Nuclear DNA content and phytohormone requirements of normal, crown gall and habituated tissues of Nicotiana tabacum var. White Burley

Determination of the nuclear DNA content of leaves and normal, habituated and Crown gall callus tissues of Nicotiana tabacum var. White Burley were performed using cytophotometry on Feulgen stained preparations. Several aspects concerning the reliability of the Feulgen technique for DNA determinations were investigated.Crown gall callus tissue used in this study had both a higher nuclear DNA content and chromosome number than normal callus (3.2C versus 2.5C). Both have a higher DNA content than the diploid tobacco leaf cells (2C).The normal callus tissue failed to grow on medium without indole acetic acid and kinetin when cultured in tubes. From this normal callus two habituated lines growing without both phytohormones were selected by culturing the normal callus first in the absence of either indole acetic acid or kinetin. Changing the culture conditions of the normal callus by using culture flasks instead of tubes resulted in a remarkably faster growth rate of the tissue. This was accompanied by an acquisition of the habituation characteristics since it was possible now to grow this tissue also directly on medium lacking both phytohormones. All habituated tissues showed a higher nuclear DNA content compared to the normal callus tissue from which they were derived. Interestingly, one of the tissues acquired a nuclear DNA content not different from that of Crown gall tissue. By changing the culture conditions of Crown gall callus tissue no concomitant change in nuclear DNA content occurred.The results suggest a correlation between the acquisition of a special chromosome complement and the loss of phytohormone requirement resulting in autonomous growth.     

Analysis of the genetic stability of <Emphasis Type="Italic">Eucalyptus globulus</Emphasis> Labill. somatic embryos by flow cytometry

Flow cytometry was used to measure the nuclear DNA content of Eucalyptus globulus Labill. somatic and zygotic embryos and leaves in order to determine if somatic embryogenesis induces DNA content and ploidy changes in this species. Mature zygotic embryos derived from open-pollination orchard families were collected from a location in the centre of Portugal. One group was kept for nuclear DNA content and ploidy analysis, and the other group was used for establishing embryogenic cultures. Mature zygotic embryos were grown on Murashige and Skoog (MS) medium supplemented with 3% (w/v) sucrose and 3 mg l^–1 -naphthaleneacetic acid (NAA) for 3 weeks and then transferred to MS medium without growth regulators. Globular somatic embryos from approximately 8-month-old embryogenic cultures were used in the assay. DNA ploidy levels and the nuclear DNA content of mature zygotic embryos, somatic embryos and leaves from the mother field tree were determined using flow cytometry combined with propidium iodide staining. Zygotic embryos had a nuclear DNA content of 1.32 pg/2C, somatic embryos had a nuclear DNA content of 1.39 pg/2C and leaves from the field tree had a nuclear DNA content of 1.40 pg/2C. The values estimated for the somatic embryos and mother plant did not differ statistically from each other (P0.05), but both differed from those of the zygotic embryos (P0.05). These results clearly indicate that no changes were induced during the embryogenic process. However, the differences found between the field plants and zygotic embryos did suggest that some aspects must be evaluated carefully, as propidium iodide fluorescence may potentially be influenced by the presence of secondary compounds (e.g. anthocyanins, tannins) in E. globulus somatic embryos and mature leaves. Therefore we believe that the somatic embryogenesis methodology used did not induce major genetic changes in the somatic embryos and that our primary goal of true-to-type propagation was assured.     

Genetic analysis of enhanced-axillary-branching-derived Eucalyptus tereticornis Smith and E. camaldulensis Dehn. plants

In a culture method for enhanced axillary branching functional plants of Eucalyptus tereticornis and E. camaldulensis are efficiently regenerated. To assess the genetic integrity among the regenerants, we employed multiple analytical tools including cytochemical and molecular assays. The 2C DNA amounts were estimated in the meristematic zones of root and shoot tips of 250 micropropagated plants, collected at various cycles of tissue culture from multiplication to field transfer, and compared to the corresponding mother plants. The culture conditions did not induce amplification or deletion of DNA sequences, nor were there drastic change(s) in chromosome number, since all the micropropagated plants of E. tereticornis (1.2 pg) and E. camaldulensis (1.4 pg) maintained the same DNA amounts as the mother plant. Total DNA of 46 micropropagated and mother plants digested with eight restriction enzymes and hybridized to 13 nuclear, mitochondrial, and synthetic oligonucleotide DNA probes yielded 82 bands. Hybridization patterns indicated that the variation observed was minor. To further confirm the genetic fidelity, 12 arbitrary 10-base primers and six synthetic oligonucleotide sequences, successfully used to amplify genomic DNA from in vivo and in vitro materials, produced 133 fragments that were monomorphic across the plants tested. The present results demonstrate that enhanced-axillary-branching culture of mature trees could be utilized commercially for mass clonal propagation of these two important Eucalyptus species that have been recalcitrant to vegetative propagation. The results also provide novel insights into the genetic differences between E. tereticornis and E. camaldulensis. Received: 8 October 1996 / Revision received: 22 July 1997 / Accepted: 30 July 1997     

