5S rDNA Sites Research Articles

Karyotype diversity of Polybia (gr. occidentalis) species complex (Hymenoptera: Vespidae): Taxonomic and evolutionary implications

AbstractPolybia is a common wasp genus found in most of South America. Polybia (gr. occidentalis) encompasses several species that are difficult to identify because of their similar morphologies. Our goal was to cytogenetically characterize three species belonging to Polybia (gr. occidentalis) and discuss the importance of cytogenetic data for integrative taxonomy. Polybia colonies were sampled from different regions of Brazil to perform classical and molecular cytogenetic analyses. Polybia paulista and Polybia sp. 1 showed 2n = 34 and 18S rDNA loci on two chromosomes, whereas Polybia sp. 2 showed 2n = 40 and 18S rDNA clusters on four chromosomes. The microsatellites GA(15), GAG(10), CAA(10), TTAGG(6), and TCAGG(6) showed similar distributions among the species, forming blocks in the euchromatic regions of the chromosomes, whereas CGG(10) did not yield any positive markings. In contrast, TAT(10) hybridized on the centromeric heterochromatin, showing differences in the number of marked chromosomes among the species. Therefore, it may potentially be a species‐specific cytotaxonomic marker in this group of wasps, but this feature needs further investigation. Fluorochromes evidenced that AT‐rich DAPI+ sequence distribution was coincident with heterochromatin, while the distribution of CG‐rich CMA3+ sequences was coincident with the 18S rDNA region. The present study revealed differences in diploid number, heterochromatin content, 18S rDNA sites, and microsatellite patterns between morphologically similar species, demonstrating the usefulness of cytotaxonomy in studying species complexes.

Chromosomal Localization and Diversity Analysis of 5S and 18S Ribosomal DNA in 13 Species from the Genus Ipomoea

Background: Sweet potato (Ipomoea batatas (L.) Lam.), a key global root crop, faces challenges due to its narrow genetic background. This issue can be addressed by utilizing the diverse genetic resources of sweet potato’s wild relatives, which are invaluable for its genetic improvement. Methods: The morphological differences in leaves, stems, and roots among 13 Ipomoea species were observed and compared. Chromosome numbers were determined by examining metaphase cells from root tips. Fluorescence in situ hybridization (FISH) was used to identify the number of 5S and 18S rDNA sites in these species. PCR amplification was performed for both 5S and 18S rDNA, and phylogenetic relationships among the species were analyzed based on the sequences of 18S rDNA. Results: Three species were found to have enlarged roots among the 13 Ipomoea species. Chromosome analysis revealed that I. batatas had 90 chromosomes, Ipomoea pes-tigridis had 28 chromosomes, while the remaining species possessed 30 chromosomes. Detection of rDNA sites in the 13 species showed two distinct 5S rDNA site patterns and six 18S rDNA site patterns in the 12 diploid species. These rDNA sites occurred in pairs, except for the seven 18S rDNA sites observed in Ipomoea digitata. PCR amplification of 5S rDNA identified four distinct patterns, while 18S rDNA showed only a single pattern across the species. Phylogenetic analysis divided the 13 species into two primary clades, with the closest relationships found between I. batatas and Ipomoea trifida, as well as between Ipomoea platensis and I. digitata. Conclusions: These results enhance our understanding of the diversity among Ipomoea species and provide valuable insights for breeders using these species to generate improved varieties.

Structure and Evolution of Ribosomal Genes of Insect Chromosomes.

