陕农138小孢子不同发育时期蛋白质差异的比较分析

为了探索小麦花药发育过程中的蛋白质代谢机理,利用SDS-PAGE电泳对陕农138花药单核早期、单核中晚期和双核期小孢子的全蛋白电泳谱带进行了分析.结果表明,双核期同时出现2条明显的条带或表达丰度大的条带,其分子量分别为50.7、48.2 kD,而这两个条带分别在单核早期和单核中晚期单独出现,认为通过电泳所获得的这些差异蛋白很可能直接或间接参与小孢子发育过程中某些关键的代谢途径;单核中晚期是小麦花药培养中出愈率最高的时期,这些差异蛋白又很可能与小麦花药培养力的表达调控和代谢机理有关.     

Rapid Swelling of Pollen Grains in Response to Floret Opening Unfolds Anther Locules in Rice (Oryza sativa L.)

Abstract

The changes in pollen grain diameter were examined in relation to the floret opening and anther dehiscence in rice (Oryza sativa L.). In the first experiment, the florets were artificially opened by excising the top of the glumes at various times before the expected natural flowering time. Pollen diameter increased rapidly in the artificially opened florets, but slowly in the controls in which the glumes were left intact. The time of anther dehiscence coincided well with the time when pollen grains reached their maximum diameter. In addition, the pollen grain diameter was significantly correlated with the percentage of dehisced anthers in the florets artificially opened at various times. These results indicate that floret opening induces the swelling of pollen grains and that the swelling is an important event for anther dehiscence. In the second experiment, anther segments with pollen left remaining intact and those with pollen removed were immersed in water. The septa in the anther segments without pollen failed to rupture in water, whereas 70% of the septa in the anther segments with pollen left remaining ruptured, indicating that the septa can be ruptured by the swelling pressure of pollen grains resulting in unfolding of anther locules. From these findings, we infer that the rapid swelling of pollen grains in response to floret opening is a driving force to rupture the septum and unfold the locules.     

Genetic variation in the sensitivity of anther dehiscence to drought stress in rice

Field-based screens for genetic variation in reproductive-stage drought tolerance are often confounded by genetic variation in root depth, flowering date and biomass at flowering. To reduce these confounding effects and to impose drought stress more precisely, we grew contrasting genotypes of rice (Oryza sativa L.) in shallow containers of flooded soil. When water was withheld from the shallow-rooted indica genotype IR64 for 6 days starting at −17, −7, 0, 10 and 20 days after heading (DAH), the impact on grain yield was greatest at −7 and 0DAH. The most sensitive yield component was filled grain percentage (spikelet fertility). Data on yield and spikelet fertility were also obtained for another shallow-rooted lowland indica (BRRI Dhan 31) and two deep-rooted upland tropical japonicas (Azucena and Moroberekan). Compared with well-watered controls, withholding of water for 6 days reduced spikelet fertility by ∼80% in IR64, BRRI Dhan 31 and Azucena but by 22% in Moroberekan. In a detailed comparison, water was withheld from IR64 for 5 days and from Moroberekan for 6 days, starting 3 days before the heading date of well-watered controls. Under these stress conditions, heading was delayed for 4 days in IR64 and 6 days in Moroberekan. Spikelet fertility in the top four rachis branches was reduced by 80% in IR64 and 16% in Moroberekan, a difference attributable principally to reduced anther dehiscence and lower stigma pollen density in IR64. Two properties of Moroberekan may contribute to high anther dehiscence after stress/re-watering: (i) constitutively superior development of fibrous structures in the endothecium at the anther apex and base and (ii) better maintenance of pollen size. All ovaries of Moroberekan received pollen—an average of 60 grains in well-watered plants and 42 grains in stressed/re-watered plants. In well-watered IR64, all ovaries received an average of 31 grains, whereas in stressed/re-watered IR64, 67% of ovaries received no pollen and the remainder received an average of eight grains. Moroberekan may, therefore, be a source of reproductive-stage drought tolerance through genes that maintain anther dehiscence during recovery from low water status.     

