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ABSTRACT Across the tree of life, many organisms are able to reproduce clonally, via vegetative spread, budding or parthenogenesis. In population genetic analyses of clonally reproducing organisms, it is common practice to retain only a single representative per multilocus genotype. Though this practice of clone correction is widespread, the theoretical justification behind it has been very little studied. Here, I use individual‐based simulations to study the effect of clone correction on the estimation of the genetic summary statistics HO, HS, FIS, FST, F′′ST and Dest. The simulations follow the standard finite island model, consisting of a set of populations connected by gene flow, but with a variable rate of sexual versus asexual reproduction. The results of the simulations show that by itself, the inclusion of replicated genotypes does not lead to a deviation in the values of the summary statistics, except when the rate of sexual reproduction is less than about one in thousand. However, clone correction can introduce a strong deviation in the values of most of the statistics, when compared to a scenario of full sexual reproduction. For HS and FIS, this deviation can be informative about the process of asexual reproduction, but for FST, F′′ST and Dest, clone correction can lead to incorrect conclusions. I therefore argue that clone correction is not strictly necessary, but can in some cases be insightful. However, when clone correction is applied, it is imperative that results for both the corrected and uncorrected data are presented. [ABSTRACT FROM AUTHOR]

AbstractApomixis, the reproduction via asexually formed seed, is taxonomically scattered in angiosperms. Here we review the major developmental pathways to apomixis and the occurrences on the level of orders and families. We provide surveys of large families for which occurrences and evolutionary pathways of apomixis are well documented (Poaceae, Ranunculaceae, Plumbaginaceae, Rosaceae, Brassicaceae, Rutaceae, and Asteraceae). Molecular phylogenetic, phylogenomic, and cytogenetic studies have confirmed that apomixis frequently arises in hybrids, with or without polyploidy. Multiple origins, different developmental pathways, and genetic/epigenetic control mechanisms confirm that apomixis is a derived trait. Pollen functions are in many taxa maintained for fertilization of polar nuclei and proper endosperm development, which allows also for maintenance of facultative sexuality and further intercrossing of lineages. This way, apomixis often results in huge and dynamic complexes of numerous hybrid genotypes and phenotypes with highly reticulate relationships. Such complexes are successful to establish in various habitats and geographical regions. Expression of apomixis appears to be in some genera influenced by environmental conditions. The diversity of evolutionary pathways is reflected in various, genus-wise taxonomic treatments. Future research is needed to understand the short-term evolutionary dynamics, the functional background for apomixis, and the long-term evolutionary fates of apomictic lineages. [ABSTRACT FROM AUTHOR]

Previous studies on the mountain plant Ranunculus kuepferi concluded that apomictic self-compatible tetraploids have experienced a niche shift toward a colder climate during the Holocene, which suggests a fitness advantage over the sexual, self-sterile diploid parents under cold and stressful high-mountain conditions. However, there is still a lack of information on whether reproductive development would be advantageous for tetraploids. Here, we report on microsporogenesis, megagametogenesis, the dynamics of flower and seed development, and the consequences for reproductive success in a common garden experiment along a 1000 m climatic elevation gradient and in natural populations. Flower buds were initiated in the year preceding anthesis and passed winter in a pre-meiotic stage. Flower morphology differed in the known cytotype-specific way in that tetraploid flowers produced about twice as many carpels and fewer petals, stamens, and pollen grains than diploid flowers. Tetraploids developed precociously aposporous embryo sacs and showed a high rate of developmental disturbances. Sexual seed formation prevailed in diploids and pseudogamous apomixis in tetraploids. Along the elevation gradient, stigma pollen load, pollen performance, and seed output decreased. Combinations of reproductive traits, namely, bypass of meiosis irregularities and uniparental reproduction, might have promoted the vast expansion of apomictic R. kuepferi lines across the European Alps. [ABSTRACT FROM AUTHOR]

Key message: The combination of a flow cytometric seed screen and genotyping of each single seed offers a cost-effective approach to detecting complex reproductive pathways in flowering plants. Reproduction may be seen as one of the driving forces of evolution. Flow cytometric seed screen and genotyping of parents and progeny are commonly employed techniques to discern various modes of reproduction in flowering plants. Nevertheless, both methods possess limitations constraining their individual capacity to investigate reproductive modes thoroughly. We implemented both methods in a novel manner to analyse reproduction pathways using a carefully selected material of parental individuals and their seed progeny. The significant advantage of this approach lies in its ability to apply both methods to a single seed. The introduced methodology provides valuable insights into discerning the levels of apomixis, sexuality, and selfing in complex Rubus taxa. The results may be explained by the occurrence of automixis in Rubus, which warrants further investigation. The approach showcased its effectiveness in a different apomictic system, specifically in Taraxacum. Our study presents a comprehensive methodological approach for determining the mode of reproduction where flow cytometry loses its potential. It provides a reliable and cost-effective method with significant potential in biosystematics, population genetics, and crop breeding. [ABSTRACT FROM AUTHOR]

In recent years, proteomics has become a powerful tool in vitro plant tissue culture research. In Miconia , the most diverse flowering plant genus in the Neotropics, these tools together were unexplored. The group is replete with cases of apomixis (asexual seed formation). Some studies indicate that apomixis may have similarities with somatic embryogenesis (SE). However, there is no investigation into whether explants from apomictic species can affect the accumulation of SE-related proteins in vitro morphogenesis processes, such as callogenesis. This study identifies differentially accumulated proteins related to SE in the callogenesis of three Miconia species with different reproductive systems and whether this can affect their embryogenic capacity. For this, we employed the shotgun comparative proteomics approach in a specific phase of callus proliferation of Miconia australis (obligate apomictic), Miconia hyemalis (facultative apomictic), and Miconia sellowiana (sexual). Comparative proteomic analysis revealed that 190 SE-related proteins were differentially accumulated and distributed among the three experimental comparisons. The calluses of apomictic species showed the highest number of up-accumulated proteins associated with SE. Histological examination of the calluses after 45 days of induction revealed the permanence of cells with meristematic characteristics in the apomictic species, however, in M. sellowiana there was greater cellular vacuolization. These results indicate that calluses of Miconia spp. originated from apomictic species may be better able to acquire embryogenic competence in vitro culture. This study is the first to explore and demonstrate the potential advantages of using proteomics to understand the molecular basis of in vitro morphogenesis in apomictic and sexual species of Miconia , highlighting new perspectives for future research in plant tissue culture. [Display omitted] • Proteomics shed light into the mechanisms underlying in vitro morphogenesis in miconia. • Calluses from apomictic miconia up-accumulated proteins related to somatic embryogenesis. • The cu/zn SOD was up-accumulated in the calluses of obligate apomictic M. australis. • Apomictic species showed a higher prevalence of callus cells with meristematic characteristics. [ABSTRACT FROM AUTHOR]

Key message: The DNA methylation status at an epigenetic quantitative trait locus in the Arabidopsis chromosome 2 is linked to the formation of apomictic-like endosperms. Seed development in most angiosperms is coupled to fertilization of the maternal gametes by two sperm cells. However, apomictic species can reproduce asexually via seeds. This trait is of great agricultural interest, as it would fix complex genotypes and allow for pollen-independent seed production. However, engineering full apomixis requires three independent processes: apomeiosis, parthenogenesis and autonomous endosperm development. While the first two have been successfully engineered in some crops, the formation of autonomous endosperms remains a challenge. Although it is known that this trait is under epigenetic control, such as of DNA methylation, the underlying mechanisms remain mostly undiscovered. Here, using epigenetic recombinant inbred lines, we identified an epigenetic quantitative trait locus in the Arabidopsis chromosome 2, which correlates with permissiveness for the formation of asexual seeds: hypomethylation at this genomic region allows the formation of larger autonomous endosperms. Importantly, the methylation at this locus only correlates with asexual seed size, and not to the size of sexual seeds or that of other organs. With this, we aim to show that screening for epialleles is a promising strategy to uncover loci underlying relevant traits and could pave the way to identifying genes necessary for the engineering of apomixis. [ABSTRACT FROM AUTHOR]

