朱顶红生殖细胞胞质分裂的两种方式

大多数植物以形成细胞板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：朱硕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)。有时尚有落后染色体出现(Fig.8)。据此认为：收缩沟的发生与核膜的重建、染色体的异常行为及微管无序有关。朱顶红生殖细胞同时存在两种方式的胞质分裂现象相当特殊,可能存在着两种胞质分裂机制。由于游离的生殖细胞在某种程度上类似于动物细胞,因而以缢缩方式完成胞质分裂是可能的。另一方面,生殖细胞对花粉管生长所处的环境极为敏感,体外培养造成生殖细胞不规剧分裂的可能性也应考虑。因此研究在柱头上萌发花粉管中的生殖细胞的胞质分裂是有意义的,此研究结果将有助于更好地理解生殖细胞胞质分裂的机制。     

朱顶兰生殖细胞的分裂和精细胞形成的观察

朱顶兰生殖细胞的分裂和精细胞形成的观察祁碧霞（广东教育学院生物学系广州５１０３０３）研究花粉和花管中各成员的状态和行为，是当前植物生殖生物学研究的前沿课题之一。过去，我们做了朱顶兰（Ａｍｙｌｉｓｖｉｔｔａｔａ）花粉在离休一活体培养下正常萌发的观察，用...     

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     

细胞有丝分裂的胞质分裂

细胞有丝分裂通常包括核分裂(karyokinesis)及胞质分裂(cytokinesis)两个方面。两者虽有一定的关系,但核分裂并不一定随之有胞质分裂,结果形成双核或多核细胞,如多核原生动物及合胞体组织。因此,应将两者的分裂机制分别理解。关于核分裂机制,在所有细胞中是十分相似的,一般的认识也比较一致。在胞质分裂方面,存在许多不同的看法。大致在动物细胞     

MAPK级联途径调控植物细胞胞质分裂

胞质分裂(cytokinesis)是细胞分裂的最后关键一步,产生2个含有完整的遗传物质和胞质细胞器的子细胞．植物胞质分裂包括细胞板的形成,这一过程是在成膜体的牵引下由一些植物特有的步骤完成的．促分裂原活化蛋白激酶(MAPK)级联途径在真核生物中是高度保守的,由MAPKs,MAPKKs,MAPKKKs组成,通过MAPKKK→ MAPKK → MAPK的逐级磷酸化传递细胞信号．近来的研究表明, NACK-MAPKKK→MAPKK→MAPK→MAP65构成的信号途径调控植物细胞的胞质分裂．本文就这一信号途径,总结了植物胞质分裂机制的研究进展,并对其中的问题进行了讨论与展望．     

SARS冠状病毒核衣壳蛋白抑制胞质分裂和细胞增值

目的:SARS冠状病毒(SARS-CoV)的核衣壳蛋白(N蛋白)能够结合病毒RNA形成螺旋状的核衣壳。在3种不同的细胞系中分别表达SARS-CoVN蛋白,研究它对转染细胞生长的影响。方法:将重组质粒pEGFP-N和pEGFP(作为对照)分别转染人胚肾HEK293细胞、成纤维细胞3T3、人宫颈癌HeLa细胞,通过激光共聚焦显微镜、荧光显微镜观察SARS-CoVN蛋白在细胞内的定位以及细胞的生长变化。结果:SARS-CoVN蛋白能定位于细胞质,并不像其他冠状病毒N蛋白那样能够定位到细胞核。同时发现SARS-CoVN蛋白能够诱导形成多核细胞,多核细胞的百分率可达33.9%。结论:SARS-CoVN蛋白抑制胞质分裂,延缓细胞生长。     

胞质分裂阻滞细胞的微核检测及方法学初步研究

培养人体外周血淋巴细胞的微核检测,是近年来常用的体外短期检测方法之一,可用来评价被检理化因子对人体细胞的遗传毒性,及监测环境、职业接触等有害因子对人类的潜在致癌作用。为了进一步提高该方法的敏感性,根据微核起源于染色体断片与落后染色体的观     

钙调素(CaM)在中体上的分布及参与胞质分裂的调控

钙调素(CaM)是细胞内Ca~(2+)的主要受体,在细胞增殖、分化、凋亡、迁移等过程中都发挥着重要的调控作用。采用GFP标记技术,我们观察了GFP-CaM在胞质分裂期HeLa细胞中的动态分布,发现在胞质分裂后期,GFP-CaM与中体紧密相连。抑制CaM的活性会阻止中体的解聚。进一步观察发现,CaM与γ-微管蛋白共分布在中体两侧,抑制CaM活性也会引起中体γ-微管蛋白解离的延迟。本实验结果说明分布在中体上的CaM很可能通过影响中体微管的稳定,参与调控胞质分裂的完成。     

