Calcium changes in ovules and embryo sacs of Plumbago zeylanica L.

Calcium was localized in ovules of Plumbago zeylanica from 1 day before anthesis to 3 days after anthesis using potassium antimonate and transmission electron microscopy in pollinated and emasculated flowers. At 1 day before anthesis, embryo sacs (containing an egg cell, a central cell and zero to three accessory cells) appear mature and contain abundant calcium precipitates (ppts), in contrast to nucellar cells. At anthesis, the vacuoles of nucellar cells have enlarged, and micropylar cells, in particular, are heavily labeled with calcium ppts. As pollen tubes elongate through ovular tissues, ppts diminish in ovular cells and become concentrated in the pollen tube cell wall. After fertilization, the calcium ppts sharply diminish in fertilized ovules; in unfertilized ovules, calcium ppts remain abundant up to 3 days after anthesis (when unfertilized ovules are shed). The distribution of calcium in the ovule changes in apparent response to fertilization, suggesting that calcium content may be related to the attraction and receipt of the pollen tube. In contrast with conventionally-organized embryo sacs with synergids, Plumbago accumulates calcium in the egg cell. Received: 30 December 1999 / Revision accepted: 24 March 2000     

Gibberellic acid impairs fertilization in Clementine mandarin under cross-pollination conditions

We investigated the effect of gibberellic acid (GA₃) in the fertilization of Clementine mandarin cv. ‘Clemenules’ (Citrus clementina Hort. ex Tan.), a parthenocarpic variety that produces seedless fruit due to its self-incompatible nature, but yields seedy fruits when grown under cross-pollination conditions.Experiments were conducted with on-tree ‘Clemenules’ flowers and ‘Fortune’ mandarin pollen (C. clementina Hort. ex Tan. × C. reticulata Blanco), which is sexually compatible with the former. Preanthesis treatment at −2 days after anthesis (−2 DAA) enhanced ovule abortion in both unpollinated and cross-pollinated (at +2 DAA) flowers. In the latter, the number of pollen tubes reaching the ovules was significantly reduced although pollen grains were not treated; thus, fertilization was partially avoided and seed set was reduced. When GA₃ was applied at anthesis (0 DAA) at the time of pollination, ovule abortion was again enhanced, and pollen tube growth was completely arrested; thus, fertilization was prevented and seed set was impeded. When GA₃ was applied 24 h after pollination (+1 DAA in flowers pollinated at anthesis), pollen tube growth was impaired but not arrested and ovule abortion was enhanced; therefore, fertilization was not prevented but impaired.We conclude that, when applied the days around anthesis, GA₃ (10 mg l⁻¹) impairs fertilization by either enhancing ovule abortion or reducing pollen tube growth, in ‘Clemenules’ flowers under cross-pollination conditions. The intensity of the response depends on the physiological flower state at the moment of treatment.     

The ponticulus: a structure bridging pollen tube access to the ovule in Pistacia vera

The path of the pollen tube has been examined in pistachio (Pistacia vera), a chalazogamous species where the pollen tube penetrate the ovule via the chalaza. Special attention was paid to the way the pollen tube gains access to the ovule. A single anatropous ovule with a big funiculus occupies the entire ovary cavity. At anthesis, a physical gap exists between the ovule and the base of the style. However, upon pollen tube arrival a protuberance, the ponticulus, develops in the uppermost area of the funiculus between the style and ovule. This structure appears to facilitate access to the ovule by the pollen tube. The pollen tube penetrates the ovule via this ponticulus. Upon penetration, callose develops in the ponticulus cells surrounding the pollen tube. After pollen tube passage, the upper layer of the ponticulus lignifies and isolates the ovule from the style. This separation is further enlarged 2 weeks later when the ovary starts to develop without expansion of the ovule and a large gap develops separating the ovule from the style. Except for the induction of callose formation by the pollen tube in the funiculus, this process is independent of pollination and appears to be developmentally regulated since it occurs in the same way and at the same time in pollinated and unpollinated flowers. The ponticulus, although by a different mechanism, appears to be playing the role of an obturator regulating access of the pollen tube to the ovule. Furthermore, this access is restricted to a particular time during the development of the ovule.     