Maize (Zea mays L.) is one of the most demanded grain crops in the world. Currently, production has exceeded one billion tons and is increasing by 3–5% annually. Such growth is due to the genetic potential of the crop and the use of heterosis F1 hybrids in production. However, the need to produce first-generation seed annually poses significant challenges and is an economically costly technology. A solution to this problem may be the transfer of the asexual (apomictic) mode of reproduction to maize from its wild relative, eastern gamagrass (Tripsacum dactyloides L.). In this work, we report the production of 56-chromosome apomictic hybrids of maize (Zea mays L.) with eastern gamagrass (T. dactyloides L.) with restored anther fertility. The mode of reproduction of the plant was confirmed by counting chromosomes and sequencing the nuclear gene (Pox3) and chloroplast tRNA-Leu (trnL) gene. These apomictic hybrids had karyotypes of 2n = 56 = [(10Zm(573MB) + 36Td) + 10Zm(611CB)] and 2n = 56 = [(10Zm(611CB) + 36Td) + 10Zm(611CB)]. The resulting hybrids can be widely used as a fodder crop. [ABSTRACT FROM AUTHOR]

Background The predominance of sex in eukaryotes, despite the high costs of meiosis and mating, remains an evolutionary enigma. Many theories have been proposed, none of them being conclusive on its own, and they are, in part, not well applicable to land plants. Sexual reproduction is obligate in embryophytes for the great majority of species. Scope This review compares the main forms of sexual and asexual reproduction in ferns and angiosperms, based on the generation cycling of sporophyte and gametophyte (leaving vegetative propagation aside). The benefits of sexual reproduction for maintenance of genomic integrity in comparison to asexuality are discussed in the light of developmental, evolutionary, genetic and phylogenetic studies. Conclusions Asexual reproduction represents modifications of the sexual pathway, with various forms of facultative sexuality. For sexual land plants, meiosis provides direct DNA repair mechanisms for oxidative damage in reproductive tissues. The ploidy alternations of meiosis–syngamy cycles and prolonged multicellular stages in the haploid phase in the gametophytes provide a high efficiency of purifying selection against recessive deleterious mutations. Asexual lineages might buffer effects of such mutations via polyploidy and can purge the mutational load via facultative sexuality. The role of organelle–nuclear genome compatibility for maintenance of genome integrity is not well understood. In plants in general, the costs of mating are low because of predominant hermaphroditism. Phylogenetic patterns in the archaeplastid clade suggest that high frequencies of sexuality in land plants are concomitant with a stepwise increase of intrinsic and extrinsic stress factors. Furthermore, expansion of genome size in land plants would increase the potential mutational load. Sexual reproduction appears to be essential for keeping long-term genomic integrity, and only rare combinations of extrinsic and intrinsic factors allow for shifts to asexuality. [ABSTRACT FROM AUTHOR]

Background and Aims Rubus ser. Glandulosi provides a unique model of geographical parthenogenesis on a homoploid (2 n  = 4 x) level. We aim to characterize evolutionary and phylogeographical patterns in this taxon and shed light on the geographical differentiation of apomicts and sexuals. Ultimately, we aim to evaluate the importance of phylogeography in the formation of geographical parthenogenesis. Methods Rubus ser. Glandulosi was sampled across its Eurasian range together with other co-occurring Rubus taxa (587 individuals in total). Double-digest restriction site-associated DNA sequencing (ddRADseq) and modelling of suitable climate were used for evolutionary inferences. Key Results Six ancestral species were identified that contributed to the contemporary gene pool of R. ser. Glandulosi. Sexuals were introgressed from Rubus dolichocarpus and Rubus moschus in West Asia and from Rubus ulmifolius agg. Rubus canescens and Rubus incanescens in Europe, whereas apomicts were characterized by alleles of Rubus subsect. Rubus. Gene flow between sexuals and apomicts was also detected, as was occasional hybridization with other taxa. Conclusions We hypothesize that sexuals survived the last glacial period in several large southern refugia, whereas apomicts were mostly restricted to southern France, whence they quickly recolonized Central and Western Europe. The secondary contact of sexuals and apomicts was probably the principal factor that established geographical parthenogenesis in R. ser. Glandulosi. Sexual populations are not impoverished in genetic diversity along their borderline with apomicts, and maladaptive population genetic processes probably did not shape the geographical patterns. [ABSTRACT FROM AUTHOR]

Parthenogenesis, the development of unfertilized egg cells into embryos, is a key component of apomixis. AtBBM (BABY BOOM), a crucial regulator of embryogenesis in Arabidopsis, possesses the capacity to shift nutritional growth toward reproductive growth. However, the mechanisms underlying AtBBM‐induced parthenogenesis remain largely unexplored in dicot plants. Our findings revealed that in order to uphold the order of sexual reproduction, the embryo‐specific promoter activity of AtBBM as well as repressors that inhibit its expression in egg cells combine to limiting its ability to induce parthenogenesis. Notably, AtRKD5, a RWP‐RK domain‐containing (RKD) transcription factor, binds to the 3′ end of AtBBM and is identified as one of the inhibitory factors for AtBBM expression in the egg cell. In the atrkd5 mutant, we successfully achieved enhanced ectopic expression of AtBBM in egg cells, resulting in the generation of haploid offspring via parthenogenesis at a rate of 0.28%. Furthermore, by introducing chimeric Arabidopsis and rice BBM genes into the egg cell, we achieved a significant 4.6‐fold enhancement in haploid induction through the atdmp8/9 mutant. These findings lay a strong foundation for further exploration of the BBM‐mediated parthenogenesis mechanism and the improvement of haploid breeding efficiency mediated by the dmp8/9 mutant. [ABSTRACT FROM AUTHOR]

Introduction: Phenotypic complexity in species complexes and recently radiated lineages has resulted in a diversity of forms that have historically been classified into separate taxa. Increasingly, with the proliferation of high-throughput sequencing methods, additional layers of complexity have been recognized, such as frequent hybridization and reticulation, which may call into question the previous morphological groupings of closely related organisms. Methods: We investigated Northern European, Asian, and Beringian populations of Ranunculus auricomus agg. with phylogenomic analysis of 736 genes and 27,586 SNPs in order to deduce the interrelatedness and hybrid origin of this phenotypically and taxonomically complicated group from Europe characterized by a history of hybridization, polyploidy, apomixis, and recent radiation. The ploidy levels and the reproductive mode of the Northern European populations were assessed via flow cytometric seed screening. In addition, in order to examine the phenotypic plasticity of the dwarf forms previously described as species and summarized as the Ranunculus monophyllus group, we conducted climate chamber experiments under cold (northern) and warm (temperate) conditions. Results: The Northern European populations are tetra- to hexaploid and propagate primarily through apomixis. The complex is characterized by highly reticulate relationships. Genetic differentiation of the main clusters has occurred between the above-mentioned geographical regions. We find evidence for the hybrid origin of the taxa in these areas with differing genomic contributions from the geographically nearest European sexual progenitor species. Furthermore, polyphyly in the taxa of the R. monophyllus group is supported. Experiments show low lability in the traits associated with the R. monophyllus group. Discussion: We conclude that multiple adaptations of hybrids to colder climates and shorter vegetation periods have shaped the phenotypes of the R. monophyllus group, and we suggest a formal classification as nothotaxa within the R. auricomus group. [ABSTRACT FROM AUTHOR]

We studied Rubus wahlbergii and related species in R. sect. Corylifolii, primarily in Sweden, using DNA microsatellites. The results show that the material can be divided into three species: R. wahlbergii s.s. which occurs along the Swedish coast from Bohuslän to Uppland, R. nordicus which occurs on the western coast all the way south to north‐western Skåne, and a previously unrecognized taxon which we here describe as R. wahlbergioides. The latter has scattered localities in Halland, Skåne, Blekinge, Öland and Småland, but is common around Kivik in Skåne and Kalmar in Småland. It is characterised by leaves that are without felt underneath, terminal leaflets with a somewhat more triangular shape and coarser serration than those of R. wahlbergii and also has more glands on the stem and the inflorescence. In the northern part of the distribution, R. nordicus is easily distinguished morphologically from R. wahlbergii, e.g. by a strong tendency to form 7‐foliolate leaves, but in Skåne and Halland, the two species are harder to distinguish. Still, genetically R. nordicus is more similar to R. sordirosanthus than to R. wahlbergii s.s. [ABSTRACT FROM AUTHOR]

