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Highlights • Origanum species and subspecies were characterized by FCM and ISSR-PCR methods. • Origanum species/subspecies possessed very small genomes. • Genetic diversity in the genus have been described. • Genome size and ISSR markers are effective in Origanum species/subspecies identification. Abstract The genus Origanum has the complex taxonomy and extensive genetic variation between species, mainly due to the presence of the significant morphological differences within plants and natural hybridization, even among distantly related species. This genus includes numerous species, subspecies, varieties and hybrids, what makes their characterization very complicated. Moreover, the existence of a large number of synonymous names makes the correct identification difficult. High degree of heterogeneity causes, that a potential for utilization of oregano has not been fully explored yet. Therefore, genome size and ISSR markers can be applied for identification and characterization of species from the Origanum genus. Genome size estimation revealed, that all Origanum species possessed very small genomes. The nuclear DNA content ranged from 1.45 pg/2C (O. vulgare) to 1.73 pg/2C (O. majorana). This is the first time the genome size have been estimated in O. heracleoticum , O. majorana , O. vulgare ssp. gracile , O. vulgare ssp. hirtum and O. vulgare ssp. vulgare. Based on the 2C-values it was possible to distinguish O. majorana from O. syriacum. For the discrimination of all accessions of Origanum genus, the ISSR markers were applied. Using primers: (GACA) 4 A, and (AG) 8 YA, it was possible to identify and distinguish species and subspecies. The relationships within tested accessions reflected by genetic distance estimation and dendrogram construction revealed the existence of two main clusters. The first cluster grouped all the accessions of O. vulgare , whereas two species, O. majorana and O. syriacum, were grouped into second cluster. The identification system composed of flow cytometry and ISSR-PCR technique demonstrated to be useful for genotypes in the Origanum genus. [ABSTRACT FROM AUTHOR]

Summary: The mitochondrial DNA of diplonemid and kinetoplastid protists is known for its suite of bizarre features, including the presence of concatenated circular molecules, extensive trans‐splicing and various forms of RNA editing. Here we report on the existence of another remarkable characteristic: hyper‐inflated DNA content. We estimated the total amount of mitochondrial DNA in four kinetoplastid species (Trypanosoma brucei, Trypanoplasma borreli, Cryptobia helicis, and Perkinsela sp.) and the diplonemid Diplonema papillatum. Staining with 4′,6‐diamidino‐2‐phenylindole and RedDot1 followed by color deconvolution and quantification revealed massive inflation in the total amount of DNA in their organelles. This was further confirmed by electron microscopy. The most extreme case is the ∼260 Mbp of DNA in the mitochondrion of Diplonema, which greatly exceeds that in its nucleus; this is, to our knowledge, the largest amount of DNA described in any organelle. Perkinsela sp. has a total mitochondrial DNA content ~6.6× greater than its nuclear genome. This mass of DNA occupies most of the volume of the Perkinsela cell, despite the fact that it contains only six protein‐coding genes. Why so much DNA? We propose that these bloated mitochondrial DNAs accumulated by a ratchet‐like process. Despite their excessive nature, the synthesis and maintenance of these mtDNAs must incur a relatively low cost, considering that diplonemids are one of the most ubiquitous and speciose protist groups in the ocean. © 2018 IUBMB Life, 70(12):1267–1274, 2018 [ABSTRACT FROM AUTHOR]

ABSTRACT Grama-tio-pedro (Paspalum oteroi Swallen) is a rare stoloniferous grass of the Plicatula group of Paspalum, well adapted to continuous grazing in areas subject to seasonal flooding in the Pantanal region, in central western Brazil. The species is a facultative apomictic (asexual reproduction by seed) tetraploid, sporadically cultivated on Pantanal farms, propagated either by cuttings or seed. Due to its potential for extensive cultivation and forage quality, Grama-tio-pedro appears as a candidate for genetic improvement within the Plicatula group through plant breeding. We used a colchicine-induced sexual autotetraploid genotype of P. plicatulum Michx. to obtain interspecific hybrids using the apomictic species, P. oteroi, as pollen donor. The very similar meiotic chromosome behavior observed in both parents, with main quadrivalent and bivalent associations, suggested that P. oteroi is a natural autotetraploid. The hybrids showed less irregular meiotic behavior with fewer quadrivalents and more bivalents than either parent. Fertility among interspecific hybrids varied from complete sterility in some of them to seed productions in others that were approximately twice as much as for either parent. The great variability of seed set performance may well be a drastic genetic consequence of joining two homologous chromosome sets of P. plicatulum together with two homologous sets of P. oteroi that, in turn, have some homeology between them. Most hybrids reproduce by sexual means, thus, they could be used as female parents in backcrosses and in crosses with other species of the Plicatula group for interspecific gene transferring in breeding programs.

Desiccation tolerance (DT) is the ability to tolerate dehydration to levels below 0.1 g(H2O) g-1(dry mass) and subsequent rehydration without lethal damage. Here, it is proposed that Leucaena leucocephala, a tree species, has potential to be model tolerant species in seed research. Using flow cytometry and transmission electron microscopy, cytological changes related to loss of DT in Leucaena primary roots were followed during germination. Leucaena seeds lost their DT at the end of germination and this coincided with an increase in cellular 4C DNA content. A negative correlation between the 8C DNA content and the capacity of germinating Leucaena seeds to tolerate desiccation was also observed. Apparently, the seeds of Leucaena underwent extra cycles of endoreduplication and accumulated a high content of DNA - an event not previously linked to DT. The ultrastructural damage imposed by drying overcame Leucaena primary root cell resilience and their ability to resume normal growth. Nuclear DNA content may be used as indicator of progress of germination and loss of DT in Leucaena.

Decrease in genome content may play a role in environmental adaptation. Many studies were reported significant correlation between genome size, weather condition and germination percentage. Relative genome content and its correlation with seedling establishment of 14 populations of tall fescue collected from various regions in Iran and two commercial tall fescue cultivars were studied under drought conditions. Results showed that except one entry diploid (Brojen = 2x), all entries were hexaploid (6x). Cluster analysis revealed that the populations fell into four groups. Isfahan (Group II: average DNA content 17.92 pg) and Ghochan (Group VІ: average DNA content 18.56 pg) with 100% and 6.7% final emergence and 8.8, 2.3 cm leaf length respectively in 40% FC soil water content wree the most tolerable and sensitive entries under drought stress. Relative genom content of the wild populations and two commercial cultivar were negatively correlated with emergence (r=-0.56) and leaf length (r=-0.61). The reduction in genome size may be a mechanism of adaptation to arid environments. The drought tolerance was observed among the entries that grouped in cluster I and II represent potentially useful germplasm for a breeding program.

