凤仙花花药发育中多糖和脂滴组织化学研究

以不同发育时期的凤仙花花药为实验材料,采用组织化学方法,对花药发育中的结构变化及多糖和脂滴物质分布进行观察。结果表明:(1)凤仙花的花药壁由6层细胞组成,包括1层表皮细胞,2层药室内壁细胞,2层中层细胞和1层绒毡层细胞。其中绒毡层细胞的形态不明显,很难与造孢细胞区分,且在小孢子母细胞时期退化。(2)在小孢子母细胞中出现了一些淀粉粒,但减数分裂后,早期小孢子中的淀粉粒消失,又出现了一些小的脂滴;随着花粉的发育,小孢子形成大液泡,晚期小孢子中的脂滴也消失;小孢子分裂形成二胞花粉后,营养细胞中的大液泡降解、消失,二胞花粉中又开始积累淀粉;接近开花时,成熟花粉中充满细胞质,其中包含了较多的淀粉粒和脂滴。(3)在凤仙花的花药发育中,绒毡层细胞很早退化,为小孢子母细胞和四分体小孢子提供了营养物质;其后的中层细胞退化则为后期花粉发育提供了营养物质。     

Differential distribution of ascorbic acid and RNA in the developing anthers ofDatura stramonium L.

The histochemical localization of ascorbic acid and RNA was studied during developmental stages ofDatura anthers. The concentration of ascorbic acid and RNA was high in primary parietal and primary sporogenous layers, sporogenous cells and pollen grains. The connective of young anther showed remarkably high concentration of ascorbic acid. The high peaks of ascorbic acid and RNA concentration correlated with the growth phases of anther. The connective and anther wall layers act as reservoirs of energy needed for developing sporogenous cells.     

Anther ontogeny in <Emphasis Type="Italic">Brachypodium distachyon</Emphasis>

Brachypodium distachyon has emerged as a model plant for the improvement of grain crops such as wheat, barley and oats and for understanding basic biological processes to facilitate the development of grasses as superior energy crops. Brachypodium is also the first species of the grass subfamily Pooideae with a sequenced genome. For obtaining a better understanding of the mechanisms controlling male gametophyte development in B. distachyon, here we report the cellular changes during the stages of anther development, with special reference to the development of the anther wall. Brachypodium anthers are tetrasporangiate and follow the typical monocotyledonous-type anther wall formation pattern. Anther differentiation starts with the appearance of archesporial cells, which divide to generate primary parietal and primary sporogenous cells. The primary parietal cells form two secondary parietal layers. Later, the outer secondary parietal layer directly develops into the endothecium and the inner secondary parietal layer forms an outer middle layer and inner tapetum by periclinal division. The anther wall comprises an epidermis, endothecium, middle layer and the secretory-type tapetum. Major documented events of anther development include the degradation of a secretory-type tapetum and middle layer during the course of development and the rapid formation of U-shaped endothecial thickenings in the mature pollen grain stage. The tapetum undergoes degeneration at the tetrad stage and disintegrates completely at the bicellular stage of pollen development. The distribution of insoluble polysaccharides in the anther layers and connective tissue through progressive developmental stages suggests their role in the development of male gametophytes. Until sporogenous cell stage, the amount of insoluble polysaccharides in the anther wall was negligible. However, abundant levels of insoluble polysaccharides were observed during microspore mother cell and tetrad stages and gradually declined during the free microspore and vacuolated microspore stages to undetectable level at the mature stage. Thus, the cellular features in the development of anthers in B. distachyon share similarities with anther and pollen development of other members of Poaceae.     

ANTHER AND POLLEN DEVELOPMENT IN RICE (ORYZA SATIVA)

Investigations of the growth of anthers and ontogeny of pollen grains of Oryza sativa (rice) IR-30 were undertaken for the purpose of 1) providing a set of growth measurements and 2) describing stable cytological features of anther and pollen development. Correlations exist between elongation of the floret and growth parameters of the anther such as its length, width, fresh and dry weights and cytological stage of pollen development. In the early ontogeny of the anther, hypodermal archesporial initials divide periclinally to form primary parietal cells and primary sporogenous cells. Each of the latter divides twice mitotically to generate four microspore mother cells, which undergo meiosis. The anther wall is formed by anticlinal and periclinal divisions of the primary parietal cells as well as of cells surrounding the primary sporogenous cells. Subsequent cytological features in the development of anther and pollen grains of rice have much in common with anther and pollen developmental biology of other members of Gramineae.     