Currently, clusters of 45S and 5S ribosomal DNA (rDNA) have been studied in about 1000 and 100 species of the class Insecta, respectively. Although the number of insect species with known 45S rDNA clusters (also referred to as nucleolus-organizing regions, or NORs) constitutes less than 0.1 percent of the described members of this enormous group, certain conclusions can already be drawn. Since haploid karyotypes with single 45S and 5S rDNA clusters predominate in both basal and derived insect groups, this character state is apparently ancestral for the class Insecta in general. Nevertheless, the number, chromosomal location, and other characteristics of both 45S and 5S rDNA sites substantially vary across different species, and sometimes even within the same species. There are several main factors and molecular mechanisms that either maintain these parameters or alter them on the short-term and/or long-term scale. Chromosome structure (i.e., monocentric vs. holokinetic chromosomes), excessive numbers of rRNA gene copies per cluster, interactions with transposable elements, pseudogenization, and meiotic recombination are perhaps the most important among them.

Karyotype Description and Comparative Chromosomal Mapping of 5S rDNA in 42 Species.

This study was conducted to evaluate the 5S rDNA site number, position, and origin of signal pattern diversity in 42 plant species using fluorescence in situ hybridization. The species were selected based on the discovery of karyotype rearrangement, or because 5S rDNA had not yet been explored the species. The chromosome number varied from 14 to 160, and the chromosome length ranged from 0.63 to 6.88 μm, with 21 species having small chromosomes (<3 μm). The chromosome numbers of three species and the 5S rDNA loci of nineteen species are reported for the first time. Six 5S rDNA signal pattern types were identified. The 5S rDNA varied and was abundant in signal site numbers (2-18), positions (distal, proximal, outside of chromosome arms), and even in signal intensity. Variation in the numbers and locations of 5S rDNA was observed in 20 species, whereas an extensive stable number and location of 5S rDNA was found in 22 species. The potential origin of the signal pattern diversity was proposed and discussed. These data characterized the variability of 5S rDNA within the karyotypes of the 42 species that exhibited chromosomal rearrangements and provided anchor points for genetic physical maps.

Cytomolecular diversity among Vigna Savi (Leguminosae) subgenera.

The genus Vigna (Leguminosae) comprises about 150 species grouped into five subgenera. The present study aimed to improve the understanding of karyotype diversity and evolution in Vigna, using new and previously published data through different cytogenetic and DNA content approaches. In the Vigna subgenera, we observed a random distribution of rDNA patterns. The 35S rDNA varied in position, from terminal to proximal, and in number, ranging from one (V. aconitifolia, V. subg. Ceratotropis) to seven pairs (V. unguiculata subsp. unguiculata, V. subg. Vigna). On the other hand, the number of 5S rDNA was conserved (one or two pairs), except for V. radiata (V. subg. Ceratotropis), which had three pairs. Genome size was relatively conserved within the genus, ranging from 1C = 0.43 to 0.70pg in V. oblongifolia and V. unguiculata subsp. unguiculata, respectively, both belonging to V. subg. Vigna. However, we observed a positive correlation between DNA content and the number of 35S rDNA sites. In addition, data from chromosome-specific BAC-FISH suggest that the ancestral 35S rDNA locus is conserved on chromosome 6 within Vigna. Considering the rapid diversification in the number and position of rDNA sites, such conservation is surprising and suggests that additional sites may have spread out from this ancestral locus.

Chromosomes of Asian cyprinid fishes: Novel insight into the chromosomal evolution of Labeoninae (Teleostei, Cyprinidae).

