The formation of cytoplasmic channels between tapetal cells inZea mays

Summary In this report we show that large cytoplasmic channels form between the tapetal cells ofZea mays (maize) during the period of tapetal cell differentiation. Tapetal cells are connected by plasmodesmata through their cellulosic cell walls prior to the first meiotic division of the meiocytes. As the tapetal cellulose wall is degraded at the onset of meiosis, both plasmodesmata and cytoplasmic channels measuring 50–200 nm are detectable between tapetal cells. By the time the meiotic tetrad is formed, the cytoplasmic channels are well-established and vary in size from 100–400 nm. The channels, with an average diameter of 200–300 nm, persist after the microspores are released from the callose wall and throughout the period of exine development in microsporogenesis. The channels could potentially allow for free exchange of cytoplasm and organelles. As the tapetal cells begin to pull apart and become vacuolate prior to microspore mitosis, the connecting channels are no longer detectable.     

Microspore and tapetal development in Echinodorus cordifolìus (Alismaceae)

In the microspore tetrad period the exine begins as rods that originate from the plasma membrane. These rods are exine units that on further development become columellae as well as part of the tectum, foot layer and “transitory endexine”. The primexine matrix is very thin in the future sites of the pores. At these sites the plasma membrane and its surface coating (glycocalyx) are without exine units and adjacent to the callose envelope. The exine around the aperture margin is characterized by units of reduced height. After the exine units and primexine matrix have become ca 0.2 μm in height a fibrillar zone forms under the aperture margin. It is the exine units around the aperture that are templates for exine processes on apertures of mature pollen. Oblique sections of the early exine show that the tectum consists of the distal portions of close-packed exine units. The exine enlarges in the free microspore period but initially its substructure (tectum, columellae, foot layer and transitory endexine) is not homogeneous and unit structures are visible until after the vacuolate microspore period. There are indications of a commissural line/plane (junction plane) which separates the foot layer from the endexine during early development. Our observations of development in Echinodorus pollen extend a growing number of reports of “transitory endexines” in monocot pollen. The exine unit-structures become 0.2 μm or more in diameter and many columellae are composed of only one exine unit. Spinules become exceptionally tall, many protruding ca 0.7 μm above the level of the tectum as units only ca 0.1 μm in diameter. The outer portion of the tectum fills in around spinules and by maturity they are microechinate with their bases spread out to ca 1 μm or more. Unit structures can be seen with SEM in mature pollen following oxidation by plasma ashing and in the tapetum these units are arranged both radially, as in spinules, and parallel with the tapetal surfaces. There are clear indications of such an arrangement of units in untreated fresh pollen. Units comprising the basal part of the exine are not completely fused by sporopollenin accumulated during development. This would seem to be a characteristic feature, based on published work, of the alismacean pollen. Our use of a tracer shows, however, that there is considerable space within or between exine structure of mature Echinodorus pollen. Based upon the ca 0.1 μm size of exine-units formed early in development and exine components seen after oxidative treatment it seems that the early (primary) accumulated sporopollenin has greater resistance to oxidation than sporopollenin added, secondarily, around and between units later in development. Both primarily and secondarily accumulated sporopollenin are resistant to acetolysis but published work indicates that acetolysis alters exine material. At the microspore tetrad time and until the vacuolate stages tapetal cells are arranged as in secretory tapetums. During early microspore stages there are orbicules at the inner surface of tapetal cells. At free microspore period tapetal cells greatly elongate into the loculus and surround the microspores. By the end of the microspore vacuolate period tapetal cells release their cellular contents and microspores are for a time enveloped by tapetal organelles and translocation material.     

MICROSPOROGENESIS IN CITRUS LIMON (RUTACEAE)

Prior to meiosis tapetal cells become binucleate, and callose deposition separates spore mother cells from each other. No cytomictic channels are present during meiosis. Cytokinesis is simultaneous, by furrowing. The primexine and a rudimentary exine are laid down while the microspores are still in tetrads. After callose dissolution the released microspores gradually become vacuolate and the exine becomes more complex and massive. During the tetrad stage tapetal walls are gradually lost and orbicules are deposited outside the plasmalemma. This continues after microspore release. Later, at the vacuolate microspore stage, the tapetal cells become amoeboid and intrude among the microspores. Tapetal dissolution occurs just prior to the appearance of large amounts of starch and lipids in the microspores.     