Environmentally induced nuclear 2C DNA content instability in Helianthus annuus (Asteraceae)

Experiments were conducted to detect developmental and environmental factors that influence nuclear DNA content in H. annuus inbred lines RHA 271 and RHA 299. DNA content (2C) was determined by laser flow cytometry of nuclei isolated from the first leaf pair of seedlings grown in a greenhouse and in growth chambers. DNA content of greenhouse grown seedlings was highly variable, ranging from 3.2 to 8.0 pg for RHA 299 and 5.2 to 8.2 pg for RHA 271. DNA content only weakly correlated to the position of the achene in the head from which the seedlings derived, and not at all to the position of the head on the plant. Experimentally varied environmental parameters of heat stress and water deficit, phosphate fertilizer levels in the substrate, and pH had little or no effect on the DNA content of seedlings. Seedlings grown with increased levels of substrate nitrogen in the form of NH₄NO₃ showed a significant increase in the mean DNA content. Plants grown in one of two types of growth chambers possessed less variability in DNA content, 6.2–8.4 pg for RHA 299 and 6.8–7.4 pg for RHA 271. Plants grown in a second growth chamber were highly variable with DNA content ranging from 3.0 to 8.6 pg for RHA 299 and 3.0 to 7.8 pg for RHA 271. Measurable physical differences between the growth chambers were irradiance level and the ratio of red to far red light. The hypothesis is presented that DNA stability of sunflowers is influenced by light quantity and/or quality.     

Nuclear DNA content of Pinus sylvestris (L.) as determined by laser flow cytometry

Nuclear DNA content was determined in nuclei isolated from needles, stems and roots of in vitro grown seedlings and from megagametophytes and embryo of mature seeds in three accessions of Pinus sylvestris L. One accession was from Inari, northern Finland at timber line, and two accessions were from the Alpine region in Italy. Nuclei were mechanically isolated by a chopping method, stained with propidium iodide, and DNA content was determined using an EPICS PROFILE laser flow cytometer. Nuclei isolated from leaves of barley (Hordeum vulgare L. cv. Sultan; 2C=11.12 pg) were used as an internal standard for measurement of pine nuclei. Mean 1C nuclear DNA content of P. sylvestris was 27.88 pg as determined from megagametophyte tissue. Mean 2C value was 52.25 pg as determined from stem and root tissue, and 55.58 pg as determined from embryo tissue. The ratio of 2C to 1C value was 1.87 and 1.99, respectively. Extracts of nuclei from needles contained propidium iodide-absorbing debris which may have interfered with measurements and resulted in lower 2C values than those obtained from stem and root.     

Nuclear DNA content in differentiated tissues of sunflower (Helianthus annuus L.)

Summary Scanning cytophotometry following Feulgen-staining was used to determine nuclear DNA content in many differentiated tissues of nine cultivars, hybrids or selfed lines ofHelianthus annuus. Apart from such ephemeral tissues as endosperm and anther tapetum, it was found that tissue differentiation in sunflower occurs in the diploid condition, cells being arrested in the DNA presynthetic phase (G₁). In certain cases, however, the nuclear DNA content of differentiated G₁ cells does not exactly match the 2C DNA content found in meristematic cells, but may be either higher or lower. In endosperm and anther tapetum cells, nuclear DNA content may be as high as 24 C and 32 C, respectively. Cytological and autoradiographic analyses after³H-thymidine incorporation reveal that polyploidy in the tapetal cells is due to chromosome endoreduplication. No detectable difference between male-fertile and male-sterile plants exists as far as occurrence and level of cell polyploidy are concerned. The results are discussed in the context of previous investigations on the nuclear condition of differentiatedHelianthus annuus tissue.     

