关于稻麦胚乳细胞发育的研究
|
摘要
稻麦的胚乳占籽粒重量的90%左右,主要贮藏物质为淀粉和蛋白质。淀粉和蛋白质分别在淀粉体和蛋白体内积累,研究稻麦胚乳淀粉体和蛋白体发育的规律,对改善稻麦籽粒的品质,提高籽粒产量具有重要意义。本文以水稻(Oryza sativa L.)品种“扬稻6号”和小麦(Triticum aestivum L.)品种“扬麦16号”为材料,采取精确标记颖果发育天数,运用组织化学、光学显微镜和电子显微镜(扫描电镜、透射电镜)相结合的观察技术,研究了稻麦胚乳细胞结构特点和发育特性,重点观察了淀粉体和蛋白体发生发育的过程,研究结果将充实细胞生物学、形态解剖学、作物生理学等学科知识,为稻麦品质改良和粮食生产提供基础理论。主要研究结果如下：
1.稻麦颖果的发育。
水稻在开花后颖果长度增长最明显,至花后10d颖果形状基本定型。小麦颖果生长较缓慢,颖果发育至成熟持续的时间较水稻长,花后约30d以后颖果形状基本维持不变。
水稻颖果在发育初期子房壁中积累的淀粉较多,淀粉粒消失的时间也较晚,胚乳I2-KI染色随着发育天数而逐渐加深,小麦颖果在花后花后第6d起胚乳开始积累淀粉,果皮厚度随开花天数的增加逐渐变薄,果皮中的淀粉随着胚乳的发育而逐渐消失。水稻颖果的维管束、胚和胚乳表层细胞脱氢酶活性较强,维持时间也较长。内胚乳在花后10d以后脱氢酶活性已经明显降低,当灌浆趋向停止时不再具有。小麦腹部维管束和果皮在颖果发育早期具有较强的呼吸活性,腹部维管束被TTC染成深红色,表明此时的呼吸活性很高,胚乳在花后17d以后TTC染色逐渐消失,在发育后期呼吸活性变弱。
2.水稻和小麦颖果鲜/干重随着花后天数,呈“S”曲线增长,即前期增长较慢,中间增长迅速,后期增长趋势逐渐减缓。
3.水稻开花后颖果呼吸速率一开始维持在较高的水平,随着发育天数的增加,呼吸速率迅速下降,15d后,慢慢停留在一个稳定的低水平上。小麦前期呼吸速率下降较快,花后5d到花后20d迅速下降,从花后20～30d经历一个缓慢下降的过程,花后30～35d呼吸速率又迅速降低。
4.不同固定和染色方法对稻麦胚乳细胞结构显示的影响。
GA-OsO4固定法能较好保存稻麦胚乳细胞的结构,能清晰的显示胚乳细胞中的淀粉体、蛋白体、内质网、液泡等细胞器,高锰酸钾固定法对细胞中膜结构细胞器的被膜湿示较好。对这两种固定方法的花后不同发育天数的水稻胚乳细胞半薄切片采用TBO,希夫(Schiffs)试剂,多色性染液,高碘酸-希夫-甲苯胺蓝-O (PAS-TBO)染色,结果发现：(1)TBO染色的GA-OsO4制样细胞组织结构清晰,淀粉体、蛋白体、细胞壁、细胞核均清晰可见,但液泡膜以及淀粉体被膜不明显。高锰酸钾固定制样的能较好的显示出细胞中内膜结构,如细胞核、液泡的被膜、淀粉体被膜。此外,淀粉体很容易被染色,而其他细胞组织不易被染色。
(2)多色性染液染色的GA-OsO4制样效果良好,而且着色均匀,反差好,组织结构清晰,细胞壁、蛋白体呈蓝色,淀粉体被膜以及淀粉粒之间的膜被染成玫红色。高锰酸钾固定制样染色细胞核呈蓝色,细胞质、间质呈玫红色,胞核、核膜及染色质清晰可辨,但淀粉体染色效果不好,着色不均匀。
(3)用PAS法染色的GA-OsO4制样能将淀粉体染成玫红色,结构较清晰,而其他细胞器很难被染色。高锰酸钾固定制样用PAS法染色能较好的显示出胚乳细胞壁、蛋白体、淀粉体、淀粉体被膜、淀粉粒之间膜结构,染色效果较GA-OsO4制样好。
(4) PAS-TBO复染的GA-OsO4固定的样品,原先没有被PAS染色的胚乳细胞壁、蛋白体、细胞基质、细胞核都被TBO染色显示。TBO的复染弥补了PAS对GA-OsO4固定样品的染色不足。PAS-TBO复染的高锰酸钾固定制样,淀粉体染色过深,无法得到清晰的照片。
总体上GA-OsO4固定制样的细胞结构要优于高锰酸钾固定液制样,其中TBO和多色性染色效果较好,呈色丰富。对小麦的半薄切片采用TBO, Schiffs试剂,多色性染液,考马斯亮蓝,PAS-TBO染色,结果发现GA-OsO4固定,TBO,多色性染液,Schiffs试剂,考马斯亮蓝染色的半薄切片染色效果比高锰酸钾固定的好,而PAS-TBO复染则是高锰酸钾固定制样的较好,不仅淀粉粒呈玫瑰红色,而且细胞壁、内质网、蛋白体、淀粉体被膜被染成蓝色,镜观丰富,照片清晰美观。在稻麦胚乳的半薄切片观察中,各染剂的着色侧重点不同,因此在染色剂的选择时则要根据观察对象的不同而选择相应的染色方法。
在这两种固定方法的超薄切片观察中,GA-OsO4固定法能较好的保存胚乳细胞超微结构,但淀粉体被膜不明显,反差弱。高锰酸钾固定制样不利于对胚乳细胞超微结构整体观察,因为其对细胞基质和各种细胞器基质固定较差,但对膜结构保存良好。如叶绿体被膜和片层结构、淀粉体被膜、小麦大淀粉体内片层膜结构等。本文利用该固定方法观察了稻麦胚乳淀粉体发育增殖及胚乳细胞发育中内质网的生理活动,获得了较满意的结果
5.稻麦胚乳淀粉体的发生发育以及增殖方式。
水稻胚乳淀粉体一般是由原生质体转化而来的。本文在观察淀粉体增殖的过程中采用戊二醛-锇酸(GA-OsO4)固定制样和高锰酸钾固定制样两种方法。GA-OsO4固定法能较好的保存胚乳细胞超微结构,但淀粉体被膜不明显,反差弱。高锰酸钾固定制样不利于对胚乳细胞超微结构整体观察,因为其对细胞基质和各种细胞器基质固定较差,但对膜结构保存良好。基于这两种不同固定方法的特性,观察淀粉体增殖的重点在于淀粉体被膜的变化,因此,高锰酸钾固定制样是此项研究的主要方法。
根据淀粉体被膜的变化,将水稻淀粉体的增殖方式划分为出芽增殖、缢缩分裂增殖、中间隔板增殖、淀粉体被膜内陷成小泡增殖、淀粉体内外膜腔膨大增殖。其中出芽增殖和缢缩分裂增殖较为常见,是胚乳细胞发育前期淀粉体增殖的主要方式。中间隔板增殖有两种方式,第一种是指某一处的淀粉体被膜向淀粉体基质中凹陷,形成单向的袋装被膜隔板,当被膜隔板触及到原淀粉体被膜时,将淀粉体分裂成两个淀粉体单元,达到增殖目的。另一种是淀粉体被膜相向内凸伸长,直到接触并融合,形成双层膜中间隔板,然后沿中间隔板将原淀粉体分裂成两个新淀粉体。在淀粉体增殖过程中,可以观察到一个淀粉体可以同时进行两种甚至多种增殖方式。
