Increasing Ploidy Level Research Articles

High Frequency of Polyploidization in Regenerated Plants of Kalanchoe blossfeldiana Cultivar 'Tetra Vulcan'.

We used flow cytometry to analyze chromosomal changes in regenerants of the Kalanchoe blossfeldiana cultivar ‘Tetra Vulcan’ (4x). About 80% of regenerants showed increased ploidy levels. Twenty-four (20.7%), 87 (75.0%), 1 (0.9%), and 4 (3.4%) regenerants had ploidy levels of 4x, 8x, 12x, and 16x, respectively. 4x and 8x regenerants grew normally and similarly to wild-type plants, but 12x and 16x plants showed remarkable delays in growth. Plant height and leaf size among the 4x and 8x plants were the same as in wild-type plants, but those of the 12x and 16x plants were dramatically reduced. Leaves of plants with ploidy levels of 8x or more were thicker than those of 4x plants, and this character increased with increasing ploidy level. Our results confirmed that regeneration from leaf segments is an efficient method of polyploidization in Kalanchoe.

Increasing ploidy level in cell suspension cultures of Doritaenopsis by exogenous application of 2,4-dichlorophenoxyacetic acid

To clarify the causal factors for ploidy variation in plant cell culture, we attempted to alter ploidy distribution in cell cultures of a tetraploid cultivar of Doritaenopsis by changing the plant growth regulators (PGRs) in the culture medium. The original suspension cultured cells, which had been maintained in medium containing 0.1 mg l-1 1-naphthaleneacetic acid and 1 mg l-1 benzyladenine, were transferred onto various gellan gum solidified media with a single application of PGRs, and the ploidy distributions of the cells were examined using flow cytometry analysis during 3 weeks of culture. Among the PGRs tested, 2,4-dichlorophenoxyacetic acid (2,4-D) and 4-amino-3,5,6-trichloropicolinic acid caused a drastic reduction in the 4C-cell proportion in cell cultures with an increased cell proportion of 8C or higher C-values. In the case of 2,4-D application, a reduction of cell viability was observed. A decreasing proportion was also observed in the 8C-cell population accumulated by 2,4-D treatment, following transfer back to the medium containing the standard PGR composition. These results suggest that the exogenous application of 2,4-D arrested the cell cycle at G2 phase in the Doritaenopsis cells, and the removal of 2,4-D might induce further endoreduplication or recover the mitotic cycle of the G2-arrested cells.

Differences between hybrids ofSolanum tuberosum L. andSolanum circaeifolium Bitt. obtained from symmetric and asymmetric fusion experiments

Potato plants obtained by symmetric and asymmetric protoplast fusion experiments withSolanum tuberosum andSolanum circaeifolium were compared and their potential use for plant breeding was investigated. For asymmetric hybridisation wild species tissue was irradiated with X-rays. Irradiation enhanced regeneration frequency. RFLP analysis showed that both techniques gave approx. 95% hybrids among the regenerates. Asymmetric protoplast fusion gave 22% regenerates with a ploidy level between 2x and 4x or 4x and 6x. None of the regenerates obtained from symmetric fusion had these ploidy levels. The tetraploid hybrids showed good male and female fertility. Male fertility was highest for the eutetraploid regenerates. With increasing asymmetry and with increasing ploidy level fertility decreased. Berry induction and seed production were observed for the eutetraploid and, to a lesser degree, for the hypotetraploid and hypohexaploid hybrids.

