DEVELOPMENTAL MORPHOLOGY OF COTTON FLOWERS AND SEED AS SEEN WITH THE SCANNING ELECTRON MICROSCOPE

This report assembles and pictorially presents observations on the timing of relatively uniform and well-defined developmental events in the cotton flower and its component parts. The first floral bud occurs on the 7–9th node approximately 35–40 days postemergence; 20–25 additional days elapse until anthesis. Floral parts are morphologically well defined by two weeks preanthesis. In about 85 % of the flowers the basal, abaxial surface of two of the three bracts contains an outer involucral nectary; occasionally, none, one, or three nectaries are found. The maximum rate of increase in floral bud length occurs during the 24 hrs preceding anthesis. Flower opening occurs at about daylight, although light is not required. Multipored pollen grains germinate in about ½ hr after deposition on the stigmatic hairs. Fertilization is accomplished, for most ovules, by the end of the first day postanthesis. Stomata are abundant, particularly at the chalazal ends of ovules. Fiber initials (epidermal cells of the ovule) begin their elongation phase on the morning of anthesis and are bounded by a thin primary wall. Areas of contrast (spots) observed through the scanning electron microscope are speculated to be organelles “seen through” the relatively amorphous fiber wall, which lacks extensive fibrillar orientation of cellulose. Fiber elongation ceases by about 24–28 days postanthesis, and by 50–70 days postanthesis fibers are mature and exhibit a thickened secondary wall and spiral twisting. Concomitant with the time of fiber maturity, the ovary wall splits and opens along locular suture lines.     

莴苣助细胞发育过程中钙的分布研究

用焦锑酸盐沉淀法对莴苣助细胞中的钙分布进行了观察。结果表明,开花前3天刚形成的助细胞中的钙颗粒很少:开花前2天助细胞壁中的钙颗粒增加;开花前1天助细胞珠孔端细胞壁加厚,其中积累了许多钙颗粒:开花当天助细胞珠孔端的丝状器中聚集了大量的钙颗粒。授粉后1h时两个助细胞的结构和钙分布发生差异,一个呈退化状,其中的钙颗粒明显增多,另一宿存助细胞中的钙分布与授粉前相似。去雄不授粉1天后两个助细胞均保持完好,且两助细胞中的钙分布没有明显差异,表明由花粉管引起一个助细胞中钙含量增加进而导致了助细胞退化。退化助细胞在卵细胞与中央细胞之间形成一薄层。助细胞退化后不同部位的钙颗粒呈现出与受精作用密切有关的变化:授粉后1h时,钙主要聚集在近合点端部位;授粉后2.5h卵细胞即将受精,这时许多细小的钙颗粒主要聚集在卵细胞与中央细胞之间的薄层中;授粉后4h精、卵细胞已融合,这时退化助细胞合点端的钙颗粒明显减少,而在其珠孔端又聚集了较多的钙。上述助细胞中的钙含量变化与吸引花粉管进入胚囊和促使精卵细胞融合密切有关。     

Chronology of the differentiation of cotton (Gossypium hirsutum L.) fiber cells

Cotton fibers are single elongated cells that develop from epidermal cells of the ovule. The chronology of fiber differentiation was investigated using cultured ovules. Epidermal cells differentiate into fiber cells approx. 3 d before anthesis. When ovules were cultured on a defined medium, fiber growth could be initiated on ovules any time between 2 d preanthesis and the time of anthesis by adding indole-3-acetic acid and gibberellic acid to the medium. In the absence of phytohormones, fibers did not grow, and when ovules between 2 d preanthesis and anthesis were cultured without hormones past the day of anthesis and hormones then added, most ovules failed to produce fibers. The results define the timing of fiber differentiation from epidermal cells, and also define a window of time when differentiated cells are capable of further development. During this window, fiber cells are latent awaiting appropriate stimulation which in the intact plant is apparently associated with anthesis.Abbreviations GA₃ gibberellic acid - IAA indole-3-acetic acid     

Development of Embryo and Endosperm and Nutrient Accumulation in Vigna sesquipedalis (L.) Fruwirth

The structure of embryo sac, fertilization and development of embryo and endosperm in Vigina sesquipedalis (L.) Fruwirth were investigated. Pollization occures 7–10h before anthesis, and fertilization is completed 10 h after anthesis. After fertilization, wall ingrowths are formed at the micropylar and chalazal ends of the embryo sac. Embryo development conforms to the Onagrad type, and passes through 2 or more celled proembryo, long stick-shaped, globular, heart shaped, torpedo, young embryo, growing and enlarging embryo and mature embryo. Wall ingrowths are formed on the walls of basal cells and outer walls of the cells at basal region of suspenser. The suspensor remains as the seed reaches maturity. The starch grains accumulate in the cells of cotyledons by 9–16 days after anthesis, and proteins accumulate by 12–18 days after. The endosperm development follows the nuclear type. The endosperm ceils form at the micropylar end, and remain free nuclear phase at chalazal end. The outer cells are transfer cells. Those cells at the micropylar end form folded cells with wall ingrowths. At heartembryo stage, the endosperm begins to degenerate and disintegrates before the embryo matures.     

