Molecular structure and chromosome distribution of three repetitive DNA families in Anemone hortensis L. (Ranunculaceae).

The structure, abundance and location of repetitive DNA sequences on chromosomes can characterize the nature of higher plant genomes. Here we report on three new repeat DNA families isolated from Anemone hortensis L.; (i) AhTR1, a family of satellite DNA (stDNA) composed of a 554-561 bp long EcoRV monomer; (ii) AhTR2, a stDNA family composed of a 743 bp long HindIII monomer and; (iii) AhDR, a repeat family composed of a 945 bp long HindIII fragment that exhibits some sequence similarity to Ty3/gypsy-like retroelements. Fluorescence in-situ hybridization (FISH) to metaphase chromosomes of A. hortensis (2n = 16) revealed that both AhTR1 and AhTR2 sequences co-localized with DAPI-positive AT-rich heterochromatic regions. AhTR1 sequences occur at intercalary DAPI bands while AhTR2 sequences occur at 8-10 terminally located heterochromatic blocks. In contrast AhDR sequences are dispersed over all chromosomes as expected of a Ty3/gypsy-like element. AhTR2 and AhTR1 repeat families include polyA- and polyT-tracks, AT/TA-motifs and a pentanucleotide sequence (CAAAA) that may have consequences for chromatin packing and sequence homogeneity. AhTR2 repeats also contain TTTAGGG motifs and degenerate variants. We suggest that they arose by interspersion of telomeric repeats with subtelomeric repeats, before hybrid unit(s) amplified through the heterochromatic domain. The three repetitive DNA families together occupy approximately 10% of the A. hortensis genome. Comparative analyses of eight Anemone species revealed that the divergence of the A. hortensis genome was accompanied by considerable modification and/or amplification of repeats.

Two classes of 5S rDNA unit arrays of the silver fir, Abies alba Mill.: structure, localization and evolution.

The structure and organization of the 5S ribosomal DNA units of the silver fir, Abies alba Mill., as well as their position in the chromosome complement were investigated. PCR amplification of the gene and nontranscribed spacer region, sequence analysis and Southern hybridization, using a homologous probe, detected DNA sequences of approximately 550 bp and 700 bp. Sequence analysis of the spacers revealed that the difference in length between the sequences occurred in the middle spacer region as a result of the amplification of a 75-bp sequence of the short unit class, which is organized in four 54- to 68-bp tandem repeats in the long spacer unit. The 5S rDNA transcribed region is 120 bp long and shows high sequence similarity with other gymnosperm species. The comparative analysis of 5' and 3' flanking sequences of 5S rRNA genes of silver fir and other gymnosperms indicates that A. alba spacer units have the same rate of evolution and are more closely related to Larix and Pseudotsuga than to Pinus and Picea. Southern hybridization and fluorescence in situ hybridization of metaphase chromosomes of A. alba suggest that the short and long spacer units are organized as separate tandem arrays at two chromosomal loci on chromosomes V and XI.

Application of the micronucleus and comet assays to mussel Dreissena polymorpha haemocytes for genotoxicity monitoring of freshwater environments.

Assessment of DNA damage is of primary concern when determining the pollution-related stress in living organisms. To monitor genotoxicity of the freshwater environments we used micronucleus (MN) and comet assay on Dreissena polymorpha haemocytes. Caged mussels, collected from the river Drava, were transplanted to four monitoring sites of different pollution intensity in the river Sava. Exposition lasted for a month. The baseline level of MN frequencies in the haemocytes of mussels from reference site (river Drava) was 0.5 per thousand. No increase in MN frequency was found in mussels from the medium-polluted site (Zagreb) in the river Sava while other, more polluted sites showed higher MN frequencies ranging from 2.7 per thousand (Lukavec) and 3.1 per thousand (Oborovo) to 5.2 per thousand (Sisak). Results from comet assay showed concordance with MN assay in indicating intensity of DNA damage. The use of haemocytes from caged, non-indigenous mussels in MN and comet assay proved to be a sensitive tool for the freshwater genotoxicity monitoring.