Effect of ploidy level on fruit morphology, seed germination and juvenile growth in scurvy grass (Cochlearia officinalis L. s.l., Brassicaceae)

AbstractThe biennial to short‐lived wintergreen perennial Cochlearia officinalis L. s.l. (basic chromosome number x = 6) encompasses a group of closely related taxa that is difficult to delimitate morphologically. Despite a lack of distinct sets of discontinuous morphological characters, at least six numerically different cytotypes have commonly been recognized, and they have different ecologic distributions. Evolution of the wide ploidy level was likely brought about by hybridization and polyploidization. Because plants are short‐lived, seed germination was established and was likely to favor subsequent establishment. Therefore, shoot growth of juveniles was also compared. Five ecologically allopatric populations of C. officinalis s.l., each supposedly representing a different cytotype, were sampled for flowering shoots and seeds to establish shoot morphological traits, germination capabilities and chromosome number. The sampled populations differed in chromosome number: 2n = 2x = 12 (diploid); 2n = 4x = 24 (tetraploid); 2n = 33; 2n = 6x = 36 (hexaploid); and 2n = 8x = 48 (octaploid), representing the taxa C. pyrenaica DC, C. officinalis L. s.s., C. danica L. ×C. officinalis L. s.s., C. x hollandica Henrard and C. anglica L., respectively. All cytotypes produced abundant seeds and seed weight was proportional to the ploidy level. Seed weight of the diploid C. pyrenaica did not follow this correlation. Germination capacity immediately after seed collection was high over a wide temperature range. Germination was non‐photoblastic. High germination capability accentuated the short‐lived life‐cycles of the five cytotypes. Seed germination of the coastal cytotypes (C. x hollandica and C. anglica) were less negatively affected by NaCl compared to the non‐coastal cytotypes (C. pyrenaica and C. officinalis s.s.) and the very rare coastal hybrid C. danica×C. officinalis s.s. The cytotypes differed markedly in growth characteristics but there was no simple relationship between them. Increases in ploidy level were not paralleled by increased shoot growth. The hexaploid hybrid C. x hollandica indicated hybrid vigor in both shoot growth (yield) and petiole length of the strongly ascending leaves. Leaves of C. anglica were arranged in a prostratous rosette and the difference in shoot architecture between both C. x hollandica and C. anglica may be one of the reasons why this species is being replaced by the hexaploid hybrid, if sympatric, in coastal regions of The Netherlands. The 2n = 33‐hybrid C. danica×C. officinalis s.s. showed no such heterosis effects in shoot growth characteristics.

Photosynthetic Rates and Ploidy Levels among Populations of Switchgrass

Photosynthetic rates reportedly increase with increasing ploidy levels among lowland and upland ecotypes of switchgrass (Panicum virgatum L.). Such reports are based on data derived from native plant materials and no investigation has tested whether a similar relationship exists among released or experimental populations. A series of studies were conducted to address this question, with rates of photosynthesis measured with an infrared gas analyzer and ploidy levels estimated with flow cytometric analysis of nuclear DNA contents. Analyses of nuclei isolated from young stems and stained with propidium iodide indicated that of the 14 populations examined, eight are tetraploids (‘Alamo’1, ‘Kanlow’, ‘NC‐116’, ‘NC‐216’, ‘PMT‐279’, ‘PMT‐785’, ‘Stuart’, and ‘Wabasso’) and six are hexaploids (‘Blackwell’, ‘Caddo’, ‘Cave‐in‐Rock’, ‘Pathfinder’, ‘Shelter’, and ‘Trailblazer’). Twelve of these populations (not including NC‐116 and NC‐216) were grown in the greenhouse and photosynthetic rates measured on the youngest fully expanded leaf blade 40 to 45 d after germination. Rates of photosynthesis did not differ among populations or between tetraploids and hexaploids. Field studies at Knoxville, TN, Stephenville, TX, and Blacksburg, VA, did indicate, however, that tetraploids often had mean photosynthetic rates 12 to 18% higher than those of hexaploids. These differences were observed at many field sites during May, June, and July, yet based on repeated sampling at Knoxville, rates of leaf photosynthesis measured in September were up to 6.5 μmol CO2m−2 s−1 higher in hexaploid compared with tetraploid populations. Contrary to earlier reports that photosynthetic rates increase with increasing ploidy level, our results indicate that such a relationship is not consistent for tetraploid and hexaploid populations of switchgrass but rather may change with season or environmental stress.