CHEMOTROPIC ACTIVITY AND THE PATHWAY OF THE POLLEN TUBE IN LILY,

A study of the pollen tube pathway in Lilium leucanthum var. centifolium and in L. regale reveals that the entire pathway from stigma to ovule is lined with cytologically unique stigmatoid cells. Assays for chemotropic activity of tissues and exudates along the pathway of pollinated or unpollinated pistils showed that onset of chemotropic activity progressed basipetally (and, when pollinated, in advance of the pollen tubes), commencing at the stigma 3-5 days before anthesis and appearing in the ovules 1-2 days after anthesis. Activity persists about 10 days in ovules of pollinated pistils and for 14-16 days in ovules of non-pollinated pistils. Attempts to localize the source of the chemotropic factor showed that gynoecial tissues bearing stigmatoid cells are chemo-tropically active while slices of style or ovary wall lacking stigmatoid cells are inactive. When ovules were sliced transversely and the micropylar and chalazal halves assayed, only the micropylar half showed activity. We suggest that the ovules and the stigmatoid tissue along the pollen tube pathway are the sources of the chemotropic factor responsible for the directional growth of the pollen tube.     

CHANGES IN MEGAGAMETOPHYTE STRUCTURE IN PAPAVER NUDICAULE L. (PAPAVERACEAE) FOLLOWING IN VITRO PLACENTAL POLLINATION

Papaver nudicaule placentae with attached ovules were dissected out of unpollinated gynoecia 1–3 days after anthesis, dusted with pollen, and cultured on modified Nitsch's growth medium at 23 C. Ovules were removed from expiants at 15, 24, 31, and 48 hr postpollination, fixed in GA-OsO₄, embedded in Spurr's resin and sectioned (1.0 μm) for light microscopy. Placentae, 15 hr after pollination, were fixed and processed for scanning electron microscopy. Pollen germinates within 1 hr. Although most pollen tube growth appears random, there is directional growth toward the micropyle. The crassinucellate ovule contains an embryo sac consisting of three antipodals, two polar nuclei, and an egg apparatus composed of two synergids and a polarized egg having a large chalazal vacuole and micropylar nucleus. Pollen tube access into the megagametophyte is through a degenerate synergid, with fertilization occurring between 24 and 31 hr after pollination. Zygote establishment is accompanied by polarity reversal in which the nucleus assumes the chalazal position subtended by a large micropylar vacuole. Fertilized ovules normally develop into germinable seeds.     

Comparative Development of Lint and Fuzz Using Different Cotton Fiber-specific Developmental Mutants in Gossypium hirsutum

A series of fiber-specific mutants, or germplasms, have been recently used in the study of fiber development. In the current study, scanning electron microscopy （SEM） was used to investigate developmental differences in lint and fuzz initiation in different genotypes （Gossypium hirsutum） of upland cotton. These fiber mutants included dominant naked seed N1, recessive naked seed n2, Xuzhou-142 lintless-fuzzless （XZ142WX）, Xinxiangxiaojilintless-fuzzless （XinWX）, Xinxiangxiaojilinted-fuzzless （XinFLM）, with TM-1, the cytogenetic and genetic experimental standard stock, as the control. Characteristics of fiber initiation were analyzed from -1 to ＋1 days post anthesis （dpa） and at 4 and 5 dpa for fuzz initiation. Our data suggested that lint initiation centered on day of anthesis （0dpa）, and elongated significantly at 1dpa, while fuzz initiation began at 4dpa, although the shape of fuzz protrusions differed from that of lint fibers. Fiber initiation occurred first on the ovule funicular crest. Compared to TM-1, there was a noted retardation in development and fiber protrusion in N1 and XinFLM. Microscopy data also demonstrated that lintless-fuzzless mutants （XZ142WX and XinWX） developed irregular protrusions during early developmental stages, which were unable to grow into fiber.     

