M. Frediani

Nuclear DNA contents, rDNAs, chromatin organization, and karyotype evolution in Vicia sect. Faba.

SummaryAutomated karyotype analyses, nuclear DNA contents, and sequences of rDNA internal transcribed spacers of the nine species inVicia sect.faba are reported. As karyomorphological parameters are used to evaluate the karyotype evolution, so the determination of the heterochromatin by Feulgen absorption at different thresholds of optical density provided further evidence on the chromatin organization withinVicia sect.faba. The comparison of sequences of rDNA spacers has enabled the definition of the phylogenetic relationships between the analyzed species.

Nuclear DNA contents, rDNAs, and karyotype evolution in Vicia subgenus Vicia: II. Section Peregrinae

Summary.Nuclear DNA contents, automated karyotype analyses, and sequences of rDNA spacers have been determined for the species of Vicia belonging to sect. Peregrinae, as well as for V. mollis. The phylogenetic data generated from the comparison of rDNA sequences and karyomorphological results would both indicate that Vicia mollis is a sister group to sect. Peregrinae. The relationships among the species belonging to the Peregrinae section and species enclosed in sections Faba, Narbonensis, and Bithynicae have been also investigated: a clade including V. mollis and sect. Peregrinae is a sister group to a clade including V. bithynica and sect. Narbonensis. With our choice of outgroup, Vicia faba (including subsp. paucijuga) is external to the above mentioned inclusive group.

Distribution of 5-methylcytosine-rich regions in the metaphase chromosomes of Vicia faba

The DNA methylation pattern ofVicia faba metaphase chromosomes was examined with a specific monoclonal antibody. 5-methylcytosine (5-mC) residues are present in different chromosomal sites, and are particularly abundant in telomeric and/or subtelomeric regions and in certain intercalary bands. Chromosomal localization of methylated regions enables a better knowledge of the lengthwise differentiation of this chromosome complement. Our results also indicate that there may be differences in monoclonal antibody binding between corresponding regions of homologous chromosomes inV. faba. This behaviour is detectable in specific regions with different frequencies. The data support results previously obtained forAllium cepa metaphase chromosomes using the same monoclonal antibody.

Nuclear DNA contents, rDNAs, and karyotype evolution in subgenus Vicia: III. The heterogeneous section Hypechusa.

Summary.Nuclear DNA contents, automated karyotype analyses, and sequences of internal transcribed spacers from ribosomal genes have been determined in the species belonging to section Hypechusa of the subgenus Vicia. Karyomorphological results and phylogenetic data generated from the comparison of rDNA (genes coding for rRNA) sequences showed that sect. Hypechusa is not monophyletic; however, some monophyletic units are apparent (one including Vicia galeata, V. hyrcanica, V. noeana, and V. tigridis, another including V. assyriaca, V. hybrida, V. melanops, V. mollis, and V. sericocarpa), which partly correspond to morphology-based infrasectional groups. The relationships among these species and the species in sections Faba, Narbonensis, Bithynicae, and Peregrinae have been also investigated.

Cytological investigation of Haplopappus gracilis (Nutt.) Gray: 5-methylcytosine-rich regions, fluorochrome banding and chromatin sensitivity to DNase I digestion

Summary.Haplopappus gracilis (Nutt.) Gray, one of the five known higher plants with a chromosome number of 2n = 4, was studied from a cytological point of view. The chromosome complement of this species was characterized by means of automated karyotype analysis. Moreover, the DNA methylation pattern and fluorochrome banding were determined and compared with cytological data present in the literature. DNA methylation distribution along metaphase chromosomes involved all chromosome territories evidenced by C-banding. Other methylated bands correlated positively with aceto-orcein-positive heterochromatic portions and/or with late replicating bands and/or fluorochrome bands. Some methylated bands showed differences between homologous chromosomes. These bands belonged partly to certain heterochromatic domains and partly to intercalary sites not defined by other standard banding techniques. Differences between the homologues were also indicated by our DNA content data obtained after DNase I digestion.

