甘蓝型油菜雌性不育突变体FS-M1乳突细胞的细胞学观察

雌性不育突变体FS-M1是从甘蓝型油菜(Brassica napus)品种宁油10号中发现的.为了从细胞学角度研究FS-M1的雌性不育机理,利用荧光显微镜、扫描和透射电子显微镜观察分析了FS-M1柱头乳突细胞的授粉行为和超微结构.结果表明:花粉粒能在FS-M1乳突细胞上附着和萌发形成花粉管,但花粉管无法穿越柱头乳突细胞;开花后的FS-M1乳突细胞迅速衰退而呈干瘪萎蔫状,在衰退过程中,FS-M1柱头乳突细胞的细胞器数量减少,细胞液泡化明显,高尔基体、内质网和线粒体等一些细胞器结构被逐渐破坏.因此,推测FS-M1的雌性不育性是由于柱头乳突细胞发育异常造成的.     

十字花科10属种与油菜萝卜胞质不育系杂交的花粉萌发情况观察

对10属种十字花科植物与油菜萝卜胞质不育系杂交时花粉在柱头上粘合、萌发、花粉管伸长等情况进行观察。结果表明：（1）海甘蓝花粉粒粘合较难;（2）48 h内无瓣焊菜〖WTBX〗(Rorippa dubia)〖WTBX〗、毛果诸葛菜(Orychophragmus violaceus)、桂竹香(Cheiranthus cheiri)、海甘蓝(Crambe abyssinica)花粉管的伸长受阻于花粉萌发启动之时,花粉壁内形成胼胝质塞;播娘蒿、紫罗兰、荠菜花粉管伸长但未进入乳突细胞;芝麻菜花粉管进入乳突细胞而未进入柱头,‘浠水白’(Brassica campestris)、蓝花子有花粉管进入柱头及花柱而未进入胚囊。     

百合远缘杂交花粉萌发及花粉管生长过程观察

利用荧光显微镜对百合远缘杂交组合Bernini×Pollyanna花粉萌发及花粉管生长过程进行观察研究,结果显示,授粉后3~30 h内花粉萌发形成花粉管,且花粉管生长速度由快到慢,48~51 h内花粉管停止生长,花粉管最后深入到花柱的1/3处,并观察到一些异常的花粉管形态;花粉管生长过程中还伴随着一系列的胼胝质反应,出现的部位依次是乳突细胞、花粉管、花柱通道细胞、胚珠中的胚囊.结果表明该杂交组合不亲和.研究认为Berni-ni柱头乳突细胞是阻碍Pollyanna花粉管生长的第一道屏障,花柱通道细胞是抑制花粉管在花柱内生长的第二道屏障.     

水稻核不育系4个柱头性状的遗传分析

为改良水稻(Oryza sativa)核不育系柱头性状提供遗传信息, 调查了粳型核不育系7001S、籼型核不育系Z913S及其杂交、自交获得的F₁、F₂和F_2:3群体的4个柱头性状, 分析了4个性状间的相关性, 并利用主基因+多基因遗传模型对2个世代4个性状进行遗传分析。结果表明, 4个性状两两间呈极显著正相关, 相关系数介于0.274-0.897之间。除F_2:3群体中花柱长度和柱头外露率分别表现出受2对加性-显性主基因和1对负等效加性-显性主基因+多基因控制外, F₂和F_2:3群体的柱头长度、花柱长度、柱头-花柱总长度以及柱头外露率均表现出受2对主基因和多基因控制, 且F_2:3群体中控制花柱长度的主基因表现出加性-显性效应, 其余均表现出加性-显性-上位性效应。2个世代中4个性状均以主基因遗传为主。     

