川百合精细胞的超微结构

川百合与朱顶红花粉管中的生殖细胞分裂行为非常不同。诸如：染色体行为微管的组织形式和分布包括着丝点微管形成的时间,纺锤体的形状及间期周质微管网络在生殖细胞分裂过程中消失与否等,但这两种细胞具有共性,包括在有丝分裂前期缺乏早前期带微管（PPB）,未其形成细胞板等,这两种植物精细胞的结构应有较大差异,我们曾报道了朱顶红精细胞的超微结构,本文详细从超微结构方面描述了川百合精细胞的特征。川百合花粉管的萌发采用半离体－活体培养方式,11－18小时后,DNA荧光染料Hoechst33258和醋酸地衣红染色检查花分管中生殖细胞和精细胞发育时期。切取含有分裂的生细胞和精细细胞的花柱部分,按曾报道的方法固定、包埋、切片、染色及观察。在所有检查的花粉管中,两精子均前后排列（Fig.1-3),营养核前导并靠近花粉管顶端（Fig.,3)。H33258染色可见两精核间以DNA联系（Fig.3)。两个新形成的精核彼此分离（Fig.1),后来又相互造近,并维持一定距离（Fig.3)偶尔一对精子与营养核靠近（Fig.2)。两精细胞被一共同的细胞壁连接,他们不仅被自己的质膜也被营养细胞的质膜包围构成周质。周质平坦光滑。共同壁横向、弯曲、网状具胞质通道（Fig.4),厚度明显大于周质。色质凝集的程度更大些（Fig.5),可能意味着一个精子发育的早些。精细胞质中具有线粒体、内质网、高尔基体、脂体和大量核糖体。无质体。线粒体具有发育完好的精细胞中,微管呈纵向束排列于随精细胞的继续发育,共同壁消失了。与朱顶红等植物的染色体行为遵循典型有丝分裂方式不同,川百合生殖细胞与紫露草相同,它的染色体在有丝分裂中期沿细胞长轴分布,胞质分裂时没有细胞板出现。可以认为：川百合象烟草一样是介于朱顶红和紫露草之间的中间类型。雄性生殖单位（MGU）在三细胞和二细胞花粉中普遍存在。尽管本工作观察到营养核与精细胞紧密联系,以及两精子与DNA联系的例子,但MGU在超薄切片中并未见到,有可能MGU是一个动态的和时间上的暂时结构。另一方面,MGU的建立是以性细胞（生殖细胞或精细胞）的突起和营养核的裂瓣相互环绕为基础的,而性细胞中的细胞骨架（即：微管）可能对维持其与营养核的附着起重要作用。缺乏微管,可能是川百合精细胞不存在MGU的原因之一。     

朱顶红生殖细胞胞质分裂的两种方式(英文)

大多数植物以形成细胞权方式完成胞质分裂过程,也有些植物以类似于动物和单细胞植物在赤道区形成收缩沟的方式而分成两部分。本工作应用电镜对朱顶红体外萌发９～１８小时花粉管中的生殖细胞胞质分裂进行了研究。结果表明：７０％的细胞表现的是第一种方式、３０％却是第二种方式。即：朱顶红生殖细胞胞质分裂同时存在两种方式。前者最初以细胞板亚单位的形式出现于有丝分裂晚后期,它们聚集于成膜体的中央区域并于分裂末期融合成一个大的连续的单位（Ｆｉｇ．１～３）。大量新的微管形成于两组染色体之间（Ｆｉｇ．１）。分裂末期,细胞板形成并具胞质通道（Ｆｉｇ．２）。成膜体微管规则排列并穿过胞质通道向新形成的末期核伸展（Ｆｉｇ．２＆３）。这些微管与构成细胞板的质膜紧密联系（Ｆｉｇ．３）。后者则在有丝分裂后期开始（Ｆｉｇ．４）,当两群染色体彼此分离时,生殖细胞质膜在中央区由两侧向内凹陷形成收缩沟。有时生殖细胞几乎被收缩沟分成两个部分（Ｆｉｇ．６）。发生缢缩的细胞中细胞器与具细胞板的无差异,但微管稀少并且排列紊乱（Ｆｉｇ．４＆５）,染色体的状态使得难以准确区分细胞分裂时期。而且核膜的形成似乎始于有丝分裂后期、出现于染色体边缘（Ｆｉｇ．７）。有时尚有落后     