Potato is the most important non-cereal crop worldwide, and, yet, genetic gains in potato have been traditionally delayed by the crop's biology, mostly the genetic heterozygosity of autotetraploid cultivars and the intricacies of the reproductive system. Novel site-directed genetic modification techniques provide opportunities for designing climate-smart cultivars, but they also pose new possibilities (and challenges) for breeding potato. As potato species show a remarkable reproductive diversity, and their ovules have a propensity to develop apomixis-like phenotypes, tinkering with reproductive genes in potato is opening new frontiers in potato breeding. Developing diploid varieties instead of tetraploid ones has been proposed as an alternative way to fill the gap in genetic gain, that is being achieved by using gene-edited self-compatible genotypes and inbred lines to exploit hybrid seed technology. In a similar way, modulating the formation of unreduced gametes and synthesizing apomixis in diploid or tetraploid potatoes may help to reinforce the transition to a diploid hybrid crop or enhance introgression schemes and fix highly heterozygous genotypes in tetraploid varieties. In any case, the induction of apomixis-like phenotypes will shorten the time and costs of developing new varieties by allowing the multi-generational propagation through true seeds. In this review, we summarize the current knowledge on potato reproductive phenotypes and underlying genes, discuss the advantages and disadvantages of using potato's natural variability to modulate reproductive steps during seed formation, and consider strategies to synthesize apomixis. However, before we can fully modulate the reproductive phenotypes, we need to understand the genetic basis of such diversity. Finally, we visualize an active, central role for genebanks in this endeavor by phenotyping properly genotyped genebank accessions and new introductions to provide scientists and breeders with reliable data and resources for developing innovations to exploit market opportunities. [ABSTRACT FROM AUTHOR]

Epigenetics studies changes in gene activity without changes in the DNA sequence. Methylation is an epigenetic mechanism important in many pathways, such as biotic and abiotic stresses, cell division, and reproduction. Eragrostis curvula is a grass species reproducing by apomixis, a clonal reproduction by seeds. This work employed the MCSeEd technique to identify deferentially methylated positions, regions, and genes in the CG, CHG, and CHH contexts in E. curvula genotypes with similar genomic backgrounds but with different reproductive modes and ploidy levels. In this way, we focused the analysis on the cvs. Tanganyika INTA (4x, apomictic), Victoria (2x, sexual), and Bahiense (4x, apomictic). Victoria was obtained from the diploidization of Tanganyika INTA, while Bahiense was produced from the tetraploidization of Victoria. This study showed that polyploid/apomictic genotypes had more differentially methylated positions and regions than the diploid sexual ones. Interestingly, it was possible to observe fewer differentially methylated positions and regions in CG than in the other contexts, meaning CG methylation is conserved across the genotypes regardless of the ploidy level and reproductive mode. In the comparisons between sexual and apomictic genotypes, we identified differentially methylated genes involved in the reproductive pathways, specifically in meiosis, cell division, and fertilization. Another interesting observation was that several differentially methylated genes between the diploid and the original tetraploid genotype recovered their methylation status after tetraploidization, suggesting that methylation is an important mechanism involved in reproduction and ploidy changes. [ABSTRACT FROM AUTHOR]

Apomixis is a common reproductive characteristic of Zanthoxylum plants, and RWP-RKs are plant-specific transcription factors known to regulate embryonic development. However, the genome-wide analysis and function prediction of RWP-RK family genes in Z. armatum are unclear. In this study, 36 ZaRWP-RK transcription factors were identified in the genome of Z. armatum, among which 15 genes belonged to the RKD subfamily and 21 belonged to the NLP subfamily. Duplication events of ZaRWP-RK genes were mainly segmental duplication, and synteny analysis revealed a close phylogenetic relationship between Z. armatum and Arabidopsis. The analysis of cis-elements indicated that ZaRWP-RK genes may be involved in the regulation of the embryonic development of Z. armatum by responding to plant hormones such as abscisic acid, auxin, and gibberellin. Results of a real-time PCR showed that the expression levels of most ZaRWP-RK genes were significantly increased from flowers to young fruits. Protein–protein interaction network analysis further revealed the potential roles of the ZaRWP-RK proteins in apomixis. Collectively, this study is expected to improve our understanding of ZaRWP-RK transcription factors and provide a theoretical basis for future investigations into the ZaRWP-RK genes and their regulatory mechanisms in the apomixis process of Z. armatum. [ABSTRACT FROM AUTHOR]

Asexual seed reproduction, apomixis, is a relatively rare reproductive strategy in angiosperms. Its rarity is probably due to evolutionary constraints that require full and synchronous expression of all the steps that modify the ancestral sexual pathway. Therefore, the documentation of any new occurrence of apomixis, especially in taxonomic groups with no previous report, should be based on thorough and solid evidence. Here, we correct the report of suggested apomictic reproduction in two endemic hexaploid species of Senecio from Australia. Based on either excess or deficiency of expected heterozygosity across loci, over-representation of several multilocus genotypes, and segregation analysis assessed using microsatellite markers, it was suggested that apomixis together with selfing could explain the observed patterns of genetic variation in perennial S. macrocarpus and annual S. squarrosus. We used flow cytometric seed screening analyses to verify the mode of reproduction of several widespread multilocus genotypes that are considered to be apomictic. These analyses revealed that all seed progeny arose by the sexual pathway, as they exhibited a constant 3:2 endosperm to embryo ploidy ratio, typical of double fertilization of a reduced embryo sac by two reduced sperm cells. Therefore, both species reproduce sexually. We suggest that the unusual genetic patterns in both allopolyploids are caused by originally low variation in the parental species, which probably also reproduce by selfing, disomic inheritance and probably also the presence of null alleles. We call for a more careful and critical interpretation of the patterns obtained, taking into account all aspects related to apomictic reproduction. [ABSTRACT FROM AUTHOR]

A geobotanical analysis of 160 steppe and ecotone plant communities with the participation of cereals in the north of the Lower Volga region (within Saratov oblast) has been carried out. The species diversity of apomictic cereals is significantly inferior to the diversity of their amphimictic relatives in the flora of the region and in the studied phytocenoses (24 apomictic and 109 amphimictic species in the flora; 17 and 34 species in the phytocenoses, respectively). At the same time, the dominants of the studied communities are equally represented by both amphimictic (14 species) and apomictic cereals (9 species). It is established that cereals with an amphimictic mode of reproduction dominate in 64 phytocenoses and apomictic cereals dominate in 48 ones. The number of phytocenoses in which apomicts are codominants significantly exceeds the number of plant communities with amphimicts codominance (46 and 22, respectively). The studied plant communities are similar in species richness and evenness. The data indicate successful competition between apomictic cereals and amphimicts and their significant contribution to the composition of steppe and ecotone phytocenoses in the northern Lower Volga region. [ABSTRACT FROM AUTHOR]

Abstract Many Melastomataceae are distributed in highland rocky fields (Campos rupestres) landscapes with their intrinsic environmental factors. These factors influence the expression of morphoanatomical characteristics and determine which adaptations are necessary for survival, including reproductive processes. The genus Miconia belongs to the Neotropical tribe Miconieae, the largest in Melastomataceae. Species of Miconieae are known to engage in apomixis, which results in much of the tribe possibly carrying out autonomous or pseudogamous reproduction. Thus, this study seeks to describe the morphoanatomy of the flowers of Miconia polystachya (Naudin) R. Goldenb. and contribute to the understanding of their reproduction. For this, flowers were collected at different stages of development in different seasons (autumn and spring) for morphoanatomical study. The flowers were fixed and cross and longitudinal sections were carried out with a rotating microtome. Pollen viability tests were also performed. The individuals of Miconia polystachya studied in highland rocky fields of the state of Paraná have a high percentage of unviable pollen, verified by the structure of pollen grains showing different degrees of degeneration. However, individuals with mature fruits were observed, corroborating the association between reproduction of this species with apomixis. The described morphological characteristics are similar to those found in previous studies for the family and genus. However, our study provides novel ecological information about the flowering and floral anatomy of M. polystachya, emphasizing the anatomy of hypanthium, fertile verticils and embryonic development of the species.