小麦胚乳有大小两种淀粉体,花后6d起,胚乳细胞中的原生质体开始积累淀粉转变成淀粉体,1个淀粉体内可以同时积累2个或多个淀粉粒,大淀粉体可以通过出芽或缢缩增殖,本文用两种固定方法都观察到了大淀粉体这两种增殖方式。
花后8～14d,大淀粉体被膜向内凹陷形成基质浓密小泡,或向细胞基质突出,然后这些突起与淀粉体分开,形成新的淀粉体,即小淀粉体。花后16d,胚乳中小淀粉体逐渐增多,小淀粉粒除了在大淀粉体内积累以外,自身还可以发生缢缩增殖和中间隔板方式分裂增殖。
6.稻麦胚乳蛋白体的形成和积累。
水稻胚乳蛋白体有两种类型,蛋白体Ⅰ (protein body Ⅰ, PbⅠ)由粗面内质网(RER)发育而来,呈球形,表面附有核糖体或多聚核糖体,在剖面上呈现同心圆状的轮纹结构,可能是包裹它的RER向内分泌本身合成的蛋白质并从内向外积累而形成。已成形的PbⅠ通常有一不连续的单层核糖体被膜包被,一些含蛋白质基质的小泡融合到Pb Ⅰ上,使Pb Ⅰ体积不断增大。蛋白体Ⅱ (protein body Ⅱ, PbⅡ)由液泡发育而来,多呈不规则块状,也有的液泡吞噬蛋白质基质小泡,在自身体内积累蛋白质。当液泡中开始积累蛋白质时,液泡便转化为蛋白贮藏液泡(PSV),在PSV中通常可见贮藏蛋白的晶体结构；花后6d的小麦胚乳细胞中富含PSV,其内出现一个或者多个圆球状的蛋白质颗粒,同时,RER在小麦胚乳蛋白体发育周期中活动也很旺盛,有些RER相互叠加形成闭合的网状RER,而有些则呈环状,在这些RER中的基质的电子致密程度要比细胞基质高,有的RER网腔或末端膨大,积累贮藏蛋白。
稻麦胚乳蛋白大多贮藏在亚糊粉层中,而内胚乳含量较少。
7.用Image J分析水稻胚乳淀粉粒表面几何特征的方法。以水稻淀粉粒光学显微镜和扫描电子显微镜图像为研究对象,介绍了图像分析软件Image J在淀粉粒表面特征分析中的应用。图像分析结果以轮廓图(outline)和拟合椭圆(Ellipse fitting)两种方式呈现,分析指标选择了面积、周长、长轴、短轴、圆度以及最大切直径。最后还对颗粒图像的获取和图像前处理进行了探讨。
8.关于水稻胚乳中是否存在单粒淀粉体的探讨。该研究分别以花后5d和成熟水稻胚乳半薄连续切片,用GA-OsO4固定法和高锰酸钾固定制样观察水稻花后不同发育天数胚乳细胞超微结构,并以垩白籽粒为对比,用显微和亚显微研究技术对水稻胚乳中是否含有单粒淀粉体进行了探讨,结果发现水稻胚乳中绝大多数的淀粉体为复粒淀粉体,少数为单粒淀粉体。
The endosperm, rich in starch and protein, is the largest part of the kernel and takes up nearly90percent of the kernel's mass. Starch and protein are stored in the form of amyloplast and protein body respectively. Elucidating their development regularity may be a great help to both theoretical and practical guiding significance.
In our study, developmental time of developing caryopsis from rice (Yangdao6) and wheat (YangMai16) is accurately tagged. Histochemical assays, light microscopy, electron microscope (SEM and TEM) were employed to investigate the structure features, developmental process, formation mechanism and physiological function of endosperm, of which the developmental formation of the amyloplast and protein body are mainly observed. Elucidation of above questions will somehow deepen the study of endosperm development, enrich the knowledge of cell biology, crop science, and provide theoretical basis for mass production and crop quality improvement.
Main results were as follows:
1. The development of rice and wheat caryopses.
The fastest growth rate of rice caryopsis appears to be within first few days after anthesis, till10days after followring (DAF) the shape of rice caryopsis is no longer changed. In wheat, the growth rate is relatively slow, till30DAF maximum volume reached.
