Rapid DNA-FISH in Arabidopsis thaliana Somatic Cells.

Fluorescence in situ hybridization (FISH) technique has been widely used to detect and localize specific DNA and RNA sequences in interphase nuclei and chromosomes in animals and plants. Here, we present a protocol for localization of genomic loci in nuclei of the model plant Arabidopsis thaliana. This protocol includes several advances and adaptations to A. thaliana, including preparation of nuclei and chromosomes without the use of liquid nitrogen, and an in situ hybridization procedure that preserves chromatin structure without the use of paraformaldehyde and formamide. Simultaneous denaturation of the BAC (bacterial artificial chromosome) probe and nuclei followed by annealing at high temperature allows hybridization in less than an hour. These hybridization conditions also provide high signal to noise ratio by a small number of washes. Thus, this simplified in situ hybridization procedure is completed in one working day.

The Influence of Edaphic Factors on DNA Damage and Repair in Wild Wheat Triticum dicoccoides Körn. (Poaceae, Triticeae).

A complex DNA repair network maintains genome integrity and genetic stability. In this study, the influence of edaphic factors on DNA damage and repair in wild wheat Triticum dicoccoides was addressed. Plants inhabiting two abutting microsites with dry terra rossa and humid basalt soils were studied. The relative expression level of seven genes involved in DNA repair pathways-RAD51, BRCA1, LigIV, KU70, MLH1, MSH2, and MRE11-was assessed using quantitative real-time PCR (qPCR). Immunolocalization of RAD51, LigIV, γH2AX, RNA Polymerase II, and DNA-RNA hybrid [S9.6] (R-loops) in somatic interphase nuclei and metaphase chromosomes was carried out in parallel. The results showed a lower expression level of genes involved in DNA repair and a higher number of DNA double-strand breaks (DSBs) in interphase nuclei in plants growing in terra rossa soil compared with plants in basalt soil. Further, the number of DSBs and R-loops in metaphase chromosomes was also greater in plants growing on terra rossa soil. Finally, RAD51 and LigIV foci on chromosomes indicate ongoing DSB repair during the M-phase via the Homologous Recombination and Non-Homologous End Joining pathways. Together, these results show the impact of edaphic factors on DNA damage and repair in the wheat genome adapted to contrasting environments.

Supernumerary B Chromosomes and Plant Genome Changes: A Snapshot of Wild Populations of Aegilops speltoides Tausch (Poaceae, Triticeae).

In various eukaryotes, supernumerary B chromosomes (Bs) are an optional genomic component that affect their integrity and functioning. In the present study, the impact of Bs on the current changes in the genome of goatgrass, Aegilops speltoides, was addressed. Individual plants from contrasting populations with and without Bs were explored using fluorescence in situ hybridization. In parallel, abundances of the Ty1-copia, Ty3-gypsy, and LINE retrotransposons (TEs), and the species-specific Spelt1 tandem repeat (TR) in vegetative and generative spike tissues were estimated by real-time quantitative PCR. The results revealed: (i) ectopic associations between Bs and the regular A chromosomes, and (ii) cell-specific rearrangements of Bs in both mitosis and microgametogenesis. Further, the copy numbers of TEs and TR varied significantly between (iii) genotypes and (iv) different spike tissues in the same plant(s). Finally, (v) in plants with and without Bs from different populations, genomic abundances and/or copy number dynamics of TEs and TR were similar. These findings indicate that fluctuations in TE and TR copy numbers are associated with DNA damage and repair processes during cell proliferation and differentiation, and ectopic recombination is one of the mechanisms by which Bs play a role in genome changes.

Long Tandem Arrays of Cassandra Retroelements and Their Role in Genome Dynamics in Plants.