朱顶红精细胞超微结构观察

精细胞是双受精作用的直接参与者,是生殖生物学中的重点研究对象之一。以往的研究表明,应用连续超薄切片和计算机辅助三维重组技术,结合免疫荧光定位,发现两个精细胞在体积和细胞器含量上存着差异,即精子的二型性,而且与营养细胞核三者构成紧密功能单位却雄性生殖单位（MGU）。微管对精细胞的性状的确定、运动和维持MGU的动态结构稳定具有重要的作用。本文应用透射电镜。详细观察了朱顶红花粉管中细胞的超微结构,并着重微管结构及其分布的观察。朱顶红成熟花粉为两细胞型。成熟花粉于26℃、黑暗条件下,在液体培养基（含10％蔗糖和100ppm硼酸）中培养13－18小时,然后收集花粉管,固定,供电观察并照相。朱顶红成熟花粉培养13小时后,生殖细胞在花粉管中完成核分裂和胞质分裂等两个过程。形成两个精细胞。形成的两个精细胞前后排列,营养核前导并靠近花粉管顶端。领头的精细胞的细胞质以很大的表面与营养核相互融合（图版1－1,2）,有时营养核与两个精细胞彼此穿插、缠绕（图版1－3）。两精细胞之间的壁上具有多胞质通道和含均质电子密度中等的基质（图版II－4）。精细胞质在核与共同壁之间的区域染色较深,经高倍放大,观察到此处含丰富的微管, 自由分布,但以纵向或斜向为主（图版II－5,6）。所有的微管构成松散的桶状网络存在于两精核之间,除此之外,其他区域无微管分布（图版II－7）。培养18小时之后,两精细胞的共同呈网状（图版II－8）,此时微管均成纵向排列,平行于细胞长轴,构成筐状结构包围状精核,但不构成紧密的束状,证明了前人的免疫荧光观察结果。从以上观察结果我们可以得出如下的结论,朱顶红花粉为两细胞型,因此在花粉管中形成的两个精细胞一般前后排列,跟在营养细胞后面,这种线状排列方式在其他植物中也观察到,可能有利于三者作为一个结构功能单位在花粉管中移动和对花粉管狭窄空间的进化适应。MGU形成得较晚,在生殖细胞和营养核进入花粉管后才形成,并一直维持到精核形成,与其他报道不同。精细胞发育过程中,微管的分布方式变化显著。精细胞中微管的分布仅限于共同的细胞壁和靠近茎核之间的区域,总体构成一个松散的桶状结构。精细胞发育后期,微管均成纵向排列,包围着精核,极似生殖细胞筐状的微管结构形式。     

朱顶红精细胞超微结构观察

精细胞是双受精作用的直接参与者,是生殖生物学中的重点研究对象之一。以往的研究表明,应用连续超薄切片和计算机辅助三维重组技术,结合免疫荧光定位,发现两个精细胞在体积和细胞器含量上存在着差异,即精子的二型性,而且与营养细胞核三者构成紧密功能单位即雄性生殖单位（ＭＧＵ）,微管对精细胞的性状的确定、运动和维持ＭＧＵ的动态结构稳定具有重要的作用。本文应用透射电镜,详细观察了朱顶红花粉管中精细胞的超微结构,并着重微管结构及其分布的观察。朱顶红成熟花粉为两细胞型。成熟花粉于２６℃、黑暗条件下,在液体培养基（含１０％蔗糖和１００ｐｐｍ硼酸）中培养１３－１８小时,然后收集花粉管,固定,供电镜观察并照相。朱顶红成熟花粉培养１３小时后,生殖细胞在花粉管中完成核分裂和胞质分裂等两个过程。形成两个精细胞。初形成的两个精细胞前后排列,营养核前导并靠近花粉管顶端。领头的精细胞的细胞质以很大的表面与营养核相互贴合（图版Ⅰ－１,２）,有时营养核与两个精细胞彼此穿插、缠绕（图版Ⅰ－３）。两精细胞之间的共同壁上具有很多胞质通道和含均质电子密度中等的基质（图版Ⅱ－４）。精细胞质在核与共同壁之间的区域染色较深,经高倍放大,观察到此处含丰富的微管,基     