Number and DNA content of nuclei in the free-living nematode Panagrellus silusiae at each stage during postembryonic development

Nuclei from the four major tissues of the nematode Panagrellus silusiae were enumerated and examined using Feulgen microspectrophotometry at each stage during postembryonic development. The number of nuclei in the hypodermis, nerve, and intestine remains fairly constant during maturation, but there is a slight increase (57%) in the number of muscle nuclei. Thus, this organism is not stringently eutelic. The total number of somatic nuclei is about 600. DNA values of hypodermis and nerve nuclei were unimodal and adult nuclei had 2C amounts of DNA. The DNA distribution of muscle nuclei reflects the pattern expected for a tissue in which a portion of the nuclei are undergoing DNA synthesis. Intestinal nuclei accumulated DNA in the absence of nuclear division and in the adult the nuclei fall into discrete DNA classes which correspond to a geometric series of the 2C value. It is concluded that chromatin diminution does not occur in this species. In addition, the relationship in the different tissues of nuclear DNA content to nuclear volume and cell size is discussed.The study was supported by the National Research Council of Canada (Grant A-3491).     

基于响应面设计茎段形成层培养猕猴桃幼苗的优化研究

为降低猕猴桃组培快繁中的污染率,提高其繁殖效率,该文以猕猴桃的幼嫩茎段为外植体,采用两步培养法进行茎段形成层的愈伤及成苗诱导研究,并利用响应面设计软件对NAA浓度、6-BA浓度、低渗处理时间进行了各条件的优化,同时通过组织切片确定愈伤的来源及幼苗的形成方式。结果表明:(1)培养过程中撕除茎段周皮能显著降低污染率,用200～400 mg·L^-1的PVP处理猕猴桃茎段可有效防止去皮茎段的褐化。(2)愈伤诱导的最佳条件为预培养28.3 h、NAA 4.45 mg·L^-1、6-BA 0.28 mg·L^-1,而幼苗形成的最佳条件为预培养26.4 h、NAA 4.84 mg·L^-1、6-BA 0.42 mg·L^-1。这表明形成层愈伤诱导需较长低渗处理时间和较高生长素,而成苗诱导则需较高生长素、激动素及较短的低渗处理时间。(3)组织切片观察结果表明猕猴桃愈伤组织源于形成层干细胞的分裂,且幼苗株源于胚状体的发育。综上结果表明,通过除去猕猴桃嫩茎周皮,外加抗氧化、低渗处理,可有效降低猕猴桃组培快繁中的污染率,提高繁殖系数和胚状体发生率,为猕猴桃种苗的规模化生产提供技术支撑。     

The relation of DNA synthesis and mitosis in tobacco pith tissue cultured in vitro

Summary DNA synthesis and mitosis were initiated in cultured tobacco pith tissue by means of IAA and kinetin. DNA classes were determined by microspectrophotometric measurements (Feulgen); autoradiographs (tritiated thymidine) served to ascertain whether or not nuclei had undergone DNA synthesis during culture.All mitoses in new cells (resulting from divisions in culture) were diploid and had been preceded by DNA synthesis in culture.Whereas many of the old cells (which had not previously divided in culture) found in diploid or polyploid mitosis had undergone DNA synthesis during culture, others had not. Such non-radioactive mitoses still occurred after 16 days.In view of this, a 4 C nucleus in differentiated tissue should be considered as potentially both diploid and tetraploid, for it appears impossible to predict whether it would, upon restoration of conditions conducive to DNA synthesis and mitosis, enter a diploid mitosis or, after undergoing DNA synthesis, a tetraploid one.A high nuclear DNA content seems to have a much more inhibiting effect on the onset of DNA doubling than on that of mitosis.Somatic polyploidization is understood as the result of two DNA doublings between which mitosis was omitted, or aborted, or in effect undone by a failure of cytokinesis leading to fusion during a later mitosis.This work has been supported by research grants to K. Patau from the U.S. Public Health Service (grant No. C-3313) and the American Cancer Society.     