Human hepatocyte polyploidization kinetics in the course of life cycle.

The processes of polyploidization in normal human liver parenchyma from 155 individuals aged between 1 day and 92 years were investigated by Feulgen-DNA cytophotometry. It was shown that polyploid hepatocytes appear in individuals from 1 to 5 years old. Up to the age of 50 years the accumulation rate of binucleate and polyploid cells is very slow, but subsequently hepatocyte polyploidization is intensified, and in patients aged 86-92 years the relative number of cells with polyploid nuclei is about 27%. Only a few hepatocytes in the normal human liver reach 16C and 8C x 2 ploidy levels for mononucleate and binucleate cells respectively. Using a mathematical modeling method, it was shown that during postnatal liver growth the polyploidization process in human liver is similar to that in the rat, and that polyploid cells are formed mainly from binucleate cells. As in rats, prior to an increase in ploidy level, diploid human hepatocytes can pass several times through the usual mitotic cycles maintaining their initial ploidy level. After birth, only one in ten hepatocytes starting DNA synthesis enters the polyploidization process. At maturity about 60% of 2C-hepatocytes starting DNA synthesis divide by conventional mitosis, the rest dividing by acytokinetic mitosis leading to the formation of binucleate cells. During ageing the probability of hepatocyte polyploidization increases and in this period there are two polyploid or binucleate cells for every diploid dividing by conventional mitosis.

On the polyploid origin ofGaltonia’Moonbeam’

Abstract Galtonia ’Moonbeam’ is shown to be a tetraploid. Details of the chromosome number and karyotype are presented in comparison with those of Galtonia candicans (Baker) Decne. The significance of increased ploidy level is discussed and contrasted with increased ploidy levels reported in other bulbous ornamental plants.

Frequency analyses of active NORs in nuclei of artificially induced triploid fishes

The correspondence between increased numbers of both chromosomal and nuclear NORs and artificially induced triploidy in three fish species (rainbow trout, Oncorhynchus mykiss; common carp, Cyprinus carpio; and tench, Tinea tinea) has been confirmed by CMA3 fluorescence and Ag-staining. The frequencies of cell nuclei with one, two and three active NORs, as revealed by Ag-staining, has been analyzed statistically to find the minimum cell number which verifies the increased ploidy level. A minimum sample size of about 80 cells exhibiting three active NORs is sufficient to confirm triploidy in all three species and may be of use for categorising other ploidy-manipulated fish species.

Evolution of parthenogenesis in the Otiorhynchus scaber complex

A new approach for estimating genetic identity values was applied to a data set on allozyme variation in the weevil, Otiorhynchus scaber (Coleoptera: Curculionidae). This species complex contains sexuals and different races of polyploid asexuals. The results were used to reanalyse the phylogenetic relationship among races and to discuss the monophyletic versus the polyphyletic origin of parthenogenesis. A polyphyletic origin of parthenogenesis was suggested based on the assumption of hybridization between related species, and the extinction of an ancestral species. The genetic diversity among the different races was compared. A decreased number of new mutations with increased ploidy level was found among current populations. This correlation was explained by a lower evolutionary rate in polyploid asexual races than in diploid sexual races.

Cytogenetic studies in the genus Chaetobromus (Poaceae: Arundineae)

The genus Chaetobromus consists of four, somewhat ill defined species. Morphological merging and the existence of intermediates render the separation of these species difficult. A basic chromosome number of x = 6 for Chaetobromus is substantiated by the presence of diploid specimens. The genus forms a polyploid complex, ranging from diploid to duodecaploid. Polyploidy occurs in the form of segmental alloploidy and occasionally as alloploidy. Meiotic abnormalities, in the form of univalents during metaphase I and anaphase laggards, were observed in some specimens. A cancerous growth form, involving the merging of different cells and the subsequent fusion of their nuclei, has been observed in two specimens, Spies 3366 and CW1. No apparent explanation could be found for this uncontrolled increase in ploidy level during microsporogenesis. Extensive morphological, anatomical, and cytogenetical variation exists in the genus, indicating hybridization and polyploidization. Numerical analysis of morphological data obtained from the specimens studied indicates two groups. Cytogenetic evidence supports the numerical analysis. In an attempt to resolve the contributor species to this polyploid hybrid swarm, attention should be given to the diploid representatives of the putative progenitor species.Key words: Chaetobromus, hybridization, meiosis, numerical taxonomy, polyploidy.

