Involvement of phenylalanine ammonia-lyase in the development of pollen in broccoli (Brassica oleracea L.)

Summary To determine whether phenylalanine ammonia-lyase (EC 4.3.1.5) is involved in the maturation of microspores to fertile pollen, anthers of a fertile strain of broccoli (Brassica oleracea L.) were studied in a comparison with anthers of a cytoplasmic male sterile strain. In the normal fertile strain, immature anthers of about 2 mm in length exhibited higher phenylalanine ammonia-lyase activity than mature anthers or those shorter than 2 mm. The 2-mm-long anthers corresponded to the mononucleate stage, just after release of the microspores during pollen development. Immunohistochemical localization of phenylalanine ammonia-lyase in the anthers indicated that the protein was present predominantly in the tapetal cells. The immature anthers of cytoplasmic male sterile broccoli had a lower phenylalanine ammonia-lyase activity than those of the normal fertile strain. The level of phenylalanine ammonia-lyase activity in the immature anthers was positively correlated with the number of fertile pollen grains at the flowering stage in both strains. It seems possible, therefore, that phenylpropanoid metabolism, which involves phenylalanine ammonia-lyase, may play an important role in the maturation of microspores in flowering plants.Abbreviations CHS chalcone synthase - CMS cytoplasmic male sterility - DAPI 4, 6-diamidmo-2-phenylindole dihydrochloride - PAL L-phenylalanine ammonia-lyase     

Qualitative inheritance of water-stress induced apical sterility in wheat (Triticum aestivum)

Grain number per unit area is an effective component of grain yield in bread wheat. Water-stress induced apical sterility (tip sterility) reduces the number of grains and, consequently, the grain yield in semi-arid regions with a shortage of available water during the pre-anthesis period. Crosses between apical sterile and apical fertile varieties and selection lines were made and F1, BC1, and F2 populations were subjected to moderate water-stress to study the inheritance of this character. The F2 and BC1 plants were qualitatively categorised into two phenotypes and tested for monohybrid and dihybrid segregation hypotheses. All the spikes of F1 plants obtained from crosses between apical fertile and apical sterile varieties were fully fertile indicating apical fertility is dominant to apical sterility. The F2 segregation Results from crosses between apical fertile lines and Y82187 suggested two complementary dominant genes segregating independently were involved in tolerance to water-stress induced apical sterility. In other words, two dominant genes determine apical fertility in these crosses and if one of these loci is homozygous, recessive waterstress will induce apical sterility. One F2 population segregated for both apical sterility and vernalisation response. Semi-winter plants had more sterile spikelets and the result of chi-square test confirmed monhybrid segregation for vernalisation response.     

Alloplasmic male-sterile <Emphasis Type="Italic">Brassica juncea</Emphasis> with <Emphasis Type="Italic">Enarthrocarpus lyratus</Emphasis> cytoplasm and the introgression of gene(s) for fertility restoration from cytoplasm donor species

A new cytoplasmic male sterility (CMS) source in Brassica juncea (2n = 36; AABB) was developed by substituting its nucleus into the cytoplasm of Enarthrocarpus lyratus (2n = 20; E(l)E(l)). Male sterility was complete, stable and manifested in either petaloid- or rudimentary-anthers which were devoid of fertile pollen grains. Male sterile plants resembled the euplasmic B. juncea except for slight leaf yellowing and delayed maturity. Leaf yellowing was due mainly to higher level of carotenoids rather than a reduction in chlorophyll pigments. Female fertility in male-sterile plants varied; it was normal in lines having rudimentary anthers but poor in those with petaloid anthers. Each of the 62 evaluated germplasm lines of B. juncea was a functional maintainer of male sterility. The gene(s) for male-fertility restoration ( Rf) were introgressed from the cytoplasm donor species through homoeologous pairing between A and E(l) chromosomes in monosomic addition plants (2n = 18II+1E(l)). The percent pollen fertility of restored F(1) ( lyr CMS x putative restorer) plants ranged from 60 to 80%. This, however, was sufficient to ensure complete seed set upon by bag selfing. The CMS ( lyr) B. juncea compared favourably with the existing CMS systems for various productivity related characteristics. However, the reduced transmission frequency of the Rf gene(s) through pollen grains, which was evident from the sporadic occurrence of male-sterile plants in restored F(1) hybrids, remains a limitation.     

