玉米花药发育的细胞生物学与分子遗传学的研究方法

雄性不育技术在玉米杂种优势利用和杂交种生产中发挥着重要作用,玉米花药发育和雄性不育的细胞生物学与分子遗传学研究是雄性不育技术利用的前提和基础。玉米花药发育是一个复杂的生物学过程,需要孢子体基因与配子体基因的协同表达调控。从玉米花药的形态结构、花药发育时期的划分、花药败育类型、花药发育的细胞学研究方法、花药发育的组学研究方法、花药发育分子遗传学研究方法等方面进行综述,以期为玉米核不育机制研究与雄性不育技术产业化应用提供方法学指导。     

水稻马协不育与可育花药ATPase定位(简报)

马协不育系系朱英国等用农家品种“马尾粘”的不育株与“协青早选”测交和回交选育成功的。超微结构研究表明马协不育花药单核边位期绒毡层提前解体,液泡膜破裂;不育花粉外壁基粒棒少,且分布不均匀;线粒体有大的电子透明区,二核期不育花粉中央大液泡崩溃。不育花药绒毡层提前解体可能影响花粉外壁合成及花粉营养供应,大量研究表明绒毡层发育异常与花粉败育密切相关,而液泡膜的破坏会导致水解酶类的泄     

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     

Chemical agents that inhibit pollen development: tools for research

Summary Several unrelated compounds are known to selectively inhibit the development of the male gametophyte. When applied at suitable dosages to plants at the appropriate stages of anther development, these substances block the formation of fertile pollen. The affected stage of pollen development is characteristic of the specific chemical structure of the compound, ranging from effects on microspore meiosis to the formation of pollen defective in the ability to germinate or fertilize. The range of effects mediated by these substances, and by known male-sterile mutants, indicates that microspore development has several critical phases that are particularly sensitive to fatal inhibition. We propose that chemical inhibitors of pollen development deserve attention as tools for elucidating the regulation of pollen development.     

Effects of water stress on male gametophyte development in plants

 Male reproductive development in plants is highly sensitive to water deficit during meiosis in the microspore mother cells. Water deficit during this stage inhibits further development of microspores or pollen grains, causing male sterility. Female fertility, in contrast, is quite immune to stress. The injury is apparently not caused by desiccation of the reproductive tissue, but is an indirect consequence of water deficit in the vegetative organs, such as leaves. The mechanism underlying this stress response probably involves a long-distance signaling molecule, originating in the organs that undergo water loss, and affecting fertility in the reproductive tissue, which conserves its water status. Much research has been focused on the involvement of abscisic acid in this regard, but the most recent evidence tends to reject a role for this hormone in the induction of male sterility. Stress-induced arrest of male gametophyte development is preceded by disturbances in carbohydrate metabolism and distribution within anthers, and an inhibition of the key sugar-cleaving enzyme, acid invertase. Since invertase gene expression can be modulated by sugar concentration, it is possible that decreased sugar delivery to reproductive tissue upon inhibition of photosynthesis by stress is the signal that triggers metabolic lesions leading to failure of male gametophyte development. Received: 31 October 1996 / Revision accepted: 18 February 1997     

Calcium distribution in fertile and sterile anthers of a photoperiod-sensitive genic male-sterile rice

Potassium antimonate was used to locate Ca²⁺ in fertile and sterile anthers of a photoperiod-sensitive genic male-sterile rice (Oryza sativa L. japonica). During the development of fertile anthers, abundant calcium precipitates accumulated in the anther walls and on the surface of pollen grains and Ubish bodies at the late developmental stage of the microspore, but not in the cytoplasm of pollen grains. Following the accumulation of starch grains in pollen, calcium precipitates on pollen walls diminished and increased in parenchymatous cells of the connective tissue. In sterile anthers, calcium precipitates were abundant in the middle layer and endothecium, but not in the tapetum, as was found in fertile anthers. A special cell wall was observed between the tapetum and middle layer of sterile anthers that appeared to relate to distinctive calcium accumulation patterns and poor pollen wall formation in the loculi. The formation of different patterns of antimonate-induced calcium precipitates in the anthers of photoperiod-sensitive genic male-sterile rice indicates that anomalies in the distribution of calcium accumulation correlate with the failure of pollen development and pollen abortion. Received: 30 May 1997 / Accepted: 5 July 1997     

Early signs of disruption of wheat anther development associated with the induction of male sterility by meiotic-stage water deficit

