Animal genomes vary widely in size, and much of their architecture and content remains poorly understood. Even among related groups, such as orders of insects, genomes may vary in size by orders of magnitude-for reasons unknown. The largest known insect genomes were repeatedly found in Orthoptera, e.g., Podisma pedestris (1C = 16.93 pg), Stethophyma grossum (1C = 18.48 pg) and Bryodemella holdereri (1C = 18.64 pg). While all these species belong to the suborder of Caelifera, the ensiferan Deracantha onos (1C = 19.60 pg) was recently found to have the largest genome. Here, we present new genome size estimates of 50 further species of Ensifera (superfamilies Gryllidea, Tettigoniidea) and Caelifera (Acrididae, Tetrigidae) based on flow cytometric measurements. We found that Bryodemella tuberculata (Caelifera: Acrididae) has the so far largest measured genome of all insects with 1C = 21.96 pg (21.48 gBp). Species of Orthoptera with 2n = 16 and 2n = 22 chromosomes have significantly larger genomes than species with other chromosome counts. Gryllidea genomes vary between 1C = 0.95 and 2.88 pg, and Tetrigidae between 1C = 2.18 and 2.41, while the genomes of all other studied Orthoptera range in size from 1C = 1.37 to 21.96 pg. Reconstructing ancestral genome sizes based on a phylogenetic tree of mitochondrial genomic data, we found genome size values of >15.84 pg only for the nodes of Bryodemella holdereri / B. tuberculata and Chrysochraon dispar / Euthystira brachyptera. The predicted values of ancestral genome sizes are 6.19 pg for Orthoptera, 5.37 pg for Ensifera, and 7.28 pg for Caelifera. The reasons for the large genomes in Orthoptera remain largely unknown, but a duplication or polyploidization seems unlikely as chromosome numbers do not differ much. Sequence-based genomic studies may shed light on the underlying evolutionary mechanisms.

• MeSH
• Biological Evolution MeSH
• Genome Size MeSH
• Phylogeny MeSH
• Genome, Insect MeSH
• Grasshoppers * genetics   MeSH
• Orthoptera * genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

The estimation of nuclear DNA content has been by far the most popular application of flow cytometry in plants. Because flow cytometry measures relative fluorescence intensities of nuclei stained by a DNA fluorochrome, ploidy determination, and estimation of the nuclear DNA content in absolute units both require comparison to a reference standard of known DNA content. This implies that the quality of the results obtained depends on the standard selection and use. Internal standardization, when the nuclei of an unknown sample and the reference standard are isolated, stained, and measured simultaneously, is mandatory for precise measurements. As DNA peaks representing G1 /G0 nuclei of the sample and standard appear on the same histogram of fluorescence intensity, the quotient of their position on the fluorescence intensity axis provides the quotient of DNA amounts. For the estimation of DNA amounts in absolute units, a number of well-established standards are now available to cover the range of known plant genome sizes. Since there are different standards in use, the standard and the genome size assigned to it has always to be reported. When none of the established standards fits, the introduction of a new standard species is needed. For this purpose, the regression line approach or simultaneous analysis of the candidate standard with several established standards should be prioritized. Moreover, the newly selected standard organism has to fulfill a number of requirements: it should be easy to identify and maintain, taxonomically unambiguous, globally available, with known genome size stability, lacking problematic metabolites, suitable for isolation of sufficient amounts of nuclei, and enabling measurements with low coefficients of variation of DNA peaks, hence suitable for the preparation of high quality samples.

• Keywords
• C-value, GC content, best practices, flow cytometry, genome size, plant sciences, plant standard species, standardization,
• MeSH
• DNA, Plant genetics   MeSH
• Genome, Plant * MeSH
• Ploidies * MeSH
• Flow Cytometry methods   MeSH
• Reference Standards MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• DNA, Plant MeSH

The first gapless, telomere-to-telomere (T2T) sequence assemblies of plant chromosomes were reported recently. However, sequence assemblies of most plant genomes remain fragmented. Only recent breakthroughs in accurate long-read sequencing have made it possible to achieve highly contiguous sequence assemblies with a few tens of contigs per chromosome, that is a number small enough to allow for a systematic inquiry into the causes of the remaining sequence gaps and the approaches and resources needed to close them. Here, we analyse sequence gaps in the current reference genome sequence of barley cv. Morex (MorexV3). Optical map and sequence raw data, complemented by ChIP-seq data for centromeric histone variant CENH3, were used to estimate the abundance of centromeric, ribosomal DNA, and subtelomeric repeats in the barley genome. These estimates were compared with copy numbers in the MorexV3 pseudomolecule sequence. We found that almost all centromeric sequences and 45S ribosomal DNA repeat arrays were absent from the MorexV3 pseudomolecules and that the majority of sequence gaps can be attributed to assembly breakdown in long stretches of satellite repeats. However, missing sequences cannot fully account for the difference between assembly size and flow cytometric genome size estimates. We discuss the prospects of gap closure with ultra-long sequence reads.