Characterization of anther differentiation in cytoplasmic male sterile maize using a specific isozyme system (esterase)

Summary During anther development, characterized in maize plants with N cytoplasm, certain esterase isozymes in non-microspore cells decrease in amount with anther age and new isozymes appear in the developing microspores. In anthers from male sterile plants with cms T or cms C cytoplasm, neither of these changes in esterase patterns occurred. In anthers from plants with cms S cytoplasm, the decrease in the esterases of non-microsporogenous cells was observed but not the appearance of microspore esterases. In lines carrying cms S cytoplasm and nuclear restorer genes, esterase changes during anther development were as in normal fertile anthers. These results are discussed with respect to the phenomenon of cytoplasmic male sterility in the different maize genotypes.     

ORGANELLE SIZE AND NUMBER IN FERTILE AND T-CYTOPLASMIC MALE-STERILE CORN

Anthers of inbred F₄₄ fertile (N) and cytoplasmic male-sterile (T) corn plants (Zea mays L.) were compared cytologically. No differences between fertile and sterile anthers were observed in size and number of mitochondria or plastids until after the start of anther degeneration. A rapid division of mitochondria was observed, however, in the tapetum and sporogenous cells of both fertile and sterile anthers during early growth stages. This rapid increase in mitochondrial numbers per cell (some 20-to 40-fold) preceded tapetal breakdown in sterile anthers and did not occur in other anther cells or in plastids. Limited observations on the megagametophyte and nucellus revealed that mitochondria in ovules remain relatively constant in size and number during gametogenesis and do not undergo degeneration.     

Effects of Petunia cytoplasmic male sterile (CMS) cytoplasm on the development of sterile and fertility-restored P. parodii anthers

The developmental defects causing cytoplasmic male sterility in Petunia parodii are described in isonuclear fertile, sterile, and fertility-restored plants using both light- and scanning electron microscopy. The aberrant development of the sporogenous tissue and tapetal layer caused by the cytoplasmic male sterile cytoplasm in both Petunia hybrida and P. parodii nuclear backgrounds is similar in onset and progression. The degeneration of the sporogenous tissue and tapetal layer of sterile anthers is first apparent late in meiosis and results in highly abnormal sterile sporogenous tissue by tetrad stage of fertile anthers. The stomium and endothecium do not show major developmental differences between fertile and sterile anthers, but the inner connective tissue of sterile anthers contained calcium crystals not found at high abundance in fertile anthers. Ovoid bodies containing magnesium and phosphorus were seen only in the vascular bundles of fertile anthers. Material prepared for the scanning electron microscope by freeze drying showed better retention of fragile morphological features, while critical-point drying permitted examination of nonvolatile structures, such as cell walls.     

Mechanism of male sterility in Petunia

Summary The free amino acid contents in the anthers of male fertile, cytoplasmic male sterile (cms) and genic male sterile (gms) petunia lines were compared at different developmental stages of the male gametophyte. Quantitative differences in the amounts of free amino acids were found between the fertile and male sterile lines and between the cms and gms lines. The differences between the sterile lines were correlated with the different developmental stages at which the breakdown in microsporogenesis occurred. In the Rosy Morn (RM) cms line, where breakdown of microsporogenesis occurred at the end of prophase 1, there was an associated increase in asparagine and decrease in the other amino acids. In the RM gms line, in which breakdown occurred at the tetrad stage, an accumulation of asparagine in the anthers corresponded with an accumulation of glutamine beginning at prophase 1. Compared with fertile anthers, the sterile anthers accumulated much proline at the early meiotic stages, but no -aminobutyric acid. Comparison of the free amino acids of the fertile and the male sterile lines indicates that certain biochemical events leading to breakdown of microsporogenesis precede the observed cytological breakdown. The results from adding asparagine and glutamine to extracts of anthers at different developmental stages suggest that the amino acid balance may contribute to the changes in pH in the fertile and male sterile anthers which we observed previously.Contribution from the Volcani Center, Agricultural Research Organization Bet Dagan, Israel. 1972 Series, No. 2083 E.     

白菜细胞核雄性不育花药的细胞化学观察

对一种由一对隐性基因控制的白菜细胞核雄性不育和可育株的花药进行了细胞学和组织化学研究。种子播种后,有1／4植株为不育株,其余的为可育株。通过对不育株和可育株花药发育的细胞学观察,确认不育花粉的败育发生在小孢子发育时期。用组织化学的方法研究了可育株和不育株花药发育过程中的多糖和脂类的分布动态,发现在减数分裂前,可育花药和不育花药的药隔细胞中都储藏了大量的淀粉粒。二者的差异仅是不育花药的绒毡层细胞液泡化明显。在减数分裂后的小孢子发育时期,可育花药的绒毡层细胞具有将药隔细胞中的淀粉粒多糖吸收并转化成脂类的功能,小孢子及以后的二胞花粉中也积累了大量的脂类储藏物质。在不育花药中,虽然减数分裂后药隔细胞中的淀粉粒也都消失,但绒毡层细胞中的脂类物质相比很少,同时绒毡层细胞显示了明显的多糖反应,表明不育花药的绒毡层细胞将糖类转化为脂类的功能受阻。在小孢子的表面有些脂类物质,但在细胞质中却没有脂类积累。这一结果暗示在该种白菜细胞核雄性不育株中,由于花药绒毡层细胞转换多糖为脂类的功能失常,导致了小孢子的败育。     

mRNA differential display between sterile and fertile anther of rice and analysis of cDNA differential fragments

