Production Of Viable Pollen Research Articles

Jasmonic Acid Plays a Pivotal Role in Pollen Development and Fertility Regulation in Different Types of P(T)GMS Rice Lines.

Two-line hybrid rice systems represent a new technical approach to utilizing the advantages of rice hybrids. However, the mechanism underlying the male sterile-line fertility transition in rice remains unclear. Peiai 64S (PA64S) is a photoperiod- and thermo-sensitive genic male sterile (PTGMS) line in which male sterility manifests at an average temperature above 23.5 °C under long-day (LD) conditions. Nongken 58S (NK58S) is a LD-sensitive genic male sterile (PGMS) rice that is sterile under LD conditions (above 13.75 h-day). In contrast, D52S is a short-day (SD)-PGMS line that manifests male sterility under SD conditions (below 13.5 h-day). In this study, we obtained fertile and sterile plants from all three lines and performed transcriptome analyses on the anthers of the plants. Gene ontology (GO) analysis suggested that the differentially expressed genes identified were significantly enriched in common terms involved in the response to jasmonic acid (JA) and in JA biosynthesis. On the basis of the biochemical and molecular validation of dynamic, tissue-specific changes in JA, indole-3-acetic acid (IAA) levels, gibberellin (GA) levels, and JA biosynthetic enzyme activities and expression, we proposed that JA could play a pivotal role in viable pollen production through its initial upregulation, constant fluctuation and leaf-spikelet signaling under certain fertility-inducing conditions. Furthermore, we also sprayed methyl jasmonate (MEJA) and salicylhydroxamic acid (SHAM) on the plants, thereby achieving fertility reversal in the PGMS lines NK58S and D52S, with 12.91–63.53% pollen fertility changes. Through qPCR and enzyme activity analyses, we identified two key enzymes—allene oxide synthase (AOS) and allene oxide cyclase (AOC)—that were produced and upregulated by 20–500-fold in PGMS in response to spraying; the activities of these enzymes reversed pollen fertility by influencing the JA biosynthetic pathway. These results provide a new understanding of hormone interactions and networks in male-sterile rice based on the role of JA that will help us to better understand the potential regulatory mechanisms of fertility development in rice in the future.

Selection based on meiotic behavior in <i>Urochloa decumbens</i> hybrids from non-shattered seed

This study aimed to evaluate the end-products of meiosis in sexual and apomictic hybrids of Urochloa decumbens, so as to identify genotypes with good production of viable pollen for use in breeding programs to increase yields of pure viable seed and reduce degree of seed shattering. From 457 intraspecific hybrids of U. decumbens arising from crosses between 3 artificially tetraploidized sexual plants and the apomictic cultivar Basilisk, 27 hybrids from non-shattered seed were selected. Slides were prepared by smearing anthers and staining to determine the presence of abnormalities. The abnormalities found were micronuclei, microcytes and polyads. The data were compared by the Scott-Knott test at P<0.05. Data obtained enabled separation of hybrids into 4 groups depending on the presence of micronuclei and formation of polyads and into 6 groups based on the presence of microcytes in the tetrads. Among the analyzed apomictic hybrids, R179 has the attributes for viable seed production to proceed to cultivar development. Among the sexual hybrids, R161, R181, R193 and S47 are recommended as female parents for use in crossing programs.

Development of a genotype-specific molecular marker associated with restoration-of-fertility (Rf) in chile pepper (Capsicum annuum)

Cytoplasmic male sterility (CMS) breeding systems expedite the production of hybrid seed in many crops. By taking advantage of an interaction between the nucleus and the cytoplasm that regulates the production of viable pollen, CMS eliminates the need for manual emasculation. Markers that can identify the status of cytoplasmic male sterility and nuclear restorer-of-fertility (Rf) genes are valuable for identification and development of CMS systems. In chile pepper (Capsicum annuum), reliable markers for the Rf trait have been elusive. To identify viable Rf markers for chile pepper, markers were created using identified Rf candidate genes and compared them to the most effective previously reported Rf markers. The previously reported cleaved amplified polymorphic sequence (CAPS) marker Co1Mod1-CAPS and PPR-derived CaRf648, had the highest rates of correctly predicting the Rf phenotype for an F2 segregating population. The Co1Mod1-CAPS marker detected the Rf genotype, and CaRf648 quickly and inexpensively identified Rf presence.