The Labeoninae subfamily is a highly diversified but demonstrably monophyletic lineage of cyprinid fishes comprising five tribes and six incertae sedis genera. This widely distributed assemblage contains some 48 genera and around 480 recognized species distributed in freshwaters of Africa and Asia. In this study, the karyotypes and other chromosomal properties of five Labeoninae species found in Thailand Labeo chrysophekadion (Labeonini) and Epalzeorhynchos bicolor, Epalzeorhynchos munense, Henicorhynchus siamensis, Thynnichthys thynnoides (´Osteochilini´) were examined using conventional and molecular cytogenetic protocols. Our results confirmed a diploid chromosome number (2n) invariably 2n = 50, but the ratio of uni- and bi-armed chromosomes was highly variable among their karyotypes, indicating extensive structural chromosomal rearrangements. Karyotype of L. chrysophekadion contained 10m+6sm+20st+14a, 32m+10sm+8st for H. siamensis, 20m+12sm+10st+8a in E. bicolor, 20m+8sm+8st+14a in E. munense, and 18m+24sm+8st in T. thynnoides. Except for H. siamensis, which had four sites of 5S rDNA sites, other species under study had only one chromosome pair with those sites. In contrast, only one pair containing 18S rDNA sites were found in the karyotypes of three species, whereas two sites were found in that of E. bicolor. These cytogenetic patterns indicated that the cytogenomic divergence patterns of these labeonine species largely corresponded to the inferred phylogenetic tree. In spite of the 2n stability, diverse patterns of rDNA and microsatellite distribution as well as their various karyotype structures demonstrated significant evolutionary differentiation of Labeoninae genomes as exemplified in examined species. Labeoninae offers a traditional point of view on the evolutionary forces fostering biological diversity, and the recent findings add new pieces to comprehend the function of structural chromosomal rearrangements in adaption and speciation.

Cytomolecular trends in Chamaecrista Moench (Caesalpinioideae, Leguminosae) diversification.

Chamaecrista is a Pantropical legume genus of the tribe Cassieae, which includes six other genera. In contrast to most of the other Cassieae genera, Chamaecrista shows significant variability in chromosome number (from 2n = 14 to 2n = 56), with small and morphologically similar chromosomes. Here, we performed a new cytomolecular analysis on chromosome number, genome size, and rDNA site distribution in a molecular phylogenetic perspective to interpret the karyotype trends of Chamaecrista and other two genera of Cassieae, seeking to understand their systematics and evolution. Our phylogenetic analysis revealed that Chamaecrista is monophyletic and can be divided into four major clades corresponding to the four sections of the genus. Chromosome numbers ranged from 2n = 14, 16 (section Chamaecrista) to 2n = 28 (sections Absus, Apoucouita, and Baseophyllum). The number of 5S and 35S rDNA sites varied between one and three pairs per karyotype, distributed on different chromosomes or in synteny, with no obvious phylogenetic significance. Our data allowed us to propose x = 7 as the basic chromosome number of Cassieae, which was changed by polyploidy generating x = 14 (sections Absus, Apoucouita, and Baseophyllum) and by ascending dysploidy to x = 8 (section Chamaecrista). The DNA content values supported this hypothesis, with the genomes of the putative tetraploids being larger than those of the putative diploids. We hypothesized that ascending dysploidy, polyploidy, and rDNA amplification/deamplification are the major events in the karyotypic diversification of Chamaecrista. The chromosomal marks characterized here may have cytotaxonomic potential in future studies.

Cyto-molecular characterization of rDNA and chromatin composition in the NOR-associated satellite in Chestnut (Castanea spp.)

The American chestnut (Castanea dentata, 2n = 2x = 24), once known as the “King of the Appalachian Forest”, was decimated by chestnut blight during the first half of the twentieth century by an invasive fungus (Cryphonectria parasitica). The Chinese chestnut (C. mollissima, 2n = 2x = 24), in contrast to American chestnut, is resistant to this blight. Efforts are being made to transfer this resistance to American chestnut through backcross breeding and genetic engineering. Both chestnut genomes have been genetically mapped and recently sequenced to facilitate gene discovery efforts aimed at assisting molecular breeding and genetic engineering. To complement and extend this genomic work, we analyzed the distribution and organization of their ribosomal DNAs (35S and 5S rDNA), and the chromatin composition of the nucleolus organizing region (NOR)-associated satellites. Using fluorescent in situ hybridization (FISH), we have identified two 35S (one major and one minor) and one 5S rDNA sites. The major 35S rDNA sites are terminal and sub-terminal in American and Chinese chestnuts, respectively, originating at the end of the short arm of the chromosome, extending through the secondary constriction and into the satellites. An additional 5S locus was identified in certain Chinese chestnut accessions, and it was linked distally to the major 35S site. The NOR-associated satellite in Chinese chestnut was found to comprise a proximal region packed with 35S rDNA and a distinct distal heterochromatic region. In contrast, the American chestnut satellite was relatively small and devoid of the distal heterochromatic region.