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Proteolytic activity in anther extracts of fertile and cytoplasmic male sterile petunia

Proteolytic activity is compared in anther extracts from Petunia parodii fertile and cytoplasmic male sterile lines. It is characterized relative to developmental stage of the anthers, effect of variable incubation times, pH of isolation buffers, and degradation of marker proteins. In fertile anthers, proteolytic activity increases at the end of microsporogenesis and peaks early in microgametogenesis. Degradation is most severe in extracts of fertile anthers and in high molecular weight proteins and reaches its maximum within 20 minutes. Degradation of marker proteins is greatest at pH 5.6 to 8.0 in fertile anther extracts and is eliminated under strong acid conditions (pH 2.8 to 4.0) in both fertile and cytoplasmic male sterile anther extracts. Marker proteins degrade more severely in extracts of fertile anthers; however, the order of substrate sensitivity—myosin > phosphorylase b > bovine serum albumin and ovalbumin > β-galactosidase—is the same in extracts from fertile and cytoplasmic male sterile anthers.     

芝麻核雄性不育系ms86-1微粉发生的细胞学观察

芝麻(Sesamum indicum)核雄性不育系ms86-1姊妹交后代表现为可育、部分不育(即微粉)及完全不育(简称不育)3种类型。不同育性类型的花药及花粉粒形态差异明显。Alexander染色实验显示微粉植株花粉粒外壁为蓝绿色, 内部为不均一洋红色, 与可育株及不育株花粉粒的染色特征均不相同。为探明芝麻微粉发生机理, 在电子显微镜下比较观察了可育、微粉、不育类型的小孢子发育过程。结果表明, 可育株小孢子母细胞减数分裂时期代谢旺盛, 胞质中出现大量脂质小球; 四分体时期绒毡层细胞开始降解, 单核小孢子时期开始出现乌氏体, 成熟花粉时期花粉囊腔内及花粉粒周围分布着大量乌氏体, 花粉粒外壁有11–13个棱状凸起, 表面存在大量基粒棒, 形成紧密的覆盖层。不育株小孢子发育异常显现于减数分裂时期, 此时胞质中无脂质小球出现, 细胞壁开始积累胼胝质; 四分体时期绒毡层细胞未见降解; 单核小孢子时期无乌氏体出现; 成熟花粉时期花粉囊腔中未发现正常的乌氏体, 存在大量空瘪的败育小孢子, 外壁积累胼胝质, 缺乏基粒棒。微粉株小孢子在减数分裂时期可见胞质内有大量脂质小球, 四分体时期部分绒毡层发生变形, 单核小孢子时期有部分绒毡层开始降解; 绒毡层细胞降解滞后为少量发育进程迟缓的小孢子提供了营养物质, 部分小孢子发育为正常花粉粒; 这些花粉粒比较饱满, 表面有少量颗粒状突起, 但未能形成覆盖层, 花粉囊腔中及小孢子周围存在少量的乌氏体。小孢子形成的育性类型与绒毡层降解是否正常有关。     

Differential Mitochondrial Electron Transport through the Cyanide-Sensitive and Cyanide-Insensitive Pathways in Isonuclear Lines of Cytoplasmic Male Sterile, Male Fertile, and Restored Petunia

Three pairs of isonuclear lines of cytoplasmic male sterile (CMS) and fertile Petunia cells (Petunia hybrida [Hook] Vilm. and Petunia parodii L.S.M.) grown in suspension culture were examined for sensitivity to inhibitors of respiratory electron transport at time-points after transfer into fresh media. Cells from CMS lines differed from cells of fertile lines in their utilization of the cyanide-insensitive oxidase pathway. Under our culture regime, after approximately 3 days of culture cells from the CMS lines exhibited much lower cyanide-insensitive, salicylhydroxamic acid-sensitive respiration than cells from the fertile lines. This respiratory difference was shown to be specific to the mitochondrial alternative oxidase pathway by using other characteristic inhibitors of mitochondrial electron transport in experiments with isolated mitochondria. Immature anthers from CMS plants also showed lower alternative oxidase activity relative to anthers from male fertile plants, but no such difference was detected in leaf tissue, ovary or perianth tissue, or anthers collected just prior to anthesis. A cell line from a fertile plant carrying a nuclear fertility restorer gene and the CMS cytoplasm exhibited increased activity of the alternative pathway compared with the CMS lines.     