Retrotransposable elements are widely distributed and diverse in eukaryotes. Their copy number increases through reverse-transcription-mediated propagation, while they can be lost through recombinational processes, generating genomic rearrangements. We previously identified extensive structurally uniform retrotransposon groups in which no member contains the gag, pol, or env internal domains. Because of the lack of protein-coding capacity, these groups are non-autonomous in replication, even if transcriptionally active. The Cassandra element belongs to the non-autonomous group called terminal-repeat retrotransposons in miniature (TRIM). It carries 5S RNA sequences with conserved RNA polymerase (pol) III promoters and terminators in its long terminal repeats (LTRs). Here, we identified multiple extended tandem arrays of Cassandra retrotransposons within different plant species, including ferns. At least 12 copies of repeated LTRs (as the tandem unit) and internal domain (as a spacer), giving a pattern that resembles the cellular 5S rRNA genes, were identified. A cytogenetic analysis revealed the specific chromosomal pattern of the Cassandra retrotransposon with prominent clustering at and around 5S rDNA loci. The secondary structure of the Cassandra retroelement RNA is predicted to form super-loops, in which the two LTRs are complementary to each other and can initiate local recombination, leading to the tandem arrays of Cassandra elements. The array structures are conserved for Cassandra retroelements of different species. We speculate that recombination events similar to those of 5S rRNA genes may explain the wide variation in Cassandra copy number. Likewise, the organization of 5S rRNA gene sequences is very variable in flowering plants; part of what is taken for 5S gene copy variation may be variation in Cassandra number. The role of the Cassandra 5S sequences remains to be established.

The Position and Complex Genomic Architecture of Plant T-DNA Insertions Revealed by 4SEE.

The integration of T-DNA in plant genomes is widely used for basic research and agriculture. The high heterogeneity in the number of integration events per genome, their configuration, and their impact on genome integrity highlight the critical need to detect the genomic locations of T-DNA insertions and their associated chromosomal rearrangements, and the great challenge in doing so. Here, we present 4SEE, a circular chromosome conformation capture (4C)-based method for robust, rapid, and cost-efficient detection of the entire scope of T-DNA locations. Moreover, by measuring the chromosomal architecture of the plant genome flanking the T-DNA insertions, 4SEE outlines their associated complex chromosomal aberrations. Applying 4SEE to a collection of confirmed T-DNA lines revealed previously unmapped T-DNA insertions and chromosomal rearrangements such as inversions and translocations. Uncovering such events in a feasible, robust, and cost-effective manner by 4SEE in any plant of interest has implications for accurate annotation and phenotypic characterization of T-DNA insertion mutants and transgene expression in basic science applications as well as for plant biotechnology.

Repetitive DNA in the Architecture, Repatterning, and Diversification of the Genome of Aegilops speltoides Tausch (Poaceae, Triticeae).

The genome's adaptability to environmental changes, especially during rapid climatic fluctuations, underlies the existence and evolution of species. In the wild, genetic and epigenetic genomic changes are accompanied by significant alterations in the complex nuclear repetitive DNA fraction. Current intraspecific polymorphism of repetitive DNA is closely related to ongoing chromosomal rearrangements, which typically result from erroneous DNA repair and recombination. In this study, we addressed tandem repeat patterns and interaction/reshuffling both in pollen mother cell (PMC) development and somatogenesis in the wild diploid cereal Aegilops speltoides, with a focus on genome repatterning and stabilization. Individual contrasting genotypes were investigated using the fluorescent in situ hybridization (FISH) approach by applying correlative fluorescence and electron microscopy. Species-specific Spelt1 and tribe-specific Spelt52 tandem repeats were used as the markers for monitoring somatic and meiotic chromosomal interactions and dynamics in somatic interphase nuclei. We found that, the number of tandem repeat clusters in nuclei is usually lower than the number on chromosomes due to the associations of clusters of the same type in common blocks. In addition, tightly associated Spelt1-Spelt52 clusters were revealed in different genotypes. The frequencies of nonhomologous/ectopic associations between tandem repeat clusters were revealed in a genotype-/population-specific manner. An increase in the number of tandem repeat clusters in the genome causes an increase in the frequencies of their associations. The distal/terminal regions of homologous chromosomes are separated in nuclear space, and nonhomologous chromosomes are often involved in somatic recombination as seen by frequently formed interchromosomal chromatin bridges. In both microgametogenesis and somatogenesis, inter- and intrachromosomal associations are likely to lead to DNA breaks during chromosome disjunction in the anaphase stage. Uncondensed/improperly packed DNA fibers, mainly in heterochromatic regions, were revealed in both the meiotic and somatic prophases that might be a result of broken associations. Altogether, the data obtained showed that intraorganismal dynamics of repetitive DNA under the conditions of natural out-crossing and artificial intraspecific hybridization mirrors the structural plasticity of the Ae. speltoides genome, which is interlinked with genetic diversity through the species distribution area in contrasting ecogeographical environments in and around the Fertile Crescent.

