BACKGROUND: The genus Pulmonaria (Boraginaceae) represents a taxonomically complex group of species in which morphological similarity contrasts with striking karyological variation. The presence of different numbers of chromosomes in the diploid state suggests multiple hybridization/polyploidization events followed by chromosome rearrangements (dysploidy). Unfortunately, the phylogenetic relationships and evolution of the genome, have not yet been elucidated. Our study focused on the P. officinalis group, the most widespread species complex, which includes two morphologically similar species that differ in chromosome number, i.e. P. obscura (2n = 14) and P. officinalis (2n = 16). Ornamental cultivars, morphologically similar to P. officinalis (garden escapes), whose origin is unclear, were also studied. Here, we present a pilot study on genome size and repeatome dynamics of these closely related species in order to gain new information on their genome and chromosome structure. RESULTS: Flow cytometry confirmed a significant difference in genome size between P. obscura and P. officinalis, corresponding to the number of chromosomes. Genome-wide repeatome analysis performed on genome skimming data showed that retrotransposons were the most abundant repeat type, with a higher proportion of Ty3/Gypsy elements, mainly represented by the Tekay lineage. Comparative analysis revealed no species-specific retrotransposons or striking differences in their copy number between the species. A new set of chromosome-specific cytogenetic markers, represented by satellite DNAs, showed that the chromosome structure in P. officinalis was more variable compared to that of P. obscura. Comparative karyotyping supported the hybrid origin of putative hybrids with 2n = 15 collected from a mixed population of both species and outlined the origin of ornamental garden escapes, presumably derived from the P. officinalis complex. CONCLUSIONS: Large-scale genome size analysis and repeatome characterization of the two morphologically similar species of the P. officinalis group improved our knowledge of the genome dynamics and differences in the karyotype structure. A new set of chromosome-specific cytogenetic landmarks was identified and used to reveal the origin of putative hybrids and ornamental cultivars morphologically similar to P. officinalis.

• Klíčová slova
• Pulmonaria, Comparative karyotyping, Genome size, Repeatome, Satellite DNA,
• MeSH
• chromozomy rostlin * genetika   MeSH
• délka genomu MeSH
• fylogeneze MeSH
• genom rostlinný * MeSH
• karyotyp MeSH
• karyotypizace * MeSH
• plicník genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Genome size variation is a crucial aspect of plant evolution, influenced by a complex interplay of factors. Repetitive elements, which are fundamental components of genomic architecture, often play a role in genome expansion by selectively amplifying specific repeat motifs. This study focuses on Amomum, a genus in the ginger family (Zingiberaceae), known for its 4.4-fold variation in genome size. Using a robust methodology involving PhyloNet reconstruction, RepeatExplorer clustering, and repeat similarity-based phylogenetic network construction, we investigated the repeatome composition, analyzed repeat dynamics, and identified potential hybridization events within the genus. Our analysis confirmed the presence of four major infrageneric clades (A-D) within Amomum, with clades A-C exclusively comprising diploid species (2n = 48) and clade D encompassing both diploid and tetraploid species (2n = 48 and 96). We observed an increase in the repeat content within the genus, ranging from 84% to 89%, compared to outgroup species with 75% of the repeatome. The SIRE lineage of the Ty1-Copia repeat superfamily was prevalent in most analyzed ingroup genomes. We identified significant difference in repeatome structure between the basal Amomum clades (A, B, C) and the most diverged clade D. Our investigation revealed evidence of ancient hybridization events within Amomum, coinciding with a substantial proliferation of multiple repeat groups. This finding supports the hypothesis that ancient hybridization is a driving force in the genomic evolution of Amomum. Furthermore, we contextualize our findings within the broader context of genome size variations and repeatome dynamics observed across major monocot lineages. This study enhances our understanding of evolutionary processes within monocots by highlighting the crucial roles of repetitive elements in shaping genome size and suggesting the mechanisms that drive these changes.

• Klíčová slova
• 5S rDNA, Zingiberaceae, genome evolution, genome size, interspecific hybridization, phylogeny, repeatome, repetitive DNA,
• Publikační typ
• časopisecké články MeSH

The banana is a staple food crop and represents an important trade commodity for millions of people living in tropical and subtropical countries. The most important edible banana clones originated from natural crosses between diploid Musa balbisiana and various subspecies of M. acuminata. It is worth mentioning that evolution and speciation in the Musaceae family were accompanied by large-scale chromosome structural changes, indicating possible reasons for lower fertility or complete sterility of these vegetatively propagated clones. Chromosomal changes, often accompanied by changes in genome size, are one of the driving forces underlying speciation in plants. They can clarify the genomic constitution of edible bananas and shed light on their origin and on diversification processes in members of the Musaceae family. This article reviews the development of molecular cytogenetic approaches, ranging from classical fluorescence in situ hybridization (FISH) using common cytogenetic markers to oligo painting FISH. We discuss differences in genome size and chromosome number across the Musaceae family in addition to the development of new chromosome-specific cytogenetic probes and their use in genome structure and comparative karyotype analysis. The impact of these methodological advances on our knowledge of Musa genome evolution at the chromosomal level is demonstrated. In addition to citing published results, we include our own new unpublished results and outline future applications of molecular cytogenetics in banana research.