Extended Abstract Background: It is important to research in the field of germplasm protection of native and medicinal tree species in northern Iran forests, especially blood hawthorn with many medicinal properties. Sexual reproduction of hawthorn is possible with apomixis due to the thick shell and two types of physical and physiological dormancy, and its asexual reproduction by rooting cuttings faces many problems. Bloody hawthorn (Crataegus atrosanguinea from the Rosaceae family) is considered one of the valuable species of the forests of the north and west of the country, which is very important due to its fruitfulness, very strong wood, and numerous medicinal properties. The medicinal properties of hawthorn have been proven in the treatment of blood pressure, arteriosclerosis, heart congestion, heart muscle weakness, and coronary insufficiency. Hawthorn contains many secondary metabolites such as flavonoids proanthocyanidin and glycosides. Optimizing the in vitro cultivation conditions of blood hawthorn as one of the natural species of Iran can be a suitable solution for its propagation. The present research was carried out to control pollution, control browning, and induce the callus of blood hawthorn (C. atrosanguinea) in vitro. Controlling the contamination of explants, seeds, and plant tissues is of great importance for the success of plant tissue culture. Since the enrichment of natural plant explants is a priority for tissue culture, this is usually associated with many problems with browning, especially for woody plants that have a thick and woody stem. Determining the type and concentration of the growth regulator of a variable factor is necessary for the growth of plants under in vitro conditions. After applying the growth regulator to the culture medium, the plant will show different reactions, but one of the appropriate ways to choose the correct path of plant regeneration is to determine the wet and dry weights of the callus, which is widely measured as a standard of callus growth. The callus growth curve can be obtained using the wet and dry weights of the callus. The results of past research indicate a wide range of changes in the type and concentration of growth regulators on callus formation and regeneration of different species of the hawthorn genus. A specific type and concentration of growth regulators should be applied according to the type of plant species. Methods: Young and healthy stems of blood hawthorn were collected from Larijan in Amol, Mazandaran province, Iran, with geographical coordinates of 38' 58° N and 10' 52° E, at the beginning of the growing season. The collected stems were immediately transferred to the plant tissue culture laboratory of the Faculty of Natural Resources at Sari University of Agriculture and Natural Resources, Iran. Disinfection of natural hawthorn stems is very necessary due to surface and tissue contamination. For this purpose, explants of 0.5-1 cm size were cut from the stems. The infection control treatments included HgCl2, H2O2, NaClO, and AgNO3, and five treatments were designed to control the browning of explants. Browning control treatments included control, running water, ascorbic acid, polyvinyl chloride, and activated charcoal. In general, the culture medium consisted of substances including mineral salts (micro- and macroelements) as providers of maximum growth, amino acids and vitamins as nitrogen sources, sugar as a source of carbon and energy, growth regulators as growth and morphogenesis stimulants, agar as the solidifier of the cultivation medium, and water, which includes 90% of the cultivation medium. In this research, the culture medium was MS, which was first created by Driver and Koniaki (1984) and prepared according to Dufussard (1976). In this study, the pH of the culture medium was set to 5.8. For callus induction, IBA, NAA, 2,4-D, BAP, TDZ, and Kin treatments at three concentration levels of 0.1, 1, and 6 mg/L and a combination of NAA (6 mg-1) + BAP (1 mg-1), and IBA (6 mg l-1) + BAP (1 mg-1) were used in the MS culture medium. Stem explants were placed horizontally on the culture medium. Among different callus treatments, callus percentage, degree of callus, callus fresh and dry weights, and cross-sectional area of callus were measured to investigate callus induction. The best treatment was obtained by calculating the mentioned characteristics and that treatment was used for planting. It was transplanted approximately every 20 days and a total of 5-6 times. After callus cultivation in the reproduction medium, the explants were placed in a culture chamber and adaptation racks under 16 hours of light and 8 hours of dark treatment at 25 ± 2 °C with a humidity of 70%. Results: HgCl2 (0.5%, 16 minutes) with an average of 100% health was the most appropriate pollution control treatment. The control treatment with 98% health was observed as the most appropriate browning control treatment. Moreover, the type, concentration, and interaction effect of the type and concentration of the growth regulator significantly affected the callus induction characteristics of blood hawthorn (P < 0.01). The highest fresh and dry weights of callus, percentage and degree of callus formation, and cross-sectional area of callus were observed among auxin and cytokinin treatments, respectively, for 6 mg/L of NAA and 1 mg/L of BAP treatments. The highest levels of the studied callus induction characteristics were observed in the culture medium containing the combination of auxin and cytokinin (6 mg-1 of IBA+1 mg-1 of BAP). Conclusion: In general, the results showed a 90% chance of controlling the browning of the explants of natural blood hawthorn branches in a normal state and without applying any treatment. It is recommended to use 0.5% mercuric chloride for 16 minutes to disinfect the explants. The combination of growth regulators in the culture medium was evaluated as very suitable for the callus induction of bloody hawthorn. The results of this research can provide laboratory instructions for the regeneration and mass reproduction of blood hawthorn.

Plant species respond to varying plant species diversity and associated changes in their abiotic and biotic environment with changes in their phenotype. However, it is not clear to what degree this phenotypic differentiation is due to genotype diversity within populations or phenotypic plasticity of plant individuals. We studied individuals of 16 populations of the clonal herb Taraxacum officinale grown in plant communities of different species richness in a 17‐year‐old grassland biodiversity experiment (Jena Experiment). We collected 12 individuals in each population to measure phenotypic traits and identify distinct genotypes using microsatellite DNA markers. Plant species richness did not influence population‐level genotype and trait diversity. However, it affected the expression of several phenotypic traits, e.g. leaf and inflorescence number, maximum leaf length and seed mass, which increased with increasing plant species richness. Moreover, population‐level trait diversity correlated positively with genotype richness for leaf dry matter content (LDMC) and negatively with inflorescence number. For several traits (i.e. seed mass, germination rate, LDMC, specific leaf area (SLA)), a larger portion of variance was explained by genotype identity, while variance in other traits (i.e. number of inflorescences, leaf nitrogen concentration, leaf number, leaf length) resided within genotypes and thus was mostly due to phenotypic plasticity. Overall, our findings show that plant species richness positively affected the population means of some traits related to whole‐plant performance, whose variation was achieved through both phenotypic plasticity and genotype composition of a population. [ABSTRACT FROM AUTHOR]

Summary: Asexual organisms often differ in their geographic distributions from their sexual relatives. This phenomenon, termed geographic parthenogenesis, has long been known, but the underlying factors behind its diverse patterns have been under dispute. Particularly problematic is an association between asexuality and polyploidy in most taxa. Here, we present a new system of geographic parthenogenesis on the tetraploid level, promising new insights into this complex topic.We used flow cytometric seed screen and microsatellite genotyping to characterise the patterns of distribution of sexuals and apomicts and genotypic distributions in Rubus ser. Glandulosi across its range. Ecological modelling and local‐scale vegetation and soil analyses were used to test for niche differentiation between the reproductive groups.Apomicts were detected only in North‐western Europe, sexuals in the rest of the range in Europe and West Asia, with a sharp borderline stretched across Central Europe. Despite that, we found no significant differences in ecological niches. Genotypic richness distributions suggested independence of the reproductive groups and a secondary contact.We argue that unless a niche differentiation (resulting from polyploidy and/or hybridity) evolves, the main factors behind the patterns of geographic parthenogenesis in plants are phylogeographic history and neutral microevolutionary processes, such as clonal turnover. [ABSTRACT FROM AUTHOR]