Variations in leaf terpenoids with ploidy level in citrus cultivars and hybrids

The leaves of six diploid and six tetraploid citrus taxa contain monoterpenes, sesquiterpenes, phytol, β-amyrin and three phytosterols. The total amounts of lower terpenoids and phytosterols per gram fresh weight of leaf decreased with increasing ploidy level. The same trend was apparent when the amounts of the individual terpenoid components were analysed, although the phytosterol composition showed less variation than in the case of the lower terpenoids.

Nuclear DNA variation in diploid and polyploid taxa of Larrea (Zygophyllaceae)

A study of nuclear DNA content was made in telophase nuclei (2C) of the root apex of germinating seed in nine populations of the following species and cytotypes of Larrea: L. nitida (2x), L. divaricata (2x), L. cuneifolia (4x) and L. tridentata (2x, 4x, 6x). There were no significant differences in DNA content per basic monoploid genome among the diploid taxa nor between the latter and the tetraploid, among tetraploids or between tetraploids and the hexaploid. On the other hand, the difference between means was significant when all diploids were compared with the hexaploid cytotype. These results would indicate: (1) Speciation at the diploid level in Larrea has not produced great differences in DNA content per basic genome. This is in contrast with the related genus Bulnesia. (2) In Larrea there is a slight diminution in DNA content per basic genome when there is an increase in ploidy level. (3) Species of Larrea, Bulnesia and Pintoa (Zygophyllaceae) that inhabit the most arid environments are the ones possessing the highest DNA content. (4) This increase is due to an increment in ploidy level in Larrea and an augment of intrachromosomal DNA in Bulnesia and Pintoa.

Ploidy levels and DNA synthesis in fat body cells of the adult mosquito, Aedes aegypti: The role of juvenile hormone

AbstractAdult females of the mosquito Aedes aegypti showed two cycles of DNA replication in the fat body based on microspectrophotometric measurement of changes in nuclear DNA. The first cycle began after emergence and resulted in 80% of diploid fat body cells becoming tetraploid and 20% becoming octoploid by the end of the third day. The second replication cycle occurred 48–72 h after a blood meal and resulted in an increase in octoploid nuclei to 67% Topical application of juvenile hormone or methoprene to abdomens isolated at emergence stimulated an increase in ploidy levels above that normally seen in situ. Synthesis of DNA, estimated by incorporation of injected [3H]‐thymidine, rose after emergence and remained high for 2 days. Synthesis increased again after a blood meal, reached a peak by 6 h, and returned to low levels by 24 h after the meal. The timing of DNA synthesis and a measurable increase in ploidy were temporally separated. The ploidy increase, but not DNA synthesis, was correlated with increases in juvenile hormone levels.

Morphological variation in relation to chromosomal variation in Saccharum bengalense Retz. complex.

Comparative morphological study of plant parts and floral parts in different chromosomal races of Saccharum bengalense Retz. complex is presented. Although variation between the cytotypes is basically of quantitative nature yet some of the characters help in distinguishing diploid (2n=20), tetraploid (2n=40) and hexaploid (2n=60) cytotypes in the field. A key has been suggested for this purpose. Hairyness of lower glume of sessile spikelet helps in distinquishing diploid from diploid with one to five accessory chromosomes, it being smooth in the latter. Stomatal size did not prove a reliable character to distinguish the diploids and the polyploids. Pollen size increased with the increase in ploidy level. Seed germination percentage was low in the polyploids compared to in the diploid. An interesting observation was that the gigas effect of ploidy existed upto tetraploid level only and in the hexaploid the effect is reversed. Karyotype was studied in different cytotypes and it was of symmetrical type.