6种番荔枝属果树雄蕊特性的研究

采用显微观察法研究6种番荔枝属果树的雄蕊特征,观测不同种的雄蕊长度、数量、花药长度和花粉数量及形态大小等雄蕊特征。结果表明:雄蕊特征随树种的不同而有变化,但普通番荔枝、阿蒂莫耶番荔枝、秘鲁番荔枝、牛心番荔枝等4种番荔枝的雄蕊特征及变化基本一致,而与山刺番荔枝、圆滑番荔枝等相差甚远;山刺番荔枝和圆滑番荔枝的一些雄蕊特征则较为接近。此外,尽管花药长短相差大,但每个花药内花粉数量的差异不显著,表明每朵花的花粉量仅由雄蕊数决定。     

栽培大豆(Glycine max)与野生大豆(G.soja)解剖学的比较研究——Ⅰ.雄配子体的发育

本文对栽培大豆(Glycine max)和野生大豆(G.soja)雄配子体的发育和同步性及精子发生进行了比较研究。大豆和野生大豆花粉母细胞的减数分裂是属于同时型的,小孢子的发育过程基本相同。花粉粒的发育在同一花药中基本上是同步的,在同花中九个连体花药的花粉粒的发育也基本上是同步的,而单个花药中花药粉粒的发育要稍落后于九个连体花药中的花粉粒。大豆的两个精子是在花粉管和花粉粒中形成的,即二、三细胞型;野生大豆的两个精子是在花粉管中形成的即二细胞型。本文讨论了这两种植物精子发生途径的类型及分类学上的意义。     

新型玉米细胞质雄性不育材料泰玉D2花粉败育的细胞学研究

通过光学显微镜观察泰玉D2不育系的花药发育进程,探明新型细胞质雄性不育材料泰玉D2花粉败育的细胞学机理。研究发现,花药绒粘层在二分体时期发生异常,小孢子在单核前期开始败育,至单核后期完全解体,花药发育后期不育系药腔中只观察到少量的小孢子残体,花药壁的4层细胞依然存在,绒粘层细胞已经彻底液泡化, 占据了药腔的整个空间,花粉成熟期不育系花药维管束消失。泰玉D2不育系花药绒粘层在二分体时期成辐射状膨大,小孢子在单核期败育,花粉败育完全,属无花粉类型。     

Fertility of doubled monoploids ofSolanum phureja

Thirteen anther-derived doubled monoploids and their heterozygous diploid anther donor were evaluated in the greenhouse for flowering, fruit set, seed set and pollen fertility. Doubled monoploids varied for number of days to flower, duration of flowering, abundance of flowers and general flower quality. Fruit set on the doubled monoploids in controlled crosses with diploid male-fertile clones varied from 0 to 73% with a mean of 33% compared to the anther donor with 58%. Seed set varied from 9 to 210 seeds per fruit and was significantly lower than the anther donor (377 seeds/fruit). Five doubled monoploids that shed low quantities of pollen exhibited significantly less pollen stainability (10-30%) than the anther donor (88%). However, pollen of doubled monoploids did not germinatein vitro. 2n pollen frequencies of 20–55% were observed for the doubled monoploids compared with 22% for the anther donor. It was concluded that doubled monoploids had lower female fertility compared to the anther donor but that they produced sufficient seed to facilitate utilization in a breeding program as female parents. However, they were considered to be male-sterile for practical purposes.     

高温条件下玉米自交系材料雄花育性的观察分析

在山西运城市7～8月份气温偏高的特殊气候环境下,对120份玉米自交系材料进行了雄花育性鉴定,并对其原因进行了分析。结果表明,高温条件下,材料间的雄花育性存在着明显的差异。不育性表现方式有:无花粉粒、花药吐不出、花药不散粉。模拟玉米螟危害雄花节打孔试验表明,即使玉米植株能得到充足的水分供应,雄花内部的水分胁迫是影响花粉正常散出的主要因素之一。     

