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PubMed2003

Cytological and breeding behavior of pentaploids derived from 3x x 4x crosses in potato.

Cytology and breeding behavior of Solanum commersonii - S. tuberosum hybrids derived from 3 x x 4 x crosses was examined. The chromosome number of hybrids ranged from hypo-pentaploid (2 n=5 x - 8=52), to hyper-pentaploid (2 n=5 x + 7=67), with the euploid pentaploid 2 n=5 x=60 class predominant. The high variability in chromosome number of the 3 x x 4 x hybrids was attributed to the fact that meiotic restitution during megasporogenesis of the 3 x female may have involved poles with various chromosome numbers, resulting in 2 n eggs with 24-48 chromosomes. Microsporogenesis analyses provided evidence that chromosome pairing between S. commersonii and S. tuberosum genomes occurred. In addition, chromosome distribution at anaphase I and anaphase II revealed an average chromosome number of 29.5 and 29.1 per pole, respectively. To further study the extent of transmission of extra genome chromosomes from pentaploids, 5 x x 4 x and 4 x x 5 x crosses were performed, and the chromosome number of resulting progeny was determined. Ploidy ranged from 2 n=4 x=48 to 2 n=5 x=60 following 5 x x 4 x crosses, and from 2 n=4 x + 1=49 to 2 n=5 x=60 following 4 x x 5 x crosses. These results provided indirect evidence that the pentaploid hybrids produced viable aneuploid gametes with a chromosome number ranging from 24 to 36. They also demonstrated that gametes with large numbers of extra chromosomes can be functional, resulting in sporophytes between the 4 x and 5 x ploidy level. Fertility parameters of crosses involving various (aneuploid) pentaploid genotypes were not influenced by chromosome number, suggesting a buffering effect of polyploidy on aneuploidy. The possibility of successfully using (aneuploid) pentaploid genotypes for further breeding efforts is discussed.

Crosses, Genetic↗
PubMed2003

An EST survey of the sugarcane transcriptome.

Its large genome and high polyploidy makes sugarcane (Saccharum spp.) a singularly challenging crop to study and improve using genetic approaches. To provide large numbers of functionally characterized candidate genes that might be tested for direct association (rather than distant linkage) with economically important traits, we sequenced the 5' ends of 9,216 clones from three cDNA libraries (apex, leaf and mature internode), representing 3,401 non-redundant sequences. About 57% of these sequences could be assigned a tentative function based on statistically significant similarity to previously characterized proteins or DNA sequences. Another 28% corresponded to previously identified, but uncharacterized, sequences. Some of the remaining unidentified sequences were predicted to be genes which could potentially be new to plants or unique to sugarcane. Comparisons of the sugarcane ESTs to a large sorghum EST database revealed similar compositions of expressed genes between some different tissues. Comparison to a detailed Arabidopsis protein database showed some highly conserved sequences, which might be useful DNA markers for pan-angiosperm comparative mapping. These EST sequences provide a foundation for many new studies to accelerate isolation of agronomically important genes from the cumbersome sugarcane genome.

Base Sequence↗
PubMed2003

Cucumis monosomic alien addition lines: morphological, cytological, and genotypic analyses.

Cucumis hystrix Chakr. (HH, 2n=24), a wild relative of the cultivated cucumber, possesses several potentially valuable disease-resistance and abiotic stress-tolerance traits for cucumber ( C. sativus L., CC, 2n=14) improvement. Numerous attempts have been made to transfer desirable traits since the successful interspecific hybridization between C. hystrix and C. sativus, one of which resulted in the production of an allotriploid (HCC, 2n=26: one genome of C. hystrix and two of C. sativus). When this genotype was treated with colchicine to induce polyploidy, two monosomic alien addition lines (MAALs) (plant nos. 87 and 517: 14 CC+1 H, 2n=15) were recovered among 252 viable plants. Each of these plants was morphologically distinct from allotriploids and cultivated cucumbers. Cytogenetic and molecular marker analyses were performed to confirm the genetic constitution and further characterize these two MAALs. Chromosome counts made from at least 30 meristematic cells from each plant confirmed 15 nuclear chromosomes. In pollen mother cells of plant nos. 87 and 517, seven bivalents and one univalent were observed at diakinesis and metaphase I; the frequency of trivalent formation was low (about 4-5%). At anaphase I and II, stochastic and asymmetric division led to the formation of two gamete classes: n=7 and n=8; however, pollen fertility was relatively high. Pollen stainability in plant no. 87 was 86.7% and in plant no. 517 was 93.2%. Random amplified polymorphic DNA analysis was performed using 100 random 10-base primers. Genotypes obtained with eight primers (A-9, A-11, AH-13, AI-19, AJ-18, AJ-20, E-19, and N-20) showed a band common to the two MAAL plants and C. hystrix that was absent in C. sativus, confirming that the alien chromosomes present in the MAALs were derived from C. hystrix. Morphological differences and differences in banding patterns were also observed between plant nos. 87 and 517 after amplification with primers AI-5, AJ-13, N-12, and N-20, suggesting that these plants may contain different C. hystrix chromosomes.