 Water deficit during meiosis in microspore mother cells of wheat (Triticum aestivum L.) induces male sterility, which reduces grain yield. In plants stressed during meiosis and then re-watered, division of microspore mother cells seems to proceed normally, but subsequent pollen development is arrested. Stress-affected anthers generally lack starch. We employed light microscopy in conjunction with histochemistry to compare the developmental anatomy of water-stress-affected and normal anthers. The earliest effects of stress, detectable between meiosis and young microspore stages, were the degeneration of meiocytes, loss of orientation of the reproductive cells, and abnormal vacuolization of tapetal cells. Other effects observed during subsequent developmental stages were deposition of starch in the connective tissue where it is normally not present, hypertrophy of the middle layer or endothecial cells, and deposition of sporopollenin-like substances in the anther loculus. The resulting pollen grains lacked both starch and intine. These results suggest that abnormal degeneration of the tapetum in water-stressed anthers coupled with a loss of orientation of the reproductive cells could be part of early events leading to abortion of microspores. Received: 19 July 1996 / Revision accepted: 6 November 1996     

Influence of the gene brevistylis on development of the flower in Oenothera

The mutant brevistylis of Oenothera (Onagraceae) is characterized mainly by malformations of the style and by female sterility. Morphological features of the style and stigmas, such as the length of the style and the number, size, and shape of the stigmatic lobes is very variable. The abscission layer between ovary and style is missing and the placenta with ovules is protruded into the style. An interpretation of the disturbances in development observed in the br/br homozygotes is presented. The primary influence of br on development is proposed to be a general delay in meristematic growth, followed by irregularities in the timing of morphogenetic events which in turn lead to the anomalies in morphology of style and stigmas.     

应用基因工程技术创造植物雄性不育系

基因工程开辟了创造植物雄性不育系的一个新的途径 ,综述了利用基因工程技术创造植物雄性不育的机制及相关启动子和基因 ;创造雄性核不育和质不育的途径 ;探讨了存在的问题和应用前景。     

雄性不育基因工程及其在蔬菜上的应用

基因工程是创造雄性不育的一种新途径。概述了雄性不育基因工程的主要方法 ,并且对利用基因工程创造蔬菜雄性不育系和恢复系的研究进展进行了综述 ,探讨了这一技术在蔬菜上的应用前景。     

白菜核雄性不育花药超微结构的研究

对白菜核雄性不育两用系的可育与不育花药进行了超微结构的比较观察。结果显示不育花药的造孢细胞核仁靠边分布;包裹小孢子母细胞的胼胝质厚薄不均匀,不完整等早期异常现象。减数分裂后．四分体细胞中常有多个细胞核。从四分体释放出的小孢子外壁的孢粉素物质不均匀沉积,呈不连续的单层异常结构。最后小孢子通过细胞质收缩方式败育。在可育花药中．绒毡层细胞在小孢子发育后期已显示出退化迹象,同时在细胞中开始积累脂类物质。但在同时期的不育花药中．绒毡层细胞没有显示出退化的迹象,也不合成脂类物质。从时间上看,败育花药中小孢子母细胞及小孢子的异常在先,绒毡层细胞的异常在后。本研究揭示了白菜核雄性不育花药的超微结构特征．对我们以前的光学显微镜观察结果予以补充和修正。     

白菜核雄性不育花药超微结构的研究

对白菜核雄性不育两用系的可育与不育花药进行了超微结构的比较观察。结果显示不育花药的造孢细胞核仁靠边分布:包裹小孢子母细胞的胼胝质厚薄不均匀,不完整等早期异常现象。减数分裂后,四分体细胞中常有多个细胞核。从四分体释放出的小孢子外壁的孢粉素物质不均匀沉积．呈不连续的单层异常结构。最后小孢子通过细胞质收缩方式败育。在可育花药中,绒毡层细胞在小孢子发育后期已显示出退化迹象,同时在细胞中开始积累脂类物质。但在同时期的不育花药中, 绒毡层细胞没有显示出退化的迹象,也不合成脂类物质。从时间上看,败育花药中小孢子母细胞及小孢子的异常在先,绒毡层细胞的异常在后。本研究揭示了白菜核雄性不育花药的超微结构特征, 对我们以前的光学显微镜观察结果予以补充和修正。     

Patterns of male sterility within and among populations of the distylous shrub Erythroxylum havanense (erythroxylaceae)