小麦 TaWRKY51基因的克隆及其参与SQ-1诱导小麦花粉败育过程的表达模式研究

为进一步深入解析化学杂交剂SQ-1诱导小麦花粉败育的机理,本研究从SQ-1诱导的生理型雄性不育系PHYMS-1376及其对照可育系CK-1376各时期花药转录组测序筛选出差异表达基因TraesCS1D02G362900.1,与小麦基因组数据库比对发现,该基因编码的氨基酸与 TaWRKY51同源度为97%,说明该差异表达基因为 TaWRKY51对其进行生物信息学分析、亚细胞定位、转录激活活性与花药表达模式研究,结果表明, TaWRKY51开放阅读框全长663 bp,可编码220个氨基酸,含有WRKY结构域和锌指结构,分子量约为23.34 kD,等电点为6.42;启动子区包含与光响应、茉莉酸甲酯响应、脱落酸响应等7类顺式作用元件,且TaWRKY51蛋白被定位在细胞核内,具有一定的转录激活活性;与CK-1376相比,PHYMS-1376植株花药中 TaWRKY51的表达量在5个发育时期均高于CK-1376,且在四分体时期、单核晚期、二核期和三核期与CK-1376的差异均达极显著水平,表明该基因与PHYMS-1376花粉败育密切相关。     

Effects of the number of pollen grains on cold tolerance at the booting stage in rice lines with QTLs for cold tolerance

This study was conducted to clarify the relationship between the number of pollen grains per anther and spikelet fertility under low temperature conditions in the rice cultivars and lines including lines with quantitative trait loci (QTL) for cold tolerance, adapted to the Tohoku region of northern Japan. Cold-water treatment decreased anther length, the number of pollen grains per anther, and spikelet fertility in all cultivars and lines. The number of pollen grains was proportional to anther length in all cultivars and lines and under all temperatures. Spikelet fertility decreased with decreasing the number of pollen grains in cold-water treatments at 18.5 and 19.3?°C. ‘Ouu 415’, with the qLTB3 QTL for cold tolerance, had 28% more pollen grains and 9% higher spikelet fertility than the recurrent parent, ‘Hitomebore’, in the 18.5?°C cold-water treatment, suggesting that qLTB3 increased both parameters. Lines with the qCTB8 QTL for cold tolerance had significantly more pollen grains in two of the three years and significantly higher spikelet fertility in all three years in the 19.3?°C cold-water treatment, suggesting that qCTB8 reinforced cold tolerance. Ctb1, a proposed cold-tolerance QTL, had no effect on the number of pollen grains or spikelet fertility. In conclusion, the cultivars and lines with more pollen grains had a higher cold tolerance. Some of QTLs were inferred to increase the number of pollen grains and reinforce cold tolerance.     

水稻无花粉型核雄性不育突变体whf41的鉴定与基因定位

【目的】水稻花药角质层和蜡质层是花粉囊发育重要的结构支撑和安全屏障。对花粉囊发育相关基因进行表型分析和遗传定位,为进一步克隆相关基因和分子机制研究提供基因资源和理论基础。【方法】从籼稻中恢8015辐射诱变(~60Co-γ)突变体库中分离鉴定出了一个无花粉型雄性不育突变体whf41。对野生型和突变体不同发育时期的花药进行半薄切片观察;并对其成熟期的花药进行扫描电镜观察。将突变体分别与中恢8015和02428杂交,构建BC_1F_1、F_1株系和BC_1F_2、F_2群体,对该表型进行遗传分析,采用图位克隆的方法精细定位目的基因。【结果】表型分析结果显示,whf41突变体的花药瘦小且呈透明乳白色,花药中不包含花粉粒细胞;半薄切片结果显示,突变体的小孢子无法形成正常的花粉外壁,绒毡层细胞异常膨大而不进行程序性死亡,最终膨胀的绒毡层和花粉细胞碎片逐渐融合并充满药室;扫描电镜结果进一步发现突变体花药内外壁均呈平滑状而缺乏脂类物质,花粉细胞逐渐破碎并降解。遗传分析表明,whf41突变体的无花粉型雄性不育性状受一对隐性核基因控制,我们将该基因精细定位于第3染色体短臂XD-5和XD-11两个标记之间,物理距离45.6 kb,其中包含9个开放阅读框。序列分析显示该区间内细胞色素P450基因LOC_Os03g07250的第4个外显子处存在1个单碱基替换和3个碱基的缺失,导致其翻译序列发生一个氨基酸的替换(天冬氨酸→甲硫氨酸)和一个氨基酸(缬氨酸)的缺失,致使其功能改变进而出现该表型。q RT-PCR检测结果表明,whf41突变体中CYP704B2和一系列花药脂质合成与转运相关基因的表达水平均发生了显著下调。【结论】根据本研究结果可推断,Os WHF41是CYP704B2的新等位基因,相关结果进一步明确CYP704B2在水稻花药脂质合成与花粉壁形成过程中的重要作用。     