Agriculture↗
PubMed2006

Allelic configuration and polysomic inheritance of highly variable microsatellites in tetraploid gynodioecious Thymus praecox agg.

Polyploidy plays a pivotal role in plant evolution. However, polyploids with polysomic inheritance have hitherto been severely underrepresented in plant population genetic studies, mainly due to a lack of appropriate molecular genetic markers. Here we report the establishment and experimental validation of six fully informative microsatellite markers in tetraploid gynodioecious Thymus praecox agg. Sequence data of 150 microsatellite alleles and their flanking regions revealed high variation, which may be characteristic for polyploids with a reticulate evolutionary history. Understanding the patterns of mutation (indels and substitutions) in microsatellite flanking-sequences was a prerequisite for the development of co-dominant markers for fragment analyses. Allelic segregation patterns among progeny arrays from ten test crosses revealed tetrasomic inheritance in T. praecox agg. No evidence of frequent double reduction was detected. Polymerase chain reaction (PCR) based dosage effects allowed for precise assignment of allelic configuration at all six microsatellite loci. The quantification of allele copy numbers in PCR was verified by comparisons of observed and expected gametic allele frequencies and heterozygosities in test crosses. Our study illustrates how PCR based markers can provide reliable estimates of heterozygosity and, thus, powerful tools for breeding system and population genetic analyses in polyploid organisms.

Alleles↗
PubMed2005

Hepatotoxic and nephrotoxic effects of cadmium in the frog Rana ridibunda.

Histological and histochemical alterations in the liver and kidneys of the frog Rana ridibunda, which was exposed to 200 ppm aqueous solutions of cadmium for 4, 10 and 30 days, respectively were investigated. In both the liver and kidneys, essential changes appeared after 10 days' exposure, the maximum changes being apparent after 30 days of exposure. In the liver, what was very characteristic was an increase in the area occupied by Kupffer cells, with the area in the animals exposed to cadmium for 30 days being the largest observed. What was also apparent was karyomegaly, polyploidy and infiltration. In addition with regard the kidneys, Hyaline Globules (HG) and apoptotic bodies occurred at a higher rate. At 30 days' exposure, most of the above changes were enhanced. In comparison with 10 days' exposure, fibrosis around the blood vessels and between hepatocytes, as well as Proliferating Cell Nuclear Antigen (PCNA) reactivity and apoptotic bodies increased lightly in the liver. In the kidneys, the most pronounced changes were the increase in numbers of apoptotic bodies, PCNA reactivity and hyaline globules. It was concluded that the variability in positive reactions for various proteins in the hyaline globules may be an indication that these (and possibly other) proteins are synthesized by tumor cells, but, most probably, that they may represent liver damage, progressive nephropathy, or progressive glomerulonephropathy.

Animals↗
PubMed2006

Occurrence of tetraploidy in Nicotiana attenuata plants after Agrobacterium-mediated transformation is genotype specific but independent of polysomaty of explant tissue.

Genotypes of Nicotiana attenuata collected from Utah and Arizona were transformed with 17 different vectors (14 unpublished vectors based on 3 new backbone vectors) using an Agrobacterium-mediated procedure to functionally analyze genes important for plant-insect interactions. None of the 51 T1-T3 transgenic Utah lines analyzed by the flow cytometry were tetraploid, as opposed to 18 of 33 transgenic Arizona lines (55%). Analysis of T0 regenerants transformed with the same vector carrying an inverted repeat (IR) N. attenuata pro-systemin construct confirmed the genotype dependency of tetraploidization: none of the 23 transgenic Utah lines were tetraploid but 31 (72%) of 43 transgenic Arizonas were tetraploid. We tested the hypothesis that the differences in polysomaty of the explant tissues accounted for genotype dependency of tetraploid formation by measuring polysomaty levels in different seedling tissues. Hypocotyls, cotyledons, and roots of Utah and Arizona genotypes contained similar percentages of 4C nuclei (61 and 60; 7 and 5; and 58 and 61%, respectively). Since we used hypocotyls as explant sources and the nonoccurrence of tetraploid Utah transformants does not correspond to the high percentage of 4C nuclei in Utah hypocotyls, we can rule out a direct relationship between tetraploid formation and polysomaty level. We hypothesize that the difference between the Utah and Arizona genotypes results from the failure of polyploid Utah callus to regenerate into fully competent plants. We propose that future work on post-transformation polyploidy concentrate on the processes that occur during callus formation and plant regeneration from callus.