Cellular responses of leaf explants of Cocos nucifera L. in vitro

Leaf explants of Cocos nucifera L. (coconut palm) were studied in vitro in order to establish whether or not rapid cellular changes contribute to the well known recalcitrance of coconut cells in tissue culture. Segments from the base of immature leaves were cultured on modified Eeuwens' medium at 30°C in darkness. The mitotic index, nuclear DNA amounts, cell and nuclear size were measured both before and during culture (from 0 to 70 days). There was no basipetal gradient of cell division in immature coconut leaves; the mitotic index never exceeded 2% and showed neither a positional nor temporal relationship with leaf development. Moreover the vast majority of cells were in G1 of the cell cycle. This cell cycle pattern was maintained for most of the period in culture although at 70 days there was an increase in the proportion of cells in S- and G2-phases consistent with low rates of callus formation. The nuclear: cell size ratio was constant in cells within the immature leaf irrespective of developmental age. However upon transfer to culture media, cell size but not nuclear size increased. We suggest that this uncoupling of cell and nuclear size disrupts cell co-ordination and is a key contributor to recalcitrant cellular behaviour of this species in vitro.     

Efficient callus induction and plant regeneration from anther of Chinese narcissus (Narcissus tazetta L. var. chinensis Roem)

Callus culture has, to date, been reported only in a few species of Narcissus. We used anthers of Chinese narcissus (Narcissus tazetta L. var. chinensis Roem) as explants for callus induction and plant regeneration. A high percentage of anthers at the early- to mid-uninucleate microspore stage were responsive on the basal MS medium supplemented with 0.5–1 mg l^–1 2,4-dichlorophenoxyacetic acid and 0.5–2 mg l^–1 6-benzyladenine under dark conditions. Calli were initiated from anther connective tissue or anther wall tissue, and no division of microspores occurred during callus formation, as determined by histological observation. Using 20 random amplified polymorphic DNA primers, we verified the genetic integrity of the anther-derived plants of Chinese narcissus with respect to the donor plants. These results suggest that anther culture in vitro can provide an efficient new micropropagation technique for Chinese narcissus as well as a new strategy for in vitro mass propagation of other daffodils.     

Changes in the amounts of nuclear RNA during interphase in Vicia faba

Variations in the amounts of nuclear RNA present during interphase in Vicia faba were studied by microphotometric, autoradiographic and chemical methods. In one series of experiments, nucleic acid estimations were carried out on root meristem nuclei isolated from cells which had been partially synchronized by treatment with 5-aminouracil at 20 and 25° C. In a second series, root meristem nuclei isolated from untreated plants growing at 4, 20 and 25° C, were separated into fractions, containing different interphase stages, by sucrose density gradient centrifugation. — The results from the two series suggest that at 4 and 20° C there is a net increase in the amount of nuclear RNA during interphase which parallels the net increase in DNA. At 25° C, however, there is less RNA per nucleus and this remains at the same level throughout interphase resulting in an average increase in the DNARNA ratio of 55%. — It is suggested that the balance between the synthesis and release of nuclear RNA may change not only within plants, at different stages of interphase, but also between plants according to the temperature at which they are grown.     

Effect of Mitomycin C on Thymidine-Methyl-H3 Utilization by Entamoeba histolytica Propagated in CLG Medium*

SYNOPSIS. Autoradiographic studies were done which tested the effect of a potent DNA inhibitor, mitomycin C (MC) on the utilization of tritium from exogenous thymidine-methyl-H³ (TMH³) in Entamoeba histolytica grown with Bacteroides sp. in CLG medium. Concentrations of MC (0.0002%) which inhibited growth of amebae by ca. 50%, caused an overall depression of tritium utilization by both associate cell and amebae. However, no reduction in percent cells with nuclear activity was apparent. The effect of MC on utilization of tritium in amebae propagated with Bacteroides which were prelabeled with TMH³ was also studied. The extent of labeling and percent amebae with cytoplasmic label was not appreciably depressed by MC. MC did, however, cause a depression of the percent amebae with nuclear label. This would indicate that the utilization of bacterial DNA products for nuclear DNA (reported in a previous communication) is reduced in the presence of MC. These data on the effect of MC on use of exogenous TMH³ and prelabeled Bacteroides provide some evidence that at least some of the nuclear DNA of amebae can be synthesized from the exogenously supplied isotope. Amebae grown with exogenous TMH³ and resuspended in unlabeled medium for 24–28 hrs. with and without MC had a considerable reduction of the extent of label whether MC was present or not. This suggests that the primary effect of MC is not to degrade DNA.     