Can ploidy levels explain the variation of Herbertia lahue (Iridaceae)?

Polyploidy is often related with phenotypic variation, as observed in Herbertia lahue, a geophyte species. This study examined the H. lahue polyploid series and departure in cytogenetic, morphometric, and pollen data. Diploids (2n=2x=14) present bimodal karyotype with two long and five short chromosome pairs, while hexaploids (2n=6x=42) and octoploids (2n=8x=56) present a gradual decrease in chromosome size. All cytotypes have CMA+/DAPI- bands co-localized with 18S rDNA sites in the satellite region (no DAPI+ bands in any cytotype). Unlike diploids and octoploids, 5S rDNA interstitial sites in hexaploids are not in a syntenic position with 18S rDNA sites. Genome size is effective as an indirect predictor of the cytotypes since 2C-values increased according to ploidy level. The reduction in the number of the rDNA sites in polyploids associated with their lower 1Cx-values compared to diploids may suggest a genome downsizing process. Morphometric analysis revealed significant differences among cytotypes, and discriminant analysis identified three morphometric groupings corresponding to the cytotypes. The phenotypic variation observed in pollen grains, bulbs, and ovary characters suggested the gigas effect. Concluding, remarkable differentiation was observed at both genomic and phenotypic characters in all the cytotypes analyzed, suggesting a possible ongoing speciation process in H. lahue.

Cytogenetic profiles of two circumglobal snake mackerel species (Scombriformes: Gempylidae) from deep waters of the São Pedro and São Paulo Archipelago

Abstract Chromosomal patterns are valuable tools in evolutionary approaches. Despite the remarkable expansion of fish cytogenetic data, they are still highly deficient concerning deep oceanic species, including the Gempylidae snake mackerels. The snake mackerels are important commercial species composed by meso- and bento-pelagic predators with very limited information available about their lifestyle and genetics patterns. This study presents the first chromosomal data of two circumglobal species of this family, Ruvettus pretiosus and Promethichthys prometheus, from the São Pedro and São Paulo Archipelago. Conventional analyses, chromosomal staining with base-specific fluorochromes, and fluorescence in situ hybridization (FISH) for mapping of repetitive DNA classes were used. Both species have 2n = 48 chromosomes, but they highly differ regarding the karyotype formula (FN = 50 and FN = 84). The 18S rDNA/Ag-NOR and the 5S rDNA sites have a syntenic bi-telomeric array in R. pretiosus, but an independent distribution in P. prometheus. The transposable elements are dispersed, while the microsatellites are also clustered in the centromeric and terminal regions of some chromosomes. It is noteworthy that despite the 2n conservation, a marked macro and microstructural diversifications, mainly mediated by pericentric inversions, differentiates the karyotypes of the species, pointing to a particular chromosomal trajectory of the gempylids among marine fish.

Are the chromosomal fusions that shaped the karyotype of Tetranematichthys wallacei (Siluriformes: Auchenipteridae) a shared feature among Ageneiosini species?