The Significance of the Obturator in the Control of Pollen Tube Entry into the Ovary in Peach (Prunus persica)

Changes in the obturator of the peach (Prunus persica) havebeen investigated and related to pollen tube growth in thisregion. At anthesis, the cells of the obturator are active andrich in starch reserves. Twelve days after anthesis these cellsproduce a secretion that stains for carbohydrates and for proteins.As the secretion is produced, starch vanishes from these cellsand they degenerate and collapse as callose is accumulated.Secretion is independent of pollination as it takes place ina similar fashion both in pollinated and in unpollinated flowers. Pollen tube growth along the obturator surface depends on thissecretion for, although pollen tubes reach the base of the styleseven days after pollination, they cannot grow on the obturatoruntil five days later, when the secretion is produced. Thisdiscontinuous secretion taking place at the obturator may providea mechanism that controls the entrance of pollen tubes intothe ovary in the peach. Prunus persica, peach, obturator, pollen tube     

Development of fertilized ovules and their role in the growth of the pea pod

The histological development of fertilized ovules during fruit-set and development in pea ( Pisum sativum L. cv. Alaska) has been investigated. Killing the ovules on day 0 (anthesis) or day 1 prevented fruit-set and resulted in ovary degeneration. When the ovules were destroyed at later stages the ovaries developed, though the rate of growth of the pod was reduced significantly. Pollination in pea occurs normally the day before anthesis, and fertilization of the egg cell 32 to 48 h later. The first divisions of the zygote and endosperm nuclei started simultaneously (ca 48 h after pollination) but the endosperm developed more rapidly than the embryo; the embryo sac cavity was lined with free endosperm nuclei at the time of beginning suspensor elongation. Extracts of endosperm and ovule coats from ovules at day 7 after anthesis showed fruit-set activity in pea, the latter material having about 3 times more activity than the former per ovule basis. These results indicate that fertilization of the ovule is necessary for fruit-set in pea, and that compounds which induce fruit-set are probably synthesized in the ovules following fertilization.     

OVULE ABORTION IN ‘NONPAREIL’ ALMOND (PRUNUS DULCIS [MILL.] D. A. WEBB)

The earliest indication of ovule abortion in almond (Prunus dulcis [Mill.] D. A. Webb ‘Nonpareil‘) is the deposition of callose (as indicated by aniline blue fluorescence) 2 days after pollination which is 2 days before clear histological symptoms of ovule degeneration are evident and 6 days before fertilization of the viable ovule. Callose deposition begins in the chalazal region of the nucellus where the funicular trace enters the ovule and ramifies into the integuments. As ovule abortion progresses, callose deposition in the inner integument extends as a ring around the nucellus. Movement of the fluorescent dye disodium fluorescein (uranin) indicated that translocation from the vascular trace into abortive ovules becomes blocked at the chalazal position. The dye freely penetrates and diffuses into viable ovules but fails to penetrate abortive ovules. Lack of, or delayed and irregular, megagametophyte development was another characteristic of abortive ovules. Biochemical and histochemical analyses of abortive and viable ovules indicated that carbohydrate depletion parallels ovule abortion. These observations lead to the conclusion that ovule abortion is accompanied by blockage in metabolite supply although whether this blockage is the primary cause or a consequence of ovule abortion is uncertain.     