Starch first accumulated in the rice ovary wall, and lasts long during the whole developmental process. I2-KI staining on the rice endosperm gets darker with DAF increases. In wheat caryopsis, starch begins to accumulate from6DAF in endosperm, thickness of the pericarp declines as DAF increases, with its starch degrading gradually. Vascular bundle region, embryo and peripheral endosperm cells have a relatively higher and long-lasting dehydrogenase activity, while the endosperm cells, shows little activity in10DAF, and no activity when grain-filling finished. In wheat, ventral vascular bundle region and pericarp both show a high dehydrogenase activity, the endosperm shows a declined activity in17DAF.
2. Freshweight and dryweight of rice and wheat caryopsis grows under a typical "S-shaped" behavior, following a first slow then fast and finally slow rate.
3. Respiration rate of rice caryopsis maintained a high level at first, then declines rapidly till15DAF; while in wheat, the rate first drops sharply from5to20DAF, from20to30DAF slowed down, and30to35DAF, increases rapidly.
4. Effects of varying fixative and staining methods on the structure display of wheat and rice endosperm cells.
Conventional glutaraldehyde-osmium tetraoxide (GA-OSO4) fixation better in preserving cell ultrastructure, such as amyloplasts, protein bodies, Endoplasmic reticulum and vacuoles, while potassium permanganate fixation proves to be better fixative for endomembrane system. Semi-thin sections staining were carried out using toluidine blue (TBO), polychromatic dye, periodic acid Schiff (PAS), coomassie brilliant blue and PAS-TBO counterstaining. The results were as follows:
(1) GA-OSO4fixation and TBO stain preserve cell structures well, amyloplasts, protein bodies, cell walls and nucleus all have a better display than amyloplast envelope and tonoplast. Samples fixed with Potassium permanganate show a good result in endomembrane system display, like nuclear membrane, tonoplast and amyloplast envelope. Besides, amyloplast stains easily, while other cellular tissues not.
(2) GA-OSO4fixation and polychromatic dye provide the uniformly color, moderate contrast, with cell wall and protein body being stained blue, amyloplast envelope and inner septum stained rose red. Samples fixed in potassium permanganate present a slight different result, in that nulear being blue, cytoplasm and stroma being rose red, and amyloplast poorly stained.
(3) Only amyloplasts from samples fixed with GA-OSO4and stained with PAS can be colored rose red and other organelles are unstainable. When fixed with potassium permanganate, results improved, in that endosperm cell wall, protein body, amyloplast and its envelope and septum are all well colored.
(4) Samples fixed with above GA-OSO4and PAS, when counterstained with TBO show a improved results in color reactions, cell wall, protein body, stroma and nuclear etc all well displayed. But the amyloplast from samples fixed with potassium permanganate can be unevenly and over stained by PAS-TBO.
In most cases, cells with GA-OSO4fixation have a better result than with potassium permanganate fixation, and of all the staining methods, TBO and polychromatic dye have the best effect. By comparing effects of2different fixation and5staining methods on wheat semisections, we found that GA-OSO4fixed samples, stained by TBO, polychromatic dye, periodic acid PAS, coomassie brilliant blue, are superior to those fixed with potassium permanganate, but in PAS-TBO counterstaining, potassium permanganate fixation made the best performance. Therefore, the selection of appropriate fixation and staining methods should be based on experimental destination.
Through observation in the ultra-thin sections, we find that the GA-OsO4fixation was a better fixative in preserving endosperm cell intrinsic feature, but lacked clarity in membranous structure display. Potassium permanganate fixation, although was able to show the structure of the cell endomembrane better and its contrast was satisfactory, but it was poor on the polysaccharide and protein colour generation. We chose the latter method to study physiological activities of endoplasmic reticulum and results are optimistic.
5. Initiation, development and proliferation of rice and wheat amyloplast.
Rice endosperm amyloplast is commonly formed from proplastids. Utilizing GA-OSO4and Potassium permanganate fixation respectively, we find that:samples fixed with GA-OSO4although have better effection preserving the microstructure of endosperm cells, show a low contrast in amyloplast envelope; while samples fixed with potassium permanganate perform better in membrane structure display, but perform poor in cell stroma and organelle stroma fixation. As amyloplasts proliferation mainly manifested in their envelope changes, the latter fixation method are our main choice.
Rice endosperm amyloplasts proliferate mainly by means of envelope protrusion, constriction, invagination, middle septum dividing, expansion of amyloplast envelop et al, among which the first two are the most common ones in earlier stage of endosperm cell development. Two middle septum formation mechanisms have been observed, first is the amyloplast envelop invaginate to amyloplast stroma, when it touched opposite envelop, two new amylopplast appeared, another is two amyloplast envelops move in opposite direction, two new amyloplasts generated when two envelops linked. In the amyloplast proliferation process, two or more ways of amyloplast proliferation could be observed in a amyloplast.
Wheat endosperm amyloplast, differ from rice, has two types of amyloplast, starch begins accumulating in proplastids from6DAF, transforming into amyloplasts. Each amyloplast contains2or more starch granules. Large amyloplasts proliferate by means of envelope protrusion and constriction, of which are observed by both fixation methods.
From8to14DAF, envelope of large amyloplasts either invaginate into stroma dense vacuoles, or protrude outwardly, forming a new type of amyloplast, called small amyloplast. From16DAF, small amyloplasts accumulate, not only by way of envelope protrusion, but by self-constriction or middle septum division as well.
6. Formation and accumulation of rice and wheat protein body.
Two types of protein bodies are found in rice endosperm, protein body I, Pbl and protein body Ⅱ, PbⅡ. PbⅠ, derives from the secretion of Rough Endoplasmic reticulum, is a spherical, concentric circle-like structure(on the section),which surface covered with ribosomes and polyribosomes, and is possibly a product accumulated through RER inward secretion layer-by layers. Matured PbⅠ usually coated by a single layer of discontinuous ribosome, and can be fused by vacuoles with protein matrix.
