Chromosome Number, Ploidy Level, and Nuclear DNA Content in 23 Species of Echeveria (Crassulaceae)

Echeveria is a polyploid genus with a wide diversity of species and morphologies. The number of species registered for Echeveria is approximately 170; many of them are native to Mexico. This genus is of special interest in cytogenetic research because it has a variety of chromosome numbers and ploidy levels. Additionally, there are no studies concerning nuclear DNA content and the extent of endopolyploidy. This work aims to investigate the cytogenetic characteristics of 23 species of Echeveria collected in 9 states of Mexico, analyzing 2n chromosome numbers, ploidy level, nuclear DNA content, and endopolyploidy levels. Chromosome numbers were obtained from root tips. DNA content was obtained from the leaf parenchyma, which was processed according to the two-step protocol with Otto solutions and propidium iodide as fluorochrome, and then analyzed by flow cytometry. From the 23 species of Echeveria analyzed, 16 species lacked previous reports of 2n chromosome numbers. The 2n chromosome numbers found and analyzed in this research for Echeveria species ranged from 24 to 270. The range of 2C nuclear DNA amounts ranged from 1.26 pg in E. catorce to 7.70 pg in E. roseiflora, while the 1C values were 616 Mbp and 753 Mbp, respectively, for the same species. However, differences in the level of endopolyploidy nuclei were found, corresponding to 4 endocycles (8C, 16C, 32C and 64C) in E. olivacea, E. catorce, E. juarezensis and E. perezcalixii. In contrast, E. longiflora presented 3 endocycles (8C, 16C and 32C) and E. roseiflora presented 2 endocycles (8C and 16C). It has been suggested that polyploidization and diploidization processes, together with the presence of endopolyploidy, allowed Echeveria species to adapt and colonize new adverse environments.

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Nuclear DNA amount in pure species and hybrid willows (Salix): a flow cytometric investigation

Canadian Journal of Botany ◽

10.1139/b97-153 ◽

1998 ◽

Vol 76 (1) ◽

pp. 157-165 ◽

Cited By ~ 6

Author(s):

Jérôme Thibault

Keyword(s):

Flow Cytometry ◽

Ploidy Level ◽

Dna Content ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

Natural Hybridization ◽

Ploidy Levels ◽

Key Words Flow Cytometry ◽

Dna Values ◽

Dna Amounts

Flow cytometry (FCM) has been used to estimate the nuclear DNA content of 11 Salix species and 5 hybrids. One hundred and sixty nine individuals were studied including 159 individuals from a sequence of 32 communities along a stretch of river in France and 10 individuals from French and English collections for comparison. Isolated nuclei were stained with propidium iodide. FCM was a significantly more practical and rapid technique than that of establishing the karyotype to survey many samples of Salix for variation in ploidy. The 2C DNA amounts for diploid species ranged from 0.76 to 0.98 pg, and tetraploid values ranged from 1.62 to 1.80 pg. The DNA values were consistent with the known ploidy levels. With the exception of a doubtful Salix xquercifolia, ploidy levels and DNA amounts of hybrids were intermediate compared with those of their parents. Intraspecific variation of nuclear DNA values including instrumental variation was low (i.e., 6-11% at the same ploidy level). FCM appeared to be an accurate tool for determination of Salix triploid hybrids. However, it remains limited concerning hybrids from crosses between species of the same ploidy level. Results suggest that natural hybridization might not be frequent in the communities studied, although they have been subject to disturbance. Previous overestimates of hybridization frequency in willows were probably due to misinterpretation of the effects of the environment on Salix spp. morphology; however, the extent and mechanisms of introgression in the genus remain to be further investigated. Key words: flow cytometry, Salix, hybridization, nuclear DNA content, riparian vegetation, disturbance.

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Determining Ploidy Level and Nuclear DNA Content in Rubus by Flow Cytometry

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.127.5.767 ◽

2002 ◽

Vol 127 (5) ◽

pp. 767-775 ◽

Cited By ~ 37

Author(s):

Rengong Meng ◽

Chad Finn

Keyword(s):