马铃薯花药与花粉粒发育的细胞形态学研究

为了更有效地通过有性杂交选育优良品种,本文对马铃薯的雄性器官进行了系统的形态学观察:1.马铃薯花药的孢原组织为新月形排列,成熟花药药室呈马蹄形。花药壁厚薄不均,药室内壁与中层细胞无明显区分;绒毡层为腺质型,其来源有二;外侧的来源于壁细胞,内侧的来源于药隔组织,并常有二型性。2.马铃薯花药中的造孢细胞产生3～4层时,进而变成小孢子母细胞。小孢子母细胞的减数分裂为同时型,四分体多呈四面体型排列,成熟花粉为2-细胞型。花粉圆球状,具3孔沟,花粉较小,花粉量较大。3.马铃薯花药的开裂,以孔裂为主兼部分纵裂。孔口边缘表皮下有纤维层状(即具条纹状加厚的细胞壁)细胞。部分纵裂部位有裂口异形细胞。花药开裂前,同侧两药室彼此勾通,有利于散粉。     

温光敏核雄性不育小麦花药中保护酶活性的变化

为了揭示温光型核雄性不育小麦花粉败有的生化机制1，比较研究了温光型细胞核雄性不育小麦C49S的不育和可育花药在不同发育时期的超氧化物歧化酶（SOD），过氧化氢酶（CAT），过氧化物酶（POD）的活性丙二醛（MDA），可溶性蛋白质含量的变化。结果表明，温光型核雄不育小麦不育花药SOD活性在花粉母细胞减数分裂期至小孢子早期的下降速率是可育花药的4．34－4．35倍，在成熟花粉期，可育花药的活性比不育花药高1－2倍，不育花药的CAT活性从花粉母细胞到小孢子中期有一定程度的下降；发育早期，不育花药MDA含量极显著高于可育花药；在花药发育的整个过程中，不育花药的POD活性一直比同一发育时期的可育花药高；可育花药中的可溶性蛋白质含量显著高于不育花药。以上结果说明，温光型核雄不育小麦C49S在早播低温短日照条件下，花药中有机自由基酶保护系统遭到破坏，花药中膜酯过氧化作用加剧，生化代谢出现异常，是其花粉败育的重要原因。     

关于玉米单倍体人工加倍方法及花粉活力测定的初步研究

以孤雌生殖诱导系高诱1号诱导的单倍体植株为材料,分别在6叶期和拔节期用不同浓度的秋水仙素配以二甲基亚砜(DMSO)注射处理茎尖生长点,对单倍体植株进行人工加倍,并对部分不散粉花药中的花粉活力进行测定.结果表明:在6叶期,以0.5%秋水仙素配以2.0%的DMSO处理效果最好,加倍率高达32.3%,是对照的4.8倍.在拔节期的处理随秋水仙素浓度的升高,药害株增多;测定的不散粉单倍体都具有活性花粉存在,花药中平均活性花粉为8.0%,最高者为20.0%。     

CRMS诱导水稻雄性不育的研究——Ⅲ.CRMS对水稻花药游离脯氨酸含量的影响及其与花药败育的关系

利用~3H-脯氨酸、~3H-谷氨酸示踪分析方法,研究了CRMS诱导水稻雄性不育的作用机理。试验结果表明,CRMS阻碍剑叶向花药运输脯氨酸,抑制花药本身合成脯氨酸能力,并降低脯氨酸合成醇不溶性大分子物质的结合率,引起花药营养匮乏,导致花药败育。     

Infection and Colonization of Pathogenic Fungus Fusarium proliferatum in Rice Spikelet Rot Disease