PbⅡ, developed from vacuoles, mostly are irregular clots like. When vacuoles start accumulating proteins, they turn into PSV, in which crystal structures of storage proteins can be detected. Enriched PSVs are found in wheat endosperm cells from6DAF, in the form of spherical particles. Meanwhile, RER retains a high level of activity (of which matrix electron density higher than cell matrix), which stacked together, and thus forming either looped or reticulated structure. Some RER's cavity or terminal may be protruding where storage protein is accumulated.
Most rice and wheat protein bodies are stored in the sub-aleurone layer, seldom seen in endosperm cells.
7. The surface of the geometric characteristics analysis for rice endosperm starch granules by using Image J.
Based on the optical microscope and scanning electron microscope images of rice starch granules, the image analysis software image J was introduced in analysis of starch granules of surface features. Image analysis presented in two ways:outline and ellipse fitting. Analysis of indicators chosen area, perimeter, major, minor, circularity and Feret. Finally, the particle image acquisition and image pre-treatment were discussed.
8. Discussion on the existence of amyloplast which contain single starch grain in rice endosperm.
5DAF and mature rice caryopsis were made to consecutive semithin sections, GA-OSO4and potassium permanganate fixation were used for different days after followering rice caryopsis, Meanwhile, chalky caryopsis was took as contrast, both of them were observed ultrastructure by TEM and SEM. The results found that the vast majority of the rice endosperm amyloplast was compound grain amyloplast, a few was single grain amyloplast.
1.稻麦颖果的发育。
水稻在开花后颖果长度增长最明显,至花后10d颖果形状基本定型。小麦颖果生长较缓慢,颖果发育至成熟持续的时间较水稻长,花后约30d以后颖果形状基本维持不变。
水稻颖果在发育初期子房壁中积累的淀粉较多,淀粉粒消失的时间也较晚,胚乳I2-KI染色随着发育天数而逐渐加深,小麦颖果在花后花后第6d起胚乳开始积累淀粉,果皮厚度随开花天数的增加逐渐变薄,果皮中的淀粉随着胚乳的发育而逐渐消失。水稻颖果的维管束、胚和胚乳表层细胞脱氢酶活性较强,维持时间也较长。内胚乳在花后10d以后脱氢酶活性已经明显降低,当灌浆趋向停止时不再具有。小麦腹部维管束和果皮在颖果发育早期具有较强的呼吸活性,腹部维管束被TTC染成深红色,表明此时的呼吸活性很高,胚乳在花后17d以后TTC染色逐渐消失,在发育后期呼吸活性变弱。
2.水稻和小麦颖果鲜/干重随着花后天数,呈“S”曲线增长,即前期增长较慢,中间增长迅速,后期增长趋势逐渐减缓。
3.水稻开花后颖果呼吸速率一开始维持在较高的水平,随着发育天数的增加,呼吸速率迅速下降,15d后,慢慢停留在一个稳定的低水平上。小麦前期呼吸速率下降较快,花后5d到花后20d迅速下降,从花后20～30d经历一个缓慢下降的过程,花后30～35d呼吸速率又迅速降低。
4.不同固定和染色方法对稻麦胚乳细胞结构显示的影响。
GA-OsO4固定法能较好保存稻麦胚乳细胞的结构,能清晰的显示胚乳细胞中的淀粉体、蛋白体、内质网、液泡等细胞器,高锰酸钾固定法对细胞中膜结构细胞器的被膜湿示较好。对这两种固定方法的花后不同发育天数的水稻胚乳细胞半薄切片采用TBO,希夫(Schiffs)试剂,多色性染液,高碘酸-希夫-甲苯胺蓝-O (PAS-TBO)染色,结果发现：(1)TBO染色的GA-OsO4制样细胞组织结构清晰,淀粉体、蛋白体、细胞壁、细胞核均清晰可见,但液泡膜以及淀粉体被膜不明显。高锰酸钾固定制样的能较好的显示出细胞中内膜结构,如细胞核、液泡的被膜、淀粉体被膜。此外,淀粉体很容易被染色,而其他细胞组织不易被染色。
(2)多色性染液染色的GA-OsO4制样效果良好,而且着色均匀,反差好,组织结构清晰,细胞壁、蛋白体呈蓝色,淀粉体被膜以及淀粉粒之间的膜被染成玫红色。高锰酸钾固定制样染色细胞核呈蓝色,细胞质、间质呈玫红色,胞核、核膜及染色质清晰可辨,但淀粉体染色效果不好,着色不均匀。
(3)用PAS法染色的GA-OsO4制样能将淀粉体染成玫红色,结构较清晰,而其他细胞器很难被染色。高锰酸钾固定制样用PAS法染色能较好的显示出胚乳细胞壁、蛋白体、淀粉体、淀粉体被膜、淀粉粒之间膜结构,染色效果较GA-OsO4制样好。
(4) PAS-TBO复染的GA-OsO4固定的样品,原先没有被PAS染色的胚乳细胞壁、蛋白体、细胞基质、细胞核都被TBO染色显示。TBO的复染弥补了PAS对GA-OsO4固定样品的染色不足。PAS-TBO复染的高锰酸钾固定制样,淀粉体染色过深,无法得到清晰的照片。