Abstract The genus Tetranematichthys has only three species, and none of them have undergone cytogenetic analyses. Therefore, this study brings for the first time the analysis of Tetranematichthys wallacei, collected from the Igarapé Apaú, Guamá River basin, municipality of Castanhal, Pará State, Brazil. The diploid number found was 52 chromosomes (32m+18sm+2st, NF = 104), in both sexes, with predominantly terminal and some interstitial heterochromatin. Telomeric sequences were observed exclusively in terminal regions. The 18S rDNA sites were found on pair 17sm of all specimens and in only one of the homologous of pair 7 in three specimens. The 5S rDNA sites were found in pairs 8m and 10m. Tetranematichthys wallacei exhibits characteristics worthy of attention regarding its current phylogenetic position, including a probable diploid number reduction. Additionally, it shares with Tympanopleura atronasus the 18S rDNA allocated in the long arm of a large sm chromosome (first pair) but does not share with Ageneiosus the large first m pair with evidence of fusion, as observed in Ageneiosus inermis. The chromosomal data generated for T. wallacei, along with the data from the other two previously studied Ageneiosini taxa, reinforces proposals from morphology-based studies suggesting that the tribe represents the most distinct clade within the family.

Karyotypic characterization of Centromochlus schultzi Rössel 1962 (Auchenipteridae, Centromochlinae) from the Xingu River basin: New inferences on chromosomal evolution in Centromochlus.

Centromochlinae is a widely diverse subfamily with more than 50 species and several taxonomic conflicts due to morphological similarity between Tatia and Centromochlus species. However, cytogenetic studies on this group have been limited to only four species so far. Therefore, here we present the karyotype of Centromochlus schultzi from the Xingu River in Brazil using classic cytogenetic techniques, physical mapping of the 5S and 18S rDNAs, and telomeric sequences (TTAGGG)n. The species had 58 chromosomes, simple NORs and 18S rDNA sites. Heterochromatic regions were detected on the terminal position of most chromosomes, including pericentromeric and centromeric blocks that correspond to interstitial telomeric sites. The 5S rDNA had multiple sites, including a synteny with the 18S rDNA in the pair 24st, which is an ancestral feature for Doradidae, sister group of Auchenipteridae, but appears to be a homoplastic trait in this species. So far, C. schultzi is only the second species within Centromochlus to be karyotyped, but it has already presented characteristics with great potential to assist in future discussions on taxonomic issues in the subfamily Centromochlinae, including the first synteny between rDNAs in Auchenipteridae and also the presence of heterochromatic ITSs that could represent remnants of ancient chromosomal fusions.

Cytogenetic analysis of sympatric Trachelyopterus Valenciennes 1840 (Siluriformes, Auchenipteridae) species reveals highly conserved karyotypes despite the geographic distance

Trachelyopterus Valenciennes 1840 species exhibit striking morphological and cytogenetic similarities, leading to persistent taxonomic challenges. This research focuses on Trachelyopterus galeatus Linnaeus 1766 and Trachelyopterus porosus Eigenmann &amp; Eigenmann 1888, both widely distributed throughout South America and often sympatric, facilitating cytogenetic comparisons. These taxonomic entities are noteworthy for their extensive geographical ranges within the genus. We examined two populations of T. galeatus and T. porosus collected from sympatric sites in the Amazon and Pantanal regions. Both species had the same diploid number and simple Ag-NORs. The 18S rDNA sites were found in only one subtelocentric chromosome pair. Meanwhile, the 5S rDNA sites were found on two distinct chromosomal pairs, with differences in the chromosomal morphology and site position among the species, constituting the most efficient chromosomal marker to distinguish them. The 5S rDNA pattern differed between species but remained consistent between populations of the same species. Minor differences were observed between the T. galeatus populations, probably related to chromosomal rearrangements. In contrast, despite the considerable geographical distance, no cytogenetic differences were detected among the T. porosus populations. Overall, the congruence between cytogenetic and morphological characteristics, combined with our findings from sympatric samples and existing data from geographically separated populations of Trachelyopterus, indicates that the cytogenetic is a promising tool for species differentiation and for delving into the cytotaxonomic and evolutionary aspects of Auchenipteridae.

