Chromosome number, karyotype formula, C-banding pattern, genome size and DNA base composition were studied in three species of Hyalidae and seven species of Talitridae. A karyotype of 25 chromosome pairs, with median centromeres (FN = 100), was found in all the species of Talitridae analysed and Apohyale prevostii. Genome size (C-value) varies among Talitrida from 0.94 pg in Apohyale crassipes to 2.81 pg in Orchestia gammarellus, and the percentage of AT-DNA in the whole genome ranges from 56.12% in A. crassipes to 68.17% in Sardorchestia pelecaniformis. In comparison with Hyalidae, Talitridae show more uniformity in chromosome number and karyotype formula, and have larger genomes. There is a direct correlation between total DNA content and the amount of C-heterochromatic DNA. The cytogenetical data on Talitrida were compared from a phylogenetic and an evolutional point of view. The increase in genome size during the evolution of the Talitrida possibly had a role in their adaptation to supralittoral life and extreme subaerial conditions.
The formation and maintenance of polyploids (via the development of various reproductive barriers) rank among the central questions of studies on polyploid evolution. However, the long time scale of most evolutionary processes makes the study of the dynamics of diploid-polyploid groups difficult. A suitable candidate for a targeted comparative study is Vicia cracca (Fabaceae), which in the late 1960s was subjected to a detailed cytotype screening in Central Europe. Re-sampling the original localities offers a unique opportunity to assess changes in the ploidy structure of the populations, which should reflect the cumulative effect of all the evolutionary forces acting on the plants. Using flow cytometry, the DNA ploidy levels of more than 6,500 individuals of V. cracca collected at 257 localities in Austria, the Czech Republic, Germany and the Slovak Republic were estimated. Three different cytotypes (2x, 3x and 4x) were detected. While tetraploids predominated in the western part of the area investigated (179 populations), the diploids had a more easterly distribution (62 populations). There is a secondary zone of cytotype contact near the boundary between the Czech and Slovak Republics. Sixteen populations (~6%) consisted of a mixture of 2x and 4x cytotypes. Triploids are very rare; only seven individuals were found in two otherwise diploid populations, indicating the existence of breeding barriers between diploids and tetraploids. The distribution of cytotypes is similar to that determined four decades ago using chromosome counts. Nevertheless, there are some discrepancies, namely the current absence of: (i) the diploid cytotype in southern Bohemia and (ii) the altitudinal segregation in the distribution of cytotypes, including two formerly recognized chromosomal races of diploids, perhaps a result of more representative sampling. Identical monoploid genome sizes (1Cx-values) of both the majority ploidy levels support an autopolyploid origin of the tetraploids.
DAPI and propidium iodide flow cytometry were used to determine the variation in genome size in 166 samples and of all taxa and ploidy levels of Fallopia section Reynoutria (knotweeds) recorded in the Czech Republic. Significant differences were detected in the amount of nuclear DNA, associated with the ploidy levels and taxonomic identity of the material. At each ploidy level, F. sachalinensis showed the lowest and F. japonica the highest fluorescence intensities. The fluorescence values for the hybridogenous F. ×bohemica were located in-between these two levels. In most cases, there was at least a four-percent gap in fluorescence values between the nearest neighbours belonging to a different taxon. Intraspecific variation in genome size was very low in all taxa except hexaploid F. ×bohemica; this could be due to the complex evolutionary history of this taxon. Our results indicate that the amount of nuclear DNA can be used as a reliable marker for the identification of homoploid knotweed species and their hybrids. Different evolutionary pathways for the origin of high polyploids and/or hybridogenous taxa are proposed based on genome size.