Microalgae are freshwater and marine unicellular photosynthetic organisms that utilize sunlight to produce biomass. Due to fast microalgal growth rate and their unique biochemical profiles and potential applications in food and renewable energy industries, the interest in microalgal research is rapidly increasing. Biochemical and genetic engineering have been considered to improve microalgal biomass production but these manipulations also limited microalgal growth. The aim of the study was the biochemical characterization of recently identified microalgal strain Planktochlorella nurekis with elevated cell size and DNA levels compared to wild type strain that was achieved by a safe non-vector approach, namely co-treatment with colchicine and cytochalasin B (CC). A slight increase in growth rate was observed in twelve clones of CC-treated cells. For biochemical profiling, several parameters were considered, namely the content of proteins, amino acids, lipids, fatty acids, β-glucans, chlorophylls, carotenoids, B vitamins and ash. CC-treated cells were characterized by elevated levels of lipids compared to unmodified cells. Moreover, the ratio of carotenoids to chlorophyll a and total antioxidant capacity were slightly increased in CC-treated cells. We suggest that Planktochlorella nurekis with modified DNA levels and improved lipid content can be considered to be used as a dietary supplement and biofuel feedstock.

Background Polyploid specimens are usually characterized by greater exuberance: they reach larger sizes and/or have a larger number of some organs. Festuca amethystina L. belongs to the section Aulaxyper. Based on morphological features, four subspecies of F. amethystina have been already identified. On the other hand, it has two cytotypes: diploid and tetraploid. The main aim of our study was to distinguish morphological differences between the cytotypes of F. amethystina, assuming that its phenotype differs significantly. Methods The nuclear DNA content was measured by flow cytometry in dry leaves from specimens originating from 13 populations of F. amethystina. Several macrometric and micrometric traits of stems, spikelets and leaf blades were taken into account in the comparative analysis of two cytotypes. Results In the case of cytotypes, specimens of tetraploids were larger than diploids. The conducted morphometric analysis of leaf cross-sections showed significant differences between the cytotypes. Discussion The research has confirmed for the first time that in the case of F. amethystina the principle of greater exuberance of polyploids is true. Differences between the cytotypes are statistically significant, however, they are not enough to make easy the distinction of cytotypes on the basis of the measurements themselves. Our findings favor the rule known in Festuca taxonomy as a whole, i.e. that the ploidy level can be one of the main classification criteria.

Abstract In this study, fig and black mulberry DNA contents were estimated using DAPI fluorescence stain in flow cytometry. The 2C DNA contents of the fig and black mulberry were found as 0.82 pg and 8.34 pg, respectively. The calculated 1C value of genome size of fig is 401.8 Mbp and that of black mulberry is 4086.6 Mbp. The ratio of 2C DNA content and 1C genome of the black mulberry was 10.17 times that of the fig although fig is diploid and black mulberry is decosaploid.

Abstract Sisyrinchium is the largest genus of Iridaceae in the Americas and has the greatest amount of cytological data available. This study aimed at investigating how genomes evolved in this genus. Chromosome number, genome size and altitude from species of sect. Viperella were analyzed in a phylogenetic context. Meiotic and pollen analyses were performed to assess reproductive success of natural populations, especially from those polyploid taxa. Character optimizations revealed that the common ancestor of sect. Viperella was probably diploid (2n = 2x =18) with two subsequent polyplodization events. Total DNA content (2C) varied considerably across the phylogeny with larger genomes detected mainly in polyploid species. Altitude also varied across the phylogeny, however no significant relationship was found between DNA content changes and altitude in our data set. All taxa presented regular meiosis and pollen viability (> 87%), except for S. sp. nov. aff. alatum (22.70%), suggesting a recent hybrid origin. Chromosome number is mostly constant within this section and polyploidy is the only source of modification. Although 2C varied considerably among the 20 taxa investigated, the diversity observed cannot be attributed only to polyploidy events because large variations of DNA content were also observed among diploids.

There is a strong positive relationship between nuclear genome size and cell size across the eukaryotic domain, but the cause and effect of this relationship is unclear. A positive coupling of cell size and DNA content has also been recorded for various bacteria, suggesting that, with some exceptions, this association might be universal throughout the tree of life. However, the link between cell size and genome size has not yet been thoroughly explored with respect to chloroplasts, or organelles as a whole, largely because of a lack data on cell morphology and organelle DNA content. Here, I speculate about a potential positive scaling of cell size and chloroplast genome size within different plastid-bearing protists, including ulvophyte, prasinophyte, and trebouxiophyte green algae. I provide examples in which large and small chloroplast DNAs occur alongside large and small cell sizes, respectively, as well as examples where this trend does not hold. Ultimately, I argue that a relationship between cellular architecture and organelle genome architecture is worth exploring, and encourage researchers to keep an open mind on this front.

In vitro induction of polyploids using colchicine causes an increase in DNA content in plants. This is of high importance especially for plants that have medicinal and commercial values. Seeds of two medicinal plants, licorice Glycyrrhiza glabra L. var.glandulifera and safflower Carthamus tinctorius were treated with different concentrations of colchicine, 0%, 0.03%, 0.05%, 0.08%, 0.1% (W/V) in vitro for 24 and 48 h. Treated seeds then were cultured on solid Murashige and Skoog (MS) media under controlled conditions. After a month, the length of the stomata was measured to study the effect of colchicine on stomata size. Cellular DNA content of the regenerated plants was measured by spectrophotometry. Flow cytometry was used for confirming the results obtained from stomata size measurement and spectrophotometry. Results suggested that treated plants have a fair amount of larger stomata, significantly in licorice plantlets that were treated with 0.1% colchicine for 24 h and safflower plantlets that were treated with 0.03%, 0.05% and 0.1% colchicine. Safflower DNA content in all treatments enhanced significantly, but in licorice only DNA content of plantlets that were treated with 0.05% colchicine for 24 h and 0.1%, 0.03% colchicine for 48 h found to be increased significantly. The morphological features of treated plantlets such as shoot and leaf thickness were found to be increased. Flow cytometry confirmed the previously mentioned results and suggested tetraploids in all treated safflower plantlets and licorice plantlets obtained from treatment with 0.08% of colchicine and mixoploids in licorice plantlets obtained from treatment with 0.1% of colchicine.