3D Imaging of Tapetal Mitochondria Suggests the Importance of Mitochondrial Fission in Pollen Growth.

Mitochondrial fission occurs frequently in plant cells, but its biological significance is poorly understood because mutants specifically impaired in mitochondrial fission do not show obvious defects in vegetative growth. Here, we revealed that the production of viable pollen was reduced in mutants lacking one of the three main proteins involved in mitochondrial fission in Arabidopsis (Arabidopsis thaliana), DYNAMIN-RELATED PROTEIN3A (DRP3A)/Arabidopsis DYNAMIN-LIKE PROTEIN2A, DRP3B, and ELONGATED MITOCHONDRIA1 (ELM1). In drp3b and elm1, young microspores contained an abnormal number of nuclei, and mature pollen had aberrant accumulation of lipids in their coat and an irregular pollen outer wall. Because the formation of the pollen wall and coat is mainly associated with tapetal function, we used 3D imaging to quantify geometric and textural features of cells and mitochondria in the tapetum at different stages, using isolated single tapetal cells in which the in vivo morphology and volume of cells and mitochondria were preserved. Tapetal cells and their mitochondria changed in the volume and morphology at different developmental stages. Defective mitochondrial fission in the elm1 and drp3b mutants caused changes in mitochondrial status, including mitochondrial elongation, abnormal mitochondrial ultrastructure, a decrease in cross-sectional area, and a slight alteration of mitochondrial distribution, as well as a large reduction in mitochondrial density. Our studies suggest that mitochondrial fission is required for proper mitochondrial status in the tapetum and possibly in pollen as well and therefore plays an important role for the production of viable pollen.

Developmental role of the tomato Mediator complex subunit MED18 in pollen ontogeny.

Pollen development is a crucial step in higher plants, which not only makes possible plant fertilization and seed formation, but also determines fruit quality and yield in crop species. Here, we reported a tomato T-DNA mutant, pollen deficient1 (pod1), characterized by an abnormal anther development and the lack of viable pollen formation, which led to the production of parthenocarpic fruits. Genomic analyses and the characterization of silencing lines proved that pod1 mutant phenotype relies on the tomato SlMED18 gene encoding the subunit 18 of Mediator multi-protein complex involved in RNA polymerase II transcription machinery. The loss of SlMED18 function delayed tapetum degeneration, which resulted in deficient microspore development and scarce production of viable pollen. A detailed histological characterization of anther development proved that changes during microgametogenesis and a significant delay in tapetum degeneration are associated with a high proportion of degenerated cells and, hence, should be responsible for the low production of functional pollen grains. Expression of pollen marker genes indicated that SlMED18 is essential for the proper transcription of a subset of genes specifically required to pollen formation and fruit development, revealing a key role of SlMED18 in male gametogenesis of tomato. Additionally, SlMED18 is able to rescue developmental abnormalities of the Arabidopsis med18 mutant, indicating that most biological functions have been conserved in both species.

Water deficit disrupts male gametophyte development in Quercus ilex

Tree species distribution, and hence forest biodiversity, relies on the reproductive capacity of trees, which is currently affected by climate change. Drought-induced pollen sterility could increase as a consequence of more intense and more frequent droughts projected for temperate and Mediterranean regions, and threaten the sexual regeneration of trees in these regions. To evaluate this possibility, we examined the effect of long-term partial rainfall exclusion (-27% precipitation) on male reproductive development in holm oak, Quercus ilex, one of the most important and widespread tree species of the Mediterranean region. We examined anther area, pollen production, pollen abortion as well as viable pollen production in control and dry treatments. Microscopic examinations revealed significant differences in pollen development between trees in the dry and the control treatments, even though anthesis occurred before the onset of annual drought. Our results demonstrate that anthers collected from Q.ilex trees in the dry treatment, which experienced long-term increased drought stress especially during the summer, were the same size as anthers in the control treatment, but displayed 25% pollen abortion and almost 20% reduction in pollen production. Subsequently, the number of viable pollen grains in anthers from dry treatment was 35% less than in control. These results suggest a carry-over effect of drought stress on pollen production that could reduce the reproductive success of Q. ilex. The results have broad implications for better understanding of the determinants of tree reproduction by masting and anticipate the outcomes of expected drought increase in the Mediterranean on forest dynamics.