水稻非花粉型细胞质雄性不育系及其保持系花药发育过程中Ca2+的分布变化

钙在高等植物中被称为第二信使,与植物的有性生殖有关。为了研究水稻（Oryza sativa L.）花药中钙的定位与花粉败育的关系,利用焦锑酸钾沉淀法研究了非花粉型细胞质雄性不育系G37A及其保持系G37B花药的发育过程及其细胞中Ca^2＋ 的分布变化。研究发现,在2个材料间花药中钙的分布存在大量差异。G37B的可育花药在花粉母细胞时期及二分体时期,很少看到有Ca^2＋的沉积;而在单核花粉时期,Ca^2＋沉积急速地增加,主要定位在绒毡层细胞、花粉外壁外层及乌氏体的表面;随后花药壁上沉积的Ca^2＋减少而花粉的外壁外层仍然有很多Ca^2＋沉积物。相反,G37A的不育花药在花粉母细胞时期和二分体时期有大量的Ca^2＋沉积在小孢子母细胞和花药壁,中间层和绒毡层特别多。在二分体时期之后,不育花药的Ca^2＋沉积减少,特别是绒毡层内切向质膜附近的Ca^2＋几乎消失。但是同时期的可育花药中,有大量的Ca^2＋沉积在绒毡层。不育花药的Ca^2＋沉积在开花几天后消失。根据研究结果推测在不育花药发育早期中更多的钙离子与花粉败育有一定的关系。     

白菜细胞核雄性不育花药的细胞化学观察

对一种由一对隐性基因控制的白菜细胞核雄性不育和可育株的花药进行了细胞学和组织化学研究。种子播种后,有1/4植株为不育株,其余的为可育株。通过对不育株和可育株花药发育的细胞学观察,确认不育花粉的败育发生在小孢子发育时期。用组织化学的方法研究了可育株和不育株花药发育过程中的多糖和脂类的分布动态,发现在减数分裂前,可育花药和不育花药的药隔细胞中都储藏了大量的淀粉粒。二者的差异仅是不育花药的绒毡层细胞液泡化明显。在减数分裂后的小孢子发育时期,可育花药的绒毡层细胞具有将药隔细胞中的淀粉粒多糖吸收并转化成脂类的功能,小孢子及以后的二胞花粉中也积累了大量的脂类储藏物质在不育花药中,虽然减数分裂后药隔细胞中的淀粉粒也都消失,但绒毡层细胞中的脂类物质相比很少,同时绒毡层细胞显示了明显的多糖反应,表明不育花药的绒毡层细胞将糖类转化为脂类的功能受阻。在小孢子的表面有些脂类物质,但在细胞质中却没有脂类积累。这一结果暗示在该种白菜细胞核雄性不育株中,由于花药绒毡层细胞转换多糖为脂类的功能失常,导致了小孢子的败育。     

矮败蓝标型小麦不育系的选育与研究

利用蓝粒太谷核不育硬粒小麦89-2343[AABB 4D(MS2)/4E]与普通小麦7739-3(2n=42)杂交、回交所产生的蓝粒可育株与白粒矮败材料杂交、回交,育成了一份矮败蓝粒小麦.选用13份遗传背景不同的白粒普通小麦与之杂交、回交,育成了13份矮败蓝粒小麦.对后代的粒色和育性分离进行分析,蓝粒矮败不育株占22.1%,白粒非矮秆可育株占77.7%,表明蓝粒基因、Ms2和Rht10均位于附加染色体上,且连锁紧密;但不同轮回亲本,矮败蓝粒的传递率有差异,477A的传递率最高,接近50%.细胞学分析表明矮败蓝粒小麦仍为单体附加系;探讨了矮败蓝粒小麦在群体改良和杂种小麦生产中的应用.     