猕猴桃花粉原位生长过程中Ca2+的超微细胞化学定位

应用扫描电镜和荧光显微镜对软枣猕猴桃同种花粉在柱头上原位萌发及花粉管生长情况进行了观察,并用焦锑酸盐沉淀法对其授粉前后柱头及花柱中Ca2+进行超微细胞化学定位.结果显示:(1)猕猴桃柱头属于干性柱头,具有一道裂沟,乳突呈长圆柱形,授粉前后形态差异不明显.(2)授粉后3 h,花粉管生长穿过柱头表面,授粉后7 h,花粉管生长到达花柱底部;(3)授粉前后,柱头接受面靠近柱头外围细胞的角质层一侧细胞器内含有丰富的钙,而柱头非接受表面钙颗粒分布很少;(4)授粉前和授粉后3 h,花柱顶端钙颗粒较少,基部钙颗粒较多;授粉后7 h,花柱顶端和基部钙分布密度无明显差别;(5)授粉前后花柱顶端钙主要均匀分布在细胞质膜位置;在花柱基部授粉前钙主要分布在引导组织胞间隙中,授粉后3 h主要分布在细胞质内,授粉后7 h主要存在于细胞质、内质网上.研究表明,猕猴桃授粉前后,柱头和花柱组织中均含有钙,授粉前和授粉后3 h花柱中的钙呈现出梯度分布,授粉后7 h钙的梯度分布现象减弱甚至消失.     

桦木科植物花柱适应风媒传粉的特征

植物体为适应自己的传粉系统, 表现出高度的适应特征。风媒花植物为适应风传播花粉, 要形成特殊的结构, 以扩大接受花粉粒的面积。利用扫描电镜观察了桦木科(Betulaceae)6属18种植物花柱的形态及花粉粒在花柱上的萌发过程, 探讨了桦木科植物花柱适应风媒传粉的特征。结果表明, 桦木科植物的二心皮(铁木、云南鹅耳枥稀为三心皮)雌蕊具柱状花柱, 柱头不发达, 花柱表皮细胞长条状, 纵向排列紧密。传粉时, 花柱表皮细胞能执行柱头的功能, 接受花粉粒, 为花粉粒萌发提供场所和萌发条件。桦木科植物花柱有2种类型: 一种是花柱表皮细胞能形成乳突, 花粉管经乳突细胞进入花柱; 另一种是花柱表皮细胞不形成乳突, 花粉管经过花柱表皮细胞或胞间隙进入花柱。无论花柱表皮细胞是否形成乳突, 乳突的形态、大小以及花粉管和乳突的结合方式等在族间、属间、属内种间存在差异。与基部被子植物相比, 桦木科植物的花柱呈现适应风媒传粉的进化特征。桦木科植物花柱表皮细胞形成的乳突与基部被子植物柱头乳突功能相同, 是桦木科植物风媒传粉的适应策略。     

高等植物受精过程中雌-雄相互作用的分子调控机制

从广义上讲,被子植物的受精过程是指花粉粒落到柱头上萌发形成花粉管,花粉管穿过柱头沿着引导组织生长进入子房内,最终在胚囊中实现精细胞与卵细胞以及中央细胞分别融合从而起始胚胎和胚乳的发育．被子植物的精细胞由于不具有鞭毛而无法自由移动,因此在受精过程中需要借助于花粉管来将精细胞运送到胚囊中．花粉管通过与雌性的孢子体组织之间的相互作用和识别将精细胞准确地运送到胚珠附近,而最终将精细胞准确地运送到胚囊内的过程则是受到了雌配子体细胞的控制．可以说,受精的成功实现有赖于雌性和雄性细胞之间的持续的识别和相互作用,这种互作具有多样性和阶段特异性．本文将主要综述被子植物受精过程中花粉粒以及花粉管与多种雌性孢子体组织以及雌配子体之间的信号互作研究．     

甘蓝油菜雌蕊中花粉管生长途径的钙离子定位

用焦锑酸钾定位 Ca~(2 )的技术,证明了甘蓝油菜(Brassica napus)柱头乳突细胞的少量分泌物、柱头表皮下的引导组织细胞间隙、花柱引导组织细胞壁外表面和胞间隙的分泌物以及子房假隔膜分泌区的表皮细胞外和内部引导组织细胞壁外表面和胞间隙的分泌物中均有 Ca~(2 )沉淀存在,而花柱皮层薄壁细胞无此种现象。花粉管生长的整个途径中 Ca~(2 )的存在与花粉管在雌蕊中的定向生长有关。     