Meiotic chromosome behavior of the male-fertile allotriploid lily cultivar 'Cocossa'.

Cytological observations of microsporogenesis in the allotriploid lily cultivar 'Cocossa' showed that viable pollen production could be attributed mainly to disoriented spindles, abnormal cytokinesis, and cytomixis during male meiosis. To identify the reasons why the allotriploid lily cultivar 'Cocossa' can produce aneuploid and euploid functional male gametes and can be used as the paternal parent in lily introgression breeding, we performed a detailed investigation of microsporogenesis using the conventional cytological methods. The allotriploid not only produced single pollen grains with variable sizes but also produced adherent pollen grains. Pollen viability was estimated at 50.1% based on staining and 30.8% based on germination. Based on the chromosomal analysis of BC2 plants derived from Oriental cultivars (♀) crossed with the OOT cultivar 'Cocossa' (♂), it was concluded that the objective allotriploid contributed haploid (x), diploid (2x), and aneuploid chromosome complements. Common meiotic abnormalities were observed, indicating the high genetic imbalance of this allotriploid. In addition to normally oriented metaphase II spindles (linear and perpendicular), abnormal spindles, such as parallel, tripolar, fused, and multiple spindles, accounted for 6.21, 6.41, 14.27, and 1.17%, respectively. Tripolar and fused spindles resulted in the production of triads and dyads, which contributed to unreduced pollen production. Some microsporocytes exhibited complete or partial absence of cytokinesis, which led to relatively high frequencies of monads, dyads, and triads. Furthermore, the phenomenon of cytomixis during microsporogenesis occurred mainly in the first meiotic prophase and early development of pollen grains, which we assume is a possible cause of unreduced gamete generation. Our study offers a new resource for lily introgression breeding.

Influence of Soil Moisture and Air Temperature on the Stability of Cytoplasmic Male Sterility (CMS) in Maize (<i>Zea mays</i> L.)

Cytoplasmic male sterility (CMS) is a maternally inherited trait that suppresses the production of viable pollen. CMS is a useful biological tool for confinement strategies to facilitate coexistence of genetically modified (GM) and non-GM crops in case where it is required. The trait is reversible and can be restored to fertility in the presence of nuclear restorer genes (Rf genes) and by environmental impacts. The aim of this study was to investigate the influence of the level of irrigation on the stability of CMS maize hybrids under defined greenhouse conditions. Additionally the combination of irrigation and air temperature was studied. Three CMS maize hybrids were grown with different levels of irrigation and in different temperature regimes. Tassel characteristics, pollen production and fertility were assessed. The CMS stability was high in hot air temperatures and decreased in lower temperatures. The level of irrigation had no major effect on the level of sterility. The extent of these phenomena was depending on the genotype of CMS maize and should be known before using CMS for coexistence purposes.

In vivo functional analysis of a nuclear restorer PPR protein.

BackgroundNuclear restorers of cytoplasmic male fertility (CMS) act to suppress the male sterile phenotype by down-regulating the expression of novel CMS-specifying mitochondrial genes. One such restorer gene is Rfo, which restores fertility to the radish Ogura or ogu CMS. Rfo, like most characterized restorers, encodes a pentatricopeptide repeat (PPR) protein, a family of eukaryotic proteins characterized by tandem repeats of a 35 amino acid motif. While over 400 PPR genes are found in characterized plant genomes and the importance of this gene family in organelle gene expression is widely recognized, few detailed in vivo assessments of primary structure-function relationships in this protein family have been conducted.ResultsIn contrast to earlier studies, which identified 16 or 17 PPR domains in the Rfo protein, we now find, using a more recently developed predictive tool, that Rfo has 18 repeat domains with the additional domain N-terminal to the others. Comparison of transcript sequences from pooled rfo/rfo plants with pooled Rfo/Rfo plants of a mapping population led to the identification of a non-restoring rfo allele with a 12 bp deletion in the fourth domain. Introduction into ogu CMS plants of a genetic construct in which this deletion had been introduced into Rfo led to a partial loss in the capacity to produce viable pollen, as assessed by vital staining, pollen germination and the capacity for seed production following pollination of CMS plants. The degree of viable pollen production among different transgenic plants roughly correlated with the copy number of the introduced gene and with the reduction of the levels of the ORF138 CMS-associated protein. All other constructs tested, including one in which only the C-terminal PPR repeat was deleted and another in which this repeat was replaced by the corresponding domain of the related, non-restoring gene, PPR-A, failed to result in any measure of fertility restoration.ConclusionsThe identification of the additional PPR domain in Rfo indicates that the protein, apart from its N-terminal mitochondrial targeting presequence, consists almost entirely of PPR repeats. The newly identified rfo allele carries the same 4 amino acid deletion as that found in the neighboring, related, non-restoring PPR gene, PPR-A. Introduction of this four amino acid deletion into a central domain the Rfo protein, however, only partially reduces its restoration capacity, even though this alteration might be expected to alter the spacing between the adjoining repeats. All other tested alterations, generated by deleting specific PPR repeats or exchanging repeats with corresponding domains of PPR-A, led to a complete loss of restorer function. Overall we demonstrate that introduction of targeted alterations of Rfo into ogu CMS plants provides a sensitive in vivo readout for analysis of the relationship between primary structure and biological function in this important family of plant proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0313-4) contains supplementary material, which is available to authorized users.