The aim of the study was a statistical evaluation of the impact of selected soil factors − water potential (pF), total organic carbon content (TOC) and land use − on the total DNA content and dehydrogenase activity (DHA) in Mollic Gleysol. Additionally, we wanted to establish the interrelations between two of the most important biological parameters in soli: activity of intracellular dehydrogenases and total DNA content. Soil originating from the surface layer of the control site displayed higher DHA (c.a. by 57%) and DNA content (c.a. by 25%) as compared to an cultiavted meadow. Our results also indicate a significant (p

The systematics of the Mentha genus is very complicated and still unclear. This is mainly due to the existence of variation in basic chromosome number, frequent interspecific hybridization, cytomixis, polymorphism in morphology and essential oil composition under different environmental conditions, colonial mutant propagation, as well as the occurrence of polyploidy, aneuploidy and nothomorphs. Morphological traits, chemical composition, and even caryological analysis failed in discrimination of Mentha species. Therefore, the aim of this study was to combine genome size estimation with ISSR markers for quick and precise species identification, determination the inter- and intraspecific variation within this genus, as well as description of relationships between genotypes. Genome size estimation revealed, that investigated Mentha species possessed very small and small genomes. Additionally, for 13 Mentha species, this is the first report on genome size. Within one species different values of genome size occurred, which indicated polyploidy and aneuploidy. The smallest genome (0.63 pg/2C) was detected in M. gattefossei , while the largest in M.   ×   verticillata (4.15 pg/2C). Flow cytometric measurements of genome size allowed to distinguish some species and cytotypes, and thus it can be used as a first step in species identification. More precise and detailed characterisation were provided by ISSR markers. All the primers used in ISSR-PCR, revealed to be useful in species identification, while primers (GACA) 4 , (GTG) 6 T, (CTC) 4 RC, (GA) 6 CC enabled to distinguish M. aquatica , M. canadensis , M. longifolia , M . pulegium and M . spicata cytotypes, and primers (GACA) 4 , (GA) 6 CC and (GA) 8 YC to identify M.   ×   gracilis and M.   ×   piperita cultivars. The phylogenetic analysis revealed the existence of seven main clusters and two species ( M. requienii and M . ×  smithiana ) being not clustered to any of the created groups within the Mentha genus. This analysis mostly confirms the latest taxonomical classification of the Mentha genus, however some exceptions are discussed. The identification system of combined genome size estimation, and ISSR markers proved to be useful for species in Mentha genus. [ABSTRACT FROM AUTHOR]

Aucuba have been cultivated for centuries and are valued as adaptable, broadleaved, evergreen shrubs that also can have attractive, spotted variegations on the foliage. Improved understanding of the cytogenetics and heritability of specific traits, for specific clones and cultivars, can provide basic information to help facilitate the breeding and improvement of aucuba. The objectives of this study were to determine ploidy level and relative genome size of a diverse collection of species and cultivars of aucuba using flow cytometry and cytology and to make additional observations on heritability of spotted leaf variegation. Chromosome counts were 2n = 2x = 16 for Aucuba chinensis (A. omeiensis), 2n = 4x = 32 for A. japonica 'Rozannie', and 2n = 6x = 48 for A. sp. 'Hosoba'. Relative 2C genome size for the 57 taxa varied from 13.8 pg for A. obcordata to 42.0 pg for A. 'Hosoba' and fell within three discrete groups consistent with cytotype. Genome size for diploid taxa (A. chinensis and A. obcordata) ranged from 13.8 to 21.0 pg, tetraploids (A. himalaica var. oblanceolata, A. japonica, and A. japonica var. borealis) ranged from 28.8 to 31.2 pg, and the first-ever reported hexaploids (A. 'Hosoba' and A. sp. - Vietnam) ranged from 40.5 to 42.0 pg. Unlike prior reports that indicated inheritance of spotted variegations were extranuclear genes that were maternally inherited, we found that the spotted leaf trait expressed in A. japonica 'Shilpot' appears to be a nuclear gene that is inherited in a quantitative fashion and not strictly maternal. These data provide an enhanced foundation for breeding improved aucuba. [ABSTRACT FROM AUTHOR]

Abstract Some plant species can cross with each other but stay nevertheless distinct with little gene flow between them. Selective herbivory could explain this pattern when hybrids are more susceptible or intermediate between their parents. We performed choice and no-choice experiments with the slug Arion lusitanicus to test this hypothesis for the crop Brassica napus , wild B. rapa and their backcross hybrids. In both experiments slugs greatly preferred B. napus over B. rapa while average herbivory on backcross hybrids was intermediate. Concentrations of aliphatic glucosinolates in the hybrids were intermediate between the parental species. Slug herbivory potentially reduces establishment of backcross hybrids. This hypothesis needs to be tested in the field. [ABSTRACT FROM AUTHOR]

Background. Polyploid specimens are usually characterized by greater exuberance: they reach larger sizes and/or have a larger number of some organs. Festuca amethystina L. belongs to the section Aulaxyper. Based on morphological features, four subspecies of F. amethystina have been already identified. On the other hand, it has two cytotypes: diploid and tetraploid. The main aim of our study was to distinguish morphological differences between the cytotypes of F. amethystina, assuming that its phenotype differs significantly. Methods. The nuclearDNAcontent was measured by flow cytometry in dry leaves from specimens originating from 13 populations of F. amethystina. Several macrometric and micrometric traits of stems, spikelets and leaf blades were taken into account in the comparative analysis of two cytotypes. Results. In the case of cytotypes, specimens of tetraploids were larger than diploids. The conducted morphometric analysis of leaf cross-sections showed significant differences between the cytotypes. Discussion. The research has confirmed for the first time that in the case of F. amethystina the principle of greater exuberance of polyploids is true. Differences between the cytotypes are statistically significant, however, they are not enough to make easy the distinction of cytotypes on the basis of the measurements themselves. Our findings favor the rule known in Festuca taxonomy as a whole, i.e. that the ploidy level can be one of the main classification criteria. [ABSTRACT FROM AUTHOR]