Alfalfa Embryogenic Cell Suspension Culture: Growth and Ploidy Level Stability

Establishment and growth characteristics of alfalfa ( Medicago sativa L.) highly embryogenic cell suspension culture are described. Flow cytometry was used to study the stability of nuclear DNA content. Somatic embryos were used as primary explant and contained only tetraploid cells, mostly in the GI phase. A slight shift to higher ploidy levels was observed during culture initiation, but polyploid cells were eliminated after a few weeks of culture and DNA histograms resembled the pattern found in primary explant. Considerable increase in ploidy levels accompanied by a loss of embryogenic capacity was observed if a long subculture interval was used. Ploidy level and embryogenic capacity remained stable with a short subculture interval in a long term culture.

Genetic and cytogenetic analyses of the A genome of Triticum monococcum. III. Cytology, breeding behavior, fertility, and morphology of autotriploids

Mature triploid seed from reciprocal (2n = 4x × 2n = 2x) crosses in Triticum monococcum was minute and shrivelled because of endosperm collapse and therefore failed to germinate. This necessitated the excision of embryos from successful pollinations and their growth in vitro to ensure subsequent germination so as to obtain viable and vigorous autotriploids. A comparison of these triploids with their diploid and tetraploid progenitors revealed that cell size, kernel weight, and pistil size increased with an increase in ploidy level. However, unlike other species, optimum expression was observed in these triploids for plant height, tillering, size of spikes, number of spikelets/spike, and leaf size. Earliness, althoughenhanced in tetraploids relative to diploids, was delayed in the triploids. Mean numbers of univalents, bivalents, and trivalents per microsporocyte were 2.65, 2.60, and 4.38, respectively. Only chains (93.5%), which formed V-shaped metaphase I (MI) configurations, frying pan (5.0%), and Y-shaped (1.5%) trivalent associations occurred. On the average, two reciprocal exchanges occurred per bivalent and trivalent. Trivalents corriented randomly at MI. At anaphase I, all sets of three homologues segreated randomly to the two poles, lagging univalents always divided equationally, and only meiocytes with such chromosomes formed micronuclei. The reasons for similarities and differences in meiotic behaviour of T. monococcum triploids with those of other species are discussed. Confirmation of the conclusions drawn with respect to the cytology of the triploids was obtained from similar cytological observations with primary single trisomics. These triploids produced euploids, primary single trisomics as well as some double and triple trisomics all of which differed phenotypically from diploids. Triticum monococcum, like most diploid species, is highly intolerant of aneuploidy. Possible reasons for the differences in levels of tolerance of aneuploidy in species like T. monococcum and those like Petunia hybrida, which are highly tolerant of aneuploidy, are discussed. Pollen fertility was high and seed fertility was very low. Reasons for the latter as well as the high fertility in species that are highly tolerant of aneuploidy and allotriploids are discussed.Key words: Triticum monococcum, autotriploid, trisomic, cytology, breeding behavior, fertility, morphology.

Zinc-resistant Cell Lines of Haplopappus gracilis

Ionic zinc (Zn2+) was completely toxic to wild type (Wt) cellsof Haplopappus gracilis (Nun.) Gray at a concentration of 7 mM in Gamborg B5 medium. On this medium stable, zinc-resistant calli developed from Wt suspension cells at a frequency of 10-7. Although most zinc-resistant cell lines were tetraploid, the resistance was not causally related with increased ploidy level. Zinc resistance was maintained after culturing the selected cell lines for two years in the absence of selection pressure. The zinc resistance was not expressed constitutively but was inducible. Zinc uptake was equal for the zinc-resistant and the Wt cell lines. An intracellularly located zinc resistance mechanismis most likely. However, a mechanism based on increased production of cellular carboxylate can be excluded. The utility of the zinc-resistant cell lines for somatic cell genetics is discussed.

