Intraspecific Genome Size Variation Research Articles

Quantifying the role of genome size and repeat content in adaptive variation and the architecture of flowering time in Amaranthus tuberculatus.

Genome size variation, largely driven by repeat content, is poorly understood within and among populations, limiting our understanding of its significance for adaptation. Here we characterize intraspecific variation in genome size and repeat content across 186 individuals of Amaranthus tuberculatus, a ubiquitous native weed that shows flowering time adaptation to climate across its range and in response to agriculture. Sequence-based genome size estimates vary by up to 20% across individuals, consistent with the considerable variability in the abundance of transposable elements, unknown repeats, and rDNAs across individuals. The additive effect of this variation has important phenotypic consequences-individuals with more repeats, and thus larger genomes, show slower flowering times and growth rates. However, compared to newly-characterized gene copy number and polygenic nucleotide changes underlying variation in flowering time, we show that genome size is a marginal contributor. Differences in flowering time are reflected by genome size variation across sexes and marginally, habitats, while polygenic variation and a gene copy number variant within the ATP synthesis pathway show consistently stronger environmental clines than genome size. Repeat content nonetheless shows non-neutral distributions across the genome, and across latitudinal and environmental gradients, demonstrating the numerous governing processes that in turn influence quantitative genetic variation for phenotypes key to plant adaptation.

Genome streamlining and clonal erosion in nutrient-limited environments: a test using genome-size variable populations.

Some selection-based theories propose that genome streamlining, favoring smaller genome sizes, is advantageous in nutritionally limited environments, particularly under P-limitation. To test this prediction, we conducted several experimental evolution trials on clonal populations of a facultatively asexual rotifer that exhibits intraspecific variation in genome size. Most trials showed a rapid decline in clonal diversity, which was accelerated in populations that were initially non-adapted. Populations consisting of three rotifer clones often became monoclonal within a few weeks, while populations starting with 120 clones eroded to ten multi-locus genotypes, of which only five were abundant in higher numbers. While P-limitation affected population growth during the experiments, it did not affect the outcome of clonal competition or the speed at which clonal diversity was lost. Common garden transplant experiments revealed that the evolved populations were better adapted to the experimental conditions than the ancestral controls. However, contrary to expectations, the evolved populations did not show an overrepresentation of small genomes. Intermediate genomes were also frequently abundant, although very large genomes were rare. Our findings suggest that fitness is more influenced by genotypic differences among clones than by differences in GS, and indicate that such differences might hinder genome streamlining during early adaptation to a new environment.

Determination of genome size variation among varieties of Ilex cornuta (Aquifoliaceae) by fow cytometry

Ilex cornuta Lindl. & Paxton is a commercially important horticultural species worldwide, and extensive cultivation and hybridization have produced many varieties. Despite the considerable breeding, selection, widespread cultivation and domestication, which may have a significant role in the composition of genomes, there are no other previous reports of intraspecific genome size variation in the different cultivars or hybrids of this species. In the present work, genome size of 12 varieties of I. cornuta was assessed and analyzed through high-resolution flow cytometry (FCM). Nuclear DNA was analyzed using nuclei isolated from young leaves, which used propidium iodide (PI) staining, with rice (Oryza sativa cv. Nipponbare) as internal reference. As a result, statistically significant differences in genome size were detected among all diploid I. cornuta varieties considered. The estimated genome size (2C value) of I. cornuta varieties ranged from 1.47 to 1.80 pg, with 1.22-fold variation and an average size of 1.65 pg. The domestication and interspecific hybridization induced variation of genome size in I. cornuta, and the genome size of hybrids exhibited a wider range of variation compared with that of cultivars. In summary, flow cytometry is a useful tool to analyze the genome size of I. cornuta. The first report of the genome sizes of varieties of this species would provide useful data for further research on I. cornuta, and enrich the C value database of Ilex L. What’s more, our findings could be the foundation in the future of I. cornuta genome sequencing and breeding programs.

Genome size variation within Crithmum maritimum: Clues on the colonization of insular environments.