Karyotypes in 16 representative taxa of the Ophrys genus are compared, based on Feulgen-stained somatic metaphase chromosomes. The karyotypes of O. omegaifera subsp. israelitica, O. ulupinara, O. lycia, O. argolica subsp. lucis, O. argolica subsp. lesbis, O. climacis and O. reinholdii subsp. reinholdii are described for the first time. Karyological analyses indicate relationships among the species with respect to their asymmetry indices. All Ophrys taxa studied were diploid with 2n = 2x = 36 chromosomes. One B chromosome has been detected among the chromosomes of O. argolica subsp. lucis. All karyotypes are symmetrical, consisting of metacentric and submetacentric chromosomes. The longest chromosomes of all the investigated specimens contain a secondary constriction. It is determined that there is a correlation between the total number of chromosomes having secondary constrictions and the evolutionary development order of the taxa. Based on nuclear DNA content, analysis was carried out by flow cytometer using propodium iodide as fluorochrome, 2C nuclear DNA content of 16 Ophrys species varying between 20.80 pg (O. argolica subsp. lucis) and 23.11 pg (O. omegaifera subsp. israelitica). Karyotype asymmetry relationships are discussed according to the bidimensional scatter plots of A1-A2, CVCL- CVCI, CVCL-MCA and CVCI-MCA. [ABSTRACT FROM AUTHOR]

Cardamine occulta, recently reported also as 'Asian C. flexuosa', is originally a native East Asian weed of paddy fields and other open habitats. It has been recorded throughout the world and is now also spreading widely in Europe. However, how this species differs morphologically from European C. flexuosa and its closest relatives in Asia (C. kokaiensis and C. scutata) is not fully known, particularly because it became widely recognized as a new taxon only recently. We used chromosome counting and flow cytometry to determine ploidies and genome size variation in these four species and morphometric analyses to ascertain their morphological differentiation. A uniformly octoploid level (2n = 64) is confirmed here for C. occulta and a tetraploid level for C. flexuosa and C. scutata (2n = 32). Here we formally describe C. kokaiensis, which is restricted to East Asia (Japan, Honshu; eastern China, Zhejiang Province; perhaps also the Russian Far East) and determine it to be a tetraploid. Considerable differences in monoploid genome size were revealed between these taxa, suggesting their mostly allopolyploid origins. Morphological differences between the species are demonstrated, and detailed morphological descriptions and an identification key are provided. [ABSTRACT FROM AUTHOR]

Abstract: Angiosperms have evolved a mechanism of double fertilization, which results in the production of a separate embryo (new individual) and endosperm (nutritive tissue). The flow cytometric seed screen (FCSS) was developed to infer plant reproduction modes based on endosperm‐to‐embryo DNA content ratio (Pind). A ratio of 1.5 indicates sexual reproduction, whereas higher values of ≥2.0 are consistent with apomixis. Although FCSS has been successfully applied to the study of sexual and asexual plants, the limits of FCSS and particularly its potential for determination of reproduction modes in hemisexual plants have not been explored. Here, we evaluated the application of FCSS to the study of reproduction modes in two asymmetrically compensating allopolyploids (ACAs), Onosma arenaria and Rosa canina. These two species are characterized by the presence of asexually inherited univalent‐forming and sexually inherited bivalent‐forming chromosome sets. They both use asymmetric meiosis, which eliminates univalent‐forming chromosome sets from the male gamete and retains them in the female gamete. Different chromosomal behavior in male and female meiosis in these plants is reflected in different theoretically derived Pind values, which deviate from a sexual 1.5 value. Here, we determined Pind FCSS‐based values in seeds of ACAs, and compared the results to sexual species. As expected, we determined that the mean Pind is 1.51, 1.52, and 1.52 in the sexual plants, that is, Capsella bursa‐pastoris, Crataegus monogyna, and O. pseudoarenaria, respectively. In the ACAs, different mean Pind values were determined for O. arenaria (1.61) and R. canina (1.82). These values are consistent with the theoretical Pind values determined based on models of chromosome inheritance. This study highlights the precision of flow cytometry in determining DNA content and it's utility in screening reproduction modes. Additionally, it advocates for more in‐depth investigations into rapid screening of accessions where the Pind ratio has deviated from the 1.5 value typical of sexual species, which may indicate meiotic irregularities. [ABSTRACT FROM AUTHOR]

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Flow cytometry analysis of propidium iodide (PI) stained nuclei isolated from leaf tissues were used to estimate the DNA content and genome size of Aquilaria malaccensis Lamk. Samples of in vitro regenerated (tissue culture-derived plantlets) and ex vitro (wild seedlings) were evaluated. The results revealed that there was no intra-specific variation in A. malaccensis DNA content (1.84 pg 2C-1) and genome sizes (899 Mbp) between in vitro and ex vitro samples, suggesting that even after numerous sub-culturing and long-term culture periods, the species was able to retain stable genome size. On the other hand, interspecific variations were determined between A. malaccensis and five other Aquilaria species in terms of DNA content (1.84 to 1.92 pg 2C-1) and genome size (894.65 to 938.88 Mbp). This study provides valuable information that will facilitate genome sequencing and subsequent molecular studies of this economically important medicinal plant. Also, as evidence that micropropagation techniques have the potential to be used as a source of planting stock, along with seeds, for restoring locally threatened A. malaccensis wild populations and for commercial cultivation to supply the growing market demand. [ABSTRACT FROM AUTHOR]

Chromosome data are fundamental in evolution. However, there has been no attempt to synthesize and evaluate the significance of such information from a phylogenetic perspective in the giant genus Solanum, which was the aim of this work. New and published information of the main cytotaxonomic features (chromosome number, polyploidy, total length of the haploid complement, mean chromosome length, mean arm ratio, karyotype formula, nuclear DNA amount, number/position of rDNA sites) was compiled and mapped onto an embracing Solanaceae phylogeny, performing Ancestral States Reconstruction. There were 506 Solanum species with chromosome counts (49.7% from an estimated total of 1,018 spp.), with x = 12 being the most frequent number (97%). Species with karyotypes represent 18.8%, while 8% have been studied with any molecular cytogenetic technique. Chromosome characters showed transitions associated with supported nodes, some of which have undergone fewer transitions than others. The common ancestor of all Solanum was a diploid with 2n = 24, a karyotype with st and/or t chromosomes, 2C DNA content of 1-1.2 pg, one locus of 18-5.8-26S rDNA and one of 5S, both loci being asyntenic. The chromosomal variables behave as homoplastic, with reversions in all branches. The analysed characters were sorted from more to less conserved: asynteny of rDNA loci; number of sites of 18-5.8-26S; chromosome number; karyotype formula; number of 5S loci. This pattern of chromosomal evolution distinguishes Solanum from closely related genera and from genera from other families with a similar number of species. [ABSTRACT FROM AUTHOR]