Molecular cytogenetics of four species of Calendula (Asteraceae)

For the first time, a comparative molecular cytogenetic study of four species of Calendula L. (Asteraceae): C. officinalis, C. stellata, C. tripterocarpa, and C. arvensis was carried out. In each species, chromosome numbers were determined: C. stellata (2n = 2х = 14), C. officinalis (2n = 4x = 32), C. tripterocarpa (2n = 2x = 30), and C. arvensis (2n = 4x = 44), and also specific chromosome localization of 45S and 5S rDNA clusters was revealed with the use of fluorescence in situ hybridization. An additional polymorphic minor 45S rDNA hybridization signal was found in the C. officinalis karyotype, which was located median in the short arm of one chromosome. The patterns of chromosome distribution of the major sites of 45S and 5S rDNA in karyotypes of the studied species confirmed the hybrid origin of C. arvensis (2n = 4x = 44) which could be a result of the introgressive hybridization of two other species: C. stellata (2n = 2x = 14) and C. tripterocarpa (2n = 2x = 30) during speciation.

Comparative Cytogenetic Analysis of Diploid and Triploid Pacific Abalone, Haliotis discus hannai

Introduction: The Pacific abalone, Haliotis discus hannai, is one of the most commercially important marine shellfish in China. Cell engineering breeding is an important tool in abalone genetic breeding, and the triploids obtained through this method have high commercial value. However, current research mainly focuses on establishing induction methods and evaluating the growth traits of triploids, while there is a lack of basic research on triploid cytogenetics. Method: In this study, Cytogenetic analysis of triploid Haliotis discus hannai larvae (produced by chemical treatment) and diploid larvae was performed. Result: The results showed that triploid H. discus hannai had a chromosome number of 3n = 54, consisting of 30 metacentric (m) and 24 submetacentric (sm) chromosomes, while the diploids had a chromosome number of 2n = 36, consisting of 20 metacentric (m) and 16 submetacentric (sm) chromosomes. Notably, both triploids and diploids displayed variation in the number of NORs and/or their diameter. The average number of NORs in triploid was significantly higher than that in diploids (p < 0.05), but the average diameter of NORs of triploid was no significant different from that of diploid (p > 0.05). Additionally, 5S rDNA localization to 3 submetacentric chromosomes was observed in triploids, compared to 2 submetacentric chromosomes in diploids. The number of 18S rDNA sites displayed positional conservancy and quantitative variability in both diploids and triploids. Specifically, 18S rDNA was found at the end of the chromosome in both groups, with triploids exhibiting a significantly higher number of loci than diploids (p < 0.01). Conclusion: This study provides valuable insights into the cytogenetic characteristics of triploid H. discus hannai, which could facilitate further research on the stability of the chromosome set in this species.

Cytological origination of the first found allotriploid tree peony, Paeonia × lemoinei ‘Oukan’ (AAB), reveled by molecular karyotype comparation

Polyploid breeding is one of the most important approaches to improve the traits of cultivated plants, but there has been no feasible method to get polyploid tree peony. In this study, we revealed a sexual triploidization approach from a diploid intersubsectional hybrid tree peony, Paeonia × lemoinei ‘High Noon’, to an allotriploid one, P. × lemoinei ‘Oukan’, by accurate karyotyping based on genomic in situ hybridization (GISH) and fluorescence in situ hybridization (FISH). The ancestor genomes from P. suffruticosa and P. delavayi, which were defined as A-genome and B-genome respectively, were clearly distinguished by GISH in ‘High Noon’ and ‘Oukan’. Nonhomologous chromosomes were successfully recognized by means of 5S and 45S rDNA FISH. Karyotype comparation based on GISH and FISH indicated that the B-genome and one of the two A-genomes in ‘Oukan’ (2n=3x=15, AAB) were highly consistent to the corresponding genomes in ‘High Noon’ (2n=2x=10, AB), implying the triploid origination via 2n gametes produced by first division restitution (FDR). In addition, it was demonstrated for the first time that the dual in series sites of 5S rDNA located on the short arm of chromosome 3 in both A and B-genomes. Besides the familiar strong 45S rDNA signals observed at terminal region of short arms, single weak 45S rDNA signal was newly detected at terminal region in the short arm of certain chromosome 1 in A-genome, and at interstitial-distal region in the long arm of chromosome 5 in both genomes. These findings provide a proof and theoretical support for tree peony polyploid breeding, and an accurate karyotyping method for genetic research of plants in Paeonia.