总体上GA-OsO4固定制样的细胞结构要优于高锰酸钾固定液制样,其中TBO和多色性染色效果较好,呈色丰富。对小麦的半薄切片采用TBO, Schiffs试剂,多色性染液,考马斯亮蓝,PAS-TBO染色,结果发现GA-OsO4固定,TBO,多色性染液,Schiffs试剂,考马斯亮蓝染色的半薄切片染色效果比高锰酸钾固定的好,而PAS-TBO复染则是高锰酸钾固定制样的较好,不仅淀粉粒呈玫瑰红色,而且细胞壁、内质网、蛋白体、淀粉体被膜被染成蓝色,镜观丰富,照片清晰美观。在稻麦胚乳的半薄切片观察中,各染剂的着色侧重点不同,因此在染色剂的选择时则要根据观察对象的不同而选择相应的染色方法。
在这两种固定方法的超薄切片观察中,GA-OsO4固定法能较好的保存胚乳细胞超微结构,但淀粉体被膜不明显,反差弱。高锰酸钾固定制样不利于对胚乳细胞超微结构整体观察,因为其对细胞基质和各种细胞器基质固定较差,但对膜结构保存良好。如叶绿体被膜和片层结构、淀粉体被膜、小麦大淀粉体内片层膜结构等。本文利用该固定方法观察了稻麦胚乳淀粉体发育增殖及胚乳细胞发育中内质网的生理活动,获得了较满意的结果
5.稻麦胚乳淀粉体的发生发育以及增殖方式。
水稻胚乳淀粉体一般是由原生质体转化而来的。本文在观察淀粉体增殖的过程中采用戊二醛-锇酸(GA-OsO4)固定制样和高锰酸钾固定制样两种方法。GA-OsO4固定法能较好的保存胚乳细胞超微结构,但淀粉体被膜不明显,反差弱。高锰酸钾固定制样不利于对胚乳细胞超微结构整体观察,因为其对细胞基质和各种细胞器基质固定较差,但对膜结构保存良好。基于这两种不同固定方法的特性,观察淀粉体增殖的重点在于淀粉体被膜的变化,因此,高锰酸钾固定制样是此项研究的主要方法。
根据淀粉体被膜的变化,将水稻淀粉体的增殖方式划分为出芽增殖、缢缩分裂增殖、中间隔板增殖、淀粉体被膜内陷成小泡增殖、淀粉体内外膜腔膨大增殖。其中出芽增殖和缢缩分裂增殖较为常见,是胚乳细胞发育前期淀粉体增殖的主要方式。中间隔板增殖有两种方式,第一种是指某一处的淀粉体被膜向淀粉体基质中凹陷,形成单向的袋装被膜隔板,当被膜隔板触及到原淀粉体被膜时,将淀粉体分裂成两个淀粉体单元,达到增殖目的。另一种是淀粉体被膜相向内凸伸长,直到接触并融合,形成双层膜中间隔板,然后沿中间隔板将原淀粉体分裂成两个新淀粉体。在淀粉体增殖过程中,可以观察到一个淀粉体可以同时进行两种甚至多种增殖方式。
小麦胚乳有大小两种淀粉体,花后6d起,胚乳细胞中的原生质体开始积累淀粉转变成淀粉体,1个淀粉体内可以同时积累2个或多个淀粉粒,大淀粉体可以通过出芽或缢缩增殖,本文用两种固定方法都观察到了大淀粉体这两种增殖方式。
花后8～14d,大淀粉体被膜向内凹陷形成基质浓密小泡,或向细胞基质突出,然后这些突起与淀粉体分开,形成新的淀粉体,即小淀粉体。花后16d,胚乳中小淀粉体逐渐增多,小淀粉粒除了在大淀粉体内积累以外,自身还可以发生缢缩增殖和中间隔板方式分裂增殖。
6.稻麦胚乳蛋白体的形成和积累。
水稻胚乳蛋白体有两种类型,蛋白体Ⅰ (protein body Ⅰ, PbⅠ)由粗面内质网(RER)发育而来,呈球形,表面附有核糖体或多聚核糖体,在剖面上呈现同心圆状的轮纹结构,可能是包裹它的RER向内分泌本身合成的蛋白质并从内向外积累而形成。已成形的PbⅠ通常有一不连续的单层核糖体被膜包被,一些含蛋白质基质的小泡融合到Pb Ⅰ上,使Pb Ⅰ体积不断增大。蛋白体Ⅱ (protein body Ⅱ, PbⅡ)由液泡发育而来,多呈不规则块状,也有的液泡吞噬蛋白质基质小泡,在自身体内积累蛋白质。当液泡中开始积累蛋白质时,液泡便转化为蛋白贮藏液泡(PSV),在PSV中通常可见贮藏蛋白的晶体结构；花后6d的小麦胚乳细胞中富含PSV,其内出现一个或者多个圆球状的蛋白质颗粒,同时,RER在小麦胚乳蛋白体发育周期中活动也很旺盛,有些RER相互叠加形成闭合的网状RER,而有些则呈环状,在这些RER中的基质的电子致密程度要比细胞基质高,有的RER网腔或末端膨大,积累贮藏蛋白。
稻麦胚乳蛋白大多贮藏在亚糊粉层中,而内胚乳含量较少。
7.用Image J分析水稻胚乳淀粉粒表面几何特征的方法。以水稻淀粉粒光学显微镜和扫描电子显微镜图像为研究对象,介绍了图像分析软件Image J在淀粉粒表面特征分析中的应用。图像分析结果以轮廓图(outline)和拟合椭圆(Ellipse fitting)两种方式呈现,分析指标选择了面积、周长、长轴、短轴、圆度以及最大切直径。最后还对颗粒图像的获取和图像前处理进行了探讨。
8.关于水稻胚乳中是否存在单粒淀粉体的探讨。该研究分别以花后5d和成熟水稻胚乳半薄连续切片,用GA-OsO4固定法和高锰酸钾固定制样观察水稻花后不同发育天数胚乳细胞超微结构,并以垩白籽粒为对比,用显微和亚显微研究技术对水稻胚乳中是否含有单粒淀粉体进行了探讨,结果发现水稻胚乳中绝大多数的淀粉体为复粒淀粉体,少数为单粒淀粉体。
The endosperm, rich in starch and protein, is the largest part of the kernel and takes up nearly90percent of the kernel's mass. Starch and protein are stored in the form of amyloplast and protein body respectively. Elucidating their development regularity may be a great help to both theoretical and practical guiding significance.
In our study, developmental time of developing caryopsis from rice (Yangdao6) and wheat (YangMai16) is accurately tagged. Histochemical assays, light microscopy, electron microscope (SEM and TEM) were employed to investigate the structure features, developmental process, formation mechanism and physiological function of endosperm, of which the developmental formation of the amyloplast and protein body are mainly observed. Elucidation of above questions will somehow deepen the study of endosperm development, enrich the knowledge of cell biology, crop science, and provide theoretical basis for mass production and crop quality improvement.