Variation in genome size in a particular taxonomic group can reflect different evolutionary processes including polyploidy, hybridization and natural selection but also neutral evolution. Using flow cytometry, karyology, ITS sequencing and field surveys, the causes of variation in genome size in the ecologically and morphologically diverse high-Andean genus Lasiocephalus (Asteraceae, Senecioneae) were examined. There was a 1.64-fold variation in holoploid genome size (C-values) among 189 samples belonging to 20 taxa. The most distinct was a group of plants with large genomes corresponding to DNA triploids. Disregarding the DNA triploids, the remaining samples exhibited a pronounced (up to 1.32-fold) and rather continuous variation. Plants with the smallest genomes most likely represent intergeneric hybrids with the closely related and sympatric Culcitium nivale, which has a smaller genome than Lasiocephalus. The variation in genome size in samples of diploid Lasiocephalus was strongly correlated with several environmental and life history traits (altitude, habitat and growth form). However, all these factors, as well as genome size itself, were correlated with phylogeny (main split into the so-called ‘forest’ and ‘páramo’ clades), which most probably represents the true cause of the differentiation in intrageneric genome size. In contrast, relationships between genome size and phylogeny were not apparent at lower divergence levels. Instead, here we suggest that ecological conditions have played a role in driving shifts in genome size between closely related species inhabiting different environments. Collectively, this study demonstrates that various evolutionary forces and processes have shaped the variation in genome size and indicates that there is a need for multi-approach analyses when searching for the causes and consequences of changes in genome size.
Liverworts are poorly represented in the record of DNA C-values. Data for not more than nine species are reported in the literature. Here we present flowcytometric measurements of genome size for 32 foliose and 11 thallose species from 22 out of 83 families. The main method used in this study was flow cytometry using propidium iodide as the DNA stain. Feulgen densitometrywas applied as a supplementary method but it proved less suitable because the rigid cellwalls of liverwort tissue are resistant to maceration and apparently often inhibit the diffusion of reagents, which results in low estimates of DNA content. The precise or approximate number of chromosomes were counted, where possible. Among the thallose liverworts, the lowest 1C-value was recorded for Marchantia polymorpha (0.293 pg) and the highest for diploid Pellia epiphylla (7.401 pg). Haploid P. epiphylla (1C = 3.803 pg) had the largest genome among the haploid thalloid liverworts. Among the foliose liverworts, Lejeunea cavifolia with a value of 0.211 pg (1C) was ranked the lowest and Mylia taylorii, a haploid with 7.966 pg (1C) and a large amount of dense heterochromatin, concentrated in one big spherical chromocentre, the highest. This 38-fold variation covers the extremes of the whole sample and exceeds the ca 12-fold variation recorded in mosses (0.174–2.160 pg, 1C). This variation is nevertheless low compared to the 2000-fold interspecific variation found in angiosperms. Several instances of intraspecific variation in DNA ploidy (x and 2x) were found – in Radula complanata, Pellia epiphylla and Metzgeria furcata. In Lophocolea heterophylla, accessions differed 3.37-fold in C-value at haploid chromosome number. This points to cryptic taxonomic differentiation and warns against premature statements about ploidy levels based only on DNA measurements. Significant intraspecific intraploidal variation in C-value was also observed in certain instances. In Frullania dilatata, female plants with two large heterochromatic sex-chromosomes have a 1.35-fold higher C-value than male plants with only one sex chromosome. In most other cases of intraspecific variation the role of sex differences remains to be clarified.
Genome size has been suggested as one of the traits associated with invasiveness of plant species. To provide a quantitative insight into the role of this trait, we estimated nuclearDNAcontent in 93 alien species naturalized in the Czech Republic, belonging to 32 families, by using flow cytometry, and compared it with the values reported for non-invading congeneric and confamilial species from the Plant DNA C-values database. Species naturalized in the Czech Republic have significantly smaller genomes than their congeners not known to be naturalized or invasive in any part of the world. This trend is supported at the family level: alien species naturalized in the Czech flora have on average a smaller genome than is the mean value for non-invading confamilials. Moreover, naturalized and non-invading species clearly differed in the frequency of five genome size categories; this difference was mainly due to very small genomes prevailing and intermediate to very large genomes underrepresented in the former group. Our results provide the first quantitative support for association of genome size with invasiveness, based on a large set of alien species across a number of plant families. However, there was no difference in the genome size of invasive species compared to naturalized but non-invasive. This suggests that small genome size provides alien plants with an advantage already at the stage of naturalization and need not be necessarily associated with the final stage of the process, i.e. invasion.