Anther and pollen development in the lodgepole pine dwarf mistletoe (Arceuthobium americanum) staminate flower

The lodgepole pine dwarf mistletoe, Arceuthobium americanum Nutt. ex Engelm., is a parasitic angiosperm that infects conifers in western Canadian forests. While production of viable pollen in anthers is critical to dwarf mistletoe reproduction, the few existing reports that examine staminate development in Arceuthobium are often incomplete or conflicting. The objective of this work was to investigate the developmental anatomy of anther and pollen of A. americanum using modern microscopy. We found that the microsporangium was toroidal from the outset and gave rise to a central peg-shaped sterile “columella” early in anther development. The endothecium was absent, the epidermis persisted as an “exothecium” fulfilling the role of the endothecium, and a primary parietal layer generated a secretory tapetum and middle layer. Thus, we suggest that a new category of anther wall development, the Arceuthobium type, be created. Microsporogenesis produced tetrahedral microspores via simultaneous cytokinesis and involved callose wall formation. Microgametogenesis resulted in round and atypical generative and vegetative nuclei. Additionally, the heterocolpate, echinate pollen grains, which were shed at the two-celled stage, were seen for the first time in their native state with environmental scanning electron microscopy. This work contributes to the understanding of A. americanum, the genus Arceuthobium, and angiosperms as a whole.

Production of a new generation of seeds through the use of somatic clones in controlled crosses of black spruce (Picea mariana)

To assess the potential to integrate somatic clones (SC) of desired characteristics in production of high genetic quality seed, controlled crosses between different SCs of black spruce (Picea mariana (Mill) B.S.P.) were used to assess their suitability for the production of viable pollen, cones, seeds and seedlings. These SC produced male and female strobili at an early stage. Pollen, cones and seeds produced were characterized (mass, size, germination); their characteristics were similar to those produced by trees in natural forests or seed orchards. A maternal effect was demonstrated for the cone size and seed mass. Although seeds had excellent germination rates, the somatic biparental crosses were divided into three distinct groups with different germination curves using the Weibull function. Seeds from controlled crosses between different SC enabled the production of high morpho-physiological quality seedlings in a forest nursery. Using black spruce as a model, we showed, for the first time, that SC can be used as seed producers. These encouraging results open new perspectives on the tangible integration of somatic embryogenesis (SE) in the chain of seed, vegetative propagation (cuttings and SE) and production of plants for high productivity plantations. Controlled crosses can be made between SC with the desired characteristics (fewer large branches, fewer nodes, good growth, high wood density, performance, improved yield, etc.), vegetative propagules produced and deployed to clonal tests. After elimination of the worst performing SC, clonal tests can be converted into seed orchards that produce a new generation of seeds of high genetic quality. This will allow the rapid introduction of new materials in elite breeding programs of forest species.