• Klíčová slova
• BAC clones, DNA repeats, chromosomes, flow cytometry, fluorescence in situ hybridization, karyotyping, oligo painting, rRNA genes,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Trifolium L. is an economically important genus that is characterized by variable karyotypes relating to its ploidy level and basic chromosome numbers. The advent of genomic resources combined with molecular cytogenetics provides an opportunity to develop our understanding of plant genomes in general. Here, we summarize the current state of knowledge on Trifolium genomes and chromosomes and review methodologies using molecular markers that have contributed to Trifolium research. We discuss possible future applications of cytogenetic methods in research on the Trifolium genome and chromosomes.

• Klíčová slova
• chromosomal markers, clover, cytogenetics, genome size, interspecific hybridization, polyploidy, synteny,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

BACKGROUND: Cultivated grasses are an important source of food for domestic animals worldwide. Increased knowledge of their genomes can speed up the development of new cultivars with better quality and greater resistance to biotic and abiotic stresses. The most widely grown grasses are tetraploid ryegrass species (Lolium) and diploid and hexaploid fescue species (Festuca). In this work, we characterized repetitive DNA sequences and their contribution to genome size in five fescue and two ryegrass species as well as one fescue and two ryegrass cultivars. RESULTS: Partial genome sequences produced by Illumina sequencing technology were used for genome-wide comparative analyses with the RepeatExplorer pipeline. Retrotransposons were the most abundant repeat type in all seven grass species. The Athila element of the Ty3/gypsy family showed the most striking differences in copy number between fescues and ryegrasses. The sequence data enabled the assembly of the long terminal repeat (LTR) element Fesreba, which is highly enriched in centromeric and (peri)centromeric regions in all species. A combination of fluorescence in situ hybridization (FISH) with a probe specific to the Fesreba element and immunostaining with centromeric histone H3 (CENH3) antibody showed their co-localization and indicated a possible role of Fesreba in centromere function. CONCLUSIONS: Comparative repeatome analyses in a set of fescues and ryegrasses provided new insights into their genome organization and divergence, including the assembly of the LTR element Fesreba. A new LTR element Fesreba was identified and found in abundance in centromeric regions of the fescues and ryegrasses. It may play a role in the function of their centromeres.

• Klíčová slova
• Centromere organization, Festuca, Illumina sequencing, Lolium, Repetitive DNA,
• MeSH
• centromera genetika   MeSH
• chromozomy rostlin * MeSH
• Festuca genetika   MeSH
• genom rostlinný genetika   MeSH
• jílek genetika   MeSH
• repetitivní sekvence nukleových kyselin * MeSH
• Publikační typ
• časopisecké články MeSH

Plant genomes vary greatly in composition and size mainly due to the diversity of repetitive DNAs and the inherent propensity for their amplification and removal from the host genome. Most studies addressing repeatome dynamics focus on model organisms, whereas few provide comprehensive investigations across the genomes of related taxa. Herein, we analyze the evolution of repeats of the 13 species in Melampodium sect. Melampodium, representing all but two of its diploid taxa, in a phylogenetic context. The investigated genomes range in size from 0.49 to 2.27 pg/1C (ca. 4.5-fold variation), despite having the same base chromosome number (x = 10) and very strong phylogenetic affinities. Phylogenetic analysis performed in BEAST and ancestral genome size reconstruction revealed mixed patterns of genome size increases and decreases across the group. High-throughput genome skimming and the RepeatExplorer pipeline were utilized to determine the repeat families responsible for the differences in observed genome sizes. Patterns of repeat evolution were found to be highly correlated with phylogenetic position, namely taxonomic series circumscription. Major differences found were in the abundances of the SIRE (Ty1-copia), Athila (Ty3-gypsy), and CACTA (DNA transposon) lineages. Additionally, several satellite DNA families were found to be highly group-specific, although their overall contribution to genome size variation was relatively small. Evolutionary changes in repetitive DNA composition and genome size were complex, with independent patterns of genome up- and downsizing throughout the evolution of the analyzed diploids. A model-based analysis of genome size and repetitive DNA composition revealed evidence for strong phylogenetic signal and differential evolutionary rates of major lineages of repeats in the diploid genomes.