杉木大孢子叶球发育过程中形态结构的变化

以福州生长的成年杉木（Cunninghamia lanceolata（Lamb.） Hook.）为实验材料,采用数码相机实地拍照、体视镜、半薄切片以及扫描电镜等方法,从形态学、解剖学系统观察了杉木大孢子叶球的发育过程。结果显示,2011年10月底至11月初,杉木大孢子叶球形成,此时大孢子叶球呈绿色,体积较小;翌年3月中下旬,大孢子叶球成熟,进入传粉期,期间大孢子叶球经历了由绿变黄的颜色转变、体积增大以及苞片开张的过程。胚珠发育过程中,胚珠原基于1月上旬发生,1月中下旬珠被和珠心组织已分化形成;2月下旬,珠心组织继续发育,形态呈椭圆型,并在其上方形成贮粉室,周围的珠被组织继续生长包围珠心组织,形成珠孔道;3月初珠孔形成,开口达到最大,胚珠的体积继续增大;3月中下旬,胚珠珠孔处开始分泌传粉滴。授粉后,传粉滴消失,珠孔上方的组织停止生长,珠孔开口亦不再增大。研究结果表明杉木大孢子叶球从分化形成到发育成熟需要约5个月的时间,胚珠的形态结构经过长期演化形成了许多适应风媒传粉的结构特征。     

Activity and localisation of sucrose synthase and invertase in ovules of sexual and apomicticBrachiaria decumbens

Summary Brachiaria decumbens has sexual and apomictic reproduction. Apomixis is facultative and of the aposporic type. In early stages of ovule development, differences in the pattern of callose deposition between sexual and apomictic plants were observed which points to possible differences in carbohydrate metabolism. Therefore, a comparative study on carbohydrate metabolism between a sexual diploid ecotype and an apomictic tetraploidB. decumbens was made. A histochemical determination of two enzymes responsible for sucrose degradation, sucrose synthase and invertase, was performed for all stages of ovule development. In addition, the concentrations of sucrose, glucose, and fructose were measured for each stage of ovule development, both for sexual and apomictic plants. The enzymes were localised by immunohistochemistry with heterologous antibodies. A distinct difference between sexual and apomictic plants was observed in the localisation of sucrose synthase activity as well as in the amount of activity, especially in the early stages of ovular development. Invertase activity localisation was comparable between ovules of the sexual and apomictic plants, but its activity is clearly higher in ovules of sexual plants. The localisation of the enzymes coincided with the place of activity. For both sexual and apomictic plants the amount of sucrose in the ovaries increased with the stage of ovule development. Differences in the amount of sucrose between sexual and apomictic plants in ovaries with ovules in comparable stages of development were detected. A delay in the onset of carbohydrate metabolism during early stages of ovule development characterises the apomictic plant.Abbreviations MMC megaspore mother cell - MC meiocyte - MS megaspore - AI apospore initial - CO coenocyte - MES mature embryo sac - SuSy sucrose synthase - INV invertase - BMM buthylmethyl methacrylate - DTT dithiothreitol - DAPI 4,6-diamidino-2-phenylindole - PBS phosphate-buffered saline     

Sexual reproduction in the cork oak (Quercus suber L.). I. The progamic phase

 Cork oak (Quercus suber L.) is a monoecious wind-pollinated species with a protandrous system to ensure cross-pollination. To the best of our knowledge, this report provides the first insight into the sexual reproduction cycle in this species. The cork oak flowering season extends from April until the end of May. Our results show that, at anthesis, the pistillate flower is not completely formed and ovules are just starting to develop. Pollen reaching the dry stigmatic surface adheres to the receptive cells, germinates and penetrates the epidermis in aproximately 24 h, and grows through the intercellular spaces of a solid transmitting tissue. In cross-pollination, a sequential arrest of pollen tubes was observed along the style, providing preliminary evidence for a pollen tube competition mechanism. As a consequence, few pollen tubes reach the basal portion of the style. Furthermore, pollen tube growth is a discontinuous process since tubes are arrested in the basal portion of the style about 10–12 days after pollination. While tubes are latent, the ovarian loculus starts to develop from an emerging mass of sporogeneous cells which later will differentiate into the placenta and ovules. One and a half months after pollination ovules complete their differentiation, tubes resume growth and fertilisation occurs. Ovular abortion is frequent at this stage, and only one ovule will successfully mature during autumn into a monospermic seed. Received: 23 July 1998 / Revision accepted: 2 November 1998     

In vitro pollination of isolated ovules of Cichorium intybus L.