POLLEN AND TAPETUM DEVELOPMENT IN DESMODIUM GLUTINOSUM AND D. ILLINOENSE (PAPILIONOIDEAE; LEGUMINOSAE)

Each of the four microsporangia has three or four wall layers, a uninucleate tapetum of various cell shapes with nuclei that remain in prophase, and 12-24 pollen mother cells (PMCs). A sterile transverse septum sometimes bisects the microsporangium. PMCs secrete callose but not uniformly, and contact among them continues through meiosis. Simultaneous cytokinesis by furrowing isolates each microspore in callose, which later disperses. The separated microspores become vacuolate, undergo mitosis to become pollen, and later become filled with food reserves. Endothecial wall thickening and tapetal dissolution occur after pollen engorgement. Calcium oxalate crystals form in tapetal cells during the sporogenous stage, reach maximum size during early meiosis, and remain prominent until tapetal dissolution.     

Chemical agents that inhibit pollen development: effects of the phenyl-cinnoline carboxylates SC-1058 and SC-1271 on the ultrastructure of developing wheat anthers (Triticum aestivum L. var Yecora rojò)

Summary Phenylcinnoline carboxylate compounds SC-1058 and SC-1271 cause complete male sterility in wheat when applied at suitable dosages at the pre-meiotic stage of anther development. Anthers from treated and untreated plants were compared using light and electron microscopy from the pre-meiotic stage through the formation of nearly mature pollen. Overall anther development is gradually slowed in treated plants and pollen development is generally arrested in the late prevacuolate or early vacuolate microspore stage, although the first pollen mitosis does sometimes occur. The sporopollenin-containing exine walls are thinner, and show abnormally developed foot and tectum layers with sparse connecting baculi. Microspore cytoplasm degenerates and the cells eventually collapse. At the early, prevacuolate, free microspore stage treated tapetal cells hypertrophy, expanding into the locule. They contain abnormally large vacuoles that appear to form from the fusion of secretory vesicles, and some vacuoles contain electrondense deposits. The sporopollenin-containing orbicular wall and Ubisch bodies are retarded in their development and are structurally deformed. Acetolysis of whole anthers and of thick sections shows that the sporopollen-in-containing structures of treated materials are greatly reduced in thickness and are less rigid than in the control. We conclude that application of these compounds causes interference with the secretory function of tapetal cells which supplies sporopollenin cell-wall polymers to the exine of the microspores and to the tapetal orbicular wall and associated Ubisch bodies. Interference with the tapetal secretion of other nutrients required for microspore development is strongly suggested.     

Sporoderm in Populus and Salix

Four phenomena were observed in a study of Populus tremula and P. tremula f. gigas microspores from before microspore mitosis through mature pollen which may have general significance in the ontogeny of pollen grains: 1) The exine and orbicules (Ubisch bodies) were covered by membranes. 2) The exine and the tapetal surfaces where orbicules form were covered by a polysaccharide (PAS positive) coat until after microspore mitosis; subsequently the tapetum became plasmodial. 3) Material having the staining characteristics of the nexine 2 (endexine in the sense of Fægri) accumulated on membranes in microspores in the space between the exine and the plasma membrane. That material was almost completely gone from the wall in mature pollen. The membranes on which material had accumulated migrated through the exine. Following passage through the exine these membranes were seen as empty fusiform vesicles in micrographs of anthers prepared by commonly used methods. 4) At about microspore mitosis when the cellulosic intine begins to form, microtubules about 240 A in diameter occurred near the plasma membrane and generally parallel with it. Positive acid phosphatase reactions in tapetal cells together with the morphology of orbicules and other tapetal organelles suggest that the wall of orbicules, which is like the pollen exine, may form as a residual product of a lysosome system.

Sections of mature Salix humilis pollen were compared with Populus.     