王百合及兰州百合细胞质遗传的细胞学研究

描述了王百合（ＬｉｌｉｕｍｒｅｇａｌｅＷｉｌｓ．）及兰州百合（Ｌ．ｄａｖｉｄｉＤｕｃｈ．）质体和线粒体在生殖细胞及精细胞中的分布和细胞质中ＤＮＡ的状况。在刚形成的王百合的生殖细胞中含有少量的质体和大量的线粒体。当生殖细胞游离在营养细胞质中时,质体在生殖细胞中完全退化消失。ＤＡＰＩ荧光技术进一步证明,在成熟花粉、花粉管中的生殖细胞及其分裂形成的两个精细胞中无任何细胞器ＤＮＡ。兰州百合在小孢子分裂时质体严格地极性分布,造成了刚形成的生殖细胞中即无质体。兰州百合、麝香百合（Ｌ．ｌｏｎｇｉｆｌｏｒｕｍＴｈｕｎｂ．）及其杂种的ＲＦＬＰ分析,也证明兰州百合质体是单亲母系传递的。虽然在不同发育时期的生殖细胞及精细胞中可以观察到线粒体,但在雄配子体时期它们的ＤＮＡ已降解,因此雄性线粒体不能被传递至后代。研究结果提供了百合属的这两个种质体和线粒体具母系遗传方式的细胞学证据,并阐明父系细胞质不能作为遗传传递的机理。     

朱顶红生殖细胞分裂过程中微管超微结构的变化

用透射电镜的方法,对朱顶红（Ａｍ ａｒｙｌｌｉｓｖｉｔｔａｔａ Ａｉｔ．）花粉管中生殖细胞的分裂过程中微管分布和结构形态变化进行了观察,获得如下主要的结果：有丝分裂前期,微管的数量较分裂前减少并变短,靠近细胞核分布。分裂前中期,微管出现于原来的核区并与染色体发生联系,形成着丝点微管。分裂中期,染色体排列于赤道面上形成赤道板,微管构成纺锤体。分裂后期,染色体分成两群,被缩短的着丝点微管拉向两极。在纺锤体两极的微管汇聚。后期的晚期,当极的微管尚未消失时,在赤道区域出现丰富的成膜体微管,在成膜体中央,细胞板前体物聚集。分裂末期,极微管和着丝点微管消失,成膜体微管在新形成的核膜和细胞板间扩展并穿过细胞板     

小麦雄配子体的超微结构 Ⅱ.精细胞的形成和发育

1.通过小孢子有丝分裂而形成的生殖细胞,在发育过程中有一系列的包括位移和变形的变化。最后生殖细胞变为纺锤体,准备另一次有丝分裂。在此时期,生殖细胞是裸露的,它只有自己的质膜和被营养细胞的质膜包围。生殖细胞的大部分为明显的椭圆形的核所占据,具高度凝集的染色质。在生殖细胞薄层的细胞质中,除核糖体外,所有的细胞器比营养细胞质中的明显的少,而且小。微管也能在纺锤形的生殖细胞的细胞质中看到,它们的排列方向是和细胞的长轴平行的。在生殖细胞中没有造粉质体。2.当生殖细胞已移位并和营养细胞再次紧密靠近后,它开始分裂。由生殖细胞分裂所形成的精细胞及其发育包括下列主要的变化:精细胞的形状由圆球形变为椭圆形,最后变为具尾延长的细胞。与此同时,细胞质的分布逐渐集中到精细胞的一端,形成细胞质的延伸,构成所谓的精细胞的尾部。有更多的细胞器,特别是线粒体集中在尾部。从生殖细胞分裂刚形成的精细胞是裸露的,所包围的质膜是不连续的。除精细胞的质膜之外,为营养细胞的质膜所包围,在后来的发育时期此双质膜变为连续的,并在一个极短的时期有壁物质——胼胝质沉积在质膜之间的空间。但在此时期之后细胞壁变为不连续的,壁物质明显降低。对生殖细胞分裂前的位置及精细胞发育中形态变化的意义进行了讨论。     