Hedychiums are multipurpose plants cultivated as ornamentals because of their multicolor and showy and scented flowers and as medicinal plants because of their essential oils that have been found to possess antimicrobial and insecticidal properties. There is often taxonomic and botanical confusion about Hedychium species and cultivars because of the apparent existence of more than one cytotype within several Hedychium species, most likely because of the ease with which these species hybridize. Nuclear DNA content (2C-DNA) was determined for the first time in 23 Hedychium species and hybrids. The 2C-values ranged from 1.70 pg to 3.98 pg. We also determined the chromosome number of one species, Hedychium stenopetalum, which was found to be diploid (2n = 2x = 34), supporting the widely held view that the basic chromosome number in Hedychium is x = 17, and the existence of different cytotypes is most probably attributable to natural hybridization. Our results should be helpful not only in clarifying the botanical and taxonomic confusion in Hedychium but also in paving the way for a sound breeding program for this increasingly important ornamental ginger. [ABSTRACT FROM AUTHOR]

Summary Canine lymphoma is a multifaceted disease encompassing numerous entities with different prognosis. Objective assessment of the proliferation rate is of importance from the pathological and clinical perspectives. Different methods have been described in the literature to assess proliferation rate, including evaluation of Ki67 expression in fresh lymph node (LN) aspirates measured by flow cytometry (FC). This test has a high accuracy in discriminating between low- and high-grade lymphomas, and provides prognostic information among high-grade B-cell lymphomas. DNA content analysis is less expensive and suitable for well-preserved samples. We describe DNA-content analysis using LN aspirates from 112 dogs with lymphoma. S-phase fraction (SPF) accurately discriminated between low- and high-grade lymphomas, with 3.15% being the best discriminating cut-off value. SPF values strongly correlated with Ki67 expression as assessed by FC. Survival analyses were restricted to 33 dogs with high-grade B-cell lymphoma receiving standardized multi-agent chemotherapy, but no significant result was obtained for SPF. We also describe a subset of aneuploid cases and their respective follow-up. We conclude that DNA content analysis may be combined with morphological examination of LN aspirates to improve the objectivity in lymphoma subtype classification in dogs. Further studies are needed to assess the possible prognostic role of SPF and ploidy status within specific lymphoma subtypes in dogs. [ABSTRACT FROM AUTHOR]

Birds exhibit small, constrained genome sizes relative to many other vertebrates. This has often been attributed to the metabolic constraints of powered flight, which requires small cells for efficient gas exchange. Small cells, in turn, are achieved in part by limiting nucleus size and thus DNA content. However, metabolism is not the only organismal trait that correlates with genome size in birds; relationships have also been reported with body size (positive) and relative brain size (inverse). In this study, a recent data set on cell numbers in the brains of birds was combined with available data on genome sizes to demonstrate a relationship between genome size and density of cells per gram of brain tissue. This suggests that small genomes are relevant not only in the evolution of avian flight but may also be associated with the extraordinary behavioural complexity shown by birds. [ABSTRACT FROM AUTHOR]

Genome size, karyotypic features and pollen size in Argentinian species of Spergula (Caryophyllaceae). The objective of this work was to estimate the DNA content (2C) in four species of Spergula that inhabit Argentina and to analyze the variation of genome size (Cx) in relation to karyotypic and palynological features. The DNA content of five taxa and the genome size of four, which varied very little between species, were analyzed for the first time from Cx= 0.458 pg (S. salina, 4x) to Cx= 0.856 pg (S. arvensis, 2x). Two new chromosomal counts were recorded for the genus and the pollen size varied from small to medium. The regression coefficients suggest that the chromosome number varies inversely proportional in regards to the genome size, while the genome size presents a low relation with the total chromosome length (TCL) of the diploid genome, although it shows a high relation with respect to the average chromosome length (ACL). Meanwhile, the pollen size varies independently regarding to the genome size. The detected karyotypic variation could be due by numerical and structural changes during the genus evolution, while the pollen size could be related with selection pressures associated with dispersal strategies. [ABSTRACT FROM AUTHOR]

Interpopulation variation of relative and absolute genome size was studied in Fagus sylvatica subsp. sylvatica and subsp. orientalis. The study included 18 populations of beech planted in a common-garden experiment in central Slovakia and three additional populations from the Caucasus. Nuclear DNA content was determined by means of flow cytometry using the AT-specific fluorochrome 4′,6-diamidino-2-phenylindole and non-specific propidium iodide, and its associations with climate, growth, phenology and physiological traits were assessed. The approximate average nuclear DNA content (2C) across all samples was 1.178 ± 0.020 pg. The lowest mean relative genome sizes were observed in the Alpine range, whereas they increased toward the range margins; no clear trend was observed for 2C values. Temperature seasonality and temperature annual range were found to be negatively associated with genome size. Among phenotypic traits, the maximum chlorophyll a fluorescence yield (Fv/Fm) was found to be negatively correlated with relative genome size, whereas phenology and some photosynthetic parameters were correlated with the 2C values. [ABSTRACT FROM AUTHOR]

Tulbaghia violacea, called ‘wild garlic’, is a native of Africa, though it is widely cultivated all over the world for its medicinal and horticultural importance. Absence of the genus from Indian floras and taxonomic surveys, as well as lack of proper molecular and cytogenetic characterizations, have instigated questions regarding origin and divergence of the Indian counterpart from its African relatives. In the present study, morphotaxonomical identification of Indian T. violacea was carried out, along with its karyomorphological characterization, 2C nuclear genome content determination, and analysis of ITS sequence. Mitotic study revealed that this species possessed 2n = 2x = 12 somatic chromosomes, two of which had secondary constrictions, whereas unstable meiotic chromosomal behavior was observed in flower buds, as reported earlier. 2C nuclear DNA content of T. violacea (44.51 ± 0.06 pg) was reported for the first time using flow cytometry, as was the ITS sequence data, from India. [ABSTRACT FROM AUTHOR]