Autotetraploid Origin of Chinese Cherry Revealed by Chromosomal Karyotype and In Situ Hybridization of Seedling Progenies.

Polyploidy is considered a driving force in plant evolution and diversification. Chinese cherry [Cerasus pseudocerasus (Lindl.) G.Don], an economically important fruit crop native to China, has evolved at the tetraploid level, with a few pentaploid and hexaploid populations. However, its auto- or allo-polyploid origin remains unclear. To address this issue, we analyzed the ploidy levels and rDNA chromosomal distribution in self- and open-pollinated seedling progenies of tetraploid and hexaploid Chinese cherry. Genomic in situ hybridization (GISH) analysis was conducted to reveal the genomic relationships between Chinese cherry and diploid relatives from the genus Cerasus. Both self- and open-pollinated progenies of tetraploid Chinese cherry exhibited tetraploids, pentaploids, and hexaploids, with tetraploids being the most predominant. In the seedling progenies of hexaploid Chinese cherry, the majority of hexaploids and a few pentaploids were observed. A small number of aneuploids were also observed in the seedling progenies. Chromosome 1, characterized by distinct length characteristics, could be considered the representative chromosome of Chinese cherry. The basic Chinese cherry genome carried two 5S rDNA signals with similar intensity, and polyploids had the expected multiples of this copy number. The 5S rDNA sites were located at the per-centromeric regions of the short arm on chromosomes 4 and 5. Three 45S rDNA sites were detected on chr. 3, 4 and 7 in the haploid complement of Chinese cherry. Tetraploids exhibited 12 signals, while pentaploids and hexaploids showed fewer numbers than expected multiples. Based on the GISH signals, Chinese cherry demonstrated relatively close relationships with C. campanulata and C. conradinae, while being distantly related to another fruiting cherry, C. avium. In combination with the above results, our findings suggested that Chinese cherry likely originated from autotetraploidy.

Comparative cytogenetic study of Plantago species

Plantago is a large genus with a worldwide distribution characterized by three basic sets, x = 6 is the most common and is considered the ancestral set from which x = 5 and x = 4 derived following mechanisms of descending dysploidy. Advanced molecular cytogenetic methods are important for understanding evolutionary relationships but in Plantago their use has been rather limited. In this study the karyotypes of Plantago lanceolata L. (2n = 12), Plantago major L. (2n = 12) and Plantago coronopus L. (2n = 10) were characterized by using rDNA FISH, GISH, fluorochrome banding and chromosome morphology to obtain markers useful for evaluating their genomic affinity. The rDNA FISH mapping produced three distinct patterns but with evident similarities such as the chromosome position of 45S rDNA sites and the colocalization of 45S and 5S rDNA. Reciprocal GISH showed a low level of genome affinity between P. lanceolata and P. major and a total absence of homology between them and P. coronopus. This result supports the exclusion of the 2n = 12 species as possible ancestors of P. coronopus. This study confirms that a systematic use of in situ hybridization techniques, and in particular GISH, provide valid information for understanding the origin of species and can resolve many taxonomic and phylogenetic problematics.

Are scattered microsatellites weak chromosomal markers? Guided mapping reveals new insights into Trachelyopterus (Siluriformes: Auchenipteridae) diversity.