生态因子对滇重楼花药开裂的影响

滇重楼为延龄草科重楼属植物,具有极高的药用价值,由于重楼传统药用部位生长缓慢、繁殖力低下,以及人们对野生重楼资源的过度采挖使其资源日趋枯竭。滇重楼的花药在整个花期中存在开裂－关闭的现象,花药的有效闭合应是保护花粉、延长花粉寿命、增强雄性适合度的一种适应机制。该研究以滇重楼为对象,通过设计正交实验和对比实验,观测其花药开裂过程中的光照强度、温度、湿度的变化,探究光照强度、温度、湿度等生态因子对滇重楼花药开裂的影响以及滇重楼花药开裂与生态因子变化的关系。结果表明：(1)在滇重楼花药开裂的过程中,光照强度增强、温度升高、相对湿度下降;(2)温度是影响滇重楼花药开裂时间的主导因子,升温促进花药开裂,降温促进花药关闭;(3)高湿度及黑暗推迟花药开裂,但并不能阻止花药开裂;(4)低温可使滇重楼花药持续关闭,而光照强度越高,花药持续关闭所需的温度越低。该研究有利于解释滇重楼花药白天开裂夜晚关闭的现象与环境因子的关系,对滇重楼的栽培育种提供理论指导。     

紫斑牡丹的花药发育和小孢子发生

紫斑牡丹的花药是在花芽发育的第３个年周期中,从雄蕊原厚基发育而来,花药壁按特有方式发生,主要特点是绒毡层与次生造孢细胞同源。花药由４个花粉囊组成,绒毡层属分泌型,中层３－４层,其中１－３层与药室内壁同步发育出纤维素壁加厚,并在花药成熟时宿存。小包子母细胞减数分裂同时开始,但不同时结束,分裂过程高度不同步。胞质分裂为同时型,四分体中小孢子排列呈正四面体形,减数分裂前期Ⅰ通常有Ｂ－染色体和染色体桥形。     

Ultrastructural analysis of the behavior of the dimorphic fungus Microbotryum violaceum in fungus-induced anthers of female Silene latifolia flowers

Summary. The development of male organs is induced in female flowers of the dioecious plant Silene latifolia by infection with the fungus Microbotryum violaceum. Stamens in a healthy female flower grow only to stage 6, whereas those in an infected female flower develop to the mature stage (stage 12), at which the stamens are filled with fungal teliospores instead of pollen grains. To investigate these host–parasite interactions, young floral buds and fungus-induced anthers of infected female flowers were examined by electron microscopy following fixation by a high-pressure freezing method. Using this approach, we found that parasitic hyphae of this fungus contain several extracellular vesicles and have a consistent appearance up to stage 8. At that stage, parasitic hyphae are observed adjacent to dying sporogenous cells in the infected female anther. At stage 9, an increased number of dead and dying sporogenous cells is observed, among which the sporogenous hyphae of the fungus develop and form initial teliospores. Several types of electron-dense material are present in proximity to some fungi at this stage. The initial teliospores contain two types of vacuoles, and the fungus cell wall contains abundant carbohydrate, as revealed by silver protein staining. The sporogenous cell is probably sensitive to infection by the fungus, resulting in disruption. In addition, the fungus accelerates cell death in the anther and utilizes constituents of the dead host cell to form the mature teliospore. Correspondence and reprints (present address): Molecular Membrane Biology Laboratory, RIKEN, 2-1, Hirosawa, Wako, Saitama 351-0198, Japan.     

Anther culture of kale (Brassica oleracea L. convar.acephala (DC.) Alef.)

The pollen development and androgenic ability of 18 kale (Brassica oleracea convar.acephala) genotypes was observed during an anther culture study. Anther culture was successful in 6 of the genotypes and the highest yield obtained was 17 embryos per 100 anthers plated. Two stages of anther development were identified as being responsive to anther culture. The first and most responsive was that corresponding to the late uninucleated stage and the second to the late binucleated stage. These stages correspond with the onset of mitotic events in the microspores. Pollen viability was studied and low viability was noted which declined to zero after 9 days of anther culture. The initial viability level however was not clearly related to androgenic ability. The significance of the production of haploid and dihaploid kale genotypes in the study and breeding of resistance to clubroot is discussed.     