Premise: Apomixis in ferns is relatively common and obligatory. Sterile hybrids may restore fertility via apomixis at a cost of long‐term genetic stagnation. In this study, we outlined apomixis as a possible temporary phase leading to sexuality and analyzed factors relating to transitioning to and away from apomixis, such as unreduced and reduced spore formation in apomict and apo‐sex hybrid ferns. Methods: We analyzed the genome size of 15 fern species or hybrids ("taxa") via flow cytometry. The number of reduced and unreduced gametophytes was established as a proxy for viable spore formation of either type. We also calculated the spore abortion ratio (sign of reduced spores) in several taxa, including the apo‐sex hybrid Dryopteris × critica and its 16 apomictically formed offspring. Results: Four of 15 sampled taxa yielded offspring variable in genome size. Specifically, each variable taxon formed one viable reduced plant among 12–451 sampled gametophytes per taxon. Thus, haploid spore formation in the studied apomicts was very rare but possible. Spore abortion analyses indicated gradually decreasing abortion (haploid spore formation) over time. In Dryopteris × critica, abortion decreased from 93.8% to mean 89.5% in one generation. Conclusions: Our results support apomixis as a transitionary phase toward sexuality. Newly formed apomicts hybridize with sexual relatives and continue to form haploid spores early on. Thus, they may get the genomic content necessary for regular meiosis and restore sexuality. If the missing relative goes extinct, the lineage gets locked into apomixis as may be the case with the Dryopteris affinis complex. [ABSTRACT FROM AUTHOR]

Research on Satyrium nepalense var. ciliatum (Lindl.) Hook. f. has primarily focused on populations in Northwestern Yunnan, with limited studies on pollination syndromes and insect behavior. In addition, it is geographically limited in its breeding system studies. Here, pollination syndromes, florivory, and breeding systems of S. nepalense var. ciliatum from Liangwang Mountain (Central Yunnan, China) were investigated through field work, microscope, scanning electron microscope (SEM), and parafin section. It was revealed that the pollination syndrome was possessing out-crossing, such as bright color, a developed rostellum, nectar glands in the spur, and food hairs at the lip base. The color and nectar attracted flower visitors, and florivory was observed. Some flower visitors pollinated their companion species. Ants were identified as floral visitors for the first time in Satyrium, although substantial pollination was not observed. Ants might be potential pollinators. S. nepalense var. ciliatum possessed a mixed breeding system, including selfing, out-crossing, and apomixis, with apomixis being predominant in nature. It is suggested that the pollination syndrome, florivory, and pollination competition would contribute to its mixed breeding systems, particularly leading to the occurrence of apomixis. [ABSTRACT FROM AUTHOR]

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In higher plants, sexual reproduction is characterized by meiosis of the first cells of the germlines, and double fertilization of the egg and central cell after gametogenesis. In contrast, in apomicts of the genus Boechera , meiosis is omitted or altered and only the central cell requires fertilization, while the embryo forms parthenogenetically from the egg cell. To deepen the understanding of the transcriptional basis underlying these differences, we applied RNA-seq to compare expression in reproductive tissues of different Boechera accessions. This confirmed previous evidence of an enrichment of RNA helicases in plant germlines. Furthermore, few RNA helicases were differentially expressed in female reproductive ovule tissues harboring mature gametophytes from apomictic and sexual accessions. For some of these genes, we further found evidence for a complex recent evolutionary history. This included a homolog of Arabidopsis thaliana FASCIATED STEM4 (FAS4). In contrast to AtFAS4 , which is a single-copy gene, FAS4 is represented by three homologs in Boechera , suggesting a potential for subfunctionalization to modulate reproductive development. To gain first insights into functional roles of FAS4 , we studied Arabidopsis lines carrying mutant alleles. This identified the crucial importance of AtFAS4 for reproduction, as we observed developmental defects and arrest during male and female gametogenesis. [ABSTRACT FROM AUTHOR]

Aim: Apomixis is a widespread trait in extreme environments worldwide, yet phylogeographical studies for species exhibiting these complex reproductive systems are still limited to temperate zones in the Northern Hemisphere. Through analyses of a combination of adult plants and seedlings, and nuclear and chloroplast DNA, we assessed the contemporary genetic outcomes of apomixis and phylogeographical patterns in an arid unglaciated landscape to understand the evolutionary trajectory of apomictic species. Location: Pilbara bioregion, north‐western Australia. Taxon: Senna glutinosa subsp. glutinosa. Materials and Methods: Spatial patterns of genetic diversity and differentiation in three chloroplast regions and 17 nuclear microsatellite markers were evaluated in 480 plants from 20 populations of S. glutinosa subsp. glutinosa across the Pilbara bioregion. We also germinated and genotyped seed for a progeny analysis to quantify polyembryony and the relative frequency of asexual versus sexual reproduction. Results: We found frequent polyembryony and a high proportion of asexual offspring (84.56%), resulting in low within‐population genotypic diversity. The nuclear dataset identified 72 multilocus lineages (MLLs) with moderate pairwise population genetic fixation and low differentiation among populations with no geographically structured genetic clustering. Chloroplast DNA showed high haplotype diversity and a widespread distribution of haplotypes. Only one peripheral population presented a single unique haplotype and MLL. Main Conclusions: Analysis of this woody perennial in an arid landscape shows the influence of apomixis on genetic patterns and species persistence. Haplotype diversity was indicative of long‐term persistence within the ancient Pilbara region. Widespread distribution of haplotypes and sharing of several nuclear MLLs among distant populations indicated extensive seed dispersal throughout the study area. The combination of polyploidy, facultative apomixis and extensive seed dispersal maintains genetic variability within local populations and promotes the spread of advantageous genotypes across wide geographical distances. Therefore, apomixis, rather than an evolutionary 'dead‐end', can be regarded as a mechanism facilitating evolutionary success of apomictic species in extreme and complex environments. [ABSTRACT FROM AUTHOR]

Introduction: Among candidate genes underlying the control components of apomixis, APOLLO is known for its strong linkage to apomeiosis in the genus Boechera. The gene has "apo alleles," which are characterized by a set of linked apomixis-specific polymorphisms, and "sex alleles." All apomictic Boechera genotypes are heterozygous for the apo/sex alleles, whereas all sexual genotypes are homozygous for sex alleles. Methods: In this study, native and synthetic APOLLO promoters were characterized by detecting the expression level of the β-glucuronidase (GUS) gene in Arabidopsis. Results: Comparing various flower developmental stages in transgenic lines containing different constructs with 2-kb native transgenic lines revealed that changes to the APOLLO promoter causes shifts in tissue and developmental stage specificity of GUS expression. Importantly, several apomixis-specific polymorphisms in the 5'UTR change the timing and location of GUS activity from somatic to reproductive tissues. Discussion: These synthetic data simulate a plausible evolutionary process, whereby apomixis-specific gene activity can be achieved. [ABSTRACT FROM AUTHOR]

Key message: High-frequency clonal seeds and near-normal fertility were obtained by engineering synthetic apomixis in hybrid rice. The one-line strategy, with the advantage of unnecessary seed production, is the final stage for the hybrid rice development and can be achieved through the fixation of heterosis via artificially inducing apomixis. Recently, synthetic apomixis has been generated in rice by combining MiMe (Mitosis instead of Meiosis) with either the ectopic expression of BABY BOOM (BBM1 or BBM4) or mutation of MATRILINEAL (MTL), resulting in over 95.00% of clonal seeds. However, the frequency of clonal seeds was only 29.20% when AtDD45 promoter was used to drive BBM1. In addition, achieving both a high frequency of clonal seeds and near-normal fertility simultaneously had been elusive in earlier strategies. In this study, using AtDD45 promoter to drive BBM1 expression in combination with the MiMe mutant resulted in the apomixis frequency as high as 98.70%. Even more, employing fusion promoters (AtMYB98_AtDD1_OsECA1-like1) to drive WUS expression in combination with pAtDD45:BBM1 and MiMe could produce clonal seeds at rates of up to 98.21%, the highest seed setting rate reached to 83.67%. Multiple-embryos were observed in clonal lines at a frequency ranging from 3.37% to 60.99%. Transmission of the high frequency of apomixis through skipped generations (atavism) was identified in two clonal lines, even though it remained stable in the majority of clonal lines. These findings significantly advance the pursuit of fixed heterosis in rice through synthetic apomixis, edging closer to its agricultural application. [ABSTRACT FROM AUTHOR]