Distyly, a reproductive system characterized by the presence of long-styled (thrum). and short-styled (pin) individuals within a population, has been repeatedly used as a model for the study of the evolution of the reproductive systems in plants. Erythroxylum havanense is a distylous species in which most thrum plants fail to develop a fertile androecium, thus behaving as male-sterile or partially male-sterile plants. Short-styled (thrum) individuals have an increased performance as female parents, thereby compensating for their loss of male fitness. Previous studies of populations within close proximity to each other suggest that E. havanense may be involved in a process of gender specialization in which, unlike other heterostylous species, thrum plants are specializing as females and pins (long-styled) as males. In this paper we describe more general patterns of male sterility, one of the first steps in the evolution of gender specialization, among populations of the distylous shrub Erythroxylum havanense. Pollen germination differed among populations (range 0.52 ± 0.03 to 0.06 ± 0.04), and between morphs. Pollen from pin plants was almost two times (1.89) as fertile as that from thrums (0.36 ± 0.03 and 0.19 ± 0.03, pin and thrums respectively). Thrums were significantly more male sterile in four out of five populations. The population where differences between the floral morphs were not apparent showed the lowest levels of pollen fertility. Accordingly, our results indicate that populations of E. havanense show marked differences in pollen fertility and higher male sterility associated with the thrum morph. We hypothesize that differences between morphs could be explained if restorers of male sterility are linked to the distyly haplotype, while differences in genes associated with male sterility could explain the variation among populations. Overall, the prevalence of thrum-biased male sterility across populations suggests that E. havanense is subject to a process of gender specialization.     

Double-stranded RNA and male sterility in rice

Summary Double-stranded RNA (dsRNA) was isolated from rice Oryza sativa ssp. japonica, but not from other subspecies. The dsRNA has been found in all of the examined cytoplasmic male-sterile (CMS) lines of BT (Chinsurah Boro II)-type rice, but was not detected in their companionate maintainer lines. It is uniquely and positivley correlated with the CMS trait in BT-type rice. Recently, the dsRNA was also found in a nuclear malesterile (NMS) rice, Nongken 58s, but was not found in its normal Nongken 58. The molecular weight of this dsRNA was estimated to be about 18 kb. Electron microscopic analysis reveals that it is linear snapped. The double strandedness of the RNA molecules was characterized by CF-11 cellulose column chromatography and nuclease treatments. It bound to CF-11 cellulose in the presence of 15% ethanol. It was sensitive to RNase A at low salt concentrations, but insensitive to DNase I, SI nuclease, and RNase A at high salt concentrations. The dsRNA was detected in both mitochondrial and cytoplasmic fractions. Dot-blot hybridization reveals that there is no sequence homology between this dsRNA and mtDNA, but there is homology between this dsRNA and nuclear genomic DNA. We have not been able to transmit this dsRNA to fertile rice.     

白菜核雄性不育两用系可育和不育花药ATPase的分布

对白菜核雄性不育两用系可育花药和不育花药的ATPase做了定位分析。可育花药的花粉母细胞核中积累了大量的ATPase反应颗粒,而细胞质中ATPase反应颗粒较少,但在线粒体中特异地聚集了一些大的ATPase反应颗粒。减数分裂后,小孢子细胞质中ATPase反应颗粒明显增加。随着小孢子发育,其细胞质中ATPase反应颗粒逐渐减少,但在线粒体中又特异性地聚集了较多的AT-Pase反应颗粒。当花药发育到二胞花粉时期,花粉和绒毡层细胞中的ATPase反应颗粒已很少了。不育花药的花粉母细胞中呈现较多的ATPase,然而在线粒体中很少。异常四分体小孢子细胞质中虽然有较多的ATPase反应颗粒,但还是通过细胞质收缩和质壁分离方式退化。对可育花药的花粉母细胞线粒体中特异出现的簇状ATPase分布现象进行了分析,讨论了不育花药中花粉母细胞线粒体ATPase的异常与花粉败育的可能关系。     

茄子雄性不育系花蕾内源激素研究

利用间接酶联免疫测定技术研究2对茄子雄性不育系及保持系不同发育阶段花蕾中IAA、GA3、ABA及ZR含量动态变化。结果表明：IAA、GA3、ABA含量变化表现为不育系高于保持系,ZR含量变化则为保持系高于不育系,在2个不育系间及2个保持系间各激素变化趋势也有所不同。     

雄性不育嵌合基因的构建及番茄转化研究

用从烟草里克隆的TA29启动子分离自Bacillus amyloliquefaciens的核糖核酸酶基因barnase,构建成雄性不育嵌合基因。再用农杆菌介导法转化番茄子叶,获得了具有雄性不育特征的转基因植株。Abstract:The utilization of heterosis based on male sterility has great significance in raising crop yeld and improving quality.A tapetum-specific promoter TA29 from tobacco young leaves,and a Rnase gene-barnase from Bacillus amyloliquefaciens have been cloned for the construction of male sterile chimeric gene.Transgenic tomato plants with male sterile property have been obtained after transformation with this gene.     

油菜细胞质雄性不育系发育进程中活性氧的代谢

为进一步认识活性氧化谢同雄性不育的联系,本文研究了油菜不同生长发育时期不育系与保持系的活性氧代谢的差异。