Presence of the anther gland is a key feature in pollination of the early‐branching papilionoids Dipteryx alata and Pterodon pubescens (Leguminosae)

• The presence of glandular appendages in the anthers is a rare condition in angiosperms. In Leguminosae it occurs in species of the Mimosoid clade and in early‐branching clades of papilionoids such as Dipterygeae. In Dipterygeae such appendages surprisingly exhibit a secretory cavity instead of secretory emergences as is the case for the Mimosoid clade. Thus, the objective of this study was to elucidate the function of anther glands in Dipteryx alata and Pterodon pubescens, species in the Dipterygeae clade that exhibit a pollen release mechanism that is intermediate between the explosive and valvular types.
• Flower buds and flowers were processed for surface, anatomical, histochemical and ultrastructural analyses.
• Anther glands consist of a cavity secreting sticky substances (oleoresins and polysaccharides) that play a key role during the flower's lifespan by aggregating pollen grains and attaching them to the floral visitor's body. Other floral features that are important for understanding the pollen release mechanism that is intermediate between the valvular and the explosive types are: (i) keel petals intertwined with tector trichomes; (ii) glandular appendages in the abaxial and lateral sepals and in petals composed of secretory ducts; and (iii) a continuous secretion process of the anther glands followed by an asynchronous dehiscence of anthers.
• The well‐adapted papilionoid flag blossom with anther glands and keel petals intertwined with trichomes provided the foundation for a successful canalisation toward a pollen release mechanism intermediate between the explosive and valvular types inside early‐branching papilionoids.

     

Changes in calcium and calmodulin levels during microsporogenesis,pollen development and germination in Gasteria verrucosa (Mill.) H. Duval

Summary The distribution of membrane calcium and calmodulin (CaM) has been fluorimetrically determined in the anther of Gasteria verrucosa with particular attention to sporogenous cells, meiocytes, microspores, pollen and stages of pollen germination and tube growth using chlortetracycline (CTC) and fluphenazine (FPZ). CTC and FPZ fluorescence in sporogenous cells is relatively higher than in the adjacent tapetal cells, indicating higher membrane calcium and CaM levels in the former cell type. However, during meiosis there is a significant increase in membrane calcium and CaM levels in the meiocytes compared to that found in the young microspores. CTC and FPZ fluorescence in the sporogenous cells, meiocytes and young microspores is punctate and slightly diffused throughout the cytoplasm. In the microspores of the tetrad and the young released microspores CTC fluorescence (CTCf) is polarized and mainly associated with the area opposite the future colporal region. FPZ fluorescence (FPZf) becomes polarized in the young microspore. Subsequently, there is a shift in the polarity, and most of the CTCf and FPZf in the old microspores and pollen is regionalized towards the colporal region, and the fluorescence is more diffused, indicating a change in the organellar-bound calcium and CaM. This final graded distribution of CTCf is maintained during pollen germination in that the growing pollen tubes invariably show a tip to base membrane-calcium gradient. In the tapetal cells a high level of Ca²⁺ is present during the microspore stage. During the preparation for anthesis the endothecium differentiation is marked by the presence of Ca²⁺. Post-treatment of labelled cells with a Ca²⁺ chelator such as EGTA resulted in a substantial decrease in diffuse and punctate CTCf. Alternatively, treatment of cells with non-ionic detergent Nonidet P-40 resulted in the total elimination of CTCf, suggesting that the observed CTC fluorescence was due to membrane-associated calcium. The cytological specification of CTC as a probe for calcium is discussed. From cytofluorometric measurements and atomic absorption, it became clear that the level of Ca²⁺ in the anther is high during the sporogenous and meiotic phases. An increase in CTCf and FPZf occurred after microspore mitosis. An interaction of Ca²⁺ transport from tapetum to the young pollen is postulated. These findings suggest that the level of Ca²⁺ in the anther during meiosis is generally relatively higher than at the sporogenous or young microspore stage. These findings are discussed in the light of available information on the role of Ca²⁺ and CaM-mediated processes such as cell division, callose synthesis and pollen-tube tip growth.     