EVIDENCE FOR ASEXUAL RESTING CYSTS IN THE LIFE CYCLE OF THE MARINE PERIDINOID DINOFLAGELLATE,SCRIPPSIELLA HANGOEI1

Scrippsiella hangoei (Schiller) Larsen is a peridinoid dinoflagellate that grows during winter and spring in the Baltic Sea. In culture this species formed round, smooth cysts when strains were mixed, indicating heterothallic sexuality and hypnozygote production. However, cysts of the same morphology were also formed in clonal strains exposed to slightly elevated temperature. To better understand the role of cysts in the life cycle of S. hangoei, cyst formation and dormancy were examined in culture experiments and the cellular DNA content of flagellate cells and cysts was compared in clonal and mixed strains using flow cytometry. S. hangoei exhibited a high rate of cyst formation in culture. Cysts produced in both clonal and mixed strain cultures were thick‐walled and underwent a dormancy period of 4 months before germinating. The S. hangoei flagellate cell population DNA distributions consisted of 1C, intermediate, and 2C DNA, indicative of respective eukaryotic cell cycle phases G1, S, and G2M. The majority (>95%) of cysts had a measured DNA content equivalent to the lower 1C DNA value, indicating a haploid nuclear phase and an asexual mode of cyst formation. A small percentage (<5%) of cysts produced in the mixed strain culture had 2C DNA, and thus could have been diploid zygotes. These findings represent the first measurements of dinoflagellate resting cyst DNA content, and provide the first quantitative evidence for dinoflagellate asexual resting cysts. Asexual resting cysts may be a more common feature of dinoflagellate life cycles than previously thought.     

ULTRASTRUCTURAL CYTOLOGY OF CALLUS CULTURES OF STREPTANTHUS TORTUOSUS AS AFFECTED BY TEMPERATURE

Tissue culture cells of Streptanthus tortuosus var. orbiculatus (Cruciferae) which have acquired a spherical viruslike particle located in their nucleoli, designated cell line STV, developed supergranal chloroplasts and lost the ability to differentiate vascular tissues. The effect of temperature on the ultrastructural cytology of one line of the STV tissue, STV-I, was compared with the effect of temperature on the ultrastructural cytology of tissue culture cells lacking the viruslike particles (control cell lines). At 4 C, the cellular and ultrastructural appearance of control tissue culture cells differed from that of tissue grown at 22 C by producing increased amounts of endoplasmic reticulum and dictyosomes and by reduction of chloroplast thylakoids. STV-I cells were generally moribund as a result of 4 C treatment. Chloroplast thylakoids were also reduced in control tissue following growth at 10 C and the apparent quantities of endoplasmic reticulum and dictyosomes were similar to those observed in control cells grown at the control temperature (22 C), but less than those observed in tissue subjected to 4 C. STV-I tissue grown at 10 C demonstrated increased endoplasmic reticulum and dictyosomes and reduction of polysomal configurations. The mitochondrial morphology was variable and the cells contained supergranal chloroplasts and proplastids. At the control temperature (22 C), the fine structural appearance of control tissue culture cells was typical of parenchyma cells, but STV-I cells contained mitochondria of variable morphology and two types of chloroplasts— normal and supergranal. Control tissue grown at 30 C also contained proplastids, but these proplastids contained starch in contrast to the proplastids in control tissue grown at low temperatures. The ultrastructural cytology of STV-I cells grown at elevated temperature (30 C) was characterized by enlarged mitochondria containing massive lipid bodies and the presence of protoplastids with starch and supergranal chloroplasts.     

Genome size and environmental factors in the genus Pinus

Nuclear 1C DNA content in haploid megagametophyte tissue of 18 North American and one exotic Pinus species was determined using scanning microspectrophotometry. The nuclear DNA content in root meristematic cells of Zea mays L. ssp. mays, inbred line Va35 (4C = 10.31 pg) was used as a standard. DNA content measured by microspectrophotometry was verified using laser flow cytometry with two additional standards, Hordeum vulgare cv. Sultan (2C = 11.12 pg) and P. eldarica (2C = 47.30 pg). DNA values obtained by both methods were significantly correlated (r = 0.987). The 1C nuclear DNA content ranged from 21 pg to 31 pg. The ratio of DNA content in embryo tissue of P. eldarica to that in megagametophyte tissue was 1.72 by scanning microspectrophotometry and 1.74 by laser flow cytometry. To date, this is the most comprehensive data set available for North American Pinus species. Relationships between genome size of 18 North American Pinus species and climatic factors and indices of growth were investigated using regression and correlation analyses. Positive correlations were observed between nuclear DNA content and growth indices, minimum seed-bearing age, and seed dimensions. Strong negative correlations were observed between nuclear DNA content and two climatic factors, the lowest mean annual and monthly precipitation (excluding January) and the highest mean monthly spring air temperature. These correlations suggest that the large genome size and its variation in Pinus are adapted responses to the habitats of these species.     