Main results were as follows:
1. The development of rice and wheat caryopses.
The fastest growth rate of rice caryopsis appears to be within first few days after anthesis, till10days after followring (DAF) the shape of rice caryopsis is no longer changed. In wheat, the growth rate is relatively slow, till30DAF maximum volume reached.
Starch first accumulated in the rice ovary wall, and lasts long during the whole developmental process. I2-KI staining on the rice endosperm gets darker with DAF increases. In wheat caryopsis, starch begins to accumulate from6DAF in endosperm, thickness of the pericarp declines as DAF increases, with its starch degrading gradually. Vascular bundle region, embryo and peripheral endosperm cells have a relatively higher and long-lasting dehydrogenase activity, while the endosperm cells, shows little activity in10DAF, and no activity when grain-filling finished. In wheat, ventral vascular bundle region and pericarp both show a high dehydrogenase activity, the endosperm shows a declined activity in17DAF.
2. Freshweight and dryweight of rice and wheat caryopsis grows under a typical "S-shaped" behavior, following a first slow then fast and finally slow rate.
3. Respiration rate of rice caryopsis maintained a high level at first, then declines rapidly till15DAF; while in wheat, the rate first drops sharply from5to20DAF, from20to30DAF slowed down, and30to35DAF, increases rapidly.
4. Effects of varying fixative and staining methods on the structure display of wheat and rice endosperm cells.
Conventional glutaraldehyde-osmium tetraoxide (GA-OSO4) fixation better in preserving cell ultrastructure, such as amyloplasts, protein bodies, Endoplasmic reticulum and vacuoles, while potassium permanganate fixation proves to be better fixative for endomembrane system. Semi-thin sections staining were carried out using toluidine blue (TBO), polychromatic dye, periodic acid Schiff (PAS), coomassie brilliant blue and PAS-TBO counterstaining. The results were as follows:
(1) GA-OSO4fixation and TBO stain preserve cell structures well, amyloplasts, protein bodies, cell walls and nucleus all have a better display than amyloplast envelope and tonoplast. Samples fixed with Potassium permanganate show a good result in endomembrane system display, like nuclear membrane, tonoplast and amyloplast envelope. Besides, amyloplast stains easily, while other cellular tissues not.
(2) GA-OSO4fixation and polychromatic dye provide the uniformly color, moderate contrast, with cell wall and protein body being stained blue, amyloplast envelope and inner septum stained rose red. Samples fixed in potassium permanganate present a slight different result, in that nulear being blue, cytoplasm and stroma being rose red, and amyloplast poorly stained.
(3) Only amyloplasts from samples fixed with GA-OSO4and stained with PAS can be colored rose red and other organelles are unstainable. When fixed with potassium permanganate, results improved, in that endosperm cell wall, protein body, amyloplast and its envelope and septum are all well colored.
(4) Samples fixed with above GA-OSO4and PAS, when counterstained with TBO show a improved results in color reactions, cell wall, protein body, stroma and nuclear etc all well displayed. But the amyloplast from samples fixed with potassium permanganate can be unevenly and over stained by PAS-TBO.
In most cases, cells with GA-OSO4fixation have a better result than with potassium permanganate fixation, and of all the staining methods, TBO and polychromatic dye have the best effect. By comparing effects of2different fixation and5staining methods on wheat semisections, we found that GA-OSO4fixed samples, stained by TBO, polychromatic dye, periodic acid PAS, coomassie brilliant blue, are superior to those fixed with potassium permanganate, but in PAS-TBO counterstaining, potassium permanganate fixation made the best performance. Therefore, the selection of appropriate fixation and staining methods should be based on experimental destination.
Through observation in the ultra-thin sections, we find that the GA-OsO4fixation was a better fixative in preserving endosperm cell intrinsic feature, but lacked clarity in membranous structure display. Potassium permanganate fixation, although was able to show the structure of the cell endomembrane better and its contrast was satisfactory, but it was poor on the polysaccharide and protein colour generation. We chose the latter method to study physiological activities of endoplasmic reticulum and results are optimistic.
5. Initiation, development and proliferation of rice and wheat amyloplast.
Rice endosperm amyloplast is commonly formed from proplastids. Utilizing GA-OSO4and Potassium permanganate fixation respectively, we find that:samples fixed with GA-OSO4although have better effection preserving the microstructure of endosperm cells, show a low contrast in amyloplast envelope; while samples fixed with potassium permanganate perform better in membrane structure display, but perform poor in cell stroma and organelle stroma fixation. As amyloplasts proliferation mainly manifested in their envelope changes, the latter fixation method are our main choice.
Rice endosperm amyloplasts proliferate mainly by means of envelope protrusion, constriction, invagination, middle septum dividing, expansion of amyloplast envelop et al, among which the first two are the most common ones in earlier stage of endosperm cell development. Two middle septum formation mechanisms have been observed, first is the amyloplast envelop invaginate to amyloplast stroma, when it touched opposite envelop, two new amylopplast appeared, another is two amyloplast envelops move in opposite direction, two new amyloplasts generated when two envelops linked. In the amyloplast proliferation process, two or more ways of amyloplast proliferation could be observed in a amyloplast.
Wheat endosperm amyloplast, differ from rice, has two types of amyloplast, starch begins accumulating in proplastids from6DAF, transforming into amyloplasts. Each amyloplast contains2or more starch granules. Large amyloplasts proliferate by means of envelope protrusion and constriction, of which are observed by both fixation methods.
From8to14DAF, envelope of large amyloplasts either invaginate into stroma dense vacuoles, or protrude outwardly, forming a new type of amyloplast, called small amyloplast. From16DAF, small amyloplasts accumulate, not only by way of envelope protrusion, but by self-constriction or middle septum division as well.