Human leukemia Jurkat T cells were analyzed for apoptosis and cell cycle by flow cytometry, using the Annexin V/propidium iodide (PI) standard assay, and a simple PI staining in Triton X-100/digitonin-enriched PI/RNase buffer, respectively. Cells treated with doxorubicin or menadione displayed a very strong correlation between the apoptotic cell fraction measured by the Annexin V/PI assay, and the weight of a secondary cell population that emerged on the forward scatter (FS)/PI plot, as well as on the side scatter (SS)/PI and FL1/PI plots generated from parallel cell cycle recordings. In both cases, the Pearson correlation coefficients were >0.99. In cell cycle determinations, PI fluorescence was detected on FL3 (620/30 nm), and control samples exhibited the expected linear dependence of FL3 on FL1 (525/40 nm) signals. However, increasing doses of doxorubicin or menadione generated a growing subpopulation of cells displaying a definite right-shift on the FS/FL3, SS/FL3 and FL1/FL3 plots, as well as decreased PI fluorescence, indicative of ongoing fragmentation and loss of nuclear DNA. By gating on these events, the resulting fraction of presumably sub-cycling cells (i.e. cells with cleaved DNA, counting sub-G0/G1, sub-S and sub-G2/M cells altogether) was closely similar to the apoptotic rate assessed by Annexin V/PI labeling. Taken together, these findings suggest a possible way to recognize the entire population of cells undergoing apoptotic DNA cleavage and simultaneously determine the cell cycle distribution of non-apoptotic cells in PI-labeled cell samples with various degrees of DNA fragmentation, using a simple and reproducible multiparametric analysis of flow cytometric recordings. [ABSTRACT FROM AUTHOR]

We expand the genome size (GS) database for Bromeliaceae, specifically for subfamily Pitcairnioideae, and verify whether GS can provide information on the diversification of the five genera in this subfamily. We also provide a phylogenetic perspective on GS evolution in the subfamily and reconstruct the ancestral state for this character. We show that the evolutionary path of GS from the origin of angiosperms to the origin of Pitcairnioideae agrees with the proportional model of GS evolution. Furthermore, we propose that the high phenotypic diversity that is found across Bromeliaceae and that is well represented in Pitcairnioideae is both correlated with high rates of GS evolution of the species and associated with a short period of diversification. The paper also highlights the value of flow cytometry as a rapid and reliable technique for generating GS data which can be analysed in conjunction with other molecular and morphological data to help elucidate patterns of evolution and phylogenetic relationships within this family. [ABSTRACT FROM AUTHOR]

The Northwestern Argentina (NWA) highland region is one of the southernmost areas of native maize cultivation. We studied variations of different cytological parameters, such as DNA contents, presence/absence of B chromosomes (Bs), and number and sequence composition of heterochromatic knobs in ten accessions of four maize landraces growing along a broad altitudinal cline in NWA. The aim of this work was to assess variations in cytological parameters and their relationship with the crop altitude of cultivation, in an adaptive context. The A-DNA content of the A chromosome complements showed 40% of difference between the lowest (4.5 pg) and the highest (6.3 pg) 2C value. This variation could be attributed to differences in number and size of heterochromatic knobs. Fluorescent in situ hybridization studies revealed the sequence composition of each knob, with a higher proportion of knobs composed of 180-bp repeats rather than TR-1 repeats, in all accessions. We also found numerical polymorphisms and the highest frequency of Bs reported in maize to this date. These results lead us to propose that the frequencies and doses of Bs are influenced by the landrace genotypical make-up. The Bs might be maintained in higher frequencies in those accessions having lower heterochromatin content, so as to preserve an optimal nucleotype. Furthermore, selective forces acting along the altitudinal gradient might be modulating the cytological parameters studied, as suggested by the significant correlations found among them. [ABSTRACT FROM AUTHOR]

•Crossings between the diploid wild Brassica rapa (AA, 2n = 20) and the tetraploid cultivar B. napus (AACC, 2n = 38) can readily be made. Backcrosses to the wild B. rapa (BC1) produce aneuploids with variable chromosome numbers between 20 and 29. How does survival and performance relate to DNA content of plants? •Growth of the BC1 plants was measured in the lab. One plant in the F1 self-pollinated spontaneously and produced abundant F2 seeds that were also examined. The number of C-chromosomes was estimated from DNA values obtained with flow cytometry. •Average DNA value of the BC1 was similar to that of the parents, which shows that Cchromosomes do not reduce success of pollen or embryos. The average DNA value in the F2 was 13% higher than in the F1, suggesting that extra C-chromosomes facilitated gamete success and/or embryo survival. Under both optimal and drought stress conditions growth and survival of BC1 hybrids was similar to that of B. rapa. No significant correlations existed between growth or survival and DNA value. •Aneuploid plants were not inferior under the conditions of the growth room and may persist in nature. We discuss other factors, such as herbivory, that could prevent hybrid establishment in the field. [ABSTRACT FROM AUTHOR]

Several species within the genus Lotus are important forage crops, and many are endangered or rare. Despite the high genetic diversity of the genus, identification of Lotus species is problematic because of the limited number of reliable morphological markers. In search of a quick, inexpensive, and steady method for species identification, genome size and cell cycle/endoreduplication intensity of 14 accessions belonging to eight Lotus species were estimated by flow cytometry. ISSR-PCR was also applied to find sensitive molecular markers for genetic diversity estimation. Genome size estimation revealed that Lotus species possess very small genomes and this characteristic enables the identification of five out of eight species. However, a flow cytometric study of cell ploidy/endopolyploidy in seeds and seedlings enabled us also to distinguish the remaining species. Thus, it is proposed here that combined flow cytometric analyses (the estimation of genome size and cell cycle/endoreduplication pattern) can be applied for screening of Lotus species. Nonetheless, ISSR markers provided a more precise identification of studied accessions, including detection of genetic diversity within a species. Most of the tested primers revealed polymorphism between species, and three primers, (GACA)4, (CA)7G, and (CTC)4RC, also revealed polymorphism between accessions within a species. The ISSR markers revealed high polymorphism between eight Lotus species and low intraspecific variation between accessions within the same species (L. maritimus and L. uliginosus). [ABSTRACT FROM AUTHOR]