Morphological development of anthers induced by the dimorphic smut fungus<Emphasis Type="Italic"> Microbotryum violaceum</Emphasis> in female flowers of the dioecious plant<Emphasis Type="Italic"> Silene latifolia</Emphasis>

When inoculated with the dimorphic smut fungus Microbotryum violaceum (Pers.) G. Deml and Oberwinkler, the female flower of the dioecious plant Silene latifolia (Miller) E.H.L. Krause develops anther-like structures filled with spores instead of pollen grains. Using natural scanning electron microscopy, Nomarski interference microscopy, and fluorescence microscopy, we investigated the morphological modifications of the host plant resulting from this parasitism and the localization of smut hyphae in the flower bud. Flowers of infected plants lasted significantly longer than those of healthy plants, probably because the infection strengthened floral organs, such as the flower base and the anther filaments. Smut hyphae were observed throughout all organs of the young flower buds of infected plants, including sepals, petals, stamens, and pistil primordia. In healthy female flowers, anthers initiated sporogenous cell formation, but lacked parietal cell layers. By contrast, the parietal cell layers of infected female flowers differentiated into tapetal tissue, middle cell layers, and endothecial layers, as in the anthers of healthy male flowers. Smut spore formation in the infected anther was initiated in intercellular regions between the sporogenous cells, resulting in degeneration of premature sporogenous cells, tapetal tissue, and middle cell layers. The development of the endothecial layers and epidermis in the infected anther were morphologically normal.Abbreviations DAPI 4,6-diamidino-2-phenylidole - i infected - PMC pollen mother cell     

Pollen and anther development in Nelumbo (Nelumbonaceae)

The Nelumbonaceae are a small family of aquatic angiosperms comprising Nelumbo nucifera and Nelumbo lutea. Historically, the genus has been considered to be closely related to Nymphaeales, however new systematic work has allied Nelumbo with lower eudicots, particularly Platanus. In recent years, studies of pollen development have contributed greatly to the understanding of phylogenetic relationships, but little has been known about these events in Nelumbo. In this paper, pollen and anther development are morphologically described for the first time in N. lutea. A comprehensive ontogenetic sequence is documented, including the sporogenous tissue, microspore mother cell, tetrad, free spore, and mature pollen grain stages. The deposition of a microspore mother cell coat and callose wall, the co-occurrence of both tetrahedral and tetragonal tetrads, the formation of a primexine in tetrads, and primexine persistence into the late free spore stage are shown. The majority of exine development occurs during the free spore stage with the deposition of a tectate-columellate ectexine, a lamellate endexine, and an unusual granular layer below and intermixed with the endexine lamellae. A two-layered intine forms rapidly during the earliest mature pollen stage. Major events of anther development documented include the degradation of a secretory-type tapetum during the free spore stage and the rapid formation of U-shaped endothecial thickenings in the mature pollen grain stage. The majority of mature pollen grains are tricolpate, however less common monosulcate and diaperturate grains also develop. Co-occurring aperture types in Nelumbo have been suggested to be an important transition in angiosperm aperture number. However, aperture variability in Nelumbo may be correlated with the lateness of aperture ontogeny in the genus, which occurs in the early free spore stage. This character, as well as other details of pollen and anther ontogeny in Nelumbo, are compared to those of Nymphaeales and Platanus in an effort to provide additional insight into systematic and phylogenetic relationships. Although Nelumbo is similar to both groups in several characters, the ontogenetic sequence of the genus is different in many ways.     

Anther ontogeny in Campsis radicans (L.) Seem. (Bignoniaceae)

In this study anther ontogeny of Campsis radicans (L.) Seem. was investigated by transmission electron microscopy and light microscopy with special reference to the development of the anther wall. The anther wall formation follows the dicotyledonous type. The differentiation in anther starts with the appearance of archesporial cells which undergo periclinal divisions to give primary parietal layer to the epidermal site and the primary sporogenous cells to the inside. The primary parietal layer also divides to form two secondary parietal layers. Later, the outer secondary parietal layer (spl1) forms the endothecium and the middle layer by periclinal division whereas the inner one (spl2) directly develops into the outer tapetum forming the inner most layer of the anther wall. The sporogenous tissue is generally organized in two rows of cells with a horseshoe-shaped outline. The remainder of the tapetum lining the sporogenous mass is derived from the connective tissue. The tapetum thus has dual origin and dimorphic. Anthers are tetrasporangiate. The wall of the anther consists of an epidermis, endothecium, middle layer, and the secretory type tapetum. Tapetal cells are usually binucleated. Epidermis and Endothecium layers of anther wall remain intact until the end of anther and pollen development; however, middle layer and tapetum disappear during development.     