In vitro flowering and production of viable pollen of cucumber

Flowers were produced on sterile cucumber (Cucumis sativus L.) plants grown in vitro from seed and micropropagated shoots from stem fragments. The highest numbers of flowers on plants from both sources were produced on Murashige and Skoog (MS) medium without plant growth regulators (PGR), as well as with 6 μM of kinetin (Kin). Plants cultured on MS medium supplemented with 8.9 μM benzyladenine (BA) and 1.1 μM 1-naphthaleneacetic acid (NAA) did not flower. In vitro grown plants produced fewer, smaller flowers compared with greenhouse-grown plants. Male and female flowers developed on plants grown in vitro from seed and were morphologically similar to flowers on greenhouse grown plants. Micropropagated shoots produced male flowers with altered morphology. The highest viability (72.9 ± 4.2%) and germination (69.5 ± 4.1%) of pollen were observed for plants grown from seed on MS medium supplemented with 6 μM Kin. Cytological observations of meiosis in anthers of male flowers from in vitro grown plants revealed abnormalities, such as monads, dyads, triads, polyads, microcytes and degeneration of tetrads, causing reduced viability and germination of pollen. The fewest meiotic irregularities in pollen mother cells were observed in plants grown on MS medium that was PGR-free (12.1 ± 0.9%) or with 6 μM Kin (20.9 ± 1.7%).

Viability, Growth, and Fertility of Knotweed Cytotypes in North America

AbstractHybridization between two introduced plant species can influence the invasion capabilities of the exotic taxa, but the role of hybridization will likely differ in different invasions, even of the same species. Until now, studies concerning the ploidy of Japanese knotweed, giant knotweed, and their hybrids have been conducted in Europe or native ranges in Asia. Here, we assess the role of hybridization and ploidy in a U.S. invasion. We use flow cytometry to characterize DNA content in (1) established families in a common garden, (2) seedlings grown from common garden parents, and (3) wild populations. We also measured fertility in the garden and the field and vegetative growth traits in the garden. Although the majority of our parental and hybrid samples had ploidy levels previously documented in Europe (4X and 8X for parental species; 6X for hybrids), we found a wider range of knotweed cytotypes established in our garden (4X, 6X, 7X, 8X, 9X, and 10X) and additionally detected 5X, 11X, 12X, and possibly 14X ploidy levels in progeny from garden seed parents. The unexpected cytotypes were not confined to the greenhouse or common garden, in that all < 11X ploidy levels were also found in field populations in Massachusetts. In several cases, these data contradicted our expectations on the basis of morphological and molecular analysis, suggesting both significant introgression and the introduction of multiple cytotypes from Asia. With one exception (14X), we found all cytotypes were capable of strong vegetative growth, seed set, and the production of viable pollen. Without barriers to sexual reproduction, introgression is expected to progress, creating a progressively more diverse swarm of invasive genotypes.

Pollen morphology and viability of some Indian mangroves

A study on pollen morphology of four common Indian mangroves from Sundarbans mangrove swamps, three species from the family Rhizophoraceae (Bruguiera sexangula Laur., Ceriops tagal Perr.and Rhizophora mucronata Lamk.) and one species from the family Avicenniaceae (Avicennia marina Forsk.) was carried out. Shape of the pollen grains were prolate, prolate spheroidal and subprolate with tricolporate apertures. Surface ornamentation of the studied taxa was reticulate, finely reticulate or scabrate. Pollen grain viability of the said taxa was determined by stain test (1% aceto-orcein) under in situ condition and percentage of viable pollen production was recorded. The result revealed that all the studied taxa produce a fairly good amount of viable pollen grains in natural condition, an indication that they are good propagule producer in the long run. Pollen grain germination and measurements of pollen tube length using different sucrose concentrations (0.2, 0.6, 1.0, 1.2, 1.6 and 2.0%). Maximum pollen fertility and pollen tube length occurred in 2% sucrose solution in case of A. marina and B. sexangula, whereas 1.6% in C. tagal and 1.0% in R. mucronata. A strong positive correlation exist between pollen germination and pollen tube length, but there was a hardly relation between sucrose concentration and pollen tube length.

Sexual dimorphism in the andromonoecious Euphorbia nicaeensis: effects of gender and inflorescence development.