中国鹅掌楸雄配子体发育的超微结构研究

中国鹅掌楸（ＬｉｒｉｏｄｅｎｄｒｏｎＣｈｉｎｅｓｅ（Ｈｅｓｍｌ．）Ｓａｒｇ．）雄配子体发育的超微结构观察结果表明,中国鹅掌楸雄配子体的发育符合大多数被子植物的发育特征,表现在：１．小孢子核在萌发孔的相对侧靠近小孢子壁处进行有丝分裂,分裂末期形成细胞板,而后进一步发育形成分隔营养细胞和生殖细胞的纤维素的壁。２．胞质分裂极不均等,营养细胞与生殖细胞大小悬殊,而且生殖细胞内不含质体。３．生殖细胞在发育过程中发生一系列位移和形态变化,生殖细胞向营养核移动,与营养核贴合时,细胞核内染色质高度凝缩;与营养核分离后,生殖细胞纤维素的壁才完全解体消失,同时质膜凹陷,细胞呈不规则形态。４．营养细胞内脂质小泡极性分布,形成生殖细胞的脂体冠。５,营养核在与生殖细胞贴合过程中形状由近圆球形变为近弯月形,但核仁、核膜界限清晰,染色质始终保持均一状态。对生殖细胞分裂前的位置变化及其与营养细胞间的关系进行了初步的探讨。     

川百合花粉管的生殖细胞分裂过程中微管骨架的分布变化

应用透射电镜辅以免疫荧光定位技术研究了川百合 (Lilium davidii Duch.)花粉管中生殖细胞分裂过程中染色体动态和微管分布的关系。在生殖细胞分裂前和有丝分裂前期 ,电镜观察一直未见微管结构 ,但免疫荧光图象显示生殖细胞中有微管蛋白存在。直到分裂的前中期—中期 ,染色体出现 ,它们沿花粉管的长轴前后排列 ,横向的着丝点对相应地一对对地纵向排列。这时 ,生殖细胞中才出现大量微管 ,它们分布于细胞周质区和染色体之间 ,并跨越染色体的整个长度。前中期—中期开始时 ,只有 1～ 2对着丝点从横向转为纵向 ,微管垂直插入着丝点形成着丝点微管 ,而非前人用免疫荧光方法观察到的微管与着丝点侧向联接的图象。随着横向的着丝点对逐渐转变成纵向的过程 ,着丝点微管数量逐渐增多 ,但不形成典型的纺锤体。分裂后期 ,染色体交错分离 ,微管的分布与前中期—中期的基本相同。晚后期 ,染色体呈明显的两群 ,除极区和细胞中央区有微管残余外 ,大部分微管消失。通过染色体长度的测量 ,间接证明了分裂后期 B的存在。分裂末期的晚期 ,核膜形成后 ,在两精核之间的区域 ,微管数量开始增多。此区可能代表用免疫荧光所观察到的微管重叠区。细胞板出现后 ,微管消失     

川百合生殖细胞及其分裂形成精细胞的观察

在植物界中,生殖细胞在花粉管中进行的分裂有两种方式,一种遵循典型的有丝分裂过程,如朱顶红、烟草等归属此类;还有一种为非典型的有丝分裂,代表植物为百合类及凤仙花属。本文介绍在离休萌发条件下,川百合（LiliumdavidiiDuchartre）萌发花粉的生殖细胞进入花粉管并在其中分裂形成精细胞的过程。1实验方法培养基萌发用液体培养基由10％蔗糖和100X10－6硼酸组成。准备一个小培养皿,将5ml液体培养基倒入其中,使之刚好覆盖培养皿底部。花粉萌发①离体培养：用镊子夹取刚开裂的花药,在培养液的液面上轻轻沾几下,使新鲜的成熟花粉均…     

Live-cell imaging reveals the dynamics of two sperm cells during double fertilization in Arabidopsis thaliana

Flowering plants have evolved a unique reproductive process called double fertilization, whereby two dimorphic female gametes are fertilized by two immotile sperm cells conveyed by the pollen tube. The two sperm cells are arranged in tandem with a leading pollen tube nucleus to form the male germ unit and are placed under the same genetic controls. Genes controlling double fertilization have been identified, but whether each sperm cell is able to fertilize either female gamete is still unclear. The dynamics of individual sperm cells after their release in the female tissue remain largely unknown. In this study, we photolabeled individual isomorphic sperm cells before their release and analyzed their fate during double fertilization in Arabidopsis thaliana. We found that sperm delivery was composed of three steps. Sperm cells were projected together to the boundary between the two female gametes. After a long period of immobility, each sperm cell fused with either female gamete in no particular order, and no preference was observed for either female gamete. Our results suggest that the two sperm cells at the front and back of the male germ unit are functionally equivalent and suggest unexpected cell-cell communications required for sperm cells to coordinate double fertilization of the two female gametes.     

