAs. Mitotic chromosomes in root-tip preparations from tetraploid M. sativa and diploids Medica go coerulea and Medicago falcata were visualized by four methods that provide new data. Fluorescent in situ hybridization using the M. sativ a 18S gene as probe localized the structural rDNA to the constricted r egions of the satellited chromosomes only. Chromomycin A(3) (CMA(3)) s taining and 4',6-diamidino-2-phenylindole (DAPI) staining identified t hese chromosomal segments as the most CC-rich regions in the alfalfa k aryotype. Medicago falcata exhibited fewer DAPI bands and chromocenter s than did M. sativa and M. coerulea. Positive silver nitrate staining showed that all four rDNA regions in M. sativa (located in two chromo some pairs) and both rDNA sites in both diploid species remain transcr iptionally active. Counts of nucleoli confirmed that all rDNA regions are independently capable of nucleolus organization. Thus, the number of active NORs in M. sativa is double the number found in M. coerulea or M. falcata. Consequently, if M. sativa originated from sexual hybri dization of 2n gametes involving one or both diploid species, no major reorganization or loss of structural or functional rDNA loci has occu rred.