In andromonoecious taxa with separate floral types along the inflorescence, architectural or plastic effects can simulate floral sexual dimorphism. Both the primary and secondary sexual characteristics of the cyathia of the protogynous andromonoecious species Euphorbia nicaeensis were analysed according to their sex and arrangement on the inflorescence. The production of male and hermaphrodite cyathia at each inflorescence level was surveyed in two natural populations. The longevity, size, pollen production and viability, and nectar secretion of both types of cyathia were checked between inflorescence levels and between sexes at the only level at which they occur together. This sampling method makes it possible to know whether differences between cyathia types are based on sex or are attributable to inflorescence development. Male cyathia were produced predominantly at the first and second inflorescence levels, whereas at levels 3-5, the cyathia were almost exclusively hermaphrodite. Viable pollen production by male cyathia at the second inflorescence level was higher than that of hermaphrodite cyathia at the third level but, when males and hermaphrodites at the same level were compared, their pollen production was similar. Male and hermaphrodite cyathia were similar in size, irrespective of the inflorescence level, although the exclusively hermaphrodite cyathia of the last level were smaller. Both cyathium types produced similar amounts of sugar. However, male cyathia produced nectar during their whole lifespans, whereas hermaphrodites produced it exclusively during their male phase. Moreover, the nectary activity of male cyathia started earlier in the day than that of hermaphrodites. An apparent floral dimorphism exists in the primary sexual characteristics of Euphorbia nicaeensis because differences in pollen production between cyathium types are due to theirs positions. Similarly, differences affecting most secondary sexual characteristics are only apparent between the two cyathium types. However, E. nicaeensis shows a true but slight floral dimorphism in some of the secondary sex characters related to nectar secretion. The lack of nectar production by the female phase of the hermaphrodite cyathia of E. nicaeensis indicates that this is a deceit-pollinated species.

Gibberellin Regulates Pollen Viability and Pollen Tube Growth in Rice

Gibberellins (GAs) play many biological roles in higher plants. We collected and performed genetic analysis on rice (Oryza sativa) GA-related mutants, including GA-deficient and GA-insensitive mutants. Genetic analysis of the mutants revealed that rice GA-deficient mutations are not transmitted as Mendelian traits to the next generation following self-pollination of F1 heterozygous plants, although GA-insensitive mutations are transmitted normally. To understand these differences in transmission, we examined the effect of GA on microsporogenesis and pollen tube elongation in rice using new GA-deficient and GA-insensitive mutants that produce semifertile flowers. Phenotypic analysis revealed that the GA-deficient mutant reduced pollen elongation1 is defective in pollen tube elongation, resulting in a low fertilization frequency, whereas the GA-insensitive semidominant mutant Slr1-d3 is mainly defective in viable pollen production. Quantitative RT-PCR revealed that GA biosynthesis genes tested whose mutations are transmitted to the next generation at a lower frequency are preferentially expressed after meiosis during pollen development, but expression is absent or very low before the meiosis stage, whereas GA signal-related genes are actively expressed before meiosis. Based on these observations, we predict that the transmission of GA-signaling genes occurs in a sporophytic manner, since the protein products and/or mRNA transcripts of these genes may be introduced into pollen-carrying mutant alleles, whereas GA synthesis genes are transmitted in a gametophytic manner, since these genes are preferentially expressed after meiosis.

MALE STERILITY1 Is Required for Tapetal Development and Pollen Wall Biosynthesis

The Arabidopsis thaliana MALE STERILITY1 (MS1) gene is critical for viable pollen formation and has homology to the PHD-finger class of transcription factors; however, its role in pollen development has not been fully defined. We show that MS1 transcription appears to be autoregulated by the wild-type MS1 transcript or protein. Using a functional green fluorescent protein (GFP) fusion to analyze the temporal and spatial expression of MS1, we demonstrate that the MS1:GFP protein is nuclear localized within the tapetum and is expressed in a developmentally regulated manner between late tetraspore and microspore release, then rapidly breaks down, probably by ubiquitin-dependent proteolysis. Absence of MS1 expression results in changes in tapetal secretion and exine structure. Microarray analysis has shown that 260 (228 downregulated and 32 upreglated) genes have altered expression in young ms1 buds. These genes are primarily associated with pollen wall and coat formation; however, a number of transcription factors and Cys proteases have also been identified as the putative primary regulatory targets of MS1. Ectopic expression of MS1 alters transcriptional regulation of vegetative gene expression, resulting in stunted plants with increased levels of branching, partially fertile flowers and an apparent increase in wall material on mature pollen. MS1 therefore plays a critical role in the induction of pollen wall and pollen coat materials in the tapetum and, ultimately, the production of viable pollen.