Plants in German states Thuringia and Bavaria previously thought to belong to Potentilla thuringiaca belong to at least three different taxa. Most populations in Thuringia and Bavaria are rather uniform and have a ploidy of 2n = 10x. These populations belong to P. thuringiaca as originally described. A lectotype and an epitype are designated for P. thuringiaca. In an isolated range in Eastern Thuringia, we found octoploids (2n = 8x), which are characterized mainly by a higher number of flowering stems. These populations are described as Potentilla saalae T. Gregor & Korsch. Decaploid (2n = 10x) plants from a very small range in the Bavarian Rhoen with more rounded central leaves are probably of independent origin. The relationship of German P. thuringiaca populations to those in the Alps, the Balkans, the Carpathians, and Eastern Europe is unresolved. [ABSTRACT FROM AUTHOR]

IntroductionPhenotypic complexity in species complexes and recently radiated lineages has resulted in a diversity of forms that have historically been classified into separate taxa. Increasingly, with the proliferation of high-throughput sequencing methods, additional layers of complexity have been recognized, such as frequent hybridization and reticulation, which may call into question the previous morphological groupings of closely related organisms.MethodsWe investigated Northern European, Asian, and Beringian populations of Ranunculus auricomus agg. with phylogenomic analysis of 736 genes and 27,586 SNPs in order to deduce the interrelatedness and hybrid origin of this phenotypically and taxonomically complicated group from Europe characterized by a history of hybridization, polyploidy, apomixis, and recent radiation. The ploidy levels and the reproductive mode of the Northern European populations were assessed via flow cytometric seed screening. In addition, in order to examine the phenotypic plasticity of the dwarf forms previously described as species and summarized as the Ranunculus monophyllus group, we conducted climate chamber experiments under cold (northern) and warm (temperate) conditions.ResultsThe Northern European populations are tetra- to hexaploid and propagate primarily through apomixis. The complex is characterized by highly reticulate relationships. Genetic differentiation of the main clusters has occurred between the above-mentioned geographical regions. We find evidence for the hybrid origin of the taxa in these areas with differing genomic contributions from the geographically nearest European sexual progenitor species. Furthermore, polyphyly in the taxa of the R. monophyllus group is supported. Experiments show low lability in the traits associated with the R. monophyllus group.DiscussionWe conclude that multiple adaptations of hybrids to colder climates and shorter vegetation periods have shaped the phenotypes of the R. monophyllus group, and we suggest a formal classification as nothotaxa within the R. auricomus group.

The article explores the development of a new type of flowering dogwood called 'Erica's Appalachian Sunrise'. It explains that this cultivar was created through a form of asexual reproduction called apomixis, resulting from a breeding attempt between two existing cultivars. The article describes the tree's structure, flower bract shape, and leaf color, highlighting the distinctions between 'Erica's Appalachian Sunrise' and another cultivar called 'Cherokee Brave™'. It also addresses the misconception that 'Erica's Appalachian Sunrise' was the product of self-pollination, providing evidence to support its apomictic origin. The study further discusses the genomic evidence suggesting the occurrence of apomixis in Cornus florida, the species to which the flowering dogwood belongs. The DNA sequences of the maternal plant and the plant derived from apomixis are identical, but there are noticeable differences in their physical characteristics. These differences are attributed to epigenetic factors, such as modifications to DNA or gene expression regulation. This discovery marks the first report of apomixis in C. florida. [Extracted from the article]

Previous studies on the mountain plant Ranunculus kuepferi concluded that apomictic self-compatible tetraploids have experienced a niche shift toward a colder climate during the Holocene, which suggests a fitness advantage over the sexual, self-sterile diploid parents under cold and stressful high-mountain conditions. However, there is still a lack of information on whether reproductive development would be advantageous for tetraploids. Here, we report on microsporogenesis, megagametogenesis, the dynamics of flower and seed development, and the consequences for reproductive success in a common garden experiment along a 1000 m climatic elevation gradient and in natural populations. Flower buds were initiated in the year preceding anthesis and passed winter in a pre-meiotic stage. Flower morphology differed in the known cytotype-specific way in that tetraploid flowers produced about twice as many carpels and fewer petals, stamens, and pollen grains than diploid flowers. Tetraploids developed precociously aposporous embryo sacs and showed a high rate of developmental disturbances. Sexual seed formation prevailed in diploids and pseudogamous apomixis in tetraploids. Along the elevation gradient, stigma pollen load, pollen performance, and seed output decreased. Combinations of reproductive traits, namely, bypass of meiosis irregularities and uniparental reproduction, might have promoted the vast expansion of apomictic R. kuepferi lines across the European Alps.

Introduction: Persian walnut (Juglans regia) is an economically important nut fruit species cultivated worldwide for its nutritious kernel and timber quality wood. Walnut trees are mostly hetero-dichogamous and, depending on the genotype, some cultivars are protogynous, while others are protandrous. Although selfing is possible when male and female blooms overlap, the dichogamy of the species promotes outcrossing. In addition to sexual reproduction, some reports indicate that elements of apomixis may occur in commercial orchards of walnut varieties and in the last two decades, nut production by apomixis has been reported in walnut. However, there are no reliable studies on the occurrence of apomictic reproduction based on cytoembryological observations and/or molecular marker-progeny tests. This study addresses the combined use of molecular and cytological analyses to gain new insights into the population genetics and reproduction systems of J. regia. Methods: We systematically analyzed the reproductive origin of individual progeny plants from 8 different cultivated walnut genotypes using microsatellite genotyping and carried out cytohistological investigations of 5 cultivated walnut genotypes arising seed sets from isolated flowers, to shed light on the mode of reproduction. Results and discussion: These cytometric and genotyping analyses did not support any asexual mode of reproduction or asexual propagation by seed and all individuals studied were identified as zygotic plants produced by crossing. Likewise, the cytological findings did not confirm completely the first component of apomixis, namely apomeiosis. On the other hand, according to histological evidence, adventitious embryony seems to take place at low frequency. Overall, our findings suggest that the occurrence of gametophytic apomixis is unlikely in J. regia, but sporophytic apomixis cannot be completely ruled out. [ABSTRACT FROM AUTHOR]

In species that primarily reproduce asexually, occasional sex can have important evolutionary consequences. Many plant species in the genus Oenothera have a genetic system called permanent translocation heterozygosity (PTH), which renders plants functionally asexual when they self-fertilize. PTH results from reciprocal translocations of chromosomes that suppress recombination and a balanced lethal mortality of homozygous gametes that prevents independent assortment of alleles. When PTH plants self-fertilize, the offspring are genetically identical to the parents, but when they outcross they can create new genotypes. Here, we sought to determine the outcrossing rate of natural populations of PTH Oenothera biennis L. (Onagraceae). We genotyped 46 parents from 14 populations in Tompkins County, NY, USA, and 754 of their offspring at highly polymorphic microsatellite loci. 99.5% of offspring were genetically identical to their seed parents, whereas 4 offspring out of 754 exhibited allelic mismatches with their seed parent. The estimated outcrossing rate ranged from 0.1% to 0.4%. Our study design also permitted us to estimate mutation rate of microsatellite loci, which ranged from 1.33 × 10−3 to 3.98 × 10−3. These results show that O. biennis typically exhibits a functionally asexual genetic system, but rare outcrossing events can generate novel genotypes that may have important evolutionary consequences. [ABSTRACT FROM AUTHOR]

Apomictic plants mostly occur at higher latitudes and higher elevations, where they tend to occupy deglaciated areas. By analysing samples of the alpine floras of different latitudinal zones of the Americas (Argentina, Bolivia, Ecuador, USA), we investigated whether the proportion of gametophytic apomixis is comparable between the temperate alpine zones of the Northern Hemisphere and the Southern Hemisphere and whether the proportion of apomictic species increases from the alpine tropics towards the temperate alpine zone. The reproductive systems of 196 species and 275 individuals were determined by a flow-cytometric seed screening of 999 seeds. Phylogenetic logistic regression was used to evaluate differences in the proportion of apomictic species in samples of regional alpine floras. The apomictic embryo:endosperm ratio was determined for 33 species (two pseudogamous species and 31 species with autonomous apomixis) belonging to 28 genera of 13 families. The probability of a species being apomictic did not significantly differ between the regions. The highest probability of a species being apomictic was estimated for the northern temperate zone (29.6%), followed by the tropical zone (12.5%), and the southern temperate zone (11.9%). Our results show that asexual plant reproduction by apomixis is not restricted to the alpine zone of the Northern Hemisphere and its representation in alpine floras of the Southern Hemisphere, including the alpine tropics, has been substantially underestimated. [ABSTRACT FROM AUTHOR]