西瓜小孢子囊发育及雄配子体发生的观察

西瓜(Citrullus lanatus)小孢子囊的孢原细胞出现在雄花原基出现后4—6天,孢原细胞数目推测只有一列;初生造孢细胞经过2—3次分裂,形成次生造孢细胞。开花前7—8天,小孢子囊发育健全,小孢子母细胞进入减数分裂期。同一花药不同花粉囊相同一药室,花粉母细胞减数分裂和小孢子的发育,并不是高度同步的。绒毡层为异型细胞,腺质绒毡层。雄配子体的发育开始于开花前6—7天,充分成熟的西瓜花粉已分裂为三细胞花粉。     

LEVER ACTION ANTHERS AND THE FORCIBLE SHEDDING OF POLLEN IN TORENIA (SCROPHULARIACEAE)

The anthers of Torenia fournieri were found to shed pollen forcibly by lever action. Anther structure was modified for this function by a flangelike outgrowth of the lateral pollen sac wall forming a lever. When pressed, this lever causes an infolding of the thinner, subadjacent, pollen sac wall forcing pollen from the stomium. A force of 1–1.5 g pressing against the four levers of an anther pair resulted in the forcible shedding of 2,000–3,000 pollen grains in two parallel rows. During the 2-day anthesis the flowers shift from functioning as males to hermaphroditic outcrossers, and yet only have a pollen:ovule ratio of 98.6, a ratio more indicative of facultative autogamy. The outermost pair of anthers functions on the first day of anthesis, while the second, inner pair functions on both days. In each flower, the 2-day anther pair produces approximately twice as many pollen grains as the 1 -day anther pair, a pollen production highly correlated with the length of their functional lives. This difference in pollen production is apparent in the larger size of the 2-day anthers, a size difference that first appears with the initiation of the anther primordia.     

Gynostemium micromorphology and pollination in Epipactis microphylla (Orchidaceae)

Self-pollination conditions have been recorded frequently in local populations of the genus Epipactis, and structural modifications have sometimes been reported, supporting taxonomic recognition of new species. As part of a survey of gynostemium micromorphology of Italian Epipactis populations, we studied an Apennine population of Epipactis microphylla (Ehrh.) Sw. a species listed as autogamous but with residual allogamous characters. In this population we observed the tendency to perform pre-anthesis cleistogamy. However, self-pollination was not a consequence of modification in column morphology. Cryptic pollen germination occurred in anther thecae or in the clinandrium, bypassing any contact with the receptive stigma. Pollen germination started from the distal pool of the pollinium, close to the rostellum-viscidium, suggesting that the rostellum-viscidium may have a function in promoting pollen-tube growth. Germinated pollen was exposed at the anthesis.     

不结球白菜同源四倍体Pol CMS及其保持系花药发育的解剖学研究

以秋水仙素人工诱变获得的不结球白菜同源四倍体Pol CMS及其保持系为材料,采用石蜡切片法研究其花药发育过程.结果显示:四倍体Pol CMS与保持系花药发育差异明显,不育系花药发育受阻于孢原细胞分化期,不形成药室,属无花粉囊型.极少数温敏型花药能产生1~2个花粉囊,但四分体时期绒毡层提早退化,导致成熟小孢子数目减少.四倍体保持系花药发育过程与二倍体基本一致,同一时期的可育小孢子四倍体比二倍体大,二倍体花粉粒都为3个萌发孔,而四倍体多为4个萌发孔.四倍体保持系同一花粉囊中花粉粒发育有不同步现象,花粉粒败育频率比二倍体高.研究表明,不结球白菜同源四倍体Pol CMS比其二倍体Pol CMS不育更彻底,更具应用性.     