Cotyledon↗
PubMed2004

Chromosomes associate premeiotically and in xylem vessel cells via their telomeres and centromeres in diploid rice ( Oryza sativa).

Studies of the meiosis of diploid plants such as Arabidopsis, maize and diploid progenitors of wheat have revealed no premeiotic association of chromosomes. Premeiotic and somatic association of chromosomes has only been previously observed in the anther tissues and xylem vessel cells of developing roots in polyploid plants such as hexaploid and tetraploid wheat, polyploid relatives of wheat and artificial polyploids made from the progenitor diploids of wheat. This suggested that this association was confined specifically to polyploids or was induced by polyploidy. However, we developed procedures for in situ hybridization on structurally well-preserved tissue sections of rice, and analysed two diploid rice species ( Oryza sativa and O. punctata). Contrary to expectation, this has revealed that centromeres and telomeres also associate both in the xylem vessel cells of developing root and in undifferentiated anther cells in these diploids. However, in contrast to wheat and related polyploids, where the initial association in undifferentiated anthers is between either non-homologous or related chromosomes, and not homologous chromosomes, the initial association of rice chromosomes seems to be between homologues. Thus, in contrast to the diploid dicot model Arabidopsis, meiotic studies on the diploid model cereal, rice, will now need to take into account the effects of premeiotic chromosome association.

Centromere↗
PubMed1999

Expression of CKS1At in Arabidopsis thaliana indicates a role for the protein in both the mitotic and the endoreduplication cycle.

Although endoreduplication is common in plants, little is known about the mechanisms regulating this process. Here, we report the patterns of endoreduplication at the cellular level in the shoot apex of Arabidopsis thaliana L. Heynh. plants grown under short-day conditions. We show that polyploidy is developmentally established in the pith, maturing leaves, and stipules. To investigate the role of the cell cycle genes CDC2aAt, CDC2bAt, CYCB1;1, and CKS1At in the process of endoreduplication, in-situ hybridizations were performed on the vegetative shoot apices. Expression of CDC2aAt, CDC2bAt, and CYCB1;1 was restricted to mitotically dividing cells. In contrast, CKS1At expression was present in both mitotic and endoreduplicating tissues. Our data indicate that CDC2aAt, CDC2bAt, and CYCB1;1 only operate during mitotic divisions, whereas CKS1At may play a role in both the mitotic and endoreduplication cycle.

Arabidopsis↗
PubMed2003

Discrimination of homoeologous gene expression in hexaploid wheat by SNP analysis of contigs grouped from a large number of expressed sequence tags.

Single-nucleotide polymorphisms (SNPs) are useful markers for gene diagnosis and mapping of genes on chromosomes. However, polyploidy, which is characteristic of the evolution of higher plants, complicates the analysis of SNPs in the duplicated genes. We have developed a new method for SNP analysis in hexaploid wheat. First, we classified a large number of expressed sequence tags (ESTs) from wheat in silico. Those grouped into contigs were anticipated to correspond to transcripts from homoeologous loci. We then selected relatively abundant ESTs, and assigned these contigs to each of the homoeologous chromosomes using a nullisomic/tetrasomic series of Chinese Spring wheat strains in combination with pyrosequencing. The ninety genes assigned were almost evenly distributed into seven homologous chromosomes. We then created a virtual display of the relative expression of these genes. Expression patterns of genes from the three genomes in hexaploid wheat were classified into two major groups: (1) genes almost equally expressed from all three genomes; and (2) genes expressed with a significant preference, which changed from tissue to tissue, from certain genomes. In 11 cases, one of the three genes in the allopolyploid was found to be silenced. No preference for gene-silencing in particular genomes or chromosomes was observed, suggesting that gene-silencing occurred after polyploidization, and at the gene level, not at the chromosome or genome level. Thus, the use of this SNP method to distinguish the expression profiles of three homoeologous genes may help to elucidate the molecular basis of heterosis in polyploid plants.

Base Sequence↗
PubMed1997

Detection of mosaic and non-mosaic chromosome abnormalities in 6- to 8-day old human blastocysts.