制备微核的胞质分裂阻断新技术

微核是染色体的无着丝点断片或细胞分裂 末期未纳人子核的落后染色体,在细胞质中形 成的阳性染色质体。人淋巴细胞微核技术是快 速简便地检测染色体损伤的细胞遗传学方 法［[t)。     

朱顶红花粉萌发过程中营养细胞质的超微结构变化

本文应用透射电镜对朱顶红成熟花粉水合、活化和萌发的动态过程中营养细胞质的结构和组成变化进行了观察。成熟花粉具质体、线粒体、内质网、高尔基体。微丝束以聚集体的形式存在。花粉活化后,细胞器的数目和结构发生显著变化：质体和线粒体的片层明显增加,内质网片层狭窄,高尔基体活跃产生小泡,脂体降解及微丝聚集体散开。花粉萌发后,细胞质中出现周质微管和被刺小泡,此期细胞器的变化不明显。微丝以纤丝状遍布整个花粉管中。     

Ultrastructural Observation on Microtubules of the Generative Cell in Amaryllis Germinated Pollens and Pollen Tubes

The organization of microtubules (MTs) in the generative cell (GC) of germinated pollen and pollen tube in Amaryllis vittata Ait. has been studied with electron microscopy. At the beginning of pollen germination, the GC is long elliptic in shape, and is surrounded by its own membrane and also by that of the vegetative cell (VC) ,both of which appear undulated. In cross section, the GC appears roundish and has many lobes. The MT system of GC is mainly organized in bundles, but single MTs can also be observed. The MT bundles are generally located in the lobes, directly beneath the plasma membrane of the cell. These MT bundles orientate along the longitudinal axis of the cell. They are formed by aggregation of 5–6 MTs at least,more often about 30 MTs. In the bundles the MTs are often linked to each other by "cross-bridge". The single tubules in the eytopiasm distribute randomly in different orientations. When the GC has migrated into the pollen tube after germination ,it becomes elongated and has cytoplasmic extensions both in the anterior and posterior end of the cell. The organization of MTs of the GC in pollen tube is similar to that in the germinated pollen grain,but the number of MTs in a bundle often increases to 50–60. In the bundle the "cross-bridges" between the MTs which always link 3–5 MTs, are still seen clearly. Positional shift between the GC and Vegetative nucleus (VN) may take place during the growth of pollen tube. The physical association between GC and VN may be demonstrated some ultrastructural figures. It may be seen that irregular cytoplasmic extensions in the anterior end of the GC is always enclosed by the VN and the projections of the cytoplasmic extensions lie within enclaves of the VN. There are many MTs sheets in the lobes or extensions in the cytoplasm of the GC. Thus the present study demonstrates that MTs have an important role in maintaining the peculiar shape of the GC and the close association between GC and VN. However, it seems that the MTs are probably also engaged in the movement of the GC during pollen growth.     

Positional Shift Between the Vegetative Nucleus and the Generative Cell in Amaryllis Pollen Tube

A modified technique, FITC-tubulin immunofluorescence and DAPI localization to demonstrate simultaneously both the generative cell (GC) and the vegetative nucleus (VN) in the pollen tube under ultra-violet excitation, was developed sucessfully. During the germination of the pollen tube of Amaryllis vittata Ait. the GC and the VN, either being the first one, entered the tube within the first 1—2 h from the pollen grain. However, before the time of GC division, the VN was always positioned distally near the tip of the tube. In case when the GC entered the tube first, then the VN must have a positional shift in order to pass over the GC. The detail processes of positional shift between the GC and the VN were observed. Three basic processes were demonstrated: 1. The anterior end of the VN first reached the vicinity of the posterior attenuated extension of the GC about 2 h following germination forming a temporal physical association. Sometimes their both ends could be inserted into one another for certain extent. 2. The whole VN moved forward and contacted in parallel with the GC until they became twisted together and 3. The VN passed over the GC and greatly elongted lengthwise. Its posterior part became inserted into the anterior end of the GC. The behavior of positional shift between the VN and the GC in the pollen tube seems to be an adjustment of their diameters to fit the narrow tube. A conclusion may be drawn that the rate of movement between the VN and the GC was apparently different during the passage through the tube. Such difference may presumably be accompanied by the independent motive mechanism and structural difference between the VN and GC themselves, which provide their motive force for movement in the 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.     