Transposable Elements in the Organization and Diversification of the Genome of Aegilops speltoides Tausch (Poaceae, Triticeae).

Repetitive DNA-specifically, transposable elements (TEs)-is a prevailing genomic fraction in cereals that underlies extensive genome reshuffling and intraspecific diversification in the wild. Although large amounts of data have been accumulated, the effect of TEs on the genome architecture and functioning is not fully understood. Here, plant genome organization was addressed by means of cloning and sequencing TE fragments of different types, which compose the largest portion of the Aegilops speltoides genome. Individual genotypes were analyzed cytogenetically using the cloned TE fragments as the DNA probes for fluorescence in situ hybridization (FISH). The obtained TE sequences of the Ty1-copia, Ty3-gypsy, LINE, and CACTA superfamilies showed the relatedness of the Ae. speltoides genome to the Triticeae tribe and similarities to evolutionarily distant species. A significant number of clones consisted of intercalated fragments of TEs of various types, in which Fatima (Ty3-gypsy) sequences predominated. At the chromosomal level, different TE clones demonstrated sequence-specific patterning, emphasizing the effect of the TE fraction on the Ae. speltoides genome architecture and intraspecific diversification. Altogether, the obtained data highlight the current species-specific organization and patterning of the mobile element fraction and point to ancient evolutionary events in the genome of Ae. speltoides.

Intraspecific and intraorganismal copy number dynamics of retrotransposons and tandem repeat in Aegilops speltoides Tausch (Poaceae, Triticeae).

Transposable elements (TE) and tandem repeats (TR) compose the largest fraction of the plant genome. The abundance and repatterning of repetitive DNA underlie intrapopulation polymorphisms and intraspecific diversification; however, the dynamics of repetitive elements in ontogenesis is not fully understood. Here, we addressed the genotype-specific and tissue-specific abundances and dynamics of the Ty1-copia, Ty3-gypsy, and LINE retrotransposons and species-specific Spelt1 tandem repeat in wild diploid goatgrass, Aegilops speltoides Tausch. Copy numbers of TEs and TR were estimated by real-time quantitative PCR in vegetative and generative tissues in original plants from contrasting allopatric populations and artificial intraspecific hybrids. The results showed that between leaves and somatic spike tissues as well as in progressive microsporogenesis of individual genotypes, the copy numbers of three TEs correlatively oscillated between 2- to 4-fold and the TR copy numbers fluctuated by 18- to 440-fold. Inter-individual and intraorganismal TEs and TR copy number dynamics demonstrate large-scale parallelism with extensive chromosomal rearrangements that were detected using fluorescent in situ hybridization in parental and hybrid genotypes. The data obtained indicate that tissue-specific differences in the abundance and pattern of repetitive sequences emerge during cell proliferation and differentiation in ontogenesis and reflect the reorganization of individual genomes in changing environments, especially in small peripheral population(s) under the influence of rapid climatic changes.

Genotype- and Cell-Specific Dynamics of Tandem Repeat Patterns in Aegilops speltoides Tausch (Poaceae, Triticeae).