Peanut, Arachis hypogaea (Linnaeus, 1753) is an allotetraploid cultivated plant with two subgenomes derived from the hybridization between two diploid wild species, A. duranensis (Krapovickas & W. C. Gregory, 1994) and A. ipaensis (Krapovickas & W. C. Gregory, 1994), followed by spontaneous chromosomal duplication. To understand genome changes following polyploidy, the chromosomes of A. hypogaea, Ipa- Dur1, an induced allotetraploid (A. ipaensis × A. duranensis)4x and the diploid progenitor species were cytogenetically compared. The karyotypes of the allotetraploids share the number and general morphology of chromosomes; DAPI+ bands pattern and number of 5S rDNA loci. However, one 5S rDNA locus presents a heteromorphic FISH signal in both allotetraploids, relative to corresponding progenitor. Whilst for A. hypogaea the number of 45S rDNA loci was equivalent to the sum of those present in the diploid species, in IpaDur1, two loci have not been detected. Overall distribution of repetitive DNA sequences was similar in both allotetraploids, although A. hypogaea had additional CMA3 + bands and few slight differences in the LTR-retrotransposons distribution compared to IpaDur1. GISH showed that the chromosomes of both allotetraploids had preferential hybridization to their corresponding diploid genomes. Nevertheless, at least one pair of IpaDur1 chromosomes had a clear mosaic hybridization pattern indicating recombination between the subgenomes, clear evidence that the genome of IpaDur1 shows some instability comparing to the genome of A. hypogaea that shows no mosaic of subgenomes, although both allotetraploids derive from the same progenitor species. For some reasons, the chromosome structure of A. hypogaea is inherently more stable, or, it has been at least, partially stabilized through genetic changes and selection. [ABSTRACT FROM AUTHOR]

Objective: To assess the cellular DNA status of epithelial ovarian cancer cells for clinical stage identification and its effect on survival. Methods: Sixty-two patients treated by primary surgery and six courses of platinum-based chemotherapy were enrolled. The surgical stage was analyzed in correlation with DNA ploidy, S-phase fraction and DNA index. DNA analysis was performed via image cytometry. Results: From the 62 cases, 38 were International Federation of Gynecology and Obstetrics (Fédération Internationale de Gynécologie et d'Obstétrique, FIGO) stage I and II, 24 - stage III and IV. In the DNA histograms obtained, the DNA index ranged from 0.85 to 3.02. Sixteen were classified as diploid and 46 as aneuploid (18 multiploid). S-phase fraction ranged from 9.8 to 51%. The aneuploid cells with DNA content above 5C ranged from 0.0 to 77.2%. Patients diagnosed with FIGO III and IV (vs. I and II) were 3.3 times more likely to die. Only in FIGO stage I and II the survival differed significantly for the different groups of ploidy. The risk of death for the multiploid (vs. diploid) group is 6.4 times and for aneuploid (vs. diploid) 2.3 times. Overall survival was better in the group with low DNA index. The low percentage compared with a high percentage of 5C cells ploidy groups showed association with mortality. The death hazard for the S-phase >33 median group is 4.9 times the hazard in relation to the S-phase <33. Conclusions: DNA ploidy, DNA index, S-phase, and 5C cells are important prognosticators for epithelial ovarian cancer mainly in early stages. [ABSTRACT FROM AUTHOR]

Cowpea ( Vigna unguiculata (L.) Walp.) is widely grown by subsistence farmers in West and East Africa where its grain and leaves are sources of highly valuable food, due to their high contents of proteins, minerals and vitamins. Therefore, cowpea could play a significant role in mitigating malnutrition such as micronutrient deficiencies. The objectives of this study were to evaluate the performance in agronomic traits of cowpea entries (released and farmers’ cultivars, and gene bank accessions) from different sources, and to assess the extent of genetic diversity in this material to provide basic information for its use in breeding programs. A total of 15 entries were evaluated. All entries were morphologically uniform, except the farmers’ local cultivars. Fresh leaf yield varied from 34.6 to 52.8 g per plant and days to 50% flowering from 64 to 82 days. Hundred seed weight ranged from 7.67 to 15.12 g. On average, the number of pods ranged from 4.8 to 15.6 pods per plant. No correlation between fresh leaf yield and other traits was detected, whereas the number of pods per plant and the hundred seed weight were negatively correlated. Genetic diversity was assessed on five genotypes per entry using 544 Amplified Fragment Length Polymorphism (AFLP) and 18 microsatellite markers. Genetic distances calculated using the Jaccard algorithm ranged from 0.002 to 0.193 among genotypes of the same entry and from 0.098 to 0.301 for genotypes from different entries. A principle coordinate analysis separated four entries from the rest. Although the consensus tree based on Neighbor Joining trees was unable to resolve the whole cluster, an assignment of most of the entries into entry-specific clades was possible. [ABSTRACT FROM AUTHOR]

There is a strong positive relationship between nuclear genome size and cell size across the eukaryotic domain, but the cause and effect of this relationship is unclear. A positive coupling of cell size and DNA content has also been recorded for various bacteria, suggesting that, with some exceptions, this association might be universal throughout the tree of life. However, the link between cell size and genome size has not yet been thoroughly explored with respect to chloroplasts, or organelles as a whole, largely because of a lack data on cell morphology and organelle DNA content. Here, I speculate about a potential positive scaling of cell size and chloroplast genome size within different plastid-bearing protists, including ulvophyte, prasinophyte, and trebouxiophyte green algae. I provide examples in which large and small chloroplast DNAs occur alongside large and small cell sizes, respectively, as well as examples where this trend does not hold. Ultimately, I argue that a relationship between cellular architecture and organelle genome architecture is worth exploring, and encourage researchers to keep an open mind on this front. [ABSTRACT FROM AUTHOR]

Chrysolaena flexuosa is a South American species of potential ornamental value (basic chromosome number of x=10). Diploid (n=10) and tetraploid (n=20) cytotypes have been reported for its distribution area, although one hexaploid (n=30-32ca.) cytotype has been reported for its most southern distribution. To investigate if ploidy and latitude are positively related in Ch. flexuosa natural populations and if sexual polyploidization could have had a role in the origin of the polyploid cytotypes, we determined chromosome numbers, DNA content, and pollen viability and size, and analyzed microsporogenesis in samples of seven Argentinian accessions. Two of the northeastern accessions were diploid and one was tetraploid, whereas the four southeastern accessions were hexaploid. Ploidy levels determined both by chromosome countings and flow cytometry coincided, although monoploid genome size significantly decreased with increasing ploidy. In all accessions, variability was observed for pollen viability and size, as well as for large (presumably 2n) pollen production. This variability was underlined by abnormal cytological events in meiosis and at the tetrad stage (lagging chromosomes, parallel spindles, triads). The results would indicate that there is, apparently, a positive relation between ploidy and latitude, and suggest a likely role of sexual polyploidization in the origin, establishment and expansion of Ch. flexuosa populations. [ABSTRACT FROM AUTHOR]