• Keywords
• CenH3, Cereba, ChIP-seq, PacBio HiFi reads, flow cytometry, nanopore, ribosomal DNA, satellite, telomeric repeats,
• MeSH
• Chromosomes, Plant genetics   MeSH
• Genome, Plant genetics   MeSH
• Hordeum * genetics   MeSH
• DNA, Ribosomal genetics   MeSH
• Sequence Analysis, DNA MeSH
• Telomere genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA, Ribosomal MeSH

The genus Phytophthora comprises many economically and ecologically important plant pathogens. Hybrid species have previously been identified in at least six of the 12 phylogenetic clades. These hybrids can potentially infect a wider host range and display enhanced vigour compared to their progenitors. Phytophthora hybrids therefore pose a serious threat to agriculture as well as to natural ecosystems. Early and correct identification of hybrids is therefore essential for adequate plant protection but this is hampered by the limitations of morphological and traditional molecular methods. Identification of hybrids is also important in evolutionary studies as the positioning of hybrids in a phylogenetic tree can lead to suboptimal topologies. To improve the identification of hybrids we have combined genotyping-by-sequencing (GBS) and genome size estimation on a genus-wide collection of 614 Phytophthora isolates. Analyses based on locus- and allele counts and especially on the combination of species-specific loci and genome size estimations allowed us to confirm and characterize 27 previously described hybrid species and discover 16 new hybrid species. Our method was also valuable for species identification at an unprecedented resolution and further allowed correct naming of misidentified isolates. We used both a concatenation- and a coalescent-based phylogenomic method to construct a reliable phylogeny using the GBS data of 140 non-hybrid Phytophthora isolates. Hybrid species were subsequently connected to their progenitors in this phylogenetic tree. In this study we demonstrate the application of two validated techniques (GBS and flow cytometry) for relatively low cost but high resolution identification of hybrids and their phylogenetic relations.

• Keywords
• Flow cytometry, GBS, Hybrid, Oomycete, Phylogeny, Polyploidy,
• Publication type
• Journal Article MeSH

Any project seeking to deliver a plant or animal reference genome sequence must address the question as to the completeness of the assembly. Given the complexity introduced particularly by the presence of sequence redundancy, a problem which is especially acute in polyploid genomes, this question is not an easy one to answer. One approach is to use the sequence data, along with the appropriate computational tools, the other is to compare the estimate of genome size with an experimentally measured mass of nuclear DNA. The latter requires a reference standard in order to provide a robust relationship between the two independent measurements of genome size. Here, the proposal is to choose the human male leucocyte genome for this standard: its 1C DNA amount (the amount of DNA contained within unreplicated haploid chromosome set) of 3.50 pg is equivalent to a genome length of 3.423 Gbp, a size which is just 5% longer than predicted by the most current human genome assembly. Adopting this standard, this paper assesses the completeness of the reference genome assemblies of the leading cereal crops species wheat, barley and rye.

• Keywords
• flow cytometry, genome size, nuclear DNA content, reference genome assembly, standardization,
• MeSH
• Genome Size * MeSH
• Genome, Human MeSH
• Genome, Plant * MeSH
• Humans MeSH
• Triticum genetics   MeSH
• Reference Standards MeSH
• Sequence Analysis, DNA * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