6. Formation and accumulation of rice and wheat protein body.
Two types of protein bodies are found in rice endosperm, protein body I, Pbl and protein body Ⅱ, PbⅡ. PbⅠ, derives from the secretion of Rough Endoplasmic reticulum, is a spherical, concentric circle-like structure(on the section),which surface covered with ribosomes and polyribosomes, and is possibly a product accumulated through RER inward secretion layer-by layers. Matured PbⅠ usually coated by a single layer of discontinuous ribosome, and can be fused by vacuoles with protein matrix.
PbⅡ, developed from vacuoles, mostly are irregular clots like. When vacuoles start accumulating proteins, they turn into PSV, in which crystal structures of storage proteins can be detected. Enriched PSVs are found in wheat endosperm cells from6DAF, in the form of spherical particles. Meanwhile, RER retains a high level of activity (of which matrix electron density higher than cell matrix), which stacked together, and thus forming either looped or reticulated structure. Some RER's cavity or terminal may be protruding where storage protein is accumulated.
Most rice and wheat protein bodies are stored in the sub-aleurone layer, seldom seen in endosperm cells.
7. The surface of the geometric characteristics analysis for rice endosperm starch granules by using Image J.
Based on the optical microscope and scanning electron microscope images of rice starch granules, the image analysis software image J was introduced in analysis of starch granules of surface features. Image analysis presented in two ways:outline and ellipse fitting. Analysis of indicators chosen area, perimeter, major, minor, circularity and Feret. Finally, the particle image acquisition and image pre-treatment were discussed.
8. Discussion on the existence of amyloplast which contain single starch grain in rice endosperm.
5DAF and mature rice caryopsis were made to consecutive semithin sections, GA-OSO4and potassium permanganate fixation were used for different days after followering rice caryopsis, Meanwhile, chalky caryopsis was took as contrast, both of them were observed ultrastructure by TEM and SEM. The results found that the vast majority of the rice endosperm amyloplast was compound grain amyloplast, a few was single grain amyloplast.
引文
Adeli K, Aleosaar I. Role of endoplasmic reticulum in biosynthesis of oat globulin precursors. Plant Physiol,1983,73:949-955.
Badenhuize N P. Encyclopaedia of plant physiology, Vol VI(Ruhland ed.). Berlin:Springer Verlag,1958:137-153.
Barlow K K, Lee J W, Vesk M. Morphological development of storage protein bodies in wheat. Mechanisms of Regulations of Plant Growth, Wellington:Royal Society of New Zealand, 1974:793.
Bechtel D B, Gaines R L, Pomeranz Y. Protein secretion in wheat endosperm:formation of the matrix protein. Cereal Chemistry,1982,59(5):336-343.
Bechtel D B, Gains R L, Pomeranz Y. Early stages in wheat endosperm formation and protein body initiation. Annu Bot,1982,50:507-518.
Bechtel D B, Gaines R L, Pomeranz Y. Early stages in wheat endosperm formation and protein body initiation. Ann Bot,1982,73:949-955.
Bechtel D B, Zayas I Y, Kaleikau L, et al. Size distribution of wheat starch granules during endosperm development. Cereal Chemistry,1990,67:59-63.
Bechtel D B, Wilson J D, Shewry R P. Immunocytochemical localization of the wheat storage protein triticin in developing endosperm tissue. Cereal Chemistry,199l.68(6):573-577.
Becraft P W, Asuncion-Crabb Y. Positional cues specify and maintain aleurone cell fate in maize endosperm development. Development,2000,127(18):4039-4048.
Beehtel D R, Juliano B O. Formation of protein bodies in the starchy endosperm of rice:a rein vestigation. Ann. Bot.1980,45:503-509.
Bell P R. Megaspore abortion:a consequence of selective apoptosis. Int J Plant Sci,1996,157: 1-7.
Briarty L G, Hughes C E, Evers A D. The developing endosperm of wheat-a stereological analysis. Ann Bot,1979,44:641-658.
Brown R C, Lemon B E, Olsen O A. Endosperm development in barley:microtubule involvement in the morphogenetic pathway. Plant cell,1994,6:1241-1252.
Brown R C, Lemmon B E, Olsen O A. Polarization prediets the pattern of cellularization in cereal endosperm. Protoplasma,1996,192:168-177.
Brown R C, Lemmon B E, Stone B A, Olsen O A. Cell wall (1→3)-and(1-3,1-4)-β-glueans during early grian development in rice(Oryza sativa L.). Planta,1997,202:414-426.
Brown R C., Lernrnon B E., Nguyen H, Olsen O-A. Development of endosperm in Arabidopsis thaliana. Sex Plant Reprod.1999,12:32-42.
Buchanan B B, Gruissem W, Jones R L. Biochemistry&molecular biology of plants.北京：科 学出版社,2002,18-19.
Buttrose M S. Submicroscopic development and structure of starch granules in cereal endosperm. J Ultrastructure Research,1960,4:231-257.
Buttrose M S. Ultrastructure of the developing wheat endosperm. Austrlian Journal of Biology Science,1963,16:305-317.
Chamberlin M A, Homer H T. Nutrition of early embryo development in Glycine max (L.)Merr. Alller J Bot,1990,77:20.
Chamberlin M A, Horner H T. An ultrastruetural study of endosperm wall formation and degeneration in Glycjne max(L.) Merr.AlnerJBot,1990,77:83.
Chamberlin M A, Horner H T, Palmer R G. Early endosperm, embryo, and ovule development in Glycine max(L.) Merr. Int J Plant Sci,1994,155(4):421-436.
Chamberlin M A, Homer H T. Palmer R G. Nuelear size and DNA content of the embryo and endosperm during the irinitial stages of development in Glycine max (Fabaceae). Amer J Bot,1993,80,(10):1209-1215.
Chitra lekha P. Bhandari N N. Cellularization of endosperm in AsPhodelus trenuifolius Cav.. Phytomorphology,1992,42(3-4):185-193.
Chung M C. Development of rice caryopsis. National Science Council Monthly,1983,11(10): 959-976.