Angiosperms present an astonishing diversity of genome sizes that can vary intra- or interspecifically. The remarkable new cytogenomic data shed some light on our understanding of evolution, but few studies were performed with insular and mainland populations to test possible correlations with dispersal, speciation, and adaptations to insular environments. Here, patterns of cytogenomic diversity were assessed among geographic samples (ca. 114) of Crithmum maritimum (Apiaceae), collected across the Azores and Madeira archipelagos, as well as in adjacent continental areas of Portugal. Using flow cytometry, the results indicated a significant intraspecific genome size variation, spanning from reduced sizes in the insular populations to larger ones in the mainland populations. Moreover, there was a tendency for an increase in genome size along the mainland populations, associated with lower temperatures, higher precipitation, and lower precipitation seasonality. However, this gradient might be the result of historic phylogeographical events associated with previous dispersal and extinction of local populations. Overall, our findings provided evidence that smaller genome sizes might play a critical role in the colonization of islands, corroborating other studies that argue that organisms with smaller genomes use fewer resources, having a selective advantage under insular environments. Although further studies are needed to improve our understanding of the mechanisms underlying genome size evolution on islands, conservation strategies must be promoted to protect the rich cytogenomic diversity found among C. maritimum populations, which occur in coastal areas that are particularly threatened by human activity, pollution, invasive species, and climate changes.

Intraspecific variation in genome size in Artemisia argyi determined using flow cytometry and a genome survey.

Different collections and accessions of Artemisia argyi (Chinese mugwort) harbour considerable diversity in morphology and bioactive compounds, but no mechanisms have been reported that explain these variations. We studied genome size in A. argyi accessions from different regions of China by flow cytometry. Genome size was significantly distinct among origins of these 42 Chinese mugwort accessions, ranging from 8.428 to 11.717pg. There were no significant intraspecific differences among the 42 accessions from the five regions of China. The clustering analysis showed that these 42 A. argyi accessions could be divided into three groups, which had no significant relationship with geographical location. In a genome survey, the total genome size of A. argyi (A15) was estimated to be 7.852Gb (or 8.029pg) by K-mer analysis. This indicated that the results from the two independent methods are consistent, and that the genome survey can be used as an adjunct to flow cytometry to compensate for its deficiencies. In addition, genome survey can provide the information about heterozygosity, repeat sequences, GC content and ploidy of A. argyi genome. The nuclear DNA contents determined here provide a new reference for intraspecific variation in genome size in A. argyi, and may also be a potential resource for the study of genetic diversity and for breeding new cultivar.

Meiotic transmission patterns of additional genomic elements in Brachionus asplanchnoidis, a rotifer with intraspecific genome size variation

Intraspecific genome size (GS) variation in Eukaryotes is often mediated by additional, nonessential genomic elements. Physically, such additional elements may be represented by supernumerary (B-)chromosomes or by large heterozygous insertions into the regular chromosome set. Here we analyze meiotic transmission patterns of Megabase-sized, independently segregating genomic elements (ISEs) in Brachionus asplanchnoidis, a planktonic rotifer that displays an up to two-fold intraspecific GS variation due to variation in size and number of these elements. To gain insights into the meiotic transmission patterns of ISEs, we measured GS distributions of haploid males produced by individual mother clones using flow cytometry and compared these distributions to theoretical distributions expected under a range of scenarios. These scenarios considered transmission biases resembling (meiotic) drive, or cosegregation biases, e.g., if pairs of ISEs preferentially migrated towards the same pole during meiosis. We found that the inferred transmission patterns were diverse and ranged from positive biases (suggesting drive) to negative biases (suggesting drag), depending on rotifer clone and its ISE composition. Additionally, we obtained evidence for a negative cosegregation bias in some of the rotifer clones, i.e., pairs of ISEs exhibited an increased probability of migrating towards opposite poles during meiosis. Strikingly, these transmission and segregation patterns were more similar among members of a genetically homogeneous inbred line than among outbred members of the population. Comparisons between early and late stages of haploid male embryonic development (e.g., young synchronized male eggs vs. hatched males) showed very similar GS distributions, suggesting that transmission biases occur very early in male development, or even during meiosis. Very large genome size was associated with reduced male embryonic survival, suggesting that excessive amounts of ISEs might be detrimental to male fitness. Altogether, our results indicate considerable functional diversity of ISEs in B. asplanchnoidis, with consequences on meiotic transmission and embryonic survival.