In wild plant populations, chromosome rearrangements lead to the wide intraspecific polymorphisms in the abundance and patterns of highly repetitive DNA. However, despite the large amount of accumulated data, the impact of the complex repetitive DNA fraction on genome reorganization and functioning and the mechanisms balancing and maintaining the structural integrity of the genome are not fully understood. Homologous recombination is thought to play a key role in both genome reshuffling and stabilization, while the contribution of nonhomologous recombination seems to be undervalued. Here, tandem repeat patterns and dynamics during pollen mother cell development were addressed, with a focus on the meiotic recombination that determines chromosome/genome repatterning and stabilization under cross-pollination and artificial hybridization in wild goatgrass, Aegilops speltoides. Native plants from contrasting allopatric populations and artificially created intraspecific hybrids were investigated using a FISH approach. Cytogenetic analysis uncovered a wide spectrum of genotype- and cell-specific chromosomal rearrangements, suggesting intensive repatterning of both parental and hybrid genomes. The data obtained provide evidence that repetitive elements serve as overabundant and ubiquitous resources for maintaining chromosome architecture/genome integrity through homologous and nonhomologous recombination at the intraorganismal level, and genotype-specific repatterning underlies intrapopulation polymorphisms and intraspecific diversification in the wild.

B chromosomes of Aegilops speltoides are enriched in organelle genome-derived sequences.

B chromosomes (Bs) are dispensable components of the genome exhibiting non-Mendelian inheritance. Chromosome counts and flow cytometric analysis of the grass species Aegilops speltoides revealed a tissue-type specific distribution of the roughly 570 Mbp large B chromosomes. To address the question whether organelle-to-nucleus DNA transfer is a mechanism that drives the evolution of Bs, in situ hybridization was performed with labelled organellar DNA. The observed B-specific accumulation of chloroplast- and mitochondria-derived sequences suggests a reduced selection against the insertion of organellar DNA in supernumerary chromosomes. The distribution of B-localised organellar-derived sequences and other sequences differs between genotypes of different geographical origins.

Chromosome evolution in marginal populations of Aegilops speltoides: causes and consequences.

BACKGROUND: Genome restructuring is an ongoing process in natural plant populations. The influence of environmental changes on the genome is crucial, especially during periods of extreme climatic fluctuations. Interactions between the environment and the organism manifest to the greatest extent at the limits of the species' ecological niche. Thus, marginal populations are expected to exhibit lower genetic diversity and higher genetic differentiation than central populations, and some models assume that marginal populations play an important role in the maintenance and generation of biological diversity. SCOPE: In this review, long-term data on the cytogenetic characteristics of diploid Aegilops speltoides Tauch populations are summarized and discussed. This species is distributed in and around the Fertile Crescent and is proposed to be the wild progenitor of a number of diploid and polyploid wheat species. In marginal populations of Ae. speltoides, numerical chromosomal aberrations, spontaneous aneuploidy, B-chromosomes, rDNA cluster repatterning and reduction in the species-specific and tribe-specific tandem repeats have been detected. Significant changes were observed and occurred in parallel with changes in plant morphology and physiology. CONCLUSIONS: Considerable genomic variation at the chromosomal level was found in the marginal populations of Ae. speltoides. It is likely that a specific combination of gene mutations and chromosomal repatterning has produced the evolutionary trend in each specific case, i.e. for a particular species or group of related species in a given period of time and in a certain habitat. The appearance of a new chromosomal pattern is considered an important factor in promoting the emergence of interbreeding barriers.

Diversity of long terminal repeat retrotransposon genome distribution in natural populations of the wild diploid wheat Aegilops speltoides.

The environment can have a decisive influence on the structure of the genome, changing it in a certain direction. Therefore, the genomic distribution of environmentally sensitive transposable elements may vary measurably across a species area. In the present research, we aimed to detect and evaluate the level of LTR retrotransposon intraspecific variability in Aegilops speltoides (2n = 2x = 14), a wild cross-pollinated relative of cultivated wheat. The interretrotransposon amplified polymorphism (IRAP) protocol was applied to detect and evaluate the level of retrotransposon intraspecific variability in Ae. speltoides and closely related species. IRAP analysis revealed significant diversity in TE distribution. Various genotypes from the 13 explored populations significantly differ with respect to the patterns of the four explored LTR retrotransposons (WIS2, Wilma, Daniela, and Fatima). This diversity points to a constant ongoing process of LTR retrotransposon fraction restructuring in populations of Ae. speltoides throughout the species' range and within single populations in time. Maximum changes were recorded in genotypes from small stressed populations. Principal component analysis showed that the dynamics of the Fatima element significantly differ from those of WIS2, Wilma, and Daniela. In terms of relationships between Sitopsis species, IRAP analysis revealed a grouping with Ae. sharonensis and Ae. longissima forming a separate unit, Ae. speltoides appearing as a dispersed group, and Ae. bicornis being in an intermediate position. IRAP display data revealed dynamic changes in LTR retrotransposon fractions in the genome of Ae. speltoides. The process is permanent and population specific, ultimately leading to the separation of small stressed populations from the main group.