The species-rich and widespread genus Taraxacum F. H. Wiggers, 1780 (Asteraceae subfamily Cichorioideae) is one of the most taxonomically complex plant genera in the world, mainly due to its combination of different sexual and asexual reproduction strategies. Polyploidy is usually confined to apomictic microspecies, varying from 3x to 6x (rarely 10x). In this study, we focused on Taraxacum sect. Taraxacum (= T.sect.Ruderalia; T.officinale group), i.e., the largest group within the genus. We counted chromosome numbers and measured the DNA content for species sampled in Central Europe, mainly in Czechia. The chromosome number of the 28 species (T.aberrans Hagendijk, Soest & Zevenbergen, 1974, T.atroviride Štěpánek & Trávníček, 2008, T.atrox Kirschner & Štěpánek, 1997, T.baeckiiforme Sahlin, 1971, T.chrysophaenum Railonsala, 1957, T.coartatum G.E. Haglund, 1942, T.corynodes G.E. Haglund, 1943, T.crassum H. Øllgaard & Trávníček, 2003, T.deltoidifrons H. Øllgaard, 2003, T.diastematicum Marklund, 1940, T.gesticulans H. Øllgaard, 1978, T.glossodon Sonck & H. Øllgaard, 1999, T.guttigestans H. Øllgaard in Kirschner & Štěpánek, 1992, T.huelphersianum G.E. Haglund, 1935, T.ingens Palmgren, 1910, T.jugiferum H. Øllgaard, 2003, T.laticordatum Marklund, 1938, T.lojoense H. Lindberg, 1944 (= T.debrayi Hagendijk, Soest & Zevenbergen, 1972, T.lippertianum Sahlin, 1979), T.lucidifrons Trávníček, ineditus, T.obtusifrons Marklund, 1938, T.ochrochlorum G.E. Haglund, 1942, T.ohlsenii G.E. Haglund, 1936, T.perdubium Trávníček, ineditus, T.praestabile Railonsala, 1962, T.sepulcrilobum Trávníček, ineditus, T.sertatum Kirschner, H. Øllgaard & Štěpánek, 1997, T.subhuelphersianum M.P. Christiansen, 1971, T.valens Marklund, 1938) is 2n = 3x = 24. The DNA content ranged from 2C = 2.60 pg (T.atrox) to 2C = 2.86 pg (T.perdubium), with an average value of 2C = 2.72 pg. Chromosome numbers are reported for the first time for 26 species (all but T.diastematicum and T.obtusifrons), and genome size estimates for 26 species are now published for the first time.

• Keywords
• Asteraceae, Taraxacum officinale, chromosome number, flow cytometry, karyology,
• Publication type
• Journal Article MeSH

Pea (Pisum sativum, L.) is a major pulse crop used both for animal and human alimentation. Owing to its association with nitrogen-fixing bacteria, it is also a valuable component for low-input cropping systems. To evaluate the genetic diversity and the scale of linkage disequilibrium (LD) decay in pea, we genotyped a collection of 917 accessions, gathering elite cultivars, landraces, and wild relatives using an array of ∼13,000 single nucleotide polymorphisms (SNP). Genetic diversity is broadly distributed across three groups corresponding to wild/landraces peas, winter types, and spring types. At a finer subdivision level, genetic groups relate to local breeding programs and type usage. LD decreases steeply as genetic distance increases. When considering subsets of the data, LD values can be higher, even if the steep decay remains. We looked for genomic regions exhibiting high level of differentiation between wild/landraces, winter, and spring pea, respectively. Two regions on linkage groups 5 and 6 containing 33 SNPs exhibit stronger differentiation between winter and spring peas than would be expected under neutrality. Interestingly, QTL for resistance to cold acclimation and frost resistance have been identified previously in the same regions.

• Keywords
• FST, Pisum sativum, genetic diversity, linkage disequilibrium,
• MeSH
• Bayes Theorem MeSH
• Ecotype MeSH
• Gene Frequency genetics   MeSH
• Genome, Plant MeSH
• Pisum sativum genetics   MeSH
• Polymorphism, Single Nucleotide genetics   MeSH
• Seasons MeSH
• Seeds genetics   MeSH
• Linkage Disequilibrium genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND AND AIMS: The genome size of an organism is determined by its capacity to tolerate genome expansion, given the species' life strategy and the limits of a particular environment, and the ability for retrotransposon suppression and/or removal. In some giant-genomed bulb geophytes, this tolerance is explained by their ability to pre-divide cells in the dormant stages or by the selective advantage of larger cells in the rapid growth of their fleshy body. In this study, a test shows that the tendency for genome size expansion is a more universal feature of geophytes, and is a subject in need of more general consideration. METHODS: Differences in monoploid genome sizes were compared using standardized phylogenetically independent contrasts in 47 sister pairs of geophytic and non-geophytic taxa sampled across all the angiosperms. The genome sizes of 96 species were adopted from the literature and 53 species were newly measured using flow cytometry with propidium iodide staining. KEY RESULTS: The geophytes showed increased genome sizes compared with their non-geophytic relatives, regardless of the storage organ type and regardless of whether or not vernal geophytes, polyploids or annuals were included in the analyses. CONCLUSIONS: The universal tendency of geophytes to possess a higher genome size suggests the presence of a universal mechanism allowing for genome expansion. It is assumed that this is primarily due to the nutrient and energetic independence of geophytes perhaps allowing continuous synthesis of DNA, which is known to proceed in the extreme cases of vernal geophytes even in dormant stages. This independence may also be assumed as a reason for allowing large genomes in some parasitic plants, as well as the nutrient limitation of small genomes of carnivorous plants.

