Chromosome analysis by fluorescence in situ hybridization of callus-derived regenerants in Allium cyaneum R.

Abstract

Investigations were performed to confirm the optimal in vitro culture condition for callus induction and plant regeneration, to observe if somoclonal variation occurs among regenerated plants at the ploidy level and to analyse the chromosomal location of 5S and 18S-26S rRNA gene families using fluorescence in situ hybridization in callus-derived plants of Allium cyaneum. High-est callus initiation was achieved with bulb explants cultured on MS medium supplemented with 2,4-D and BAP at 1 mg l^–1 each. A total of 195 plants was obtained when using MS medium supplemented with 1 mg l^–1 NAA and 5 mg l^–1 BAP; about 92% were diploid having 2n=16; 8% showed a variation in ploidy level. Using digoxigenin-labelled 5S rRNA and biotin-labelled 18S-26S rRNA gene probes, we compared the fluorescence in situ hybridization patterns of autotetraploid plants with the A. cyaneum wild type. The 5S rRNA gene sites were detected on the interstitial region in the short arm of chromosome 4 and on the interstitial region in both arms of chromosome 7. The 18S-26S rRNA gene sites were detected on the terminal region of the short arm, including the satellite of chromosome 5, as well as on a part of chromosome B. The chromosomal location of both rRNA genes in regenerated autotetraploid plants corresponded to those of the wild species.