Microsporogenesis in a normal line and in the ogu cytoplasmic male-sterile line of Brassica napus

Summary In the ogu cytoplasmic male-sterile (CMS) line of Brassica napus, stamen morphology was influenced by temperature conditions. Under a high temperature regime (27° C/23° C; day/ night) CMS stamens had a near-normal morphology, but microsporogenesis proceeded to a maximum of the microspore stage. However, compared to the normal stamens, the occurrence of sporopollenin-like deposits in the tapetum and deposition of exine on the microspores was sparse. Also, the tapetal cells of the CMS line were often highly vacuolate and failed to degenerate at the same stage as the normal. Ultrastructural changes in the mitochondrial matrix and cristae plus dilation of the endoplasmic reticulum, which occurred during development in sporogenous tissues of the normal line, were often lacking or mistimed in the mutant. Due to extensive variation, even between adjacent locules, the cytological differences between the normal and CMS anthers cannot be ascribed as the cause of male sterility in the ogu CMS line of B. napus, rather they may be the consequence of it.     

Two new male-sterile mutants of Zea mays (Poaceae) with abnormal tapetal cell morphology

Two new recessive male-sterile mutants of Zea mays (Poaceae), or maize, were studied to identify the timing of pollen abortion and to examine the involvement of anther wall cell layers. The results of test crosses indicated that these mutants were not allelic with any known male-sterile mutants of maize. Light and transmission electron microscopy were used to compare pollen development in homozygous male-sterile mutants to that in fertile heterozygous siblings. In both mutants, microspores abort soon after release from the meiotic tetrad. However, the two mutations have strikingly different phenotypes. Large lipid bodies accumulate in the tapetal cells as the microspores vacuolate and die in the mutant ms25. Large vacuoles appear in both the tapetal cells and the young microspores as they begin to disintegrate in the mutant ms26. Because abnormal tapetal cell morphology is detected in both mutants, it is possible that both of these mutations affect the expression of genes in tapetal cells.     

Cell differentiation in microsporangia of Pinus sylvestris. III. Late pachytene

Cycles of hyperactivity were observed in tapetal and microspore mother cells of Pinus sylvestris L. during the pachytene stage of meiosis in microspore mother cells. Hyperactive periods were characterized by dilated rough ER, hypersecretory dictyosomes, autophagic vesicles having one to two sequestration envelopes, and maze-like whor-les of the endomembrane system. The extent and sequence of differentiation differed between the two cell types. During distinct phases of development there was either fi-brillar flocculent or lipoidal material or both within dilations and at cell surfaces. Tapetal cell transfer of material involved endocytotic and exocytotic vesicles and channels opening directly to the cell surface. Dilations of the nuclear envelope of microspore mother cells in late pachytene intruded into the nucleus and, in conjunction with dilated ER, dominated cell profiles. Cellular morphogenesis in a microspo-rangium was seldom synchronous except for intervals of dedifferentiation when plas-modesma-like connections were formed between tapetal cells. Cycles of differentiation and dedifferentiation were correlated each spring season for five years with the progressive change of pachytene chromosomes, suggesting control by a genetic program rather than annual variations in the environment.     

Secretory tapetum of Brassica oleracea L.: polarity and ultrastructural features

Summary The ultrastructure of the secretory, binucleate tapetum of Brassica oleracea in the micro spore mother cell (MMC) stage through to the mature pollen stage is reported. The tapetal cells differentiate as highly specialized cells whose development is involved in lipid accumulation in their final stage. They start breaking down just before anther dehiscence. Nuclei with dispersed chromatin, large nucleoli and many ribosomes in the cytoplasm characterize the tapetal cells. The wall-bearing tapetum phase ends at the tetrade stage. The dissolution of tapetal walls begins from the inner tangential wall oriented towards the loculus and proceeds gradually along the radial walls to the outer tangential one. The plasmodesmata transversing the radial walls between tapetal cells persist until the mature microspore, long after loss of the inner tangential wall. After wall dissolution, the tapetal protoplasts retain their integrity and position within the anther locule. The tapetal cell membrane is in direct contact with the exine of the microspores/pollen grains and forms tubular evaginations that increase its surface area and appear to be involved in the translocation of solutes from the tapetal cells to the microspores/ pollen grains. The tapetal cells exhibit a polarity expressed by spatial differentiation in the radial direction.     