Tandem repeats on an eco-geographical scale: outcomes from the genome of Aegilops speltoides.

The chromosomal pattern of tandem repeat fractions of repetitive DNA is one of the most important characteristics of a species. In the present research, we aimed to detect and evaluate the level of intraspecific variability in the chromosomal distribution of species-specific Spelt 1 and Aegilops-Triticum-specific Spelt 52 tandem repeats in Aegilops speltoides and in closely related diploid and polyploid species. There is a distinct eco-geographical gradient in Spelt 1 and Spelt 52 blocks abundance in Ae. speltoides. In marginal populations, the number of Spelt 1 chromosomal blocks could be 12-14 times lower than in the center of the species distribution. Also, in related diploid species, the abundance of Spelt 52 correlates with evolutionary proximity to Ae. speltoides. Finally, the B- and G-genomes of allopolyploid wheats have Spelt 1 chromosomal distribution patterns similar to those of the types of Ae. speltoides with poor and rich contents of Spelt 1, respectively. The observed changes in numbers of blocks of Spelt 1 and Spelt 52 tandem repeats along the eco-geographical gradient may due to their depletion in the marginal populations as a result of increased recombination frequency under stressful conditions. Alternatively, it may be accumulation of tandem repeats in conducive climatic/edaphic environments in the center of the species' geographical distribution. Anyway, we observe a bidirectional shift of repetitive DNA genomic patterns on the population level leading to the formation of population-specific chromosomal patterns of tandem repeats. The appearance of a new chromosomal pattern is considered an important factor in promoting the emergence of interbreeding barriers.

Transposable elements in a marginal plant population: temporal fluctuations provide new insights into genome evolution of wild diploid wheat.

BACKGROUND: How new forms arise in nature has engaged evolutionary biologists since Darwin's seminal treatise on the origin of species. Transposable elements (TEs) may be among the most important internal sources for intraspecific variability. Thus, we aimed to explore the temporal dynamics of several TEs in individual genotypes from a small, marginal population of Aegilops speltoides. A diploid cross-pollinated grass species, it is a wild relative of the various wheat species known for their large genome sizes contributed by an extraordinary number of TEs, particularly long terminal repeat (LTR) retrotransposons. The population is characterized by high heteromorphy and possesses a wide spectrum of chromosomal abnormalities including supernumerary chromosomes, heterozygosity for translocations, and variability in the chromosomal position or number of 45S and 5S ribosomal DNA (rDNA) sites. We propose that variability on the morphological and chromosomal levels may be linked to variability at the molecular level and particularly in TE proliferation. RESULTS: Significant temporal fluctuation in the copy number of TEs was detected when processes that take place in small, marginal populations were simulated. It is known that under critical external conditions, outcrossing plants very often transit to self-pollination. Thus, three morphologically different genotypes with chromosomal aberrations were taken from a wild population of Ae. speltoides, and the dynamics of the TE complex traced through three rounds of selfing. It was discovered that: (i) various families of TEs vary tremendously in copy number between individuals from the same population and the selfed progenies; (ii) the fluctuations in copy number are TE-family specific; (iii) there is a great difference in TE copy number expansion or contraction between gametophytes and sporophytes; and (iv) a small percentage of TEs that increase in copy number can actually insert at novel locations and could serve as a bona fide mutagen. CONCLUSIONS: We hypothesize that TE dynamics could promote or intensify morphological and karyotypical changes, some of which may be potentially important for the process of microevolution, and allow species with plastic genomes to survive as new forms or even species in times of rapid climatic change.

En/Spm-like transposons in Poaceae species: transposase sequence variability and chromosomal distribution.