Comparative Development of Lint and Fuzz Using Different Cotton Fiber-specific Developmental Mutants in Gossypium hirsutum

A series of fiber-specific mutants, or germplasms, have been recently used in the study of fiber development. In thecurrent study, scanning electron microscopy (SEM) was used to investigate developmental differences in lint and fuzzinitiation in different genotypes (Gossypium hirsutum) of upland cotton.These fiber mutants included dominant nakedseed N1, recessive naked seed n2, Xuzhou-142 lintless-fuzzless (XZ142WX), Xinxiangxiaojilintless-fuzzless (XinWX),Xinxiangxiaojilinted-fuzzless (XinFLM), with TM-1, the cytogenetic and genetic experimental standard stock, as the control.Characteristics of fiber initiation were analyzed from -1 to 1 days post anthesis (dpa) and at 4 and 5 dpa for fuzz initiation.Our data suggested that lint initiation centered on day of anthesis (Odpa), and elongated significantly at 1dpa, while fuzzinitiation began at 4dpa, although the shape of fuzz protrusions differed from that of lint fibers. Fiber initiation occurred firston the ovule funicular crest. Compared to TM-1, there was a noted retardation in development and fiber protrusion in N1and XinFLM. Microscopy data also demonstrated that lintless-fuzzless mutants (XZ142WX and XinWX) developed irregularprotrusions during early developmental stages, which were unable to grow into fiber.     

ADVENTIVE EMBRYOGENESIS IN CITRUS (RUTACEAE) II. POSTFERTILIZATION DEVELOPMENT

Cytological and histological studies on postfertilization development of ovules were carried out in six facultatively apomictic Citrus cultivars. At the time of anthesis, adventive embryo initial cells (AEICs) were detected mainly in the cell layers of the nucellus around the chalazal half of the embryo sac. During the approximately 40 days rest period of the AEICs after fertilization, rapid cell division and enlargement in the endosperm and the chalazal half of the nucellus resulted in the split of AEICs into several separated areas forming the micropylar, lateral and chalazal islands surrounding the enlarging embryo sac. Both in diploid seeds with triploid endosperm and triploid seeds with pentaploid endosperm, the AEICs located in the micropylar half successfully developed into adventive embryos. In diploid seeds, almost all AEICs located in the chalazal half did not develop beyond the initial-celled stage, while in the triploid seeds, those located in the chalazal half occasionally developed into cotyledonary embryos. In seeds with aborted endosperm, the AEICs located in the chalazal half often developed into cotyledonary embryos. The chalazal expiants from normal seeds produced a large number of embryos in vitro. Four results can be summarized from these studies on adventive embryogenesis as follows: 1) All AEICs are initiated prior to anthesis. 2) Whether or not the AEICs successfully developed into adventive embryos is dependent upon their position in the seed. 3) The farther the AEICs are located from the micropylar end, the more adventive embryogenesis is suppressed by endosperm. 4) The degree of adventive embryogenesis in the chalazal half is affected by time and extent of malfunction of the endosperm. Under natural conditions, these regulatory systems of adventive embryogenesis contribute to high production of zygotic seedlings in apomictic Citrus species and cultivars.     

EMBRYOLOGY OF CALYPSO BULBOSA. I. OVULE DEVELOPMENT

Calypso bulbosa is a terrestrial orchid that grows in north temperate regions. Like many orchids, the Calypso has ovules that are not fully developed at anthesis. After pollination, the ovule primordia divide several times to produce a nucellar filament which consists of five to six cells. The subterminal cell of the nucellar filament enlarges to become the archesporial cell. Through further enlargement and elongation, the archesporial cell becomes the megasporocyte. An unequal dyad results from the first meiotic division. A triad of one active chalazal megaspore and two inactive micropylar megaspores are the end products of meiotic division. Callose is present in the cell wall of the megaspore destined to degenerate. In the mature embryo sac the number of nuclei is reduced to six when the chalazal nuclei fail to divide after the first mitotic division. The chalazal nuclei join the polar nucleus and the male nucleus near the center of the embryo sac subsequent to fertilization.     

Spatial distribution of mRNA during pre-fertilization ovule development in Capsella bursa-pastoris

Summary In situ hybridization of sections of the ovary and ovule of Capsella bursa-pastoris with ³H-polyuridylic acid [³H-poly(U)] showed the presence of polyadenylic acid-containing RNA [poly(A) + RNA] in the cells of the placenta and nucellus. During megasporogenesis there was a decrease in ³H-poly(U) binding activity of the nucellar cells concomitant with the appearance of poly(A) + RNA in the integuments. As the typical eight-nucleate embryo sac was formed, ³H-poly(U) binding was not apparent around the quartet of nuclei at the chalazal end, while it persisted at the micropylar end. Both the egg and synergids as well as the chalazal proliferating tissue showed high concentrations of poly(A) + RNA in their cytoplasm. The results suggest a role for transient localizations of poly(A) + RNA during female sporogenesis and gametogenesis in C. bursa-pastoris.     