The loculus content and tapetum during pollen development in Lilium

 The ratio of loculus volume to the volume of the entire anther began to increase from the microspore mother cell stage and reached 32.3% at anthesis. The content of the loculus was examined in Lilium during pollen development and two waves could be distinguished. From the premeiotic stage until the vacuolated microspore stage, the loculus consisted of neutral polysaccharides, pectins and proteins. These substances originated from tapetal activity from the premeiotic stage until the young microspore stage. Dictyosomes and rough endoplasmic reticulum seemed to be involved in tapetal secretion, although, in some mitochondria, vesicles progressively developed as early as premeiosis and increased until the young microspore stage, which could reveal their involvement in the secretion process. At this stage, numerous cytoplasmic vesticles containing material similar to the locular material fused with the plasma membrane of the tapetum so that vesicle content was in contact with the loculus. It seems that tapetal and callose wall degradation at the late tetrad stage may also have contributed to the production of material in the loculus. From pollen mitosis to anthesis, the anther loculus contained mainly the pollenkitt which was synthesized in the tapetum between the young microspore stage and the vacuolated microspore stage. At the young microspore stage, proplastids divided and developed into elaioplasts and smooth endoplasmic reticulum (SER) increased dramatically. Pollenkitt had a double origin: some droplets were extruded directly from the plastid stroma through the plastid envelopes; the others were unsaturated lipid globules, which presumably derived from the interaction between SER saccules and plastids. Received: 2 September 1997 / Revision accepted: 12 March 1998     

Genetics and cytology of a genic male-sterile, female-sterile mutant from a transposon-containing soybean population

A male-sterile, female-sterile soybean mutant (w4-m sterile) was identified among progeny of germinal revertants of a gene-tagging study. Our objectives were to determine the genetics (inheritance, allelism, and linkage) and the cytology (microsporogenesis and microgametogenesis) of the w4-m sterile. The mutant was inherited as a single recessive nuclear gene and was nonallelic to known male-sterile, female-sterile mutants st2 st2, st3 st3, st4 st4, st5 st5, and st6 st6 st7 st7. No linkage was detected between the w4-m sterile and the w4w4, y10 y10, y11 y11, y20 y20, fr1 fr1, and fr2 fr2 mutants. Homologous chromosome pairing was complete in fertile plants. Chromosome pairing, as observed in squash preparation, was almost completely absent in sterile plants. Developmentally microsporogenesis proceeded normally in both the fertile and the w4-m sterile through the early microspore stage. Then the tapetal cells of the w4-m sterile surrounding the young microspores developed different-size vacuoles. These tapetal cells became smaller in size and separated from each other. Some of the microspores of the w4-m sterile also became more vacuolate prematurely and sometimes they collapsed, usually by the late microspore stage. In the w4-m sterile the microspore walls remained thinner and structurally different from the microspore walls of fertile plants. No pollen was formed in the mutant plants, even though some of the male cells reached the pollen stage, although without normal filling. The w4-m sterile was designated st8st8 and assigned Soybean Genetic Type Collection number T352.     

Differentiation of the Tapetum During Microsporogenesis in Tomato (Lycopersicon esculentum Mill.), with Special Reference to the Tapetal Cell Wall

During microsporogenesis and pollen maturation, the tapetumin anthers of tomato (Lycopersicon esculentum) underwent severalultrastructural changes and ultimately degenerated. The changesobserved related to the secretory function of the tapetum andto the transfer of materials from the cytoplasm to the surfaceof tapetal cells. Electron dense deposits, initially in thevacuoles, disappeared coincident with the appearance of orbiculeson the cell wall. The fibrillar wall of the tapetal cells loosened,presumably to facilitate transfer of materials through the wall.In Addition, membranous fragments were a consistent featurein the tapetum wall and may play a role in transport of materials.The cells of the inner tapetum (towards the connective) andouter tapetum (towards the epidermis) had different ultrastructuralfeatures. The cytoplasm of the outer tapetum was more electrondense and had a higher proportion of dictyosomes and mitochondriathan the inner tapetum, indicating the greater secretory natureof the outer tapetum. The plastids and mitochondria also differedin morphology between the two regions. Degenerations of thetapetal cytoplasm began by the vacuolate microspore stage. Atanthesis, cytoplasm was absent but the orbicular wall of thetapetum remained appressed to the wall of the middle layer ofthe anther.Copyright 1993, 1999 Academic Press Lycopersicon esculentum, microsporogenesis, pollen development, tapetum development, tomato, ultrastructure     

Microsporogenesis and tapetal development in fertile and cytoplasmic male-sterile sugar beet (Beta vulgaris L.)