Belonging to Class II of transposable elements, En/Spm transposons are widespread in a variety of distantly related plant species. Here, we report on the sequence conservation of the transposase region from sequence analyses of En/Spm-like transposons from Poaceae species, namely Zingeria biebersteiniana, Zingeria trichopoda, Triticum monococcum, Triticum urartu, Hordeum spontaneum, and Aegilops speltoides. The transposase region of En/Spm-like transposons was cloned, sequenced, and compared with equivalent regions of Oryza and Arabidopsis from the gene bank database. Southern blot analysis indicated that the En/Spm transposon was present in low (Hordeum spontaneum, Triticum monococcum, Triticum urartu) through medium (Zingeria bieberstiana, Zingeria trichopoda) to relatively high (Aegilops speltoides) copy numbers in Poaceae species. A cytogenetic analysis of the chromosomal distribution of En/Spm transposons revealed the concurence of the chromosomal localization of the En/Spm clusters with mobile clusters of rDNA. An analysis of En/Spm-like transposase amino acid sequences was carried out to investigate sequence divergence between 5 genera--Triticum, Aegilops, Zingeria, Oryza and Arabidopsis. A distance matrix was generated; apparently, En/Spm-like transposase sequences shared the highest sequence homology intra-generically and, as expected, these sequences were significantly diverged from those of O. sativa and A. thaliana. A sequence comparison of En/Spm-like transposase coding regions defined that the intra-genomic complex of En/Spm-like transposons could be viewed as relatively independent, vertically transmitted, and permanently active systems inside higher plant genomes. The sequence data from this article was deposited in the EMBL/GenBank Data Libraries under the accession nos. AY707995-AY707996-AY707997-AY707998-AY707999-AY708000-AY708001-AY708002-AY708003-AY708004-AY708005-AY708005-AY265312.

Ac-like transposons in populations of wild diploid Triticeae species: comparative analysis of chromosomal distribution.

Data are presented on the intra- and interspecific differences/similarities in chromosomal patterns of Ac-like elements (hAT family) in ecologically contrasted populations of three Triticeae species - Aegilops speltoides, Triticum urartu, and Hordeum spontaneum. Application of original computer software made it possible to precisely map transposon clusters and to link them to known chromosomal markers (rDNA sites, centromeres, and heterochromatin regions). From our data we can specify the most visible features of Ac-like elements chromosomal distribution: preferential concentration in chromosomal proximal regions; high percentage of clusters on the border between euchromatin and heterochromatin; complementary chromosomal arrangement towards En/Spm transposons (CACTA); population-specific insertions into centromeres; more differences in total cluster numbers between populations of self-pollinated species than between populations of cross-pollinated species. The application of statistical simulation (Resampling) method to analysis of data indicates that ecology may play a certain role in dynamics of Ac-like elements. Comparison of real Ayala distances, as well as real chromosomal distribution of Ac-like elements in populations of two species with different mating systems with the same but randomly simulated parameters, revealed that non-random population structure in the Mediterranean floral zone suffers and becomes chaotic in the Irano-Turanian zone.

Quantum speciation in Aegilops: molecular cytogenetic evidence from rDNA cluster variability in natural populations.

Data are presented on quantum speciation in the Sitopsis section of the genus Aegilops (Poaceae, Monocotyledones). Two small, peripheral, isolated, wild populations of annual cross-pollinated Ae. speltoides and annual self-pollinated Ae. sharonensis are located 30 m apart on different soil types. Despite the close proximity of the two species and their close relatedness, no mixed groups are known. Comparative molecular cytogenetic analysis based on the intrapopulation variability of rRNA-encoding DNA (rDNA) chromosomal patterns of individual Ae. speltoides geno-types revealed an ongoing dynamic process of permanent chromosomal rearrangements. Chromosomal mutations can arise de novo and can be eliminated. Analysis of the progeny of the investigated genotypes testifies that inheritance of de novo rDNA sites happens frequently. Heterologous recombination and/or transposable elements-mediated rDNA transfer seem to be the mechanisms for observed chromosomal repatterning. Consequently, several modified genomic forms, intermediate between Ae. speltoides and Ae. sharonensis, permanently arise in the studied wild population of Ae. speltoides, which make it possible to recognize Ae. sharonensis as a derivative species of Ae. speltoides, as well as to propose rapidness and canalization of quantum speciation in Sitopsis species.