A reliable technique has been developed for the production of good quality G-banded chromosome preparations from 6- to 8-day-old human blastocysts (20-800 cell stage) from an in vitro fertilization programme. The technique involves a thymidine cell division synchronization step to reduce the exposure time to colcemid, in conjunction with a simple 70% acetic acid disaggregation procedure to produce discrete metaphases for analysis. Of 105 blastocysts processed by this technique, 9 were lost during handling and 10 showed no dividing cells. The remaining 86 produced useful separate metaphases with a mean mitotic activity of 6.5%. A full G-banded karyotype was obtained from 1-6 cells in 55 blastocysts (64%), incomplete G-banded analysis but with full information of ploidy was obtained from 18 blastocysts (21%), with 13 (15%) producing no useful cytogenetic results. Abnormalities observed included polyploidy, diploid/polyploid mosaicism, non-mosaic trisomy 16 (2 cases), 46,Xdel(X)-(q21)/46,XX (1 case) and several single cells with trisomies or structural anomalies in otherwise normal blastocysts. Variable levels of structural chromosome damage, with apparent interchanges, chromosome branching and anomalous chromatid pairing were also seen.

Aneuploidy↗
PubMed2006

Colchicine-induced polyploidization depends on tubulin polymerization in c-metaphase cells.

The microtubule cytoskeleton plays a crucial role in the cell cycle and in mitosis. Colchicine is a microtubule-depolymerizing agent that has long been used to induce chromosome individualization in cells arrested at metaphase and also in the induction of polyploid plants. Although attempts have been made to explain the processes and mechanisms underlying polyploidy induction, the role of the cytoskeleton still remains largely unknown. Through immunodetection of alpha-tubulin, different concentrations (0.5 or 5 mM) of colchicine were found to produce opposite effects in the organization of the cytoskeleton in rye (Secale cereale L.). A low concentration (0.5 mM) induced depolymerization of the microtubular cytoskeleton in all phases of the cell cycle. In contrast, a high concentration (5 mM) was found to induce the polymerization of new tubulin-containing structures in c-metaphase cells. Furthermore, both treatments also showed contrasting effects in the induction of polyploid cells. Flow cytometric analysis and quantitative assessments of nucleolus-organizing regions revealed that only the high-concentration colchicine treatment was effective in the formation of polyploid cells. Our studies indicate that spindle disruption alone is insufficient for the induction of polyploid cells. The absence of any tubulin structures in plants treated with colchicine at the low concentration induced cell anomalies, such as the occurrence of nuclei with irregular shape and/or (additional) micronuclei, 12 h after recovery, pointing to a direct effect on cell viability. In contrast, the almost insignificant level of cell anomalies in the high-concentration treatment suggests that the presence of new tubulin-containing structures allows the reconstitution of 4C nuclei and their progression into the cell cycle.

Cell Nucleus↗
PubMed2005

Ploidy, geographical distribution and morphological differentiation of Parasenecio auriculata (Senecioneae; Asteraceae) in Japan.

Parasenecio auriculata is a woodland perennial herb widely distributed in Northeastern Asia, constituted by a poorly understood polyploidy with a diploid (2n = 2x = 60) and a tetraploid (2n = 4x = 120). In this study, for a better understanding of the polyploidal evolution, cytogeography and morphological variation were analyzed in Japanese P. auriculata, including two varieties; var. bulbifera endemic to central Hokkaido and var. kamtschatica widely distributed in northern Honshu and Hokkaido. The occurrence of two polyploidal levels was reconfirmed. While var. bulbifera is predominantly tetraploid, var. kamtschatica is comprised of diploid and tetraploid. Morphological variation among 22 quantitative characteristics is continuous and not distinctive among cytotypes or varieties, but plant size tended to be larger in the order, diploid of var. kamtschatica, var. bulbifera, and tetraploid of var. kamtschatica. The cytotype distribution showed a conspicuous geographical pattern. Besides var. bulbifera endemic to the central Hokkaido, the diploid of var. kamtschatica is mainly found in Southern Hokkaido, and the tetraploid has a disjunct distribution in eastern and northern Hokkaido and northern Honshu. Such a geographical pattern is possibly attributable to the differentiation of climatic preference among cytotypes and varieties, and may have been established in association with the climatic cline along the Japanese archipelago.

Analysis of Variance↗
PubMed2006

Germination characteristics of native Japanese dandelion autopolyploids and their putative diploid parent species.