荞麦亲和与不亲和授粉后柱头和花粉管中ATP酶的超微细胞化学定位

利用磷酸铅沉淀技术对荞麦(Fagopyrum esculentum Month.)pin型植株分别进行亲和授粉和不亲和授粉后的柱头、花粉粒、花粉管进行了ATPase的超微细胞化学定位。结果表明(1)亲和授粉和不亲和授粉后0.5h,柱头细胞的ATPase活性反应水平较低或基本无酶活性;柱头表面、柱头上附着的花粉粒内ATPase活性在不亲和授粉时较低,亲和授粉时较高,花粉粒内ATPase主要定位于线粒体和精子细胞;(2)授粉后1.5h,不亲和授粉的柱头细胞及花粉管的ATPase活性均较低,花粉管停止生长,细胞质开始解体;而亲和授粉的柱头细胞及花粉管的ATPase活性均较高,ATPase主要定位于柱头细胞的质膜、胞基质以及花粉管的壁、质体的膜、高尔基体、线粒体上。根据不同时期不同部位ATPase活性的差异,我们认为荞麦发生自交不亲和时,花粉管在花柱中停止生长不仅是因为花粉管得不到花柱中的营养物质而引起的,可能也与花粉管自身物质代谢发生障碍有关。     

青花菜自交不亲和特性的荧光鉴定

利用亲和指数和荧光显微测定法对青花菜高代自交材料进行自交不亲和特性的快速鉴定。利用蕾期和花期授粉进行亲和指数测定。荧光显微观察发现自交亲和材料与不亲和材料蕾期授粉花粉能够正常萌发,花粉管穿越柱头乳突细胞到达子房完成授粉受精过程;自交不亲和材料花期授粉后花粉与柱头乳突细胞表面发生强烈胼胝质反应,花粉在柱头上不能萌发,即使有少量萌发,其花粉管生长也不正常,最终不能伸入柱头。荧光显微观察法结果准确、方便,受外界环境条件影响较小,且与亲和指数法鉴定结果相一致,可有效应用于青花菜自交不亲和特性的快速鉴定,从而加速新品种选育进程。     

Changes in pollen tube behaviour induced by carbon dioxide and their role in overcoming self-incompatibility in Brassica

Summary Comparative studies on self-pollination after the application of high concentrations of CO₂ gas, which is known to overcome the self-incompatibility reaction in Cruciferous species, have revealed significant changes in the pollen tubes of germinating grains prior to their penetration into the papilla cells. A remarkable increase in the width of the pollen tube was induced by treating the stigmatic papillae with high CO₂ concentrations. The width of the pollen tube appeared to be greatest with CO₂ concentrations ranging from 3% to 5%; these concentrations were also optimal for tube penetration. Callose accumulation was extensively induced in the stigmatic papilla with 10%–20% of CO₂, although a typical callosic reaction remained through the ranges appropriate for blocking self-incompatibility. Observations using the scanning electron microscope (SEM) after pollination revealed that compatible pollen tubes in cross-pollinations fused completely to the papular surface during tube penetration, while in self-pollination, pollen tubes remained on the papilla with some additional diffusate. In the case of CO₂ treatment for self-pollination, some pollen tubes behaved very similarly to the incompatible or compatible ones already described, while others were different from both of them: they showed a complete fusion, similar to compatible ones, with additional diffusate, similar to incompatible ones. These responses of the pollen and stigma to high CO₂ concentrations are discussed with respect to their effect upon the expression of self-incompatibility.     

Sperm cells of the pollen tubes of Brassica: Ultrastructure and isolation

Summary Sperm cells of pollen tubes grown both in vivo and in vitro form a male germ unit. Extensions from both sperm cells of each pollen tube are closely associated with the tube nucleus. A high yield (2.7 × 10⁴. 20 mg^–1 pollen grains germinated) of intact sperm cells was obtained following release by osmotic shock from pollen tubes grown in vitro. Structural integrity of isolated sperm was maintained by isolation at low temperature in an osmotically balanced medium. At 4° C many isolated sperm pairs were still enclosed within the pollentube inner plasma membrane. Sperm cells not enclosed within this membrane no longer remained connected as a pair. During isolation vesicles formed on the sperm cell surface from disruption of the fibrillar components bridging the periplasmic space. Both in the pollen tube and after isolation the sperm nucleus is in close association with at least one region of the sperm plasma membrane. Sperm isolated at room temperature showed the presence of nucleopores, and nuclei were euchromatic, instead of heterochromatic as in intact sperm in the pollen tube.     