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     

朱顶红鳞茎芽诱导及植株再生体系的建立

以朱顶红杂交品种红孔雀(Amaryllis vittatacv. Red Peacock)鳞茎盘切块为外植体,对影响鳞茎芽直接萌发增殖及生根的主要因素进行了研究,以建立其高效再生体系.结果表明:(1)1/2MS基本培养基有利于鳞茎芽的萌发和生长;切分方式不同对朱顶红鳞茎增殖具显著效应,偏离切分鳞茎可显著提高增殖系数,最高增殖系数可达6,平均增殖系数为3.6;而对半切分或未切分的增殖系数较低.(2)1/2MS 0.1 mg/L NAA 1.5 mg/L 6-BA 2.0 g/L活性炭的培养基利于鳞茎增殖;活性碳可显著提高鳞茎生根率,在1/2MS 0.1 mg/L NAA 2.0 g/L活性炭的培养基上生根率可达100%,而未加活性炭的生根率较低;再生植株经25 d的自然光照驯化后进行移栽,成活率达90%以上.     

朱顶红花粉管中营养核的纤丝状内含体

用透射是镜研究了朱顶红生殖细胞发育过程中花粉管的营养核中的纤丝状内含体的结构形态。此内含体在花粉管生长早期即可观察到,并在生殖细胞分裂过程中频繁发现。它们具各种方向,未见与其它细胞结构发生联系。通过比较此内含体与花粉管中肌动蛋白纤丝的形态及两者对细胞松弛素Ｄ（ＣＤ）处理的反应,我们倾向认为营养核的纤丝状内含体可能由肌动蛋白组成。讨论了此结构在建立营养核运动机制上的作用。     

被子植物生殖细胞和精细胞

随着被子植物精细胞分离技术的突破和细胞生物学以及分子生物学技术的发展,对被子植物精细胞的研究不断深入。在以前细胞生物学研究的基础上结合近年来的分子生物学研究结果对被子植物雄性生殖细胞的产生、精细胞的形成和发育以及有关精细胞的表面蛋白质、精细胞的特异启动子、精细胞cDNA文库的构建等分子生物学研究进展和今后的发展趋势进行了综述。     

一种新的甘露糖结合凝集素--朱顶兰凝集素基因的克隆及序列分析

运用同源克隆的方法设计简并引物,通过3′和5′RACE技术,从石蒜科植物朱顶兰(Amaryllis vittata Ait)总RNA中克隆了编码此凝集素(AVA)的全长cDNA序列.该基因全长686 bp,起始密码子位于第41～43 bp,终止密码子位于515～517 bp处,开放阅读框长474 bp,编码158个氨基酸,包含信号肽序列、成熟蛋白序列和C-末端剪切序列的前体蛋白.成熟蛋白由109个氨基酸残基组成,分子量为11.9kD.成熟蛋白在氨基酸水平上与雪花莲凝集素、水仙凝集素、石蒜凝集素和君子兰凝集素分别有73.4%、85.3%、80.7%和83.5%的同源性;朱顶兰凝集素的分子模式显示其与雪花莲凝集素有极其相似的三维结构;在Blocks数据库中检索AVA蛋白氨基酸序列的结构域,发现有3个凝集素功能结构域,并具有3个典型的甘露糖专一结合位点盒(QDNY).     

一种新的甘露糖结合凝集素——朱顶兰凝集素基因的克隆及序列分析

运用同源克隆的方法设计简并引物,通过3′和5′RACE技术,从石蒜科植物朱顶兰(Amaryllis vittata Ait)总RNA中克隆了编码此凝集素(AVA)的全长cDNA序列。该基因全长686 bp,起始密码子位于第41～43 bp,终止密码子位于515～517bp处,开放阅读框长474 bp,编码158个氨基酸,包含信号肽序列、成熟蛋白序列和C-末端剪切序列的前体蛋白。成熟蛋白由109个氨基酸残基组成,分子量为11.9kD。成熟蛋白在氨基酸水平上与雪花莲凝集素、水仙凝集素、石蒜凝集素和君子兰凝集素分别有73.4％、85.3％、80.7％和83.5％的同源性;朱顶兰凝集素的分子模式显示其与雪花莲凝集素有极其相似的三维结构;在Blocks数据库中检索AVA蛋白氨基酸序列的结构域,发现有3个凝集素功能结构域,并具有3个典型的甘露糖专一结合位点盒(QDNY)。