Effect of phytohormones on fiber initiation of cotton ovule

In order to study the effect of phytohormones on cotton fiber initiation, contents of four endogenous phytohormones and activities of four related enzymes in ovules (in vivo) of a fuzzless–lintless mutant (fl) and its wild-type (FL) line were measured from 4 days before anthesis (day −4) to 4 days after anthesis (day 4). The results showed that contents of indole-3-acetic acid, gibberellic acid (GA), and zeatin riboside in fl ovules were lower than those in FL ovules. Therefore, indole-3-acetic acid, GA, and zeatin riboside were thought to be the promoters of fiber initiation. Although abscisic acid (ABA) content in fl ovule was slightly higher than that in FL ovule on day 0, which might imply that ABA inhibited fiber initiation. Fiber initiation could also be influenced by enzyme through regulating synthesis and degradation of related phytohormones since fl ovules were significantly higher in activities of indole-3-acetic acid oxidase, cytokinin oxidase and peroxidase, but lower in activity of tryptophan synthetase than those in FL ovules. To test the above hypothesis, exogenous plant growth regulators were also applied for the culture of ovules from fl and FL in vitro. When no regulators were added, no fiber was induced on fl ovule, but a few fibers were induced in FL ovule. Higher total fiber units (TFU) were observed when indole-3-acetic acid and gibberellic acid (GA₃) were applied either separately or in combination to media. TFU did not increased with zeatin riboside alone, but the highest TFU was achieved when zeatin riboside was applied together with indole-3 acetic acid and GA₃, which implied that fiber initiation could be promoted by them as additive.     

Suppression of sucrose synthase gene expression represses cotton fiber cell initiation,elongation, and seed development

Cotton is the most important textile crop as a result of its long cellulose-enriched mature fibers. These single-celled hairs initiate at anthesis from the ovule epidermis. To date, genes proven to be critical for fiber development have not been identified. Here, we examined the role of the sucrose synthase gene (Sus) in cotton fiber and seed by transforming cotton with Sus suppression constructs. We focused our analysis on 0 to 3 days after anthesis (DAA) for early fiber development and 25 DAA, when the fiber and seed are maximal in size. Suppression of Sus activity by 70% or more in the ovule epidermis led to a fiberless phenotype. The fiber initials in those ovules were fewer and shrunken or collapsed. The level of Sus suppression correlated strongly with the degree of inhibition of fiber initiation and elongation, probably as a result of the reduction of hexoses. By 25 DAA, a portion of the seeds in the fruit showed Sus suppression only in the seed coat fibers and transfer cells but not in the endosperm and embryo. These transgenic seeds were identical to wild-type seeds except for much reduced fiber growth. However, the remaining seeds in the fruit showed Sus suppression both in the seed coat and in the endosperm and embryo. These seeds were shrunken with loss of the transfer cells and were <5% of wild-type seed weight. These results demonstrate that Sus plays a rate-limiting role in the initiation and elongation of the single-celled fibers. These analyses also show that suppression of Sus only in the maternal seed tissue represses fiber development without affecting embryo development and seed size. Additional suppression in the endosperm and embryo inhibits their own development, which blocks the formation of adjacent seed coat transfer cells and arrests seed development entirely.     