CAUSAS Y CONSECUENCIAS DE LA VARIACIÓN DEL CONTENIDO DE ADN EN ZEA

Cytogenetic evidence indicates that Zea, which comprises maize (Z. mays ssp. mays) and its wild relatives, is an allopolyploid genus. Our research group has carried out numerous cytogenetic studies on Zea species, mainly focused on native Argentinian and Bolivian maize landraces. We found a wide inter- and intraspecific genome size variation in the genus, with mean 2C-values ranging between 4.20 and 11.36 pg. For the maize landraces studied here, it varied between 4.20 and 6.75 pg. The objectives of this work are to analyze the causes of genome size variation and to discuss their adaptive value in Zea. This variation is mainly attributed to differences in the heterochromatin located in the knobs and to the amount of interspersed DNA from retrotransposons. Polymorphisms in presence or absence of B-chromosomes (Bs) and the population frequency of Bs are also a source of genome size variation, with doses ranging between one and eight in the landraces analyzed here. Correlation analysis revealed that the percentage of heterochromatin is positively correlated with genome size. In addition, populations cultivated at higher altitudes, which are known to be precocious, have smaller genome sizes than do those growing at lower altitudes. This information, together with the positive correlation observed between the length of the vegetative cycle and the percentage of heterochromatin, led us to propose that it has an adaptive role. On the other hand, the negative relationship found between Bs and heterochromatic knobs allowed us to propose the existence of an intragenomic conflict between these elements. We hypothesize that an optimal nucleotype may have resulted from such intranuclear conflict, where genome adjustments led to a suitable length of the vegetative cycle for maize landraces growing across altitudinal clines. Key words: B chromosomes, heterochromatin, intragenomic conflict, knobs, maize landraces

Genome Size Variation in Dianthus sylvestris Wulfen sensu lato (Caryophyllaceae).

Genome size (GS) is an important characteristic that may be helpful in delimitation of taxa, and multiple studies have shown correlations between intraspecific GS variation and morphological or environmental factors, as well as its geographical segregation. We estimated a relative GS (RGS) of 707 individuals from 162 populations of Dianthus sylvestris with a geographic focus on the Balkan Peninsula, but also including several populations from the European Alps. Dianthus sylvestris is morphologically variable species thriving in various habitats and six subspecies have been recognized from the Balkan Peninsula. Our RGS data backed-up with chromosome counts revealed that the majority of populations were diploid (2n = 30), but ten tetraploid populations have been recorded in D. sylvestris subsp. sylvestris from Istria (Croatia, Italy). Their monoploid RGS is significantly lower than that of the diploids, indicating genome downsizing. In addition, the tetraploids significantly differ from their diploid counterparts in an array of morphological and environmental characteristics. Within the diploid populations, the RGS is geographically and only partly taxonomically correlated, with the highest RGS inferred in the southern Balkan Peninsula and the Alps. We demonstrate greater RGS variation among the Balkan populations compared to the Alps, which is likely a result of more pronounced evolutionary differentiation within the Balkan Peninsula. In addition, a deep RGS divergence within the Alps likely points to persistence of the alpine populations in different Pleistocene refugia.

The nature of intraspecific and interspecific genome size variation in taxonomically complex eyebrights.

Background and aimsGenome size varies considerably across the diversity of plant life. Although genome size is, by definition, affected by genetic presence/absence variants, which are ubiquitous in population sequencing studies, genome size is often treated as an intrinsic property of a species. Here, we studied intra- and interspecific genome size variation in taxonomically complex British eyebrights (Euphrasia, Orobanchaceae). Our aim is to document genome size diversity and investigate underlying evolutionary processes shaping variation between individuals, populations and species.MethodsWe generated genome size data for 192 individuals of diploid and tetraploid Euphrasia and analysed genome size variation in relation to ploidy, taxonomy, population affiliation and geography. We further compared the genomic repeat content of 30 samples.Key resultsWe found considerable intraspecific genome size variation, and observed isolation-by-distance for genome size in outcrossing diploids. Tetraploid Euphrasia showed contrasting patterns, with genome size increasing with latitude in outcrossing Euphrasia arctica, but with little genome size variation in the highly selfing Euphrasia micrantha. Interspecific differences in genome size and the genomic proportions of repeat sequences were small.ConclusionsWe show the utility of treating genome size as the outcome of polygenic variation. Like other types of genetic variation, such as single nucleotide polymorphisms, genome size variation may be affected by ongoing hybridization and the extent of population subdivision. In addition to selection on associated traits, genome size is predicted to be affected indirectly by selection due to pleiotropy of the underlying presence/absence variants.

Plant material selection, collection, preservation, and storage for nuclear DNA content estimation.