Life History Studies on the Genus Trillium (Liliaceae) II. Reproductive Biology and Survivorship of Four Eastern North American Species

Abstract The stage class structures and the reproductive characteristics as well as energy allocation to reproductive activities were studied in four North American Trillium species (Liliaceae)—T. grandiflorum, T. erectum, T. undulatum and T. sessile. In addition, a comparison was also made of the life history characteristics of these four North American species with those of Japanese species.As a result, the stage class structures of these four species were discriminated for each species based on leaf area. The study revealed that they all possess similar depletion curves characterized by a conspicuous decrease of individuals belonging to the juvenile stages (from stage class 0 to 2). The switchover of stages, i.e, from one‐leaf to three‐leaved sterile stages, from three‐leaved sterile to fertile stages, or vice versa, was found to occur in different size ranges for each species. In all four species examined, reproductive allocation to total reproductive structures (RA) at the mature fruiting stage was almost independent of biomass, but the number of propagules produced per plant (Pn) was clearly dependent on the size of plants (individual biomass). Three species, T. grandiflorum, T. erertum and T. sessile, showed a proportional increase in Pn in response to the increase in RA, as found in four Japanese Trillium species, and also in a number of other perennial herbs growing in closed, stable and predictable environments with an outbreeding system. This trend, however, was not obvious in T. undulatum, i.e., Pn was independent of RA. With respect to seed characteristics, these species exhibited relatively high seed setting rates per ovule. Trillium undulatum showed very high fecundity, 92.40% and 85.38% in two years' samples, suggesting that this species is a typical inbreeder. Contrary to Japanese Trillium species, which showed proportional increase in seed weight in response to an increase in ploidy levels (from 2x to 6x), these diploid species showed wide variation in seed weight, ranging from 4.15 mg to 7.80 mg.

Somatic cell fusion as a source of genetic rearrangement leading to metastatic variants.

Tumor cell populations displaying metastatic properties often have higher gene dosage than their less malignant progenitor tumors, as shown by increased ploidy levels, chromosome duplication and gene amplification. The acquisition by tumor cells of high chromosome numbers may be due to endoreduplication or somatic hybridization either between tumor cells or between tumor and host cells. All such mechanisms increase genetic variability and instability in tumor cells since they trigger a polyploidization-segregation cycle. Among the wide variety of segregants which may emerge from high-ploidy cells, variants with increased malignancy can be positively selected in vivo. Evidence for in vivo fusion of tumor and normal host cells has been reported in different tumor systems. However the attainment by tumor-host hybrids of a higher degree of malignancy has only been observed following substantial chromosome segregation. The involvement of a cell of bone marrow origin as preferential host partner in the fusion process has been proved both by studies on tumor-host hybrids in bone marrow radiation chimeras and in vitro hybridization experiments between non-metastatic tumors and normal lymphoreticular cells which have led to the establishment of metastatic variants. Several different segregational mechanisms may bring about homozygosity or hemizygosity of recessive alleles in tumor-host hybrids, leading to their expression. The marked chromosome dynamics of tumor-host hybrids are also responsible for extensive chromosome rearrangements. At the molecular level these may represent mechanisms causing altered oncogene activity. The activation of new oncogenes by transposition or amplification as well as the amplification of previously activated oncogenes are the mechanisms most likely to be responsible for transition from low to high malignancy, occurring through ploidy changes, such as those produced by somatic mating.

Effect of ploidy variation on amino acid composition ofDatura innoxia (Solanaceae)

In haploid, diploid, triploid, tetraploid and hexaploid cytotypes of the scopalamine-yieldingDatura innoxia, the amino acid profiles were studied. The results suggest that the amino acid composition is influenced quantitatively by increasing ploidy levels, but in an irregular way.