Cryptic dioecy in Mussaenda pubescens (Rubiaceae): a species with stigma-height dimorphism

Background and Aims

Evolutionary transitions from heterostyly to dioecy have been proposed in several angiosperm families, particularly in Rubiaceae. These transitions involve the spread of male and female sterility mutations resulting in modifications to the gender of ancestral hermaphrodites. Despite sustained interest in the gender strategies of plants, the structural and developmental bases for transitions in sexual systems are poorly understood.

Methods

Here, floral morphology, patterns of fertility, pollen-tube growth and floral development are investigated in two populations of the scandent shrub Mussaenda pubescens (Rubiaceae), native to southern China, by means of experimental and open-pollinations, light microscopy, fluorescence microscopy and scanning electron microscopy combined with paraffin sectioning.

Key Results

Mussaenda pubescens has perfect (hermaphroditic) flowers and populations with two style-length morphs but only weak differentiation in anther position (stigma-height dimorphism). Experimental pollinations demonstrated that despite morphological hermaphroditism, the species is functionally dioecious. The long-styled (L) morph possesses sterile pollen and functions as a female, whereas the short-styled (S) morph is female sterile and functions as a male. Self- and intra-morph pollinations of the S-morph were consistent with those expected from dimorphic incompatibility. The two populations investigated were both S-morph (male) biased. Investigations of early stages of floral development indicated patterns typical of hermaphroditic flowers, with no significant differences in organ growth between the floral morphs. Meiosis of microspore mother cells was of the simultaneous type with tetrads isobilateral in shape. The tapetal cells in anther walls of the L-morph became vacuolized during meiosis I, ahead of the uninucleate microspore stage in the S-morph. In the L-morph, the microspore nucleus degenerated at the tetrad stage resulting in male sterility. Microsporogenesis and male gametophyte development was normal in the S-morph. Failure in the formation of megaspore mother cells and/or the development of megagametophytes resulted in female sterility in the S-morph, compared with normal megasporogenesis in the L-morph.

Conclusions

In M. pubescens, cryptic dioecy has evolved from stigma-height dimorphism as a result of morph-specific sterility mutations.     

Electron-Microscope Observation on the Pollen Development of Male Sterility Wheat Induced by Ethrel

The pollen development of male sterility wheat induced by Ethrel was studied in comparison with that of normal wheat by transmission and scanning electron-microscope. The results obtained are summarized as follows: 1. The primary morphological changes of abortive wheat pollen after treatment with Ethrel took place in the vacuole stage. The materials of cytoplasm were rarefied. All kinds of the cell organelles and vacuoles became degenerated and disorganized. Inside a small number of cell nuclei, chromatin granules coagulated irregularly. The number and the activity of the Ubisch bodices became reduced evidently. 2. In the mature stage, the differences between the treated wheat and normal wheat became even more striking. The normal pollens were spherical in sharp, and full of starch granules. The treated pollen were monstrous. There was a large empty vacuole in every abortive pollen cell, in which the starch granules were smaller and fewer than these in the normal ones. The nuclei and cytoplasm in some of the abortive pollens were degenerated, leaving only the cell walls. 3. Under scanning electron microscope, it was discovered that the normal wheat pollen were spherical or oval. The germinated pores jutted. The round-openings of the pores could be clearly seen. The abortive pollen induced by Ethrel looked like some shrunken balls, and became deformed and were blocked up. 4. On discussion of the mechanism of the Ethrel-induced male sterility in wheat, the authors suggest that special attention should be paid to the changes of the Ubisch bodies and the vacuoles.     