Flow Cytometry ◽

Pacific Northwest ◽

Ploidy Level ◽

Dna Content ◽

Chromosome Numbers ◽

Nuclear Dna ◽

Diploid Species ◽

Nuclear Dna Content ◽

Dna Flow Cytometry ◽

Ploidy Levels

Nuclear DNA flow cytometry was used to differentiate ploidy level and determine nuclear DNA content in Rubus. Nuclei suspensions were prepared from leaf discs of young leaves following published protocols with modifications. DNA was stained with propidium iodide. Measurement of fluorescence of 40 genotypes, whose published ploidy ranged from diploid to dodecaploid, indicated that fluorescence increased with an increase in chromosome number. Ploidy level accounted for 99% of the variation in fluorescence intensity (r2 = 0.99) and variation among ploidy levels was much higher than within ploidy levels. This protocol was used successfully for genotypes representing eight different Rubus subgenera. Rubus ursinus Cham. and Schldl., a native blackberry species in the Pacific Northwest, which has been reported to have 6x, 8x, 9x, 10x, 11x, and 12x forms, was extensively tested. Genotypes of R. ursinus were predominantly 12x, but 6x, 7x, 8x, 9x, 11x, and 13x forms were found as well. Attempts to confirm the 13x estimates with manual counts were unsuccessful. Ploidy level of 103 genotypes in the USDA-ARS breeding program was determined by flow cytometry. Flow cytometry confirmed that genotypes from crosses among 7x and 4x parents had chromosome numbers that must be the result of nonreduced gametes. This technique was effective in differentiating chromosome numbers differing by 1x, but was not able to differentiate aneuploids. Nuclear DNA contents of 21 diploid Rubus species from five subgenera were determined by flow cytometry. Idaeobatus, Chamaebatus, and Anaplobatus were significantly lower in DNA content than those of Rubus and Cylactis. In the Rubus subgenus, R. hispidus and R. canadensis had the lowest DNA content and R. sanctus had the highest DNA content, 0.59 and 0.75 pg, respectively. Idaeobatus had greater variation in DNA content among diploid species than the Rubus subgenus, with the highest being from R. ellipticus (0.69 pg) and lowest from R. illecebrosus (0.47 pg).

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Nuclear DNA Contents of Phalaenopsis sp. and Doritis pulcherrima

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.126.2.195 ◽

2001 ◽

Vol 126 (2) ◽

pp. 195-199 ◽

Cited By ~ 34

Author(s):

Sandy Lin ◽

Hsiao-Ching Lee ◽

Wen-Huei Chen ◽

Chi-Chang Chen ◽

Yen-Yu Kao ◽

...

Keyword(s):

Flow Cytometry ◽

Genome Analysis ◽

Dna Content ◽

Chromosome Numbers ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

Dna Contents ◽

Nuclear Dna Contents ◽

Different Levels ◽

Dna Amounts

Nuclear DNA contents were estimated by flow cytometry in 18 Phalaenopsis Blume species and Doritis pulcherrima Lindl. DNA amounts differed 6.07-fold, from 2.74 pg/diploid nuclear DNA content (2C) in P. sanderiana Rchb.f. to 16.61 pg/2C in P. parishii Rchb.f. Nuclear DNA contents of P. aphrodite Rchb.f. clones, W01-38 (2n = 2x = 38), W01-41 (2n = 3x = 57), and W01-22 (2n = 4x = 76), displayed a linear relationship with their chromosome numbers, indicating the accuracy of flow cytometry. Our results also suggest that the 2C-values of the Phalaenopsis sp. correlate with their chromosome sizes. The comparative analyses of DNA contents may provide information to molecular geneticists and systematists for genome analysis in Phalaenopsis. Endoreduplication was found in various tissues of P. equestris at different levels. The highest degree of endoreduplication in P. equestris was detected in leaves.

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Karyotype analysis, DNA content and molecular screening in Lippia alba (Verbenaceae)

Anais da Academia Brasileira de Ciências ◽

10.1590/s0001-37652011005000012 ◽

2011 ◽

Vol 83 (3) ◽

pp. 993-1006 ◽

Cited By ~ 18

Author(s):

Patrícia M.O. Pierre ◽

Saulo M. Sousa ◽

Lisete C. Davide ◽

Marco A. Machado ◽

Lyderson F. Viccini

Keyword(s):

Genetic Variation ◽

Dna Content ◽

Chromosome Numbers ◽

Rapd Markers ◽

Nuclear Dna ◽

Pollen Viability ◽

Flow Cytometric Analysis ◽

Nuclear Dna Content ◽

Ploidy Levels ◽

Lippia Alba

Cytogenetic analyses, of pollen viability, nuclear DNA content and RAPD markers were employed to study three chemotypes of Lippia alba (Mill.) (Verbenaceae) in order to understand the genetic variation among them. Different ploidy levels and mixoploid individuals were observed. This work comprises the first report of different chromosome numbers (cytotypes) in L. alba. The chromosome numbers of La2-carvone and La3-linalool chemotypes suggested that they are polyploids. Flow cytometric analysis showed an increase of nuclear DNA content that was not directly proportional to ploidy level variation. A cluster analysis based on RAPD markers revealed that La3-linalool shares genetic markers with La1-citral and La2-carvone. The analysis showed that the majority of genetic variation of La3-linalool could be a consequence of ixoploidy. ur data indicates that sexual reproduction aong those three chemotypes is unlikely and suggests the beginning of reproductive isolation. The results demonstrated that chromosome analysis, nuclear DNA content estimation and RAPD markers constitute excellent tools for detecting genetic variation among L. alba chemotypes.