五唇兰精细胞的分离

五唇兰(Doritis pulcherrima)具二胞花粉,授粉后1 d即有花粉开始萌发,授粉后5 d开始有生殖细胞完成有丝分裂形成一对精细胞。通过人工授粉使花粉管在子房内发育,再利用花粉管直接爆破,成功分离出五唇兰的精细胞。成对的2个精细胞在直径大小、荧光强弱上均显示出较大差异,预示2个精细胞具有不同的前途。     

Cytological Studies of the Cytoplasmic Inheritance in Lilium regale and L. davidii

The distribution and characteristics of plastids and mitochondria in the generative and sperm cells of Lilium regale Wils. and L. davidii Duch. were described. In L. regale there were few plastids and abundant mitochondria in the newly formed generative cell. When the generative cell became free in the vegetative cytoplasm, the plastids degenerated completely within the generative cell. It was further proved by DAPI fluorescent technique that there was no organell DNA in the generative cell within the mature pollen grain or the pollen tube. However, distribution of the plastids was strictly polarizable during the division of the micmspore in L. davidii, resulting the lack of plastids in the newly formed generative cell. Data of RFLP analysis comparable between L. davidii, L. longifiorum and their interspecific hybrid have also proved the plastid inheritance in L. davidii to be of uniparental maternal transmission. Although the mitoehondria were observed both in the generative and sperm cells of L. regale and L. davidii but their DNA was decomposed in the male gametophyte stage. Therefore the mitochondda in the sperm cell could not be transmitted into the offspring. The results provided the detail, cytological evidence that organelles in the microgametophyte are incapable of genetic transmission in the two species of Lilium.     

从水稻花粉管分离精细胞(简报)

精细胞的分离是植物生殖工程的一个重要组成部分,是目前被子植物有性生殖研究的一个活跃领域[1,2]。随着精细胞分离技术的完善和分离出精细胞的植物类型的增加,目前对精细胞的分子生物学研究已有一些进展,主要是精细胞特异蛋白的分离[3,4]和cDNA文库的构建以及一些精细胞特异基因的分离[5,6]。     

She's the boss: signaling in pollen tube reception

In angiosperms, the sperm cells are carried within the pollen tubes (male gametophytes) to the female gametophyte so that double fertilization can occur. The female gametophyte exerts control over the male, with specialized cells known as synergids guiding the pollen tubes and controlling their behavior when they enter the female gametophyte so that the sperm cells can be delivered to the egg and central cell. Upon pollen tube arrival at the ovule, signal transduction cascades mediated by receptor-like kinases are initiated in both the synergid and the tip of the pollen tube, leading to synergid cell death and pollen tube rupture. In this review, we discuss the role of these receptors and of newly discovered members of the pollen tube reception pathway.     

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.     

Ultrastructural Changes of the Microtubule in the Generative Cell of Amaryllis vittata Ait. During Mitosis

Structural changes of microtubules (MTs) in the generative cell (GC) of Amaryllis vittara Alt. during mitosis in pollen tube have been investigated with electron microscopy. The division cycle was completed approximately within 12 h. During prophase, the MTs bundles distributed in the cortex of the GC, they were less and shorter than that before mitosis, some of which beginning to be near the nucleus. When the chromatin condensed and the GC entered metaphase, the MTs increased in number and distributed among the chromosomes (CHs) in the original nuclear zone, but they were not arranged in distinct bundlesed. Some of them connected with the CHs to form kinetochore MTs (KMTs), where as the cortical MTs in prophase still remained there. During metaphase, the CHs were arranged on the equartor forming a metaphase plate, and all the MTs formed a diffuse spindle. When the GC entered anaphase, the KMTs were shortened and they were involved in the segregation of the CHs into two groups. The MTs were much more and focused in the two polar regions. In late anaphase, while the MTs still existed at the poles, rich phragmoplast MTs appeared in the equator zone and the precusors of cell plate (CP) aggregated in the middle of the phragmoplast. When the GC entered telophase, the CHs diffused as chromatin, and phragmoplast MTs extended between the two newly formed nuclear envelops and even through the CP While the polar MTs and KMTs disappeared, the MTs in the newly formed sperm cells were different from that of the GC.