Floral micromorphology and microsporogenesis of the gynodioecious herb Glechoma longituba (Lamiaceae)

Gynodioecy, the phenomenon of having both hermaphrodite and female (i.e. male‐sterile) individuals within the same population, is an important intermediate step in the evolution of separate sexes in flowering plants. In this study, we investigated the floral micromorphology and microsporogenesis of the gynodioecious herb Glechoma longituba from four natural populations in Korea. The floral micromorphological characters of the different sex morphs were examined and compared using scanning electron microscopy (SEM), and the ultrastructure of microspores during microsporogenesis was studied. We also examined the development of anthers and pollen grains in the three sexual morphs (i.e. hermaphrodites, females, and gynomonoecious, i.e. individuals with a mixture of female and hermaphroditic flowers) by embryological investigation. The major difference in anther development between the three phenotypes was the early disintegration of the tapetal cells in the anthers of female flowers. While mature fertile pollen grains were found in both hermaphrodite and gynomonoecious phenotypes, females did not produce any pollen grains. In addition, both fertile and sterile pollen grains in gynomonoecious phenotypes were frequently observed. The results of the present study indicate that floral micromorphological characters were not distinct between sexual morphs of G. longituba, except for the structure of the inner cell surfaces of the anther. The observed tapetum abnormalities and degeneration of pollen grains in both gynomonoecious phenotypes and females may be the consequence of inbreeding depression in hermaphrodites.     

Production of microspore-derived plants by anther culture of an interspecific F1 hybrid between Cyclamen persicum and C. purpurascens

Anthers of a F1 hybrid (2n=41) between Cyclamen persicum (2n=2x=48) and C. purpurascens (2n=2x=34) were cultured to produce microspore-derived plants. Embryoids were produced when anthers, containing microspores at the early uninucleate stage of pollen development, were cultured in B5 medium containing sucrose (90 g l-1) and NAA (0.1, 1 mg l-1) or 2,4-D (0.1 mg l-1) in the dark at 5 °C for 4 days, then at 25 °C for 60 days. The embryoids usually developed into plantlets when cultured in B5 medium containing sucrose (30 g l-1) in the dark at 25 °C. At meiosis, the F1 hybrid, used as source for anther culture, formed some cells with restitution nuclei at telophase and dyads at the tetrad stage, which resulted in the production of viable pollen grains as unreduced gametes. Plants produced by anther culture were grouped into sterile plants with 2n=41 chromosomes and fertile plants with 2n=82 chromosomes. The present findings suggested that the sterile plants were polyhaploids originating from unreduced microspores (n=41) of the F1 hybrid and that the fertile plants were amphidiploids induced by a spontaneous doubling during culture of chromosomes of such unreduced microspores. This revised version was published online in June 2006 with corrections to the Cover Date.     

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     

辣椒胞质雄性不育系和保持系内源激素含量的比较

以2个辣椒品系(199807、199803)的胞质雄性不育系和相应保持系为实验材料,采用酶联免疫吸附法(ELISA)测定IAA、(Z ZR)、GA3和ABA等内源激素含量,用气相色谱分析仪测定乙烯(ETH)释放量,对辣椒胞质雄性不育系和相应保持系内源激素含量变化规律进行研究.实验结果表明:在四分小孢子之前,花药中的IAA含量不育系显著高于保持系,在四分小孢子时期花药和花蕾中的IAA含量出现转折,到花粉粒成熟期的花蕾和花药以及开花期叶片中的IAA含量均是不育系显著低于保持系;小孢子各发育时期花药以及花期叶片中GA3含量均是不育系高于保持系,但花粉粒成熟期化蕾中GA3含量为不育系低于保持系;小孢子不同发育时期的花药以及花期叶片中ABA含量始终足不育系显著高于保持系,而花粉粒成熟期花蕾中ABA含量不育系与保持系没有显著差异;花粉粒成熟期的花蕾和花期叶片中ETH释放量表现为不育系显著高于保持系.同时,花粉粒成熟期的花蕾、花药和叶片中IAA/ABA、(Z ZR)/ABA、GA3/ABA、IAA/GA3、(Z ZR)/GA3等5个激素的比值均有不育系低于保持系的趋势.本实验结果说明辣椒的育性表现与花器和叶片等组织中内源激素的含量变化有关,花药和花期叶片中IAA亏缺、GA3和ABA增加以及化蕾和叶片中ETH过度产生,都有可能导致辣椒雄性不育.     

Cytoplasmic Sterility Factors in VICIA FABA L

Tissues of cytoplasmic male sterile, maintainer, restorer, and restored lines, and sterile plants which reverted to fertility in Vicia faba were examined in ultrathin sections. Cytoplasmic spherical bodies (CSB), ca. 70 nm in diameter, were observed in tissues of all sterile plants but not in tissues of maintainer, restorer or restored sterile plants. No CSB were observed in a reverted fertile branch of a tiller-sterile plant, nor in 5 of 6 reverted fertile plants. One reverted fertile plant contained CSB in ovules. It is proposed that the CSB are the sites of, or possibly, products of, sterility factors in Vicia faba.