The utility of the nontranscribed spacer of 5S rDNA units grouped into unit classes assigned to haplomes - a test on cultivated wheat and wheat progenitors.

Data is presented on the evolutionary dynamics of non-transcribed spacers (NTSs) of 5S rRNA genes in some diploid and polyploid Triticum and Aegilops species. FISH experiments with probes representing different unit classes revealed presence and (or) absence of these sequences in genomes or separate chromosomes of the species. Among the three diploid species only Aegilops speltoides has all of the different unit classes in ribosomal clusters as detected by the probes. Triticum urartu does not have the long D1 signals and Aegilops tauschii does not have the long A1 signals. Both polyploids possess all types of sequences, but because of genome rearrangements after polyploidization there is significant repatterning of single different rDNA unit classes in chromosomal positions when compared with those in diploid progenitors. Additional refined work is needed to ascertain if the sequences in the polyploids are mixed or are located in mini clusters in close proximity to each other. Mantel tests for association between the presence of the FISH signals of the A, B, and D genomes together and separately with the unit class data of the material, i.e., the probes used in FISH, indicated that all signals were associated with their respective probe material, but that there was no association of the unit classes found and the signals to each haplome. All combinations of the partial Mantel tests, e.g., between the A and B haplomes while controlling the effect of the all probes signals, with correlations ranging from 0.48 to 0.79 were all significant. Principal coordinate analysis showed that the signals of most unit class specific probes were more or less equally distant except for the long (S1 and short G1 signals, which were not different, and that the short A1 signals were closely related to the former two, whereas the signals of the long G1 were even less related.

Activity of the En/Spm-like transposons in meiosis as a base for chromosome repatterning in a small, isolated, peripheral population of Aegilops speltoides Tausch.

Chromosomal repatterning is considered to be one of the main mechanisms for plant genome evolution. Here, we report the first cytogenetic evidence for the involvement of En/Spm transposons in ongoing chromosomal repatterning leading to the rise of new fertile genomic forms in a small, isolated, peripheral plant population. Cytogenetical screening of original individual plants of Aegilops speltoides Tausch. with different phenotypes revealed a wide spectrum of chromosomal abnormalities including extra chromosomes, chromosomal rearrangements, and variability in chromosomal position/number of 45S and 5S rDNA sites. Cytogenetic analysis of the dynamics of En/Spm transposons in meiosis indicates that: (i) this type of transposon is active during male gametogenesis; (ii) separately or in conjunction with rDNA they form clusters in the hot spots of large chromosomal rearrangements; (iii) appearance of at least part of the mobile rDNA sites in genome of Ae. speltoides are connected with meiotic activity of En/Spm transposons. PCR screening for the site-selected transposon insertions confirm the presence of combined fragments that consist partly of the sequence of En/Spm transposon and partly of 5S rDNA sequence.

Repetitive DNas of wild emmer wheat (Triticum dicoccoides) and their relation to S-genome species: molecular cytogenetic analysis.

We have analyzed the chromosomal GISH molecular banding patterns of three populations of the wild allopolyploid wheat Triticum dicoccoides in an attempt to unravel the evolutionary relationships between highly repetitive DNA fractions of T. dicoccoides and proposed diploid progenitors of the B genome. Aegilops speltoides showed almost complete affinity of its repetitive DNA to C-heterochromatin of T. dicoccoides, whereas other S-genome species demonstrated relatedness only to distal heterochromatin. This substantiates the priority of Ae. speltoides as the most similar to the wheat B-genome donor in comparison with other Sitopsis species. Using molecular banding technique with DNA of different Aegilops species as a probe permits tracing of the origin of each heterochromatin cluster. Molecular banding analysis reveals polymorphism between three wild emmer wheat populations. Comparison of molecular banding patterns with chromosomal distribution of the Ty1-copia retrotransposons, which constitute a large share of T. dicoccoides genome, makes it possible to propose that the activity of transposable elements may lie in the background of observed intraspecific polymorphism.