Cytological and molecular characterization of Vicia esdraelonensis Warb. & Eig: a rare taxon

Summary.Vicia esdraelonensis, a rare taxon belonging to section Hypechusa of subgenus Vicia, was recovered and analyzed by cytological, karyological, and molecular methods, with the aim of both characterizing this species and furthering our knowledge of the phylogeny of subgenus Vicia. Automated karyotype analysis, nuclear DNA content, and chromatin organization were determined by the Feulgen reaction, as well as chromosome banding after double staining with 4′,6-diamidino-2-phenylindole (DAPI) and chromomycin A3. The chromosome number and the nuclear DNA content were in agreement with the values of the species of section Hypechusa. The GC- and AT-rich preferential sites were determined by chromomycin A3 and DAPI staining. Karyomorphological parameters indicated that V. esdraelonensis is in an intermediate position in the spatial representation of the species of section Hypechusa on the basis of symmetry indices, as well as in the dendrogram of linkage distance constructed on 37 chromosome parameters. Molecular data based on internal transcribed spacer sequences show that V. esdraelonensis can doubtlessly be included in section Hypechusa and document its closeness to V. noeana. A cladistic analysis combining the molecular data set with karyological characters is also reported. Karyological, cytological, and molecular data allow characterization of the V. esdraelonensis genome and provide information about the phylogenetic position of this species within the Hyrcanicae series of section Hypechusa.

Cytology of Vicia species. X. Karyotype evolution and phylogenetic implication in Vicia species of the sections Atossa, Microcarinae, Wiggersia and Vicia

Automated karyotype analyses and sequence of rDNA spacers have been analysed for the species belonging to sections Atossa, Microcarinae, Wiggersia and Vicia. Karyomorphological parameters, based on Rec, Syi and TF% indices, have been determined and evidenced that, in term of symmetry, the karyotype of Vicia lathyroides was the most asymmetric one. A multivariate analysis using 34 karyological parameters, in addition to the symmetry indices, has been carried out and the corresponding dendrogram of linkage distances showed six different groups. Molecular investigations on the inclusive group in study by employing ITS DNA sequences indicated a different pattern of relationships. The cladistic analysis combining the molecular data set with karyological parameters evidenced that the species of sections Vicia and Atossa join closely to each other in a paraphyletic group, which includes the monophyletic section Wiggersia. Therefore, our karyological and molecular data provide information about the phylogenetic position of the analysed species inside the subgenus Vicia and are discussed in relation to previous results obtained by morphology, isozymes and ribosomal genes analyses.

Repetitive DNA sequences as probes for phylogenetic analysis in Vicia genus

Abstract In the process of further characterizing phylogenetic relationships among Vicia species of the subgenus Vicia, four different molecular DNA markers were used: a tandemly repeated DNA sequences about 60 bp in length (FokI), a 336 bp element (pVf7) homologous to the IGS repeats (but that does not reside in the policistronic rDNA units); a family of repeated DNA sequences (VfB) of about 1200 bp in length (that might be derived from a mobile DNA element) and a family of repeated DNA sequences (VfM) of about 60 bp that can be considered a minisatellite-like sequence (EMBL accession nr. AJ242773). The comparison of the obtained results has enabled the definition of the phylogenetic relationships among the analysed species confirming that V. faba, V. bithynica and the species of Narbonensis section, represent three distinct taxonomic groups according to the Maxted’s classification (1993). VfB and VfM marker discriminate inside the sect. Narbonensis, too, evidencing a greater affinity between some species.

CHy-banding patterns and chromatin organization inAegilops andTriticum species (Poaceae)

The distribution of CHy-banded heterochromatin was studied in the chromosomes ofAegilops longissima, Ae. speltoides, Triticum monococcum, andT. turgidum. Interphase nuclei were measured after Feulgen staining at different thresholds of optical density; the curves so obtained indicated the relationship among the species with respect to the different fractions of the genomic DNA. The karyological and cytophotometric analyses indicate differences betweenAe. speltoides andAe. longissima, the latter species being enriched in heterochromatin. Similar results were demonstrated for the genusTriticum, in whichT. turgidum showed more heterochromatin when compared withT. monococcum. The results suggest that the B genome of the cultivated wheats possesses a type of heterochromatin that resembles the type present inAe. longissima.

Cytological characterization of Vicia oroboides Wulfen in Jacq

Vicia oroboides, a rare taxon belonging to section Atossa of subgenus Vicia, was recovered and analysed by means of cytological and karyological methods with the aim of both characterising this species and integrating our knowledge on phylogeny of subgenus Vicia. Automated karyotype analysis and nuclear DNA content have been determined after Feulgen’s reaction; chromosome banding was performed by fluorochrome staining to evidence heterochromatic blocks along the chromosome complement. The chromosome number is in line with the values of the species of section Atossa; the GC- and AT-rich sites were identified by CMA and DAPI staining. Karyomorphological parameters, based on symmetry indices, provide information about the phylogenetic position of this species inside the subgenus Vicia. DNA content is reported for the first time.