Background: It is important to research in the field of germplasm protection of native and medicinal tree species in northern Iran forests, especially blood hawthorn with many medicinal properties. Sexual reproduction of hawthorn is possible with apomixis due to the thick shell and two types of physical and physiological dormancy, and its asexual reproduction by rooting cuttings faces many problems. Bloody hawthorn (Crataegus atrosanguinea from the Rosaceae family) is considered one of the valuable species of the forests of the north and west of the country, which is very important due to its fruitfulness, very strong wood, and numerous medicinal properties. The medicinal properties of hawthorn have been proven in the treatment of blood pressure, arteriosclerosis, heart congestion, heart muscle weakness, and coronary insufficiency. Hawthorn contains many secondary metabolites such as flavonoids proanthocyanidin and glycosides. Optimizing the in vitro cultivation conditions of blood hawthorn as one of the natural species of Iran can be a suitable solution for its propagation. The present research was carried out to control pollution, control browning, and induce the callus of blood hawthorn (C. atrosanguinea) in vitro. Controlling the contamination of explants, seeds, and plant tissues is of great importance for the success of plant tissue culture. Since the enrichment of natural plant explants is a priority for tissue culture, this is usually associated with many problems with browning, especially for woody plants that have a thick and woody stem. Determining the type and concentration of the growth regulator of a variable factor is necessary for the growth of plants under in vitro conditions. After applying the growth regulator to the culture medium, the plant will show different reactions, but one of the appropriate ways to choose the correct path of plant regeneration is to determine the wet and dry weights of the callus, which is widely measured as a standard of callus growth. The callus growth curve can be obtained using the wet and dry weights of the callus. The results of past research indicate a wide range of changes in the type and concentration of growth regulators on callus formation and regeneration of different species of the hawthorn genus. A specific type and concentration of growth regulators should be applied according to the type of plant species. Methods: Young and healthy stems of blood hawthorn were collected from Larijan in Amol, Mazandaran province, Iran, with geographical coordinates of 38' 58° N and 10' 52° E, at the beginning of the growing season. The collected stems were immediately transferred to the plant tissue culture laboratory of the Faculty of Natural Resources at Sari University of Agriculture and Natural Resources, Iran. Disinfection of natural hawthorn stems is very necessary due to surface and tissue contamination. For this purpose, explants of 0.5-1 cm size were cut from the stems. The infection control treatments included HgCl2, H2O2, NaClO, and AgNO3, and five treatments were designed to control the browning of explants. Browning control treatments included control, running water, ascorbic acid, polyvinyl chloride, and activated charcoal. In general, the culture medium consisted of substances including mineral salts (micro- and macroelements) as providers of maximum growth, amino acids and vitamins as nitrogen sources, sugar as a source of carbon and energy, growth regulators as growth and morphogenesis stimulants, agar as the solidifier of the cultivation medium, and water, which includes 90% of the cultivation medium. In this research, the culture medium was MS, which was first created by Driver and Koniaki (1984) and prepared according to Dufussard (1976). In this study, the pH of the culture medium was set to 5.8. For callus induction, IBA, NAA, 2,4-D, BAP, TDZ, and Kin treatments at three concentration levels of 0.1, 1, and 6 mg/L and a combination of NAA (6 mg-1) + BAP (1 mg-1), and IBA (6 mg l-1) + BAP (1 mg-1) were used in the MS culture medium. Stem explants were placed horizontally on the culture medium. Among different callus treatments, callus percentage, degree of callus, callus fresh and dry weights, and cross-sectional area of callus were measured to investigate callus induction. The best treatment was obtained by calculating the mentioned characteristics and that treatment was used for planting. It was transplanted approximately every 20 days and a total of 5-6 times. After callus cultivation in the reproduction medium, the explants were placed in a culture chamber and adaptation racks under 16 hours of light and 8 hours of dark treatment at 25 ± 2 °C with a humidity of 70%. Results: HgCl2 (0.5%, 16 minutes) with an average of 100% health was the most appropriate pollution control treatment. The control treatment with 98% health was observed as the most appropriate browning control treatment. Moreover, the type, concentration, and interaction effect of the type and concentration of the growth regulator significantly affected the callus induction characteristics of blood hawthorn (P < 0.01). The highest fresh and dry weights of callus, percentage and degree of callus formation, and cross-sectional area of callus were observed among auxin and cytokinin treatments, respectively, for 6 mg/L of NAA and 1 mg/L of BAP treatments. The highest levels of the studied callus induction characteristics were observed in the culture medium containing the combination of auxin and cytokinin (6 mg-1 of IBA+1 mg-1 of BAP). Conclusion: In general, the results showed a 90% chance of controlling the browning of the explants of natural blood hawthorn branches in a normal state and without applying any treatment. It is recommended to use 0.5% mercuric chloride for 16 minutes to disinfect the explants. The combination of growth regulators in the culture medium was evaluated as very suitable for the callus induction of bloody hawthorn. The results of this research can provide laboratory instructions for the regeneration and mass reproduction of blood hawthorn. [ABSTRACT FROM AUTHOR]

Introduction: Persian walnut (Juglans regia) is an economically important nut fruit species cultivated worldwide for its nutritious kernel and timber quality wood. Walnut trees are mostly hetero-dichogamous and, depending on the genotype, some cultivars are protogynous, while others are protandrous. Although selfing is possible when male and female blooms overlap, the dichogamy of the species promotes outcrossing. In addition to sexual reproduction, some reports indicate that elements of apomixis may occur in commercial orchards of walnut varieties and in the last two decades, nut production by apomixis has been reported in walnut. However, there are no reliable studies on the occurrence of apomictic reproduction based on cytoembryological observations and/or molecular marker-progeny tests. This study addresses the combined use of molecular and cytological analyses to gain new insights into the population genetics and reproduction systems of J. regia. Methods: We systematically analyzed the reproductive origin of individual progeny plants from 8 different cultivated walnut genotypes using microsatellite genotyping and carried out cytohistological investigations of 5 cultivated walnut genotypes arising seed sets from isolated flowers, to shed light on the mode of reproduction. Results and discussion: These cytometric and genotyping analyses did not support any asexual mode of reproduction or asexual propagation by seed and all individuals studied were identified as zygotic plants produced by crossing. Likewise, the cytological findings did not confirm completely the first component of apomixis, namely apomeiosis. On the other hand, according to histological evidence, adventitious embryony seems to take place at low frequency. Overall, our findings suggest that the occurrence of gametophytic apomixis is unlikely in J. regia, but sporophytic apomixis cannot be completely ruled out. [ABSTRACT FROM AUTHOR]

Key message: Our results indicate the existence of interploidy gene flow in Cystopteris fragilis, resulting in sexual triploid and diploid gametophytes from pentaploid parents. Similar evolutionary dynamics might operate in other fern complexes and need further investigation. Polyploidization and hybridization are a key evolutionary processes in ferns. Here, we outline an interploidy gene flow pathway operating in the polyploid Cystopteris fragilis complex. The conditions necessary for the existence of this pathway were tested. A total of 365 C. fragilis individuals were collected covering representatives of all three predominant ploidy levels (tetraploid, pentaploid, and hexaploid), cultivated, had their ploidy level estimated by flow cytometry, and their spores collected. The spores, as well as gametophytes and sporophytes established from them, were analysed by flow cytometry. Spore abortion rate was also estimated. In tetraploids, we observed the formation of unreduced (tetraploid) spores (ca 2%). Collected pentaploid individuals indicate ongoing hybridization between ploidy levels. Pentaploids formed up to 52% viable spores, ca 79% of them reduced, i.e. diploid and triploid. Reduced spores formed viable gametophytes, and, in the case of triploids, filial hexaploid sporophytes, showing evidence of sexual reproduction. Some tetraploid sporophytes reproduce apomictically (based on uniform ploidy of their metagenesis up to filial sporophytes). Triploid and diploid gametophytes from pentaploid parents are able to mate among themselves, or with "normal" reduced gametophytes from the sexual tetraploid sporophytes (the dominant ploidy level in the sporophytes in this populations), to produce tetraploid, pentaploid, and hexaploid sporophytes, allowing for geneflow from the pentaploids to both the tetraploid and hexaploid populations. Similar evolutionary dynamics might operate in other fern complexes and need further investigation. [ABSTRACT FROM AUTHOR]