Summary The development of sporogenous and tapetal cells in the anthers of male-fertile and cytoplasmic male-sterile sugar beet (Beta vulgaris L.) plants was studied using light and transmission electron microscopy. In general, male-sterile anthers showed a much greater variability in developmental pattern than male-fertile anthers. The earliest deviation from normal anther development was observed to occur in sterile anthers at meiotic early prophase: there was a degeneration or irregular proliferation of the tapetal cells. Other early aberrant events were the occurrence of numerous small vesicles in the microspore mother cells (MMC) and a disorganized chromatin condensation. Deviations that occurred in sterile anthers at later developmental stages included: (1) less distinct inner structures in the mitochondria of both MMC and tapetal cells from middle prophase onwards. (2) dilated ER and nuclear membranes at MMC prophase, in some cases associated with the formation of protein bodies. (3) breakdown of cell walls in MMCs and tapetal cells at late meiotic prophase. (4) no massive increase in tapetal ER at the tetrad stage. (5) a general dissolution of membranes, first in the MMC, then in the tapetum. (6) abortion of microspores and the occurrence of a plasmodial tapetum in anthers reaching the microspore stage. (7) no distinct degeneration of tapetal cells after microspore formation. Thus, it seems that the factors that lead to abortive microsporogenesis are structurally expressed at widely different times during anther development. Aberrant patterns are not restricted to the tetrad stage but occur at early prophase.     

扁豆绒毡层发育的超微结构研究

应用透射电镜对扁豆绒毡层发育过程进行了研究,主要结果如下：１）首次发现扁豆绒毡层在发育过程中,经历了二交胞质重组（第一次始于减数分裂末期Ⅱ,第二次始于小孢子发育早期）,使绒毡层细胞的活动呈现３个高峰期（即小孢子母细胞减数分裂期、小孢子四分体期一小孢子早期、小孢子晚期－二胞花粉中期）。２绒毡层细胞的分泌作用有３种形式（渗透分泌、胞吐分泌和自溶）。３．首次观察到绒毡层细胞的内切向壁和径向壁经历了两个周     

Cytochemical investigation of genic male-sterility in Chinese cabbage

A genic male sterile Chinese cabbage, Brassica campestris L. ssp. chinensis Makino, was examined using cytological and cytochemical methods to characterize the process of pollen abortion in this plant. Thick sections of both fertile and sterile anthers at different developmental stages were stained using Toluidine Blue O, Periodic Acid-Schiff’s (PAS) reaction and Sudan Black B to detect cytochemical changes that may occur in the distribution of insoluble polysaccharide and lipid storage bodies. Pollen abortion in sterile anthers occurs at an early stage of microspore development. During early microspore development, reductions in the number of starch grains in the connective tissue of fertile anthers coincide with the accumulation of starch grains in cells of the anther wall. In the late microspore stage, a large vacuole forms in the microspore, and tapetal cells synthesize and accumulate lipid droplets. The cellular organization of tapetal cells in sterile anthers appears similar to that in fertile anthers, except for the absence of lipid droplets in cells of sterile anthers and diffusely labeled tapetal polysaccharides, suggesting defects in nutrient storage. Supported by National Natural Science Foundation of CHINA (30170060)     

Cell differentiation in microsporangia of Pinus sylvestris with special attention to the tapetum. I. The pre–and early–meiotic periods