大车前胚乳发育的超微结构研究

采用透射电镜技术对大车前(Plantago major L.)胚乳发育的超微结构进行了研究。结果表明:(1)大车前为细胞型胚乳;初生胚乳核经一次横分裂产生1个珠孔室细胞和1个合点室细胞;珠孔室两次纵向分裂一次横向分裂形成2层8个细胞,位于上层的4个细胞发育为4个珠孔吸器,位于下层的4个细胞发育为胚乳本体;合点室细胞进行一次核分裂,发育为两核的合点吸器。(2)珠孔吸器呈管状插入珠被组织,珠孔端细胞壁加厚呈现少量分支并具有壁内突,壁内突周围细胞质里分布着大量线粒体、粗面内质网、高尔基体、质体等,细胞核与核仁明显,细胞质浓厚,代谢活动旺盛;球胚期,珠孔吸器的体积呈现最大值,珠孔吸器周围的珠被组织均被水解,形成明显的空腔。珠孔吸器从珠被组织吸收并转运营养物质至胚乳本体,参与胚乳的构建与营养物质的贮藏。球胚后期,珠孔吸器逐渐退化。(3)4个胚乳本体原始细胞具旺盛的分生能力,经不断的平周与垂周分裂增加胚乳细胞数目,使胚乳本体呈现圆球体状,并将胚包围其中;珠孔吸器、合点吸器以及珠被绒毡层吸收转运的营养物质贮存在胚乳本体;球胚后期,随着胚柄的退化,胚体周围的胚乳细胞被水解,为发育的胚所利用。(4)合点吸器的2个细胞核与核仁巨大,线粒体、质体、高尔基体、内质网主要绕核分布,液泡化明显;胚体与胚乳本体的体积增大,逐渐将合点吸器向胚珠合点部位挤压,合点吸器周围的合点组织逐渐被水解,形成巨大空腔。合点吸器自珠心组织吸收并转运营养物质至胚乳本体,参与胚乳的结构构建与营养物质的贮藏。球胚后期,合点吸器逐渐失去功能,呈现退化状态。     

Spatiotemporal manipulation of auxin biosynthesis in cotton ovule epidermal cells enhances fiber yield and quality

The capacity of conventional breeding to simultaneously improve the yield and quality of cotton fiber is limited. The accumulation of the plant hormone indole-3-acetic acid (IAA) in cotton fiber initials prompted us to investigate the effects of genetically engineering increased IAA levels in the ovule epidermis. Targeted expression of the IAA biosynthetic gene iaaM, driven by the promoter of the petunia MADS box gene Floral Binding protein 7 (FBP7), increased IAA levels in the epidermis of cotton ovules at the fiber initiation stage. This substantially increased the number of lint fibers, an effect that was confirmed in a 4-year field trial. The lint percentage of the transgenic cotton, an important component of fiber yield, was consistently higher in our transgenic plants than in nontransgenic controls, resulting in a >15% increase in lint yield. Fiber fineness was also notably improved.     

Influence of intraovular reserves on ovule fate in apricot (Prunus armeniaca L.)

 In many plant species with multiovulate ovaries, a considerable reduction in the number of ovules takes place. However, the underlying physiological causes are not clear. In Prunus spp., although flowers present two ovules, usually only one seed is produced. We have followed the development and degeneration of the two ovules in apricot (Prunus armeniaca L.) and examined the extent to which carbohydrates within the ovule might be involved in determining the fate of the ovule. While the primary ovule grows in the days following anthesis, growth of the secondary ovule is arrested. Starch distribution along the different ovular tissues exhibits several changes that are different in the two ovules. Primary ovule growth is inversely related to starch content and this growth takes place independently of pollination since it occurs in the same way in pollinated and unpollinated flowers. In the secondary ovule, starch disappears simultaneously from all ovular structures and callose is layered at the chalazal end of the nucellus. The size of the secondary ovule does not change significantly from anthesis to degeneration, and callose starts to accumulate 5 days after anthesis. Likewise, this process occurs independently of pollination. These results are discussed in terms of the implications of the starch content of ovules in fertilization success and ovule fate. Received: 26 August 1997 / Revision accepted: 17 December 1997     

Localization of Callose during the Occurrence of Megasporocyte in Gastrodia elata Blume

The structure of ovule in Gastrodia elata Blume was very simple. Functional megaspore occurred at the chalazal end. Callose was absent at megasporocyte stage. It first appeared at the chalazal wall during the first meiotic prophase and exhibited continuous fluorescence. Soon later callose fluorescence disappeared in some part of the chalazal wall and many noncallosic dark areas took place, subsequently these nonfluorescence areas became larger and the callose fluorescence appeared discontinuous granulose distribution. This fluorescence maintained until the megaspore formed. The callose of micropylar wall appeared later and usually disappeared before megaspore formation. In the cross walls between the functional and the two degenarated megaspore callose fluorescence was very strong, continued and kept for a long time. But the side walls usually lacked callose. Accoding to the morphological character of simple ovule in G. eiata and the localization of acid phosphatase and polysaecharide grains, the transfer of vegetative materials from surrounding tissues into megasporocyte mainly passing through the chalazal end of megasporocyte. Thus a continuous callose wall deposited at the ehalazal end of megasporocyte, and it in reality caused the “isolation” of meiocyte. It was possible that a reduced form of callose disposition existed in parasetic orchids.     

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。