Abstract Plant species respond to varying plant species diversity and associated changes in their abiotic and biotic environment with changes in their phenotype. However, it is not clear to what degree this phenotypic differentiation is due to genotype diversity within populations or phenotypic plasticity of plant individuals. We studied individuals of 16 populations of the clonal herb Taraxacum officinale grown in plant communities of different species richness in a 17‐year‐old grassland biodiversity experiment (Jena Experiment). We collected 12 individuals in each population to measure phenotypic traits and identify distinct genotypes using microsatellite DNA markers. Plant species richness did not influence population‐level genotype and trait diversity. However, it affected the expression of several phenotypic traits, e.g. leaf and inflorescence number, maximum leaf length and seed mass, which increased with increasing plant species richness. Moreover, population‐level trait diversity correlated positively with genotype richness for leaf dry matter content (LDMC) and negatively with inflorescence number. For several traits (i.e. seed mass, germination rate, LDMC, specific leaf area (SLA)), a larger portion of variance was explained by genotype identity, while variance in other traits (i.e. number of inflorescences, leaf nitrogen concentration, leaf number, leaf length) resided within genotypes and thus was mostly due to phenotypic plasticity. Overall, our findings show that plant species richness positively affected the population means of some traits related to whole‐plant performance, whose variation was achieved through both phenotypic plasticity and genotype composition of a population.

Polyploid species of Hieracium s.str. are near-obligatory apomicts capable of producing non-reduced pollen grains and intrageneric crossing with sexual diploid species. Most probably, such a mating system leads to the emergence of separate lineages of apomictic plants with increased ploidy levels, each of which has a complete set of genomes of its putative (most probably triploid) apomictic parent. The correct identification of such lineages could facilitate taxonomic and phylogenetic research in Hieracium . In this paper, we analyzed the distribution of the 5S and 45S rDNA segments in the karyotypes of six selected Hieracium taxa, representing all ploidy levels reported in the genus: H. alpinum 2x, H. alpinum 3x, H. schustleri 4x, H. chrysostyloides 5x ( H . sect. Alpina ) and H. bifidum 3x, H. levicaule 3x ( H . sect. Bifida ). The analyzed rDNA markers suggest that two taxa of the higher ploidy level (4x and 5x) belonging to H . sect. Alpina inherited three genomes from the triploid H. alpinum , which has 2:1 genomic composition with two genomes from diploid H. alpinum and one genome with an untypical 5S rDNA-bearing chromosome of unknown origin. H. bifidum and H. levicaule differ from each other in the rDNA distribution pattern, which suggests that H . sect. Bifida may be a less homogenous group of species.

Introducing asexual reproduction through seeds - apomixis - into crop species could revolutionize agriculture by allowing F1 hybrids with enhanced yield and stability to be clonally propagated. Engineering synthetic apomixis has proven feasible in inbred rice through the inactivation of three genes (MiMe), which results in the conversion of meiosis into mitosis in a line ectopically expressing the BABYBOOM1 (BBM1) parthenogenetic trigger in egg cells. However, only 10-30% of the seeds are clonal. Here, we show that synthetic apomixis can be achieved in an F1 hybrid of rice by inducing MiMe mutations and egg cell expression of BBM1 in a single step. We generate hybrid plants that produce more than 95% of clonal seeds across multiple generations. Clonal apomictic plants maintain the phenotype of the F1 hybrid along successive generations. Our results demonstrate that there is no barrier to almost fully penetrant synthetic apomixis in an important crop species, rendering it compatible with use in agriculture.

Maize (Zea mays L.) is one of the most demanded grain crops in the world. Currently, production has exceeded one billion tons and is increasing by 3–5% annually. Such growth is due to the genetic potential of the crop and the use of heterosis F1 hybrids in production. However, the need to produce first-generation seed annually poses significant challenges and is an economically costly technology. A solution to this problem may be the transfer of the asexual (apomictic) mode of reproduction to maize from its wild relative, eastern gamagrass (Tripsacum dactyloides L.). In this work, we report the production of 56-chromosome apomictic hybrids of maize (Zea mays L.) with eastern gamagrass (T. dactyloides L.) with restored anther fertility. The mode of reproduction of the plant was confirmed by counting chromosomes and sequencing the nuclear gene (Pox3) and chloroplast tRNA-Leu (trnL) gene. These apomictic hybrids had karyotypes of 2n = 56 = [(10Zm(573MB) + 36Td) + 10Zm(611CB)] and 2n = 56 = [(10Zm(611CB) + 36Td) + 10Zm(611CB)]. The resulting hybrids can be widely used as a fodder crop.

This review considers the processes of morphogenesis used in the development of propagation methods and the creation of a new starting material for sugar beet. It has been demonstrated that methods of particulation, in vitro microcloning and cell breeding that reflect non-sexual forms of plant reproduction increase the effectiveness of breeding experiments. The review describes the in vitro culture methods maintaing a tendency in plants for vegetative propagation and stimulating increase in genetic variability of properties when mutagens such as ethyl methanesulfonate, alien genetic structures with mf2 and mf3 bacterial genes in Agrobacterium tumefaciens strains, and selective agents (Сd++ ions and abscisic acid) are incorporated into plant cells. It presents the results of using fluorescent microscopy, cytophotometry, biochemical analysis and determining the level of phytohormones and content of nucleic acids in nuclei for forecasting the seed setting ability. It has demonstrated that long self-pollination of plants causes decrease in fertility of pollen grains, resulting in the sterilization of male gametes and the appearance of pistillody flowers. Self-fertile plants isolated from these lines serve as sterility fixers, while the apomixis elements increased the ovule number, additional embryo sacs and embryos. A role of apomixis in contributing to variability in the ontoand phylogenetic development of plants have been substantiated. The review reflects the morphological features of the in vitro development of sexual and somatic cells in embryos during the formation of seedlings based on floral and vegetative embryoidogeny. Use of the SNP and SSR (Unigenes) molecular-genetic markers having a high polymorphism level has appeared effective to characterize the developed breeding material and hybrid components when carrying out crossings. The study of sugar beet starting materials for the presence of TRs mini-satellite loci making it possible to reveal O-type plants-pollinators (sterility fixing agent) and MS-form plants are of interest for breeding as well. The selected material can be widely used in breeding to produce hybrids, allowing for a 2–3-fold reduction of the development period. The review also discusses the prospects for the development and implementation of new methods and original schemes in sugar beet genetics, biotechnology and breeding.

IntroductionAmong candidate genes underlying the control components of apomixis, APOLLO is known for its strong linkage to apomeiosis in the genus Boechera. The gene has “apo alleles,” which are characterized by a set of linked apomixis-specific polymorphisms, and “sex alleles.” All apomictic Boechera genotypes are heterozygous for the apo/sex alleles, whereas all sexual genotypes are homozygous for sex alleles.MethodsIn this study, native and synthetic APOLLO promoters were characterized by detecting the expression level of the β-glucuronidase (GUS) gene in Arabidopsis.ResultsComparing various flower developmental stages in transgenic lines containing different constructs with 2-kb native transgenic lines revealed that changes to the APOLLO promoter causes shifts in tissue and developmental stage specificity of GUS expression. Importantly, several apomixis-specific polymorphisms in the 5′UTR change the timing and location of GUS activity from somatic to reproductive tissues.DiscussionThese synthetic data simulate a plausible evolutionary process, whereby apomixis-specific gene activity can be achieved.