 A method for the in vitro pollination of chicory (Cichorium intybus L.) ovules was developed. The purpose was to avoid self-incompatibility after in vitro self-pollination of ovules isolated from flower buds before anthesis and from open flower heads. Seedlings were obtained at a low percentage (0.76%), and the results were explained in terms of pollen viability, pollen germination on the ovule, embryogenesis studies and ploidy analysis. Received: 9 April 1997 / Revision received: 23 July 1997 / Accepted: 6 September 1999     

Prolongation of Embryo Sac Viability in Pear (Pyrus communis) Following Pollination or Treatment with Gibberellic Acid

Embryo sac development has been investigated in unpollinated,cross pollinated and gibberellic acid (GA₂) treated flowersof Pyrus communis L. While pollination and GA₃ treatments donot alter embryo sac development, they prolong embryo sac viability.In untreated unpollinated flowers, ovules degenerate between12 and 21 d after anthesis, while in cross pollinated and GA₃treated flowers this degeneration is postponed by about 10 d.Thus, in a cross pollinated flower this extends the period overwhich a successful fertilization can take place. This increasedperiod of viability is accompanied by an elongation of the embryosac itself. Elongation takes place two weeks prior to fertilizationin cross pollinated flowers. The extension of life span of embryo sacs following pollinationand treatment with gibberellic acid indicates that a stimulusinduced by ‘pollination’ could be mediated by GA₃Whatever its mechanism of operation, the prolongation of embryosac viability by pollination represents a selective advantage,in that the period at which the ovules are receptive to fertilizationmust be significantly extended. Embryo sac, gibberellic acid, Pyrus communis, pear, pollination     

The evolution of ovule number and flower size in wind-pollinated plants

In angiosperms, ovules are "packaged" within individual flowers, and an optimal strategy should occur depending on pollination and resource conditions. In animal-pollinated species, wide variation in ovule number per flower occurs, and this contrasts with wind-pollinated plants, where most species possess uniovulate flowers. This pattern is usually explained as an adaptive response to low pollen receipt in wind-pollinated species. Here, we develop a phenotypic model for the evolution of ovule number per flower that incorporates the aerodynamics of pollen capture and a fixed resource pool for provisioning of flowers, ovules, and seeds. Our results challenge the prevailing explanation for the association between uniovulate flowers and wind pollination. We demonstrate that when flowers are small and inexpensive, as they are in wind-pollinated species, ovule number should be minimized and lower than the average number of pollen tubes per style, even under stochastic pollination and fertilization regimes. The model predicts that plants benefit from producing many small inexpensive flowers, even though some flowers capture too few pollen grains to fertilize their ovules. Wind-pollinated plants with numerous flowers distributed throughout the inflorescence, each with a single ovule or a few ovules, sample more of the airstream, and this should maximize pollen capture and seed production.     

Floral development of Cardiopteris,with emphasis on gynoecial structure and ovular morphology

Cardiopteris is unique in the expanded Cardiopteridaceae for several distinctive features, including its gynoecial structure and ovular morphology. We studied the floral development of Cardiopteris to clarify floral morphology and document floral development. Cardiopteris has three carpel primordia, which are separate at their tips but congenitally fused at their bases. The synascidiate zone (the fused proximal part) develops into the unilocular ovary; the three discrete carpel apices diverge in development: the apex of the adaxial carpel differentiates into a style and stigma, while the apices of the two lateral-abaxial carpels elongate and develop into a fleshy appendage only after fertilization. The ovules are attached to the lateral-abaxial carpels. At anthesis, the ovules are ategmic and orthotropous without funicles (morphologically undifferentiated). Functional differentiation occurs in the three carpels of Cardiopteris: the adaxial one is the site of pollination, while the lateral-abaxial two produce ovules. The ategmic orthotropous ovule is unusual in Cardiopteridaceae and is an apomorphy of Cardiopteris.     