Germination characteristics of native Japanese Taraxacum lineages of Taraxacum platycarpum (diploid), T. venustum (triploid and tetraploid), and T. albidum (pentaploid) have been studied at different temperatures. Taraxacum platycarpum ssp. hondoense is the putative diploid parent of T. venustum. Diploid T. platycarpum ssp. hondoense and the polyploids T. venustum and T. albidum are found in different areas of Japan, and distribution differences may reflect divergent ecological and physiological traits among ploidy levels. In this study, to prevent mixing of seeds of different polyploidy we used flow cytometry to examine the ploidy level of the plants from which seeds were collected. Results from seed-germination experiments showed that dependence on temperature of final percentage germination was qualitatively similar for both autopolyploids T. venustum and diploids T. platycarpum--germination was suppressed at high and low temperatures. It was also shown that seed germination of autopolyploids was suppressed more than that of the ancestral diploid at low temperatures and that seed germination for polyploids was higher than for the diploid. Threshold variations at low temperatures might affect the distribution of native dandelions. Taraxacum venustum, which occurs in cool climates, might have developed a distinctly lower germination threshold at low temperatures whereas T. albidum, which is native to warm climates, might have developed an adaptive threshold at high temperatures.

Adaptation, Physiological↗
PubMed2026

Optimization of functional genetics tools for a model tetraploid Capsella bursa-pastoris, with focus on homoeolog-aware gene editing.

Capsella bursa-pastoris is a recent allotetraploid and a promising model for studying early consequences of polyploidy. One of the intriguing questions in polyploid research is how new functions arise from initially identical or nearly identical homoeologous genes. Functional genetics tools, including genetic editing, can help to understand this process, but they have not been developed for C. bursa-pastoris yet. We present here the results of our study aimed at filling this gap. In particular, we compared the efficiency of floral dip transformation in six accessions of C. bursa-pastoris representing distant populations. The Asian clade accession PGL0025 had the highest efficiency of transformation (~ 1.1%). Comparison of Agrobacterium tumefaciens strains EHA105 and GV3101 (pMP90) showed that the latter is more effective. Also, we created a genome-wide gRNA database for all pairs of homoeologs of the PGL0001 accession of C. bursa-pastoris and integrated it into publicly available genome browser: https://t2e.online/igv_capsella_bursa-pastoris/ . We assessed the possibility of differential editing for two pairs of homoeologous genes with high sequence similarity (> 90%) both in vitro and in silico. Despite the test results that indicated off-target activity, we have succeeded in obtaining lines of plants with homozygous frameshift mutations in each of the homoeologs separately in vivo. We expect that these findings and resources will promote the use of C. bursa-pastoris as a model in functional genetics experiments, in particular, the studies of the fate of duplicated gene after polyploidization event.

Capsella↗
PubMed1992

Cytogenetic analysis of spontaneous abortions with direct analysis of chorionic villi.

A total of 224 cases of spontaneous abortions were studied by analysing chromosomes directly from chorionic villi. Abnormal karyotypes were found in 51.3% of the investigated cases. Among the chromosome abnormalities the most frequent were the trisomies (55.7%), followed by polyploidy (23.5%), monosomy X (15.6%) and structural anomalies (5.2%). The rapid and relatively simple method is suitable for studying the causes of spontaneous abortions, and this information may be helpful also for genetic counselling.

Abortion, Spontaneous↗
PubMedDate not supplied

The induction of numerical chromosome aberrations in human lymphocyte cultures and V79 Chinese hamster cells by diethyl sulfate.

Human lymphocyte cultures were treated with increasing concentrations of diethyl sulfate (DES) at different times after stimulation with phytohemagglutinin (0, 24 and 45 h) and were scored for numerical chromosomal aberrations at different culture times (52, 72 and 96 h). A total of 3500 metaphases were analyzed. A statistically significant (p less than 0.01) increase in hypodiploid and hyperdiploid metaphases was found throughout the tested dose range (0.1-3 mM DES); the increase in polyploid metaphases was statistically significant at 1 mM (p less than 0.05) and 3 mM (p less than 0.01) DES. In human lymphocytes treated in Go, DES also induced chromatid breaks as well as micronuclei. In V79 Chinese hamster cells, DES induced micronuclei and polyploidy.

Aneuploidy↗
PubMed1991

Clastogenic effect of pesticides in peripheral lymphocytes of cotton-field workers.

We studied clastogenic effects in peripheral lymphocytes of cotton-field workers who were exposed to different pesticides. All the cells were grown in RPMI 1640 medium for 48 and 72 h. The type of aberrations observed in the exposed group are gaps, breaks, dicentrics, exchanges, rings and polyploidy. The frequency of total chromosomal aberrations increased significantly in male pesticide applicators when compared to controls. A significant decrease in mitotic index was observed in the exposed group as compared to the control group. The 48-h cultures showed high incidence of chromosomal aberrations and low mitotic index when compared to 72-h cultures. The difference in chromosomal aberrations between 48- and 72-h cultures was not significant. 24 out of 26 individuals showed ill health effects such as severe giddiness and nervous disorders.

Adolescent↗