Genome size (GS) diversity is of fundamental biological importance. The occurrence of giant genomes in angiosperms is restricted to just a few lineages in the analyzed genome size of plant species so far. It is still an open question whether GS diversity is shaped by neutral or natural selection. The genus Lilium, with giant genomes, is phylogenetically and horticulturally important and is distributed throughout the northern hemisphere. GS diversity in Lilium and the underlying evolutionary mechanisms are poorly understood. We performed a comprehensive study involving phylogenetically independent analysis on 71 species to explore the diversity and evolution of GS and its correlation with karyological and environmental traits within Lilium (including Nomocharis). The strong phylogenetic signal detected for GS in the genus provides evidence consistent with that the repetitive DNA may be the primary contributors to the GS diversity, while the significant positive relationships detected between GS and the haploid chromosome length (HCL) provide insights into patterns of genome evolution. The relationships between GS and karyotypes indicate that ancestral karyotypes of Lilium are likely to have exhibited small genomes, low diversity in centromeric index (CVCI) values and relatively high relative variation in chromosome length (CVCL) values. Significant relationships identified between GS and annual temperature and between GS and annual precipitation suggest that adaptation to habitat strongly influences GS diversity. We conclude that GS in Lilium is shaped by both neutral (genetic drift) and adaptive evolution. These findings will have important consequences for understanding the evolution of giant plant genomes, and exploring the role of repetitive DNA fraction and chromosome changes in a plant group with large genomes and conservation of chromosome number.

The objective of the present work was to induce chromosome doubling in genotypes of Brachiaria ruziziensis with potential for production of good-quality biomass and tolerance/resistance to stress caused by aluminum and spittlebugs. Germinated seeds of B. ruziziensis were immersed in 0.1% colchicine solution for 2 or 3 hours. Ploidy level was determined by flow cytometry and confirmed by chromosome count. Fertility of the tetraploid plants was assessed through evaluation of pollen viability by staining. There was no statistical difference with regard to seedling survival and number of duplicate seedlings between the treatments. Survival rate was 8%, of which 11.45% were tetraploid genotypes. The mean viability of pollen grains from the evaluated tetraploid plants ranged from 51.75 to 55.50%. So, the tetraploids plants obtained from genotypes of a bred population of ruzigrass produce fertile pollen, in percentages that render crosses viable in genetic breeding programs.

Flow cytometry is a technique that yields rapid results in analyses of cell properties such as volume, morphological complexity and quantitative DNA content, and it is considered more convenient than other techniques. However, the analysis usually generates histograms marked by variations that can be produced by many factors, including differences between the software packages that capture the data generated by the flow cytometer. The objective of the present work was to evaluate the performance of four software products commonly used in flow cytometry based on quantifications of DNA content and analyses of the coefficients of variation associated with the software outputs. Readings were obtained from 25 ‘NBA’ (AA) banana leaf samples using the FACSCalibur (BD) flow cytometer, and 25 histograms from each software product (CellQuest™, WinMDI™, FlowJo™ and FCS Express™) were analyzed to obtain the estimated DNA content and the coefficient of variation (CV) of the estimates. The values of DNA content obtained from the software did not differ significantly. However, the CV analysis showed that the precision of the WinMDI™ software was low and that the CV values were underestimated, whereas the remaining software showed CV values that were in relatively close agreement with those found in the literature. The CellQuest™ software is recommended because it was developed by the same company that produces the flow cytometer used in the present study.

Mitotic activity and nuclear DNA content in endosperm of Zea mays cv. Złota Karłowa were examined. DNA content was cytophotometrically measured on squashed preparations after Feulgen procedure. Mitotic activity in endosperm was determined till the stage when embryo sack reached 4.5 mm in length. Some of mitotic figures show multiplied DNA content. Endosperm nuclei have various DNA contents which increase throughout endosperm development. DNA content enhancement indicates endoreduplication in progress. Some nuclei with high DNA content display changes in chromatin structure, which are expressed by the presence of strands and aggregates of chromatin characterised by high staining intensity. A conclusion has been drawn that mitotic divisions and the endoreduplication phase of nuclear DNA may occur simultaneously and dominate one over another at different phases of endosperm development.

The DNA content after the Feulgen reaction in the endosperm of three monocotyledonous plant species (Asparagus officinalis, Muscari comosom, Haemanthus kurharinae) differing in their 2C DNA content, was cytophotometrically measured. During endosperm development 1-6 endoreplication cycles take place, depending on the species. Differences in nuclear DNA endoreplication dynamics in the tested species are similar to those occurring in root parenchyma, but the endoreplication level in the endosperm is higher.

DNA, NYS and DNFB protein contents were measured cytophotometrically using the Feulgen method in the nuclei of the epidermis from. the basal zone of young leaves and from the basal and apical zones of old leaves in two perennial monocotyledonous species, Clivia miniata and Rhoeo discolor. Dry mass was determined interferometrically. It was shown that nuclei with a 2C DNA content dominated in both zones of old leaves, and that a significant percentage of cells with a DNA content below 2C were present. The ratio between euchromatin DNA and heterochromatin DNA indicates a greater decrease in euchromatin during aging. Changes in DNA due to real DNA loss are accompanied by decreases in NYS and DNFB stained proteins and a decrease in dry mass content correlated mainly with the decrease in the amount of NYS proteins.

Using cytophotometric procedures, following Feulgen-NYS staining, the measurements of DNA and nuclear, nucleolar and cytoplasmic protein contents in cortex cells of pea roots growing for 144 h in calcium and/or heavy metals (Cd2+, Cr3+, Pb2+) presence were made. All tested metals treatment reduced the number of nuclei in 4C DNA class and induced appearance of nuclei with DNA amount below 2C, that was expressed in diminished DNA content. The level of NYS proteins in cells underwent also reduction. In lead presence protein content diminished in nucleus. On the other hand, increased amount of nuclear, nucleolar and cytoplasmic proteins was observed in material treated with cadmium while only of nucleolar protein content in chromium presence. In root cells treated with tested metals protein content in nucleus was related with ploidy level, disturbances in this relation appeared in nucleolus and mostly in cytoplasm. Calcium added to chromium and mostly to lead solutions diminished the toxic effect of these metals that was demonstrated by an increase in DNA content, although calcium alone reduced DNA amount in nucleus due to lower number of 4C nuclei accompanied by appearance of 1C and 1-2C DNA classes. Calcium in different ways affected protein content changed by metal treatment. Present in cadmium solution it caused a further reduction in protein content in nucleus, nucleolus and cytoplasm but increased nuclear and cytoplasmic protein when added to lead, and nucleolar proteins - in chromium solution. Moreover, calcium ions presence in metal solutions did not restore the relationship between ploidy level and nucleolar and cytoplasmic NYS stained proteins and it did not disturbe the relation existing in nucleus.