Sexual reproduction of cordgrass hybrids (Spartina foliosa x alterniflora) invading tidal marshes in San Francisco Bay

ABSTRACTWe genetically analysed cordgrass plants and seedlings throughout the San Francisco, California, USA, estuary and found that hybrids between exotic Spartina alterniflora and native Spartina foliosa are the principal cordgrass invaders and colonizers. We hypothesized that this was due to higher seed set and siring ability by hybrids relative to the native species; too few alien parents remained in San Francisco Bay for our comparative studies. Hybrid seed comprised 91% to 98% of that set in the marsh study plants over the 2 years of the study. Total viable pollen production by hybrid plants was 400 times that of the native plants. Seed and pollen production were highly skewed towards a few hybrid genotypes. In addition to seed produced by hybrid plants, hybrid seed was produced by S. foliosa due to hybrid backcrossing. While the greatest advantage for hybrids was in pollen and seed production, hybrid seeds germinated, and seedlings survived and grew as well or better than the native species. As native S. foliosa becomes increasingly rare, hybrid seed floating on the tides will predominate, overwhelming recruitment sites and resulting in further colonization by hybrids. In an evolutionary context, hybrids with exceptional pollen and seed production will be initially favoured by natural selection, leading to the evolution of even more fertile hybrid genotypes.

INDUCTION OF 2N GAMETES FOR OVERCOMING F1-STERILITY IN LILY AND TULIP

For overcoming F1-sterility in interspecific hybrids, mitotic and meiotic polyploidisation is applied in lily and can result in fertile allopolyploids. The mechanism of viable pollen production of mitotic and meiotic polyploidisation is quite different. Mitotic polyploids are obtained by artificial chromosome doubling and results in normal pairing of homologous chromosome set, which enables the formation of 2x gametes during the meiosis. Meiotic polyploidisation, on the other hand, leads in rare cases of restitutional chromosome division, to the formation of unreduced gametes (2n gametes). In contrast to mitotic doubling, homoeologous recombination can occur in these gametes. Genomic in situ hybridization (GISH) was used to discriminate parental chromosomes in the F1 hybrids and backcross progenies to detect homoeologous recombination. Mitotic polyploidisation showed no homoeologous recombinations between the parental genomes whereas in meiotic polyploids it was detected in a high frequency. The use of 2n gametes is therefore the most promising approach for the introgression of desirable characters in using interspecific hybrids in breeding. In both cases, the frequency of viable gametes appeared to be low and limited to a few F1-hybrids. A new and promising method is developed in which 2n-gametes can be induced by the application of laughing gas (N2O). This method proved to be successful in lily and the occurrence of homoeologous recombination, a characteristic of meiotic polyploidisation, was detected. In tulip where the production of mitotic polyploids requires at least five years, it will speed up breeding with interspecific hybrids enormously

INTERSPECIFIC HYBRIDISATION AND POLYPLOIDISATION AS TOOLS IN ORNAMENTAL PLANT BREEDING

Interspecific hybridisation and polyploidy are recognized as the most impor-tant sources of evolution and domestication of flowering plants. In ornamental plant breeding these phenomena go hand in hand and can be observed in the breeding his-tory of many ornamental crops (Rosa, Chrysanthemum, Gladiolus, Alstroemeria, Lil-ium, orchids etc). With lily as model crop techniques developed for overcoming pre- and post-fertilisation barriers are reviewed. For overcoming F1-sterility mitotic and meiotic polyploidisation are applied and can result in fertile allopolyploids. The mechanism of viable pollen production of mitotic and meiotic polyploidisation is quite different. Mitotic polyploidisation possess one homologous chromosome set. They undergo normal meiotic division like diploid cells. However, meiotic poly-ploidisation often show irregular chromosome division resulting in two of unreduced chromosome number instead of reduced chromosome number in tetrads. In contrast to mitotic doubling homoeologous recombination can occur. There are two signifi-cant mechanisms, FDR and SDR, for the formation of 2n-gametes. The FDR-gamete increase heterozygosity while SDR-gamete increase homozygosity. Genomic in situ hybridisation (GISH) has been used to discriminate parental chromosomes and to detect homoeologous recombination. Mitotic polyploidisation showed no homoeolo-gous recombinations between the parental genomes whereas in meiotic polyploids it can detected frequently. The use of 2n-gametes is therefore the most promising way for the introgression of desirable characters in the breeding with interspecific hy-brids. Although spontaneous occurring in the domestication of many ornamental crops the systematically detected unreduced gametes proved to be highly efficient tool for introgression of characters.