Rice spikelet rot disease (RSRD), caused by Fusarium proliferatum, is an emerging disease. So far, the effects of diseased rice floral organs as well as the primary infection sites and stages of this pathogen are not determined. We investigated changes in the floral organs, along with the infection processes of the pathogen in plants inoculated with F. proliferatum and labelled with a green fluorescent protein during different growth stages of rice. The results showed that RSRD is not a systemic infectious disease, which has negative effects on the fertility of the infected rice. F. proliferatum caused brown colored anthers, crinkled pistils and ovaries, pollen grain deformities and anther indehiscence. The number of pollen grains on the stigmas decreased significantly in the infected spikelets, and the anther dehiscence and seed-setting rate successively declined by 69% and 73%, respectively, as a result of the infection. The initial infection stage occurred at the pollen cell maturity stage, and the primary invasion sites were determined to be the anthers of rice. It was noted that the pathogen mainly damaged the pollen cells, and with the exception of the filaments, proceeded to colonize the pistils and endosperm.     

黄花蒿小孢子的发生和雄配子体的形成

黄花蒿是当今优良抗疟药青蒿素的药源植物。采用石蜡切片法观察了黄花蒿小孢子发生及雄配子体发育的细胞学过程。黄花蒿花药为4室结构;横切面呈蝴蝶状排列;花药壁发育为典型单子叶型;花药壁中层在花药发育的较早期解体,其绒毡层由腺质型绒毡细胞组成,在花粉成熟前绒毡层细胞原位解体;花粉母细胞在减数分裂过程中,细胞分裂同步相差1～2个细胞周期,其细胞质分裂则为同时型;经2次分裂后形成的四分体为正四面体型;花药开裂前的成熟花粉粒为2细胞型,含1个营养细胞和1个具2个细胞核的生殖细胞。     

CRMS诱导水稻雄性不育的研究：Ⅲ ．CRMS对水稻花药游离脯氯酸含量的影响及其与花药败育的关系

 利用³H-脯氟酸、³H-谷氨酸示踪分析方法，研究了CRMS诱导水稻雄性不育的作用机理。试验结果表明，CRMS阻碍剑叶向花药运输脯氨酸，抑制花药本身合成脯氨酸能力，并降低脯氨酸合成醇不溶性大分子物质的结合率，引起花药营养匮乏，导致花药败育。     

Susceptibility to Coolness at the Young Microspore Stage under High Nitrogen Supply in Rice (Oryza Sativa L.). Proteome Analysis of Mature Anthers

Abstract

In vitro pollen germination experiment using agar plates showed that the growth under high nitrogen conditions enhanced the damage to pollen germination ability caused by the cooling at the young microspore stage. To clarify the physiological factors related to this damage to pollen germination, we performed the comparative proteome analysis of mature anthers and identified proteins that were changed by high nitrogen conditions or high nitrogen plus cooling conditions. Proteins were extracted from mature anther samples and separated by two-dimensional polyacrylamide gel electrophoresis. By comparing anther protein maps of the samples collected from the plants grown under standard nitrogen conditions, high nitrogen conditions and high nitrogen plus cooling conditions, we found 11 protein spots, which varied with the treatment. These protein spots were identified based on the rice proteome database and/or peptide mass fingerprinting (PMF) analysis after digestion with trypsin. Digested samples were analyzed by matrix-assisted laser desorption/ionization-time flight mass spectrometry to produce PMF data. Database searches using these PMF data revealed the identities of 9 proteins. Seven of these proteins were polypeptides involved in cell elongation, stress responses and sugar metabolism. The relation between the fluctuations of these proteins and the decrease in pollen germination are discussed.     

芝麻细胞色素P450超家族的成员鉴定及SiCYP703A2的功能验证

细胞色素P450(CYP450)超家族广泛参与初生和次生代谢物质的生物合成,其中CYP703家族基因参与拟南芥、水稻等的花药发育.本研究利用生物信息学方法,从芝麻基因组中鉴定出303个CYP450基因,发现它们分为9个家族簇(clan),43个基因家族,随机分布于16个连锁群中.CYP703家族是71 Clan的成员之...