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NUCLEAR DNA CONTENT IN BLUEBERRY DETERMINED BY FLOW CYTOMETRY

HortScience ◽

10.21273/hortsci.27.6.580e ◽

1992 ◽

Vol 27 (6) ◽

pp. 580e-580

Author(s):

Rodomiro Ortiz ◽

D.E. Costich ◽

T.P. Meagher ◽

N. Vorsa

Keyword(s):

Flow Cytometry ◽

Dna Content ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

Haploid Genome ◽

Dna Flow Cytometry ◽

Dna Amount ◽

Ploidy Levels ◽

Polyploid Evolution ◽

Haploid Genome Size

DNA flow cytometry was used to determine nuclear DNA content in diploid blueberry species, and 3x, 4x, 5x, and 6x ploidy levels. Relative fluorescence intensity of stained nuclei measured by flow cytometry was a function of the number of chromosome sets (X): Y = 3.7X – 2.3 (r2 = 95.1%). DNA flow cytometry should be useful for ploidy level determination in the seedling stage. A significant linear relationship was established between nuclear DNA content and number of chromosomes (x); DNA (pg) = 0.52 x1 (r2 = 99.8%). Based on this equation the haploid genome DNA amount (1C) was calculated as 0.62 ± 0.08 pg, with an approximate haploid genome size of 602 Mbp/1C. The results indicate that conventional polyploid evolution occured in the section Cyanococcus, genus Vaccinium: the increase in DNA was concurrent with increase in chromosome number. DNA content differences among 2x species were correlated with Nei's genetic distance estimates based on 20 isozyme markers. Most of the variation was among species (49%), with 26% between populations within species, and 25% within populations.

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Flow cytometry-based determination of ploidy from dried leaf specimens in genomically complex collections of the tropical forage grass Urochloa s. l.

10.1101/2021.03.26.437252 ◽

2021 ◽

Author(s):

Paulina Tomaszewska ◽

Till K. Pellny ◽

Luis Miguel Hernandez ◽

Rowan A. C. Mitchell ◽

Valheria Castiblanco ◽

...

Keyword(s):

Flow Cytometry ◽

Dna Content ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

Germplasm Collection ◽

Forage Grass ◽

Ploidy Levels ◽

Sustainable Improvement ◽

Tropical Forage ◽

Relative Differences

We aimed to develop an optimized approach to determine ploidy for dried leaf material in a germplasm collection of a tropical forage grass group, including approaches to collect, dry and preserve plant samples for flow cytometry analysis. Urochloa (including Brachiaria, Megathyrus and some Panicum) tropical grasses are native to Africa and are now, after selection and breeding, planted worldwide, particularly in South America, as important forages with huge potential for further sustainable improvement and conservation of grasslands. The methods enable robust identification of ploidy levels (coefficient of variation, CV, typically <5%). Ploidy of some 353 forage grass accessions (ploidy range from 2 to 9), from international genetic resource collections, showing variation in basic chromosome numbers and reproduction modes (apomixis and sexual), were determined using our defined standard protocol. Two major Urochloa agamic complexes used in the current breeding programs at CIAT and EMBRAPA: the ' brizantha' and 'humidicola' agamic complexes are variable, with multiple ploidy levels and DNA content. U. brizantha has odd level of ploidy (x=5), and the relative differences in nuclear DNA content between adjacent cytotypes is reduced, thus more precise examination of this species is required. Ploidy measurement of U. humidicola revealed some aneuploidy.