Analysis of gene expression profile in pollen development of recessive genic male sterile <Emphasis Type="Italic">Brassica napus</Emphasis> L. line S45A

Male sterility in a near-isogenic line S45AB after 25 generations of subcrossing is controlled by two pairs of duplicate genes. The genotype of S45A is Bnms1Bnms1Bnms2Bnms2, and that of S45B is BnMs1Bnms1Bnms2Bnms2, respectively. Histological observations revealed that abnormal anther development appeared in the tapetum and pollen exine during the tetrad stage. This male sterility was characterized by hypertrophy of the tapetal cells at the tetrad stage and a complete lack of microspore exine after the release of microspores from the tetrads. To elucidate the mechanism of this recessive genic male sterility, the flower bud expression profiles of the S45A and S45B lines were analyzed using an Arabidopsis thaliana ATH1 oligonucleotide array. When compared with the S45B line, 69 genes were significantly downregulated, and 46 genes were significantly upregulated in the S45A line. Real-time polymerase chain reaction (PCR) was then used to verify the results of the microarray analysis, and the majority of the downregulated genes in the S45A line were abundantly and specifically expressed in the anther. The results of the real-time PCR suggest that Bnms1 might be involved in the metabolism of lipid/fatty acids, and the homologous mutation of Bnms1 may either block the biosynthesis of sporopollenin or block sporopollenin from being deposited on the microspore surface, thus, preventing pollen exine formation. The role of Bnms1 in the regulatory network of exine formation is also discussed as well. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Cytological and ultra-structural study on microsporogenesis of cytoplasmic male sterility in <Emphasis Type="Italic">Raphanus sativus</Emphasis>

The cytological development of microspores and tapetum in cytoplasmic male sterile (CMS) line A₁₄ and its maintainer B₁₄ in radish were studied using light- and transmission electron microscopy (LM and TEM). The microspores of the CMS line began to abort soon after they were released from tetrads in pollen sacs with light microscopy investigation, while abnormal behavior of pollen mother cells (PMC) were observed during its meiotic stage in its ultra-structural study, including degeneration of organelles and irregularity of nuclear membrane. At the same time, development of tapetal cells was similar to that of the maintainer. With further development of the anther, the tapetal cells of CMS line showed an abnormal increase in size and other appearances, such as fewer organelles and indistinct cytoplasm. The microspores of the CMS line were always distinguishable from the maintainer line with irregular structure, more osphilic deposits and abnormal exine. It is inferred that abortion of microspores is attributed to mutation of genes controlling male sterility, which further leads to hypertrophy of tapetum and destruction of ultra-structure.     

Intergeneric hybridization between Diplotaxis siifolia,a wild species and crop brassicas

Summary Attempts were made to obtain intergeneric hybrids between Diplotaxis siifolia, a wild species, and cultivars of Brassica (B. campestris, B. juncea, and B. napus). The crosses showed unilateral incompatibility. When the wild species was used as female parent, pollen germination and pollen tube growth were normal, but hybrid seeds aborted due to post-fertilization barriers. Reciprocal crosses (cultivars as female parent) showed strong pre-fertilization barriers; although pollen grains showed germination, pollen tubes failed to enter the stigma. Hybrids were realized in two of the crosses, D. siifolia x B. juncea and D. siifolia x B. napus, through ovary culture. The hybrids were multiplied in vitro by multiplication of axillary shoots, or somatic embryogenesis. Detailed studies were carried out on the hybrid D. siifolia x B. juncea. F₁ hybrids had shrivelled anthers and were pollen sterile. Amphiploids of this hybrid showed 60% pollen fertility and produced seeds upon self-pollination as well as backcross pollination with the pollen of B. juncea.     

Wide hybridization in Brassica

Summary Crosses were made to obtain interspecific hybrids between B. fruticulosa (wild species , 2n = 16) × B. campestris (cultivar , 2n = 20). Although many pollen grains germinated and their tubes entered the style, only about 30% of the ovules received pollen tubes. Fertilized ovules aborted at various stages of development. A few hybrid seeds resulted from hand pollinations in the field, and they showed poor germination and seedling establishment. The in vitro culture of ovaries, ovules, and seeds increased the frequency of obtaining hybrid seeds and plants: the most effective method was ovary culture followed by ovule culture. The hybrid nature of the plants was confirmed through morphological, cytological, and electrophoretic studies. A meiotic analysis of F₁ hybrids (2n = 18) showed that they had 0–5 bivalents and were completely pollen sterile. Electrophoretic analysis of leaf esterases and acid phosphatases of F₁ hybrids revealed bands derived from each parent. Induced amphidiploids of F₁ hybrids contained mostly bivalents, and had about 50% fertile pollen.