Plant Genome Size Estimation by Flow Cytometry: Inter-laboratory Comparison

Doležel, Jaroslav; Greilhuber, Johann; Lucretti, Sergio; Meister, Armin; Lysak, Martin A.; Nardi, L.; Obermayer, Renate

doi:10.1093/oxfordjournals.aob.a010312

Cited by 431 publications

(396 citation statements)

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“…For the primary standard Pisum sativum, a value of 2C ϭ 9.07 pg and an AT frequency of 61.5% were used (see "Genome Size and AT Frequency of the Standards"). The 2C values and AT frequencies of the secondary standards were calculated on the base of the primary standard and the results obtained from Doležel et al (11), Tables 4 and 7 and laboratory L1. The family-attached code is used for the identification of the families in Figure 1.…”

Section: Results and Discussion Estimation Of Genome Size And At Freqmentioning

confidence: 99%

“…Some of the standards proposed by Doležel et al (11) were used as references: Allium cepa, Glycine max, Pisum sativum, Raphanus sativus, Secale cereale, and Vicia faba. The calculation of reference values is described in the Results and Discussion section.…”

Section: Materials and Methods Plant Materialsmentioning

confidence: 99%

“…In some cases, 0.5-1% Triton (Sigma, Steinheim, Germany) for reducing adhesion of cellular debris, 5% polyvinylpyrrolidone 25 (PVP; Serva, Heidelberg/New York) for binding phenolic compounds, and/or 50 -100 mM potassium metabisulfite (Riedel-de Haën, Seelze, Germany) as an antioxidant were added in order to obtain acceptable coefficient of variation (CV) values. Different buffers result only in minor variation of peak ratios in experiments with internal standardization (11).…”

Section: Preparation Of Nuclear Suspensionsmentioning

confidence: 99%

“…For the genome size, a value of 2C ϭ 9.07 pg is used. The 2C values of the other references were calculated on the basis of this primary standard and the results obtained by Doležel et al (11), Table 4, laboratory L1.…”

Section: Genome Size and At Frequency Of The Standardsmentioning

confidence: 99%

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Lack of correlation between AT frequency and genome size in higher plants and the effect of nonrandomness of base sequences on dye binding

Barow

Meister

2001

Cytometry

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Background: Different plant species vary as to the ratio of nucleotide base pairs of genomic DNA. A correlation between genome size and base pair ratio has been claimed. Base composition can be analyzed by base-specific dyes. Methods: Genome size is determined by flow cytometry of suspensions of nuclei stained by the base independent dye, PI. For estimation of the AT frequency, the ATspecific dyes 4,6-diamidino-2-phenylindole, dihydrochloride (DAPI) and Hoechst 33342 (HO) were used. We define a dye factor (DF) as the ratio of the two estimates (peak ratios) of nuclear fluorescence intensities of sample relative to reference plant nuclei using a given dye and an intercalating fluorochrome. Results: No significant correlation between genome size and the DF for DAPI was found when 54 plant species were investigated. However, similarities within and differ-

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Section: Results and Discussion Estimation Of Genome Size And At Freqmentioning

confidence: 99%

Section: Materials and Methods Plant Materialsmentioning

confidence: 99%

Section: Preparation Of Nuclear Suspensionsmentioning

confidence: 99%

Section: Genome Size and At Frequency Of The Standardsmentioning

confidence: 99%

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Lack of correlation between AT frequency and genome size in higher plants and the effect of nonrandomness of base sequences on dye binding

Barow

Meister

2001

Cytometry

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“…Mycelium was collected with a needle and added to a Petri dish with 1 mL of modified woody plant buffer (WPB; 0.2 M Tris-HCl, 4 mM MgCl 2 ·6H 2 O, 2 % Triton X-100, 2 mM disodium EDTA, 86 mM NaCl, 20 mM metabisulfite, 4 % PVP-10, [pH 7.5]; Loureiro et al 2007). Nuclei were released following the procedure of Galbraith et al (1983) by chopping approximately 10 mg of mycelium with a razor blade together with 50 mg of fresh leaf tissue of Raphanus sativus (internal reference standard with 2C= 1.16 pg or 1,135 Mbp; Doležel et al 1998) to ensure that nuclei of both species were exposed to identical chemical and mechanical conditions.…”

Section: Genome Size Estimationmentioning

confidence: 99%

Large and variable genome size unrelated to serpentine adaptation but supportive of cryptic sexuality in Cenococcum geophilum

et al. 2013

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Estimations of genome size and its variation can provide valuable information regarding the genetic diversity of organisms and their adaptation potential to heterogeneous environments. We used flow cytometry to characterize the variation in genome size among 40 isolates of Cenococcum geophilum, an ectomycorrhizal fungus with a wide ecological and geographical distribution, obtained from two serpentine and two non-serpentine sites in Portugal. Besides determining the genome size and its intraspecies variation, we wanted to assess whether a relationship exists between genome size and the edaphic background of the C. geophilum isolates. Our results reveal C. geophilum to have one of the largest genome sizes so far measured in the Ascomycota, with a mean haploid genome size estimate of 0.208 pg (203 Mbp). However, no relationship was found between genome size and the edaphic background of the sampled isolates, indicating genetic and demographic processes to be more important for shaping the genome size variation in this species than environmental selection. The detection of variation in ploidy level among our isolates, including a single individual with both presumed haploid and diploid nuclei, provides supportive evidence for a possible cryptic sexual or parasexual cycle in C. geophilum (although other mechanisms may have caused this variation). The existence of such a cycle would have wide significance, explaining the high levels of genetic diversity and likelihood of recombination previously reported in this species, and adds to the increasing number of studies suggesting sexual cycles in previously assumed asexual fungi.

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Estimation of Relative Nuclear DNA Content in Dehydrated Plant Tissues by Flow Cytometry

Suda

Trávníček

2006

CP Cytometry

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The power of flow cytometry in field plant research may be greatly enhanced by analysis of nonfresh tissues. Previous attempts to use chemical fixatives, however, received only little attention because of protocol complexity and limited time, after which successful assays were achieved. This unit describes simple and rapid procedures for relative nuclear DNA content estimation in dehydrated tissues of vascular plants by DAPI flow cytometry. Histograms with reasonable resolution can be obtained in several‐year‐old specimens. The approach here is particularly suitable for reliable determination of DNA ploidy level, although detection of small variation in nuclear DNA content is also possible in many cases. Retrospective ploidy inference in silica‐dry or herbarium vouchers and simplified transport of plant material from remote areas are among the principal benefits for plant biosystematics, ecology, and population biology.

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Plant Genome Size Estimation by Flow Cytometry: Inter-laboratory Comparison

Cited by 431 publications

References 0 publications

Lack of correlation between AT frequency and genome size in higher plants and the effect of nonrandomness of base sequences on dye binding

Lack of correlation between AT frequency and genome size in higher plants and the effect of nonrandomness of base sequences on dye binding

Large and variable genome size unrelated to serpentine adaptation but supportive of cryptic sexuality in Cenococcum geophilum

Estimation of Relative Nuclear DNA Content in Dehydrated Plant Tissues by Flow Cytometry

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