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MICROPROPAGATION AND PLOIDY STABILITY OF Lippia lacunosa Mart. & Schauer: AN ENDANGERED BRAZILIAN MEDICINAL PLANT

JOURNAL OF NEOTROPICAL AGRICULTURE ◽

10.32404/rean.v6i1.3203 ◽

2019 ◽

Vol 6 (1) ◽

pp. 1-7

Author(s):

Diego Pandeló José ◽

José Marcello Salabert De Campos ◽

Lyderson Facio Viccini ◽

Emilly Ruas Alkimim ◽

Marcelo De Oliveira Santos

Keyword(s):

Flow Cytometry ◽

Medicinal Plant ◽

Dna Content ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

Naphthalene Acetic Acid ◽

Flow Cytometry Analysis ◽

Ploidy Levels

Lippia lacunosa is a Brazilian savanna plant that belongs to the Verbenaceae family. It has been used in folk medicine as a treatment for different diseases. This species represents an endangered Brazilian medicinal plant, and this is the first report documenting a reliable protocol for the in vitro propagation and regeneration of L. lacunosa. Axenic explants were cultivated in MS medium containing different concentrations of naphthalene acetic acid (NAA) to induce root growth. The mean shoot length and the number of roots were highest with 0.06 mg·L-1 NAA. The highest number of buds in shoot regeneration was induced with 2 mg·L-1 6-benzylaminopurine (BA). To obtain a long-term culture, the dwarf shoots were elongated on MS media containing 0.5 mg·L-1 BA alternated with MS containing 2 mg·L-1 BA every 40 days. In the present protocol, the long-term shoots retained the ability to root even after long periods of BA treatment. In addition, we evaluated the nuclear DNA content and ploidy levels, including the occurrence of endopolyploidy, in long-term micropropagated plant leaves using flow cytometry analysis. The plants propagated in vitro over several years possessed nuclear DNA contents ranging from 2.940 to 3.095 pg, and no differences in DNA content were found among in vitro plants or between these plants and the control (L. lacunosa from a greenhouse with a DNA content of 3.08 pg). The flow cytometry analysis also demonstrated that there was no polyploidization. The present study will be useful for biotechnological approaches and provides the first estimate of the nuclear DNA content of this species using flow cytometry.

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Karyotype characterization and nuclear DNA content measurement in Bromeliaceae: State of the art and future perspectives

Anais da Academia Brasileira de Ciências ◽

10.1590/0001-3765201420140224 ◽

2014 ◽

Vol 86 (4) ◽

pp. 1849-1862 ◽

Cited By ~ 1

Author(s):

ANDREI C.P. NUNES ◽

WELLINGTON R. CLARINDO

Keyword(s):

Flow Cytometry ◽

Chromosome Number ◽

Dna Content ◽

Nuclear Dna ◽

State Of The Art ◽

Nuclear Dna Content ◽

Basic Chromosome Number ◽

Basic Chromosome ◽

Content Measurement ◽

2C Dna Content

In Bromeliaceae, cytogenetic and flow cytometry analyses have been performed to clarify systematic and evolutionary aspects. Karyotyping approaches have shown the relatively high chromosome number, similar morphology and small size of the chromosomes. These facts have prevented a correct chromosome counting and characterization. Authors have established a basic chromosome number of x = 25 for Bromeliaceae. Recently, one karyomorphological analysis revealed that x = 25 is no longer the basic chromosome number, whose genome may have a polyploid origin. Besides cytogenetic characterization, the 2C DNA content of bromeliads has been measured. Nuclear DNA content has varied from 2C = 0.60 to 2C = 3.34 picograms. Thus, in relation to most angiosperms, the 2C DNA content of Bromeliaceae species as well as their chromosome size can be considered relatively small. In spite of some advances, cytogenetic and flow cytometry data are extremely scarce in this group. In this context, this review reports the state of the art in karyotype characterization and nuclear DNA content measurement in Bromeliaceae, emphasizing the main problems and suggesting prospective solutions and ideas for future research.

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Genome Size and Karyotype Studies in Five Species of Lantana (Verbenaceae)

HortScience ◽

10.21273/hortsci15603-20 ◽

2021 ◽

pp. 1-5

Author(s):

S. Brooks Parrish ◽

Renjuan Qian ◽

Zhanao Deng

Keyword(s):