In theory, any plant tissue providing intact nuclei in sufficient quantity is suitable for nuclear DNA content estimation using flow cytometry (FCM). While this certainly opens a wide variety of possible applications of FCM, especially when compared to classical karyological techniques restricted to tissues with active cell division, tissue selection and quality may directly affect the precision (and sometimes even reliability) of FCM measurements. It is usually convenient to first consider the goals of the study to either aim for the highest possible accuracy of estimates (e.g., for inferring genome size, detecting homoploid intraspecific genome size variation, aneuploidy, among others), or to decide that histograms of reasonable resolution provide sufficient information (e.g., ploidy level screening within a single model species). Here, a set of best practices guidelines for selecting the optimal plant tissue for FCM analysis, sampling of material, and material preservation and storage are provided. In addition, factors potentially compromising the quality of FCM estimates of nuclear DNA content and data interpretation are discussed.

Genome size variation in natural populations of wild potato species from west Argentina

Wild potato species (Solanum, section Petota) have great ecological adaptability and represent essential genetic resources for the improvement of the third most important food crop worldwide. From more than 100 species described for this section, the genome size has been established for only 12. The aims of this work were to: contribute data on the C-values for Solanum species; assess the degree of interspecific and intraspecific genome size variation; investigate correlations between genome size and bioclimatic variables; and determine whether the data for genome size are helpful for delineating taxa. The DNA contents of 97 genotypes from 28 populations and two accessions of seven wild potato species, including diploids, triploids, and tetraploids, were measured by flow cytometry. Statistical genome size differences within and among species were found. The natural interploidal hybrid Solanum × rechei had the highest intraspecific genome size variation. The tetraploid S. acaule presented the smallest monoploid genome size (0.72 pg), whereas the triploid cytotype of S. microdontum was the largest (0.82 pg). Correlations among bioclimatic and geographic variables and genome size were found in most species. The roles of hybridisation and polyploidisation events on genome size variability are discussed.

Substantial intraspecific genome size variation in golden-brown algae and its phenotypic consequences.

While nuclear DNA content variation and its phenotypic consequences have been well described for animals, vascular plants and macroalgae, much less about this topic is known regarding unicellular algae and protists in general. The dearth of data is especially pronounced when it comes to intraspecific genome size variation. This study attempts to investigate the extent of intraspecific variability in genome size and its adaptive consequences in a microalgal species. Propidium iodide flow cytometry was used to estimate the absolute genome size of 131 strains (isolates) of the golden-brown alga Synura petersenii (Chrysophyceae, Stramenopiles), identified by identical internal transcribed spacer (ITS) rDNA barcodes. Cell size, growth rate and genomic GC content were further assessed on a sub-set of strains. Geographic location of 67 sampling sites across the Northern hemisphere was used to extract climatic database data and to evaluate the ecogeographical distribution of genome size diversity. Genome size ranged continuously from 0.97 to 2.02 pg of DNA across the investigated strains. The genome size was positively associated with cell size and negatively associated with growth rate. Bioclim variables were not correlated with genome size variation. No clear trends in the geographical distribution of strains of a particular genome size were detected, and strains of different genome size occasionally coexisted at the same locality. Genomic GC content was significantly associated only with genome size via a quadratic relationship. Genome size variability in S. petersenii was probably triggered by an evolutionary mechanism operating via gradual changes in genome size accompanied by changes in genomic GC content, such as, for example, proliferation of transposable elements. The variation was reflected in cell size and relative growth rate, possibly with adaptive consequences.

A genome size and phylogenetic survey of Mediterranean Tripleurospermum and Matricaria (Anthemideae, Asteraceae).

The study of genome size variation can contribute valuable information on species relationships as well as correlate to several morphological or ecological features, among others. Here we provide an extensive report on genome sizes on genus Tripleurospermum and its closely related genus Matricaria, which are two typically Mediterranean genera particularly widespread and diverse in Turkey, the origin of most of the populations here studied. We analyse and discuss genome size variation in the first relatively complete molecular phylogenetic framework of Tripleurospermum (based on ITS and ETS ribosomal DNA–rDNA–regions). We find cases of intraspecific genome size variation, which could be taxonomically significant. Genome downsizing is also detected as the typical response to polyploidisation in Tripleurospermum taxa, being most conspicuous at the tetraploid level. Several positive correlations with genome size, including those with pollen and stomatal size or cypsela length, among others, are also found. Remarkably, taxa presenting rhizomes tend to present higher genome sizes, confirming a trend to accumulate nuclear DNA in such species, which could be explained by the nutrient reserves availability in their storage organs, allowing genome expansion, or by the lower rates of sexual reproduction in rhizomatous taxa.