Detection of protoplast-derived DNA tetraploid Lisianthus (Eustoma grandiflorum) plants by leaf and flower characteristics and by flow cytometry

Plants of lisianthus (Eustoma grandiflorum (Griesbach)Schinners=Lisianthus russellianus Hook.) were regenerated from protoplasts and grown in pots until flowering. Vegetative and floral characteristics were measured and compared with parent plants. Larger leaves and petals and longer guard cells, sepals and filaments were recorded from protoplast-derived plants suggestive of polyploidy. The nuclear DNA contents of protoplast-derived and parental plants were determined by flow cytometry. Protoplast-derived plants were confirmed as DNA tetraploid by flow cytometry with a DNA index of 1.95. Their nuclear DNA content was measured as 6.33±0.04 pg DNA per 2C nucleus compared with 3.26±0.10 pg DNA per 2C nucleus from parental plants. Polyploidisation induced during protoplast regeneration offers an alternative to that of colchicine treatment.     

The genetic stability of Penicillium chrysogenum transformants in a fermentor

Summary A number of transformants of Penicillium chrysogenum have been obtained with the plasmid vector p3SR2. Southern analysis showed that transformation had occurred by integration of vector sequences into the nuclear DNA of the fungus. A number of transformants were tested for stability of the transformed phenotype in agar medium and some were found to be unstable. Two transformants, shown to be stable in agar culture, were grown in 5-l batch fermentors for further stability tests. Over periods of up to 312 h in the fermentor both transformants were 100% stable with respect to the transformed phenotype. In addition Southern analysis of DNA extracted from the spent mycelium showed that no change had occurred in the position of the integrated vector sequences within the transformant nuclear DNA.     

ISSR and RAPD based evaluation of genetic fidelity and active ingredient analysis of regenerated plants of Picrorhiza kurroa

Genetic stability and phytochemical analysis of in vitro established plants of Picrorhiza kurroa Royle ex Benth, have been carried out. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) markers were used to assess the genetic fidelity of tissue culture products including three adventitious shoots from three calli and 6 months old tissue culture raised plants growing in green house condition with mother plant. Apparent genetic variation was detected in the five types of plant materials. The percentage of polymorphic bands in the RAPD and ISSR analysis were 16.25 and 14.54 %, respectively. The genetic similarity was calculated on the basis of RAPD and ISSR data among the five types of plant materials and were ranged from 0.5 to 1.0 (mean 0.75) and 0.47 to 1.0 (mean 0.73), respectively. The similarity coefficient by both RAPD and ISSR analysis revealed that differences between tissue culture raised plants and mother plant was not remarkable, but notable differences were observed among three adventitious shoots regenerated from three calli. The phytochemical analysis of tissue culture raised products showed higher secondary metabolite (picrotin and picrotoxinin) content as compare to mother plant. The information gained on genetic stability/variability will be valuable for the large scale propagation and secondary metabolite production of P. kurroa.     

Flow cytometric measurement of nuclear DNA content inCapsicum (Solanaceae)

Nuclei were isolated from leaf tissue of differentCapsicum species and the relative fluorescence intensity was measured by flow cytometry after propidium iodide staining.Pisum sativum nuclei with known nuclear genome size (9.07 pg) were used as internal standard to determine nuclear DNA content of the samples in absolute units. The 2C DNA contents ranged between 7.65 pg inC. annuum and 9.72 pg inC. pubescens, and the general mean of the genus was 8.42 pg. These values correspond, respectively, to 1C genome size of 3.691 (C. annuum), 4.690 (C. pubescens) and 4.063 (general mean) Mbp. In general, white-flowered species proved to have less DNA, with the exception ofC. praetermissum, which displayed a 2C DNA content of 9.23 pg. It was possible to divide the studied species into three main groups according to their DNA content, and demonstrate differences in DNA content within two of the three species complexes established on the basis of morphological traits.