Flow cytometry measurements confirmed the occurrence of Polypodium ×mantoniae (P. interjectum × P. vulgare) at three localities in the eastern part of the Czech Republic (Blansko and Rudice N of Brno and Javoříčko WNW of Olomouc). Nuclear DNA contents (± Sx) were determined for P. vulgare (2C = 29.00 ± 0.32 pg), P. ×mantoniae (2C = 37.18 ± 0.38 pg) and P. interjectum (2C = 45.24 ± 0.31 pg) using a PAS Partec GmbH flow cytometer (PI staining / standard Vicia faba, 2C = 26.9 pg). The relative DNA content ratio was measured in all pairs of taxa (± Sx range), i.e. P. ×mantoniae : P. vulgare = 1.340 ± 0.008; P. interjectum : P. vulgare = 1.681 ± 0.003; P. interjectum : P. ×mantoniae = 1.255 ± 0.008. Six new localities for Polypodium interjectum were found in the region of Moravský Kras (= Moravian Karst, N of Brno). From the PI/DAPI index it can be inferred that the AT/GC ratio (or heterochromatin occurrence) is 1.05× bigger in P. ×mantoniae than in P. vulgare and 1.08× bigger in P. interjectum than in P. vulgare. Anatomical data (number of thick- walled cells in the anulus, spore length and stomata length) of selected specimens and live samples from the Czech Republic were in good agreement with the range of variation of these features published by earlier authors from other European countries. A brief historical survey of the knowledge of P. interjectum in the Czech Republic is included.
This paper reviews recent use of flow cytometry in studies on apomictic plant taxa. The most of apomictic angiosperms are polyploid, often differing in ploidy level from their sexual counterparts within the agamic complex. Flow cytometry is widely used for screening the ploidy levels of mature plants and their seed generated both in the field and in experiments. Routine ploidy screening often accompanied by molecular markers distinguishing individual genotypes are used to reveal novel insights into the biosystematics and population biology of apomictic taxa. Apomixis (asexual seed formation) is mostly facultative, operating together with other less frequent reproductive pathways within the same individual. The diversity in modes of reproduction in apomicts is commonly reflected in the ploidy structure of their progeny in mixed-cytotype populations. Thus, flow cytometry facilitates the detection and quantification of particular progeny classes generated by different reproductive pathways. The specific embryo/endosperm ploidy ratios, typical of the different reproductive pathways, result from modifications of double fertilization in sexual/apomictic angiosperms.Thus, the reproductive origin of seed can be identified, including autonomous or pseudogamous apomixis, haploid parthenogenesis and sexual reproduction, involving either reduced or unreduced gametes. Collectively, flow cytometry has been used to address the following research topics: (i) assessing the variation in ploidy levels and genome sizes in agamic complexes, (ii) detection and quantification of different reproductive modes in facultative apomicts, (iii) elucidation of processes in populations with coexisting sexual and apomictic biotypes, (iv) evolution of agamic complexes, and (v) genetic basis of apomixis. The last topic is of paramount importance to crop breeding: the search for candidate gene(s) responsible for apomixis is the main objective of many research programmes. A list of the angiosperm taxa that could provide model systems for such research is provided.
Over the last decade there has been a tremendous increase in the use of flow cytometry (FCM) in studies on the biosystematics, ecology and population biology of vascular plants. Most studies, however, address questions related to differences in genome copy number, while the value of FCM for studying homoploid plant groups has long been underestimated. This review summarizes recent advances in taxonomic and ecological research on homoploid plants that were made using FCM. A fairly constant amount of nuclear DNA within each evolutionary entity together with the often large differences between species means that genome size is a useful character for taxonomic decision-making. Regardless of the number of chromosomes, genome size can be used to delimit taxa at various taxonomic levels, resolve complex low-level taxonomies, assess the frequency of interspecific hybridization or infer evolutionary relationships in homoploid plant groups. In plant ecology and evolutionary biology, variation in genome size has been used for prediction purposes because genome size is associated with several phenotypic, physiological and/or ecological characteristics. It is likely that in the future the use ofFCM in studies on taxonomy, ecology and population biology of homoploid plants will increase both in scope and frequency. Flow cytometry alone, but especially in combination with other molecular and phenotypic approaches, promises advances in our understanding of the functional significance of variation in genome size in homoploid plants.