• Keywords
• Cx-value, Genome size evolution, energy reserves, ephemeroids, flow cytometry, life form, spring geophytes, storage organ,
• MeSH
• Genome Size genetics   MeSH
• DNA, Plant genetics   MeSH
• Genetic Variation MeSH
• Genome, Plant genetics   MeSH
• Plant Roots genetics   MeSH
• Magnoliopsida genetics   MeSH
• Evolution, Molecular MeSH
• Polyploidy MeSH
• Flow Cytometry MeSH
• Retroelements MeSH
• Seasons MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH
• Names of Substances
• DNA, Plant MeSH
• Retroelements MeSH

BACKGROUND AND AIMS: The genus Carex exhibits karyological peculiarities related to holocentrism, specifically extremely broad and almost continual variation in chromosome number. However, the effect of these peculiarities on the evolution of the genome (genome size, base composition) remains unknown. While in monocentrics, determining the arithmetic relationship between the chromosome numbers of related species is usually sufficient for the detection of particular modes of karyotype evolution (i.e. polyploidy and dysploidy), in holocentrics where chromosomal fission and fusion occur such detection requires knowledge of the DNA content. METHODS: The genome size and GC content were estimated in 157 taxa using flow cytometry. The exact chromosome numbers were known for 96 measured samples and were taken from the available literature for other taxa. All relationships were tested in a phylogenetic framework using the ITS tree of 105 species. KEY RESULTS: The 1C genome size varied between 0·24 and 1·64 pg in Carex secalina and C. cuspidata, respectively. The genomic GC content varied from 34·8 % to 40·6 % from C. secalina to C. firma. Both genomic parameters were positively correlated. Seven polyploid and two potentially polyploid taxa were detected in the core Carex clade. A strong negative correlation between genome size and chromosome number was documented in non-polyploid taxa. Non-polyploid taxa of the core Carex clade exhibited a higher rate of genome-size evolution compared with the Vignea clade. Three dioecious taxa exhibited larger genomes, larger chromosomes, and a higher GC content than their hermaphrodite relatives. CONCLUSIONS: Genomes of Carex are relatively small and very GC-poor compared with other angiosperms. We conclude that the evolution of genome and karyotype in Carex is promoted by frequent chromosomal fissions/fusions, rare polyploidy and common repetitive DNA proliferation/removal.

• MeSH
• Carex Plant genetics   MeSH
• Chromosomes, Plant genetics   MeSH
• Genome Size genetics   MeSH
• Phylogeny MeSH
• Genome, Plant genetics   MeSH
• Markov Chains MeSH
• Monte Carlo Method MeSH
• Evolution, Molecular * MeSH
• Polyploidy MeSH
• Base Composition genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND AND AIMS: Genome size is known to affect various plant traits such as stomatal size, seed mass, and flower or shoot phenology. However, these associations are not well understood for species with very large genomes, which are laregly represented by geophytic plants. No detailed associations are known between DNA base composition and genome size or species ecology. METHODS: Genome sizes and GC contents were measured in 219 geophytes together with tentative morpho-anatomical and ecological traits. KEY RESULTS: Increased genome size was associated with earliness of flowering and tendency to grow in humid conditions, and there was a positive correlation between an increase in stomatal size in species with extremely large genomes. Seed mass of geophytes was closely related to their ecology, but not to genomic parameters. Genomic DNA GC content showed a unimodal relationship with genome size but no relationship with species ecology. CONCLUSIONS: Evolution of genome size in geophytes is closely related to their ecology and phenology and is also associated with remarkable changes in DNA base composition. Although geophytism together with producing larger cells appears to be an advantageous strategy for fast development of an organism in seasonal habitats, the drought sensitivity of large stomata may restrict the occurrence of geophytes with very large genomes to regions not subject to water stress.

• MeSH
• Genome Size * MeSH
• DNA, Plant analysis   genetics   MeSH
• Ecology MeSH
• Ecosystem MeSH
• Genome, Plant * MeSH
• Evolution, Molecular MeSH
• Plant Stomata anatomy & histology   MeSH
• Seasons MeSH
• Plants anatomy & histology   genetics   MeSH
• Seeds anatomy & histology   MeSH
• Base Composition MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA, Plant MeSH