Pollen tube growth and early ovule development following self- and cross-pollination in<Emphasis Type="Italic"> Eucalyptus nitens</Emphasis>

Controlled self- and cross-pollinations were conducted on flowers of five mature Eucalyptus nitens trees. Levels of self-sterility of the trees ranged from 25.8 to 93.6%. Pollen tube numbers in styles and ovule penetration by pollen tubes was investigated 2 weeks after pollination by fluorescence microscopy. There were no significant differences between treatments in the number of pollen tubes present in styles or in the percentage of ovules penetrated by pollen tubes. Embryology of material harvested 2 and 4 weeks after pollination was investigated by bright-field microscopy. Fertilisation had taken place by 2 weeks after pollination with nearly every ovule showing evidence of fertilisation. Cross-pollination resulted in a greater proportion of healthy, developing ovules, at both 2 and 4 weeks after pollination, compared with self-pollination. The proportion of degenerating ovules increased from 2 to 4 weeks after pollination. The reduced ability of E. nitens to set self-pollinated seed compared with cross-pollinated seed appears to be controlled by a post-zygotic mechanism. Differences in ovule size may potentially assist in the identification of trees incapable of setting self-pollinated seed.     

Expression of female sterility in alfalfa (<Emphasis Type="Italic">Medicago sativa</Emphasis> L.)

Female sterility associated with the presence of callose in the nucellus at anthesis was studied in an F₁ progeny of two alfalfa plants displaying 5 and 81% ovule sterility. Transgressive segregation was observed and 100% sterile plants were obtained. Two of the sterile plants were used for cytological analyses on sectioned and stain-cleared whole ovules, in comparison to a 100% fertile full sib plant. The first sign of sterility was callose deposition in the nucellus cell walls surrounding the sporogenous cells of the young ovules. At the same stage, no trace of callose was present in ovule primordia of the fertile plant. Megaspore mother cells differentiated in both fertile and sterile ovules and meiosis was initiated, as indicated by chromatin patterning typical of a zygotene stage. However, meiosis was never completed in the sterile plants. In the control, callose was deposited around the meiocyte and as sects between the cells of the dyads and tetrads during meiosis, and disappeared after the completion of meiosis; an embryo sac developed and female fertility was normal. In the sterile ovules, some nucellus cells enlarged and callose accumulation continued forming thick deposits. At anthesis, the sterile ovules lacked an embryo sac and showed massive callose accumulation in the nucellus. Male fertility was normal in female-sterile plants, thus a female-specific arrest of sporogenesis appears to be the cause of sterility. Pistil development was aberrant in some sterile genotypes, even with arrested pistil growth in early flower buds.     

Ovule differences between single-kernelled and double-kernelled fruits in almond (Prunus dulcis)

In order to detect differences between almond cultivars that commonly produce one or two seeds, we compared ovule sizes and developmental stages at anthesis and three days after anthesis. The results showed that the presence of only small differences in size between primary and secondary ovules during flowering seemed to be a necessary but not sufficient condition for the production of double‐kernelled fruits. In contrast, differences in the developmental stage of both ovules did not prevent the production of double kernels. Large differences in ovule size at anthesis or in the following days resulted in fruits with single seeds. The behaviour of the secondary ovules in cultivars producing fruit with single seeds was very different, even in closely related cultivars. The degeneration of the secondary ovule seems to be genetically programmed and follows a genotype‐dependent pattern.     