Anther starch variations in Lilium during pollen development

Starch was cytologically localized and biochemically assayed in different anther cell layers of Lilium cv. Enchantment during pollen development and its presence was correlated with anther growth. Two phases could be distinguished: the first, the growth phase, extends from the beginning of meiosis to the vacuolated microspore stage and corresponds to maximum increase in anther size and weight. During this period, microspores lack amyloplasts and starch is degraded in the outer staminal wall layers. The tapetum does not contain starch reserves but accumulates a PAS-positive substance in its vacuole. The second phase, the maturation phase, begins with the late vacuolated microspore stage and lasts until pollen maturation. Anther growth is slowed during this phase. A wave of amylogenesis/ amylolysis occurs first in the late vacuolated-microspores and young pollen grains and, next, in the staminal envelopes. In the pollen grain, the cytoplasm of the vegetative cell is filled with starch, but amyloplasts are not detected in the generative cell. When pollen grains ripen, amylaceous reserves are replaced with lipids. In the staminal envelopes, the second amylogenesis is particularly evident in the endothecium and the middle layers; the peak of starch is reached at the young bicellular pollen grain stage; starch disappears from the anther wall early during the maturation phase. The wave of amylogenesis/amylolysis occurring in the staminal envelopes during the maturation phase is peculiar to Lilium. It is interpreted as a sudden increase in carbohydrate level caused by lower anther needs when the growth is completed. Staminal envelopes may act as a physiological buffer and regulate soluble sugar level in the anther. Stages of anther growth correlate with starch content variations and this suggests that during the growth phase, products of starch hydrolysis in the staminal envelopes may be consumed partly by anther cell layers and partly by microspores.     

关苍术两性花与雌花花药的解剖学研究

苍术属(Atractylodes DC.)是菊科菜蓟族(Cynareae)刺苞亚族(Carlininae O. Hoffm.)的一个东亚特有属,世界上仅有7种,其中我国有5种。该研究以关苍术为材料,采用石蜡切片法比较研究了两性花和雌花的花药及雄配子体发育进程,并进一步探讨了其雌花产生花药退化的时期及原因。结果表明：(1)关苍术小孢子发育与花蕾长度间存在相关性,当花蕾长度在5 mm时进入花粉母细胞时期,花药壁已分化,在7~9 mm时处于四分体时期,大于11 mm时开始进入花粉粒时期。(2)关苍术花药5个,花粉囊4个,减数分裂属同时型,四分体以正四面体为主,属3-细胞型,萌发沟3个。(3)关苍术花粉囊壁发育属双子叶型,从外层的表皮、药室内壁,到内层的中层和绒毡层均由一层细胞构成,关苍术绒毡层为腺质绒毡层。(4)关苍术雌花花药退化发生在花药发育早期至四分体时期,表现为花药发育早期畸形、药壁分化异常、小孢子母细胞发育停滞在前期、绒毡层增生4个原因。该研究结果为苍术属植物的系统发育、物种形成和进化提供胚胎学依据。     

毛竹的花药发育研究

竹类植物因有着较长的开花周期,其生殖生物学研究的报道相对较少。该研究采用石蜡切片与野外观察的方法,对毛竹花药的发育以及花药发育与花序的关系进行了研究。结果表明:毛竹的花药壁结构包括4层细胞:表皮细胞、药室内壁细胞、中层细胞和绒毡层细胞。药室内壁和中层都只有一层细胞,而且细胞形状较扁,花药发育后期药室内壁会逐渐降解,而中层则会完全解体消失。花药壁的发育为单子叶型,绒毡层为腺质型,而且只有一层,细胞径向较长,最后也会消失。小孢子母细胞减数分裂时,胞质分裂方式为连续型。形成的小孢子经一次有丝分裂后逐渐形成成熟花粉粒,大多为二细胞型,很少产生三细胞型。此外,还发现毛竹花药的发育与花序形态变化存在着相对应的关系。野外连续观察和切片发现,随着花序形态的不断发育变化,首先花药开始形成并不断分化,药壁备层也逐渐形成;接着小孢子逐渐成熟,备层也慢慢随之解体、消失;最后花药逐渐开裂并开始散粉。该研究结果不仅丰富了毛竹和竹类生殖生物学的研究内容,而且对毛竹种质的研究也具有重要意义。