The slowdown of Y chromosome expansion in dioecious Silene latifolia due to DNA loss and male-specific silencing of retrotransposons

BackgroundThe rise and fall of the Y chromosome was demonstrated in animals but plants often possess the large evolutionarily young Y chromosome that is thought has expanded recently. Break-even points dividing expansion and shrinkage phase of plant Y chromosome evolution are still to be determined. To assess the size dynamics of the Y chromosome, we studied intraspecific genome size variation and genome composition of male and female individuals in a dioecious plant Silene latifolia, a well-established model for sex-chromosomes evolution.ResultsOur genome size data are the first to demonstrate that regardless of intraspecific genome size variation, Y chromosome has retained its size in S. latifolia. Bioinformatics study of genome composition showed that constancy of Y chromosome size was caused by Y chromosome DNA loss and the female-specific proliferation of recently active dominant retrotransposons. We show that several families of retrotransposons have contributed to genome size variation but not to Y chromosome size change.ConclusionsOur results suggest that the large Y chromosome of S. latifolia has slowed down or stopped its expansion. Female-specific proliferation of retrotransposons, enlarging the genome with exception of the Y chromosome, was probably caused by silencing of highly active retrotransposons in males and represents an adaptive mechanism to suppress degenerative processes in the haploid stage. Sex specific silencing of transposons might be widespread in plants but hidden in traditional hermaphroditic model plants.

Genome size variation among and within Ophiopogoneae species by flow cytometric analysis

Genome size variation in a taxonomic group reflects evolutionary processes. DNA contents of Ophiopogoneae (40 populations of 31 species) were estimated by flow cytometry. Ploidy levels of Ophiopogon (ten species), Liriope (two species), and Peliosanthes (three species) were determined based on the DNA contents. The genus Peliosanthes showed significant larger genome sizes than Ophiopogon (P < 0.01), and Ophiopogon also significant larger than Liriope (P < 0.05). Intraspecific variation in genome size was mainly chromosome difference. The ITS sequence phylogeny splitted Ophiopogon into two clades, clade I comprising sect. Ophiopogon with diploids and tetraploids, and clade II including transitional species and sects. Ophiopogon and Peliosanthoides with diploids. The trend seemed to increase in genome size from Ophiopogon sect. Peliosanthoides (13.45 pg) to Ophiopogon sect. Ophiopogon (14.27 pg). Polyploidization may be evolutionary direction of Ophiopogon. Our results also suggested that the ‘increase’ hypothesis for genome size evolutionary may hold true in the genus Ophiopogon.

Quick cytogenetic screening of breeding bulls using flow cytometric sperm DNA histogram analysis.

The aim of the present study was to test the FXCycle PI/RNase kit for routine DNA analyses in order to detect breeding bulls and/or insemination doses carrying cytogenetic aberrations. In a series of experiments we first established basic DNA histogram parameters of cytogenetically healthy breeding bulls by measuring the intraspecific genome size variation of three animals, then we compared the histogram profiles of bulls carrying cytogenetic defects to the baseline values. With the exception of one case the test was able to identify bulls with cytogenetic defects. Therefore, we conclude that the assay could be incorporated into the laboratory routine where flow cytometry is applied for semen quality control.

Patterns of genome size variation in snapping shrimp.

Although crustaceans vary extensively in genome size, little is known about how genome size may affect the ecology and evolution of species in this diverse group, in part due to the lack of large genome size datasets. Here we investigate interspecific, intraspecific, and intracolony variation in genome size in 39 species of Synalpheus shrimps, representing one of the largest genome size datasets for a single genus within crustaceans. We find that genome size ranges approximately 4-fold across Synalpheus with little phylogenetic signal, and is not related to body size. In a subset of these species, genome size is related to chromosome size, but not to chromosome number, suggesting that despite large genomes, these species are not polyploid. Interestingly, there appears to be 35% intraspecific genome size variation in Synalpheus idios among geographic regions, and up to 30% variation in Synalpheus duffyi genome size within the same colony.