蚕豆胚珠发育过程中淀粉动态的观察

蚕豆胚珠发育过程中淀粉动态变化如下:1.发育早期,整个胚珠中未见淀粉粒。其后首先在合点区出现淀粉,而后从合点向珠孔逐渐扩大分布范围。2.珠心和内、外珠被中均含有淀粉粒,尤以内珠被的淀粉增长迅速,数量多、个体大。受精后,内珠被解体,淀粉出现在外珠被细胞中,推测营养物质可通过整个胚囊表面进入其中。3.合点与胚囊之间的珠心细胞特化或长形。可能有助于营养物质进入胚囊。4.功能大孢子中贮存丰富的淀粉粒,它和珠心细胞一起是胚囊发育时的营养来源。5.卵细胞受精后,所含淀粉粒的数量和大小明显增长,随着合子和胚细胞的分裂,其中贮存的淀粉逐渐被消耗,到多细胞球形胚时完全消失。6.胚乳核周围始终未出现淀粉粒。7.胚器官分化之后,子叶和胚轴等处逐渐出现淀粉粒,其中生长活跃的结构如生长点、维管束等不贮存淀粉。8.子叶中的淀粉粒含量迅速增加,颗粒特大,是种子内营养物质的最终贮存场所。     

Origin of gynogenetic embryos of Beta vulgaris L.

Haploid plants of Beta vulgaris were obtained by gynogenesis from ovules isolated from male-fertile and annual and biennial male-sterile plants. We show that on a N6 basal medium, supplemented with 2.85 M IAA and 0.94 M Kinetin or 0.88 M BAP, haploids originate directly through embryogenesis. In order to determine the optimal developmental stage of the ovule of Beta vulgaris for gynogenesis, we carried out a histological study of whole ovules from open male-sterile flowers (collected 1 to 5 days after flowering) and unopened male-fertile flowers (collected 1 to 3 days before anthesis). In all cases, the gametophyte appeared completely differentiated. These results suggest that maturity of the gametophyte is reached a few days before anthesis and therefore ovules from unopened flowers are already suitable for plating. A developmental study of the haploid cells of the sugarbeet embryo sac during the first week of in vitro culture showed that the viable gynogenetic embryo originated only from the egg cell.     

Abscisic Acid Levels during Early Seed Development in Sechium edule Sw

The time-course growth of single tissues in pollinated and unpollinated ovules of Sechium edule Sw. is described in relation to the endogenous levels of abscisic acid. Quantitation of abscisic acid (ABA) in the minute amounts of material obtained after ovule dissection has been performed by using a highly specific and sensitive solid-phase radioimmunoassay based on a monoclonal antibody raised against free (S)-ABA. While the absolute amount of ABA rises in both types of ovules, only in unpollinated ones does this leads to an increase in the hormone concentration. Infact in pollinated ovules the rapid growth following pollination prevents, through a dilution effect, the increase in ABA concentration. Growth patterns and endogenous ABA levels are similar for integuments and nucellus tissues either in pollinated or unpollinated ovules. It is suggested that the growth inhibition induced by the increase in ABA concentration after anthesis could be counteracted by the pollination triggered fast ovule growth.     

Does successful ovule development depend on its position within the pod? Examples from Neotropical Fabaceae

Previous studies have shown a nonrandom pattern of ovule fate probabilities according to ovule position in legume pods. Here, we tested how ovule position within the pods of two Fabaceae affects its fate. We expected higher proportion of well‐formed seeds near the fruit tips and of unfertilized and aborted ovules near fruit bases. We collected pods of Poincianella pyramidalis and Anandenanthera colubrina in a seasonal dry forest in northeastern Brazil and recorded total pod length and ovule number, position, and fate (unfertilized, well formed, aborted, and predated). The proportion of well‐formed ovules at fruit tips was significantly higher than at fruit bases in P. pyramidalis. The opposite pattern was found for unfertilized and aborted ovules, thus corroborating our hypothesis. However, the probability of seed predation in A. colubrina was significantly higher in pod tips, thus providing moderate support for our hypotheses. Interspecific differences in the patterns of ovule fate are likely to be driven by species pollination systems.