The scattered distribution pattern of microsatellites is a challenging problem in fish cytogenetics. This type of array hinders the identification of useful patterns and the comparison between species, often resulting in over-limited interpretations that only label it as "scattered" or "widely distributed". However, several studies have shown that the distribution pattern of microsatellites is non-random. Thus, here we tested whether a scattered microsatellite could have distinct distribution patterns on homeologous chromosomes of closely related species. The clustered sites of 18S and 5S rDNA, U2 snRNA and H3/H4 histone genes were used as a guide to compare the (GATA)n microsatellite distribution pattern on the homeologous chromosomes of six Trachelyopterus species: T. coriaceus and Trachelyopterus aff. galeatus from the Araguaia River basin; T. striatulus, T. galeatus and T. porosus from the Amazonas River basin; and Trachelyopterus aff. coriaceus from the Paraguay River basin. Most species had similar patterns of the (GATA)n microsatellite in the histone genes and 5S rDNA carriers. However, we have found a chromosomal polymorphism of the (GATA)n sequence in the 18S rDNA carriers of Trachelyopterus galeatus, which is in Hard-Weinberg equilibrium and possibly originated through amplification events; and a chromosome polymorphism in Trachelyopterus aff. galeatus, which combined with an inversion polymorphism of the U2 snRNA in the same chromosome pair resulted in six possible cytotypes, which are in Hardy-Weinberg disequilibrium. Therefore, comparing the distribution pattern on homeologous chromosomes across the species, using gene clusters as a guide to identify it, seems to be an effective way to further the analysis of scattered microsatellites in fish cytogenetics.

SCREENING FOR BIOMORPHOLOGICAL AND CYTOGENETIC FEATURES OF POLEMONIUM CAERULEUM L. AFTER COLCHICINE APPLICATION

Background. Currently, it is quite relevant in breeding to create varieties with high yields of raw materials and the content of biologically active compounds.&#x0D; This work aim is to identify promising forms of P. caeruleum L. is for further inclusion in the breeding process based on biomorphological, anatomical, cytogenetic studies and the bioproductivity of colchicine-induced and wild-growing forms in comparison with the Lazur variety.&#x0D; Materials and methods. To obtain polyploids in 2018, the seeds of the P. caeruleum L. the Lazur variety were germinated in Petri dishes on filter paper at room temperature. When the roots reached the length of the seed, they were placed in different concentrations of colchicine solution 0.05 %, 0.1 %, 0.2 % and 0.52 % for 12 and 24 hours, then washed in running water for three hours.&#x0D; Results. The article for the first time presents data on a comparative study of three forms of Polemonium caeruleum L. in terms of morphological characteristics, microscopy of the leaf epidermis and cytogenetic analysis. It was established that the raw materials productivity of the underground part of the plants tetraploid forms in the C1 generation significantly exceeded the diploid and wild forms of plants by 32 %, as well as the yield of seeds by 22 %. In the C2 generation, the yield of raw materials (rhizomes with roots) in tetraploid plants increases in relation to the control by 56 %, seeds by 34 %. Quantitative determination of the amount of triterpene saponins in terms of β-escin in the plant material of three forms of P. caeruleum L. showed an excess of this indicator in the colchicine form by 24 % in the grass and by 36 % in rhizomes with roots, in the wild-growing form - by 15 % in the aerial part and decrease in the underground part by 9 % in relation to the control. It was found that plants of colchicine and wild forms differ in the presence of larger stomata compared to the control. Based on the pattern of DAPI-bandin and the distribution of 45S rDNA and 5S rDNA genes, a generalized species idiogram of blue cyanosis was constructed, taking into account polymorphic variants of DAPI-bandin patterns and indicating the location of the sites of 45S rDNA and 5S rDNA genes. The cytogenetic stability of the resulting tetraploid form was revealed.&#x0D; Conclusion. The article presents comparative studies on morphological characteristics, bioproductivity, microscopy of the leaf epidermis and cytogenetic analysis of the tetraploid form (colchicine) of P. caeruleum L. in comparison with the variety Lazur (diploid form) and the wild form of P. caeruleum L. plants.