Impact of dysploidy and polyploidy on the diversification of high mountain Artemisia (Asteraceae) and allies

Molecular cytogenetics and the study of genome size have been used to understand evolutionary and systematic relationships in many species. However, this approach has seldom been applied to alpine plants. A group of dysploid–polyploid high mountain Artemisia species, distributed from the European Sierra Nevada to Central Asian mountains, through the Pyrenees, the Alps and the Caucasus, is a good model to consider changes at the genome and chromosome levels. These small perennial Artemisia, found frequently in isolated populations, present highly disjunct distributions. Some are considered rare or even endangered. Here, we show results for nine species and 31 populations, including genome size (2C-values), fluorochrome banding and fluorescent in situ hybridisation of ribosomal RNA genes (rDNA). Significant intraspecific genome size variation is found in certain populations of A. eriantha and A. umbelliformis, but without taxonomic significance due to the absence of morphological or ecological differentiation. The number and position of GC-rich DNA bands is mostly coincidental with rDNA although there is an expansion of GC-rich heterochromatin in centromeres in some taxa. Ancestral character states have been reconstructed and x = 9 is inferred as the likely ancestral base number, while the dysploid x = 8 has appeared repeatedly during the evolution of Artemisia. On the basis of cytological observations, Robertsonian translocations are proposed for the appearance of dysploidy in the genus. A remarkable presence of x = 8-based species has been detected in the clade including high mountain species, which highlights the important role of dysploidy in the diversification of high mountain Artemisia. Conversely, polyploidy, though present in the alpine species, is more common in the rest of the genus, particularly in arctic species. Hypotheses on the mechanisms underpinning the relative abundance of dysploids and scarcity of polyploids in high mountain Artemisia are discussed.

Intra-specific variation in genome size in maize: cytological and phenotypic correlates.

Genome size variation accompanies the diversification and evolution of many plant species. Relationships between DNA amount and phenotypic and cytological characteristics form the basis of most hypotheses that ascribe a biological role to genome size. The goal of the present research was to investigate the intra-specific variation in the DNA content in maize populations from Northeastern Argentina and further explore the relationship between genome size and the phenotypic traits seed weight and length of the vegetative cycle. Moreover, cytological parameters such as the percentage of heterochromatin as well as the number, position and sequence composition of knobs were analysed and their relationships with 2C DNA values were explored. The populations analysed presented significant differences in 2C DNA amount, from 4.62 to 6.29 pg, representing 36.15 % of the inter-populational variation. Moreover, intra-populational genome size variation was found, varying from 1.08 to 1.63-fold. The variation in the percentage of knob heterochromatin as well as in the number, chromosome position and sequence composition of the knobs was detected among and within the populations. Although a positive relationship between genome size and the percentage of heterochromatin was observed, a significant correlation was not found. This confirms that other non-coding repetitive DNA sequences are contributing to the genome size variation. A positive relationship between DNA amount and the seed weight has been reported in a large number of species, this relationship was not found in the populations studied here. The length of the vegetative cycle showed a positive correlation with the percentage of heterochromatin. This result allowed attributing an adaptive effect to heterochromatin since the length of this cycle would be optimized via selection for an appropriate percentage of heterochromatin.

Miscanthus: Inter- and Intraspecific Genome Size Variation Among M. × Giganteus, M. Sinensis, M. Sacchariflorus Accessions

AbstractSinceM. sinensisAnderss.,M. sacchariflorus(Maxim.) Hack. andM. ×giganteusJ.M.Greef &amp; Deuter ex Hodk. and Renvoize have considerably the highest potential for biomass production amongMiscanthusAnderss. species, there is an urgent need to broaden the knowledge about cytological characteristics required for their improvement. In this study our objectives were to assess the genome size variation among eighteenMiscanthusaccessions, as well as estimation of the monoploid genome size (2C and Cx) of theM. sinensiscultivars, which have not been analyzed yet. The characterization of threeMiscanthusspecies was performed with the use of flow cytometry and analysis of the stomatal length. The triploid (2n = 3x = 57)M. sinensis‘Goliath’ andM. ×giganteusclones possessed the highest 2C DNA content (8.34 pg and 7.43 pg, respectively). The intermediate 2C-values were found in the nuclei of the diploid (2n = 2x = 38)M. sinensisaccessions (5.52–5.72 pg), whereas they were the lowest in the diploid (2n = 2x = 38)M. sacchariflorusecotypes (4.58–4.59 pg). The presented study revealed interspecific variation of nuclear DNA content (P&lt;0.01) and therefore allowed for recognition of particular taxa, inter- and intraspecific hybrids and prediction of potential parental components. Moreover, intraspecific genome size variation (P&lt;0.01) was observed inM. sinensiscultivars at 3.62%. The values of the stomatal size obtained for the triploidM. ×giganteus‘Great Britain’ (mean 30.70 μm) or ‘Canada’ (mean 29.67 μm) and diploidM. sinensis‘Graziella’ (mean 29.96 μm) did not differ significantly, therefore this parameter is not recommended for ploidy estimation.