United States ◽

Genome Size ◽

Dna Content ◽

Chromosome Numbers ◽

Nuclear Dna ◽

Nuclear Dna Content ◽

The United States ◽

Ploidy Levels ◽

Chromosome Group ◽

Group 3

Lantana species are an important component of the U.S. environmental horticulture industry. The most commonly produced and used species are L. camara and, on a smaller scale, L. montevidensis. Both were introduced to the United States from Central and/or South America. Lantana species native to the continental United States include L. canescens, L. depressa, L. involucrata, etc. and most of them have not been well exploited. This study was conducted to obtain information about somatic chromosome numbers, karyotypes, and genome size of these five species. Nuclear DNA content in these species ranged from 2.74 pg/2C (L. involucrata) to 6.29 pg/2C (L. depressa var. depressa). Four chromosome numbers were observed: 2n = 2x = 22 in L. camara ‘Lola’ and ‘Denholm White’, 2n = 4x = 44 in L. depressa var. depressa, 2n = 2x = 24 in L. canescens and L. involucrata, and 2n = 3x = 36 in L. montevidensis. Two basic chromosome numbers were observed: x = 11 in L. camara and L. depressa var. depressa, and x = 12 in L. canescens, L. involucrata, and L. montevidensis. Analysis of somatic metaphases resulted in formulas of 20m + 2sm for L. camara ‘Lola’ and ‘Denholm White’, 12m + 12sm for L. canescens, 44m for L. depressa var. depressa, 10m + 14sm for L. involucrata, and 32m + 4sm for L. montevidensis. Satellites were identified in all five species, but were associated with a different chromosome group in different species. L. depressa var. depressa had the longest total chromatin length (146.78 µm) with a range of 1.88 to 4.41 µm for individual chromosomes. The maximum arm ratio was observed in L. canescens, with a ratio of 2.5 in chromosome group 3. L. depressa var. depressa was the only species that had all of its centromeres located in the median region of the chromosome. The results show significant differences in nuclear DNA content, chromosome number, and karyotype among three native and two introduced lantana species and will help to identify, preserve, protect, and use native lantana species. The information will be helpful in assessing the ploidy levels in the genus by flow cytometry.

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Nuclear DNA Content and Ploidy Level of Apple Cultivars Including Polish ones in Relation to Some Morphological Traits

Acta Biologica Cracoviensia s Botanica ◽

10.1515/abcsb-2016-0008 ◽

2016 ◽

Vol 58 (1) ◽

pp. 81-93 ◽

Cited By ~ 7

Author(s):

Małgorzata Podwyszyńska ◽

Dorota Kruczyńska ◽

Aleksandra Machlańska ◽

Barbara Dyki ◽

Iwona Sowik

Keyword(s):

Chromosome Number ◽

Flowering Time ◽

Ploidy Level ◽

Dna Content ◽

Nuclear Dna ◽

Pollen Viability ◽

Nuclear Dna Content ◽

Pollen Grain ◽

Clear Correlation ◽

Dna Amount

Abstract Apple species and cultivars differ in nuclear (2C) DNA content and ploidy level. The majority of these genotypes are diploids, but there are some triploids and a few tetraploids. Nuclear DNA content is a specific feature and its flow cytometric evaluation can be helpful in differentiating taxa. For many apple genotypes – including all the Polish ones, these characteristics are not known. 2C DNA was evaluated in relation to leaf, flower, fruit, pollen grain and stomata sizes as well as to the flowering time for seventy genotypes (including 46 Polish cultivars) gathered in the gene bank of the Research Institute of Horticulture, Skierniewice, Poland. For standard cultivars with the known chromosome number, 2C value was 1.71 pg for diploid cultivar ‘Alwa’ (2n=2x=34), 2.55 pg for triploid ‘Boskoop’ (3x=51), and 3.37 pg for tetraploid genome (4x=68) of mixoploid ‘McIntosh 2x+4x’. In 61 cultivars (including 41 Polish ones), the nuclear DNA content ranged from 1.58 to 1.78 pg indicating their diploid chromosome number. Five cultivars were identified as triploids (‘Bursztówka Polska’, ‘Pagacz’, ‘Rapa Zielona’, ‘Rarytas Śląski’ and ‘Witos’) owing to their nuclear DNA amount ranging between 2.42 and 2.58 pg. Leaf, flower, fruit, stomata and pollen grain sizes were on average significantly larger in triploids. Thus, in 3x plants the mean leaf surface was 49.1 cm2, flower diameter – 52.4 mm, fruit weight – 204.7 g, stomata length – 32.1 μm and pollen grain diameter – 33.7 μm, whereas in diploids – 36.0 cm2, 46.1 mm, 162.7 g, 28.4 μm and 30.7 μm, respectively. Pollen grain viability was on average significantly higher in diploids (75.6%), compared to triploids (22%). These results confirm that in apple, as in many other plant species, the higher ploidy level of triploids is generally associated with increased sizes of pollen grains, stomata, flowers, fruits and leaves but decreased pollen viability. No clear correlation between ploidy level and flowering time was found. In the case of mixoploid apple genotypes possessing diploid and tetraploid genomes, some phenotype observation is helpful in describing the ploidy level of the histogenic layers, L1 and L2. Small stomata sizes (similar to diploid) indicate diploid L1 and larger leaf sizes, compared to diploid counterparts, show tetraploid L2. The results will be used for breeding, in which it is important to determine maternal and paternal genotypes as well as the direction of the crossing that is of great importance in obtaining seeds and materials for further selection.

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