• Klíčová slova
• Bayesian analysis, Melampodium, ancestral state reconstruction, genome size, phylogenetics, repetitive DNA, tandem repeats, transposable elements,
• Publikační typ
• časopisecké články MeSH

Oligo painting FISH was established to identify all chromosomes in banana (Musa spp.) and to anchor pseudomolecules of reference genome sequence of Musa acuminata spp. malaccensis "DH Pahang" to individual chromosomes in situ. A total of 19 chromosome/chromosome-arm specific oligo painting probes were developed and were shown to be suitable for molecular cytogenetic studies in genus Musa. For the first time, molecular karyotypes of diploid M. acuminata spp. malaccensis (A genome), M. balbisiana (B genome), and M. schizocarpa (S genome) from the Eumusa section of Musa, which contributed to the evolution of edible banana cultivars, were established. This was achieved after a combined use of oligo painting probes and a set of previously developed banana cytogenetic markers. The density of oligo painting probes was sufficient to study chromosomal rearrangements on mitotic as well as on meiotic pachytene chromosomes. This advance will enable comparative FISH mapping and identification of chromosomal translocations which accompanied genome evolution and speciation in the family Musaceae.

• Klíčová slova
• Musa, banana, chromosome identification, fluorescence in situ hybridization, molecular karyotype, oligo painting FISH,
• Publikační typ
• časopisecké články MeSH

In many plant species, somatic cell differentiation is accompanied by endoreduplication, a process during which cells undergo one or more rounds of DNA replication cycles in the absence of mitosis, resulting in nuclei with multiples of 2C DNA amounts (4C, 8C, 16C, etc.). In some orchids, a disproportionate increase in nuclear DNA contents has been observed, where successive endoreduplication cycles result in DNA amounts 2C + P, 2C + 3P, 2C + 7P, etc., where P is the DNA content of the replicated part of the 2C nuclear genome. This unique phenomenon was termed "progressively partial endoreplication" (PPE). We investigated processes behind the PPE in Ludisia discolor using flow cytometry (FCM) and Illumina sequencing. In particular, we wanted to determine whether chromatin elimination or incomplete genome duplication was involved, and to identify types of DNA sequences that were affected. Cell cycle analysis of root tip cell nuclei pulse-labeled with EdU revealed two cell cycles, one ending above the population of nuclei with 2C + P content, and the other with a typical "horseshoe" pattern of S-phase nuclei ranging from 2C to 4C DNA contents. The process leading to nuclei with 2C + P amounts therefore involves incomplete genome replication. Subsequent Illumina sequencing of flow-sorted 2C and 2C + P nuclei showed that all types of repetitive DNA sequences were affected during PPE; a complete elimination of any specific type of repetitive DNA was not observed. We hypothesize that PPE is part of a highly controlled transition mechanism from proliferation phase to differentiation phase of plant tissue development.

• Klíčová slova
• DNA replication, EdU, Ludisia discolor, cell cycle, endoreduplication, orchids,
• MeSH
• buněčné jádro genetika   MeSH
• endoreduplikace genetika   MeSH
• genom rostlinný MeSH
• listy rostlin genetika   MeSH
• mitóza genetika   MeSH
• Orchidaceae genetika   MeSH
• polyploidie MeSH
• průtoková cytometrie metody   MeSH
• replikace DNA genetika   MeSH
• vysoce účinné nukleotidové sekvenování metody   MeSH
• Publikační typ
• časopisecké články MeSH

The differential accumulation and elimination of repetitive DNA are key drivers of genome size variation in flowering plants, yet there have been few studies which have analysed how different types of repeats in related species contribute to genome size evolution within a phylogenetic context. This question is addressed here by conducting large-scale comparative analysis of repeats in 23 species from four genera of the monophyletic legume tribe Fabeae, representing a 7.6-fold variation in genome size. Phylogenetic analysis and genome size reconstruction revealed that this diversity arose from genome size expansions and contractions in different lineages during the evolution of Fabeae. Employing a combination of low-pass genome sequencing with novel bioinformatic approaches resulted in identification and quantification of repeats making up 55-83% of the investigated genomes. In turn, this enabled an analysis of how each major repeat type contributed to the genome size variation encountered. Differential accumulation of repetitive DNA was found to account for 85% of the genome size differences between the species, and most (57%) of this variation was found to be driven by a single lineage of Ty3/gypsy LTR-retrotransposons, the Ogre elements. Although the amounts of several other lineages of LTR-retrotransposons and the total amount of satellite DNA were also positively correlated with genome size, their contributions to genome size variation were much smaller (up to 6%). Repeat analysis within a phylogenetic framework also revealed profound differences in the extent of sequence conservation between different repeat types across Fabeae. In addition to these findings, the study has provided a proof of concept for the approach combining recent developments in sequencing and bioinformatics to perform comparative analyses of repetitive DNAs in a large number of non-model species without the need to assemble their genomes.

• MeSH
• délka genomu * MeSH
• Fabaceae klasifikace   genetika   MeSH
• fylogeneze MeSH
• genetická variace * MeSH
• genom rostlinný * MeSH
• genomika * metody   MeSH
• koncové repetice MeSH
• molekulární evoluce MeSH
• repetitivní sekvence nukleových kyselin * MeSH
• reprodukovatelnost výsledků MeSH
• sekvenční analýza DNA MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH