Cytogenetic analysis of starting materials
The starting plant material was analyzed in order to exclude any cytogenetic variation that could influence further analysis. The levels of ploidy and endopolyploidy in rosette leaves from 3 week old plants were estimated.The flow cytometry measurements were conducted on 20 plants, and two samples from each plant were analyzed. All plants were found to be diploids. The young rosette leaves exhibited two cycles of endoreduplication and the maximum DNA content was 16C (Fig. 1a), which is consistent with the organ specific endopolyploidy pattern in A. thaliana
(Galbraith et al. 1991). The chromosome number analysis was performed on root apical meristems of at least 20 seedlings as well as on flower buds collected individually from 20 plants. In the root apical meristems all metaphases had an exact diploid chromosome number (2n = 10), while in flower buds the diploid number occurred in 65 % of the metaphases and the number of chromosomes in the remaining cells varied from 7 to 9 (Table 2). This phenomenon was defined as aneusomaty by Duncan (1945) and has been described for different types of organs and tissues in many species (D’Amato 1995).Although it was suggested that aneusomaty is more frequent in polyploids, for example in Artemisia czekanowskiana and A. macrantha (Pellicer et al. 2007), it was also
noticed in species with a lower chromosome number, e.g.Deschampsia antarctica (Cardone et al. 2009). The rDNA sequences are robust chromosome markers for Arabidopsis thaliana and after applying FISH with these sequences as probes all chromosomes are easily distinguishable (Fransz
et al. 1998). The distribution of 25S and 5S rDNA loci in the karyotype of control plants was analyzed in root tips and in flower buds. The first pair of chromosomes does not possess any rRNA genes, the second pair is characterized by the presence of 45S rDNA site, the chromosomes of the third and the fifth pair bear 5S rRNA genes and the fourth pair has both 45S and 5S rDNA (Fig. 3a). These results confirm that the plants used as the starting material were uniform in the ploidy level, chromosome number and rDNA distribution.