The tapetal layer becomes distinct from the other layers of parietal cells about three days prior to the meiosis in the microspore mother cells. Differentiation of the tapetal cells includes an increased relative volume for dictyosomes, mitochondria and plas–tids, the appearance of autophagic vacuoles in the cytoplasm, and periplasmic spaces between the plasma membrane and the cell wall. About one day before the meiosis the basophilia in tapetal cells is elevated; there are numerous nonaggregated ribo–somes, nuclei are intensely stainable, and the rough ER is dilated. There is also a partial digestion of the cell walls around microspore mother cells and tapetal cells including the adaxial wall of the adjacent parietal cell layer. A wedge–shaped portion of the wall system between this parietal cell layer and tapetal cells is not lysed. A lamellation in the middle lamellar position is also spared. That lamellation remains prominent as the extratapetal lamellation. By the initiation of meiosis the surfaces of both tapetal and microspore mother cells are entirely free of cell walls. During that period the intense basophilia of tapetal cells recedes and there are many polyribosomes, an extensive system of rough ER, dictyosomes with vesicles containing fibrils, multivesicular bodies, and autophagic vacuoles. Microtubules occur close to the plasma membrane. The plasma membrane–glycocalyx differs in portions of the surface facing the extratapetal lamellation from the Iocular facing surface. We presume that the abaxial portion of tapetal cells with cavations containing glycocalyx–like filaments is a region of uptake and that the adaxial surface with detached glycocalyx is secretory.     

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。     

Ultrastructure of microsporogenesis and microgametogenesis inArabidopsis thaliana (L.) Heynh. ecotype Wassilewskija (Brassicaceae)

Summary The process of microsporogenesis and microgametogenesis was studied at the ultrastructural level in wild-typeArabidopsis thaliana ecotype Wassilewskija to provide a basis for comparison with nuclear male-sterile mutants of the same ecotype. From the earliest stage studied to mature pollen just prior to anther dehiscence, microsporocyte/microspore/pollen development follows the general pattern seen in most angiosperms. The tapetum is of the secretory type with loss of the tapetal cell walls beginning at about the time of microsporocyte meiosis. Wall loss exhibits polarity with the tapetal protoplasts becoming located at a distance from the inner tangential walls first, followed by an increase in distance from the radial walls beginning at the interior edge and progressing outward. The inner tangential and radial tapetal walls are completely degenerated by the microspore tetrad stage. Unlike other members of the Brassicaceae that have been studied, the tapetal cells ofA. thaliana Wassilewskija also lose their outer tangential walls, and secretion occurs from all sides of the cells. Exine wall precursors are secreted from the tapetal cells in a process that appears to involve dilation of individual endoplasmic reticulum cisternae that fuse with the tapetal cell membrane and release their contents into the locule. Following completion of the exine, the tapetal cell plastids develop membranebound inclusions with osmiophilic and electron-transparent regions. The plastids undergo ultrastructural changes that suggest breakdown of the inclusion membranes followed by release of their contents into the locule prior to the complete degeneration of the tapetal cells.     

Genetic control of male fertility in Arabidopsis thaliana: structural analyses of postmeiotic developmental mutants

Seven new male-sterile mutants (ms7–ms13) of Arabidopsis thaliana (L.) Heynh. (ecotype columbia) are described that show a postmeiotic defect of microspore development. In ms9 mutants, microspores recently released from the tetrad appear irregular in shape and are often without exines. The earliest evidence of abnormality in ms12 mutants is degeneration of microspores that lack normal exine sculpturing, suggesting that the MS12 product is important in the formation of pollen exine. Teratomes (abnormally enlarged microsporocytes) are also occasionally present and each has a poorly developed exine. In ms7 mutant plants, the tapetal cytoplasm disintegrates at the late vacuolate microspore stage, apparently causing the degeneration of microspores and pollen grains. With ms8 mutants, the exine of the microspores appears similar to that of the wild type. However, intine development appears impaired and pollen grains rupture prior to maturity. In ms11 mutants, the first detectable abnormality appears at the mid to late vacuolate stage. The absence of fluorescence in the microspores and tapetal cells after staining with 4′,6-diamidino-2-phenylindole (DAPI) and the occasional presence of teratomes indicate degradation of DNA. Viable pollen from ms10 mutant plants is dehisced from anthers but appears to have surface abnormalities affecting interaction with the stigma. Pollen only germinates in high-humidity conditions or during in-vitro germination experiments. Mutant plants also have bright-green stems, suggesting that ms10 belongs to the eceriferum (cer) class of mutants. However, ms10 and cer6 are non-allelic. The ms13 mutant has a similar phenotype to ms10, suggesting is also an eceriferum mutation. Each of these seven mutants had a greater number of flowers than congenic male-fertile plants. The non-allelic nature of these mutants and their different developmental end-points indicate that seven different genes important for the later stages of pollen development have been identified. Received: 14 August 1997 / Accepted: 7 October 1997