Dang J M, Copeland L. Studies of the fracture surface of rice grains using environmental scanning electron microscopy. J Sci Food Agric,2004,84:707-713.
Dellaporta S L, Calderon-Urrea A. The sex determination proeess in maize. Science,1994,266: 1501-1505.
Dominique Cledat, Serge. Battu, Redouane. Mokrini, Philippe. J. P. Cardot. Rice starch granule charact-erization by flow cytometry scattering techniques hyphenated with sedimentation field-flow fractionation. Journal of chromatography A.2004,1049: 131-138.
Dute R R, Peterson C M. Early endosperm development in ovules of soybean, Glycine max(L.) Merr.AnnBot,1992,69:263-271.
Fineran B A, Wild D J, Ingerfeld M. Initial wall formation in the endosperm of wheat. Triticum aestivum:a reevaluation.CAN J BOT,1982,60:1776-1795.
Folsom M W, Cass D D. Embryo sac development in soybean:the central cell and aspects of fertilization. Amer J Bot,1992,79(12):1407-1017.
Fujita N, Yoshida M, Kondo T. et al. Characterization of SSIIla-Deficient Mutants of Rice: The Function of SSIlIa and Pleiotropic Effects by SSIIIa Deficiency in the Rice Endosperm. Plant Physiology.2007,144:2009-2023.
Gerhard P. Schwall, Richard Safford, Roger J. Westcott, et al. Production of very-high-amylose potato starch by inhibition of SBE A and B. Nature Biotechnology.2000,18: 551-554.
G. E. Vandeputte, J. A. Delcour. From sucrose to starch granule to starch physical behaviour:a focus on rice starch. Carbohydrate Polymers.2004,58:245-266.
Goldberg R B, Beals T P, Sanders P M. Anther development:basic principles and practical applications, Plant Cell,1993,5:1217-1228.
Gray J, Johal G S. Programmed cell death in plants. In:Anderson M, Roberts J A eds. Arabidopsis. Annual reviews, Boca Raton:Sheffield Academic Press,1998,360-394.
Greenberg J T. Programmed cell death:a way of lifef or plants. Proc Natl Acad Sci USA,1996, 93:12094-12097.
Groover A, Jones A M. Tracheary element differentiation uses a novel mechanism coordinating programmed cell death and secondary cell wall synthesis. Plant Physiol, 1999,119:375-384.
Hepler P K, Palevitz B A, Lancelle S A, et al. Cortical endoplasmic reticulum in plants J Cell Sci,1990,96:355-373.
Hiffins T J V. synthesis and regulation of major protein in seeds. Ann Rev plant Physiol,1984, 35:191-221.
Hughes C E. The developing endosperm of Triticum aestivum (L.):An ultrastructural and morphometric study. PhD Dissertation of University of Nottingham,1976.
Jensen W A., Shlz P. and Aston M.E. An ultrastruetoral study of early endosperm development and synergid chandges inunfertilized cotton ovuies. Planta,1977,133:179-189.
Jensen C G. Dynamic of spindle microtubule organization:kinetoehore fiber mierotubules of plant endosperm. J Cell Bio,1982,92:540-557.
Johnson K L, Degnan K A. Ross Walker J. Ingram GC AtDEKl is essential for specification of embryonic epidermal cell fate. Planta,1971,101:189-209.
Juliano, B.O. Rice in human nutrition/prepared in collaboration with FAO. Rome Published with the Cooperation of the International Rice Research Institute, Food and Agriculture Organization of the United Nations,1993.
Kim W T, Franceschi V R, Krishnan H B, et al. Formation of wheat protein bodies: involvement of the Golgi apparatus in gliadin transport. Plants,1988,173:173-182.
Kluth A, Sprunck S, Becker D. et al.5'deletion of a gbssl promoter region from wheat leads to changes in tissue and developmental specificities. Plant Molecular Biology.2002,49: 669-682.
Krishnan H B, White J A, Pueppke S G. Immunocytochemical localization of wheat prolamins in the lumen of the rough endoplasmic reticulum. Cananian Journal of Botany,1991:39: 2574-2577.
Kumamaru T, Ogawa M, Satoh H, et al. Protein body biogenesis in cereal endosperms//In: Olsen O A. Plant cell Monogr(8):Endosperm. Heidelberg:Springer-Verlag Berlin,2007: 141-158.
Lammeren V, M A A. Structure and function of the microtubular cytoskeleton during endosperm development in wheat:an immunofluorescence study. Protoplasm,1988, 14(5):613-622.
Langeveld S M J, van Wijk R, Stuurman N,et al. B-type granule containing protrusions and interconnection between amyloplasts in developing wheat endosperm revealed by transmission electron microscopy and GFP expression. Journal of Experimental Botany, 2000,51:1357-1361.
Levanony H, Rubin R, Altsehuler, Y. Evidence for a novel roate of wheat storage protein to vacuoles. J Cell Biology,1992,119:117-128.
Loussert C, Popineau Y, Mangavel C. Protein bodies ontogeny and localization of prolamin components in the developing endosperm of wheat caryopses. J Cereal Sci,2008,47: 445-456.
Lyon H O, Schulte E K, Prento P, et al. Standardized staining methods:feulgen-rossenbeck reaction for desoxyribonucleic acid and periodic acid-Schiff (PAS) procedure. Biotech Histochem,2002,77 (3):121-125.
Mares D.J, Seone B.A., Jeffery C. Early stages in the development of wheat endosperm Ⅱ. Ultrastructural observationon cell wall formation. Aust.J.Bot.1977,25:599-613.
M.S. Buttrose. Ultrastructure of the developing wheat endosperm. Aust J Biol Sci,1963.16: 305-317.
Niedeu U Z, Weber E, Neurnann D. Dictyosomes partieipate in the intracellular pathway of storage protein in developing vicia faba cotyledons.European J Cell Biol,1984,34:9-17.
Ogawa M,Tanaka K.Kasai Z. Energy-dispersive X-ray analysis of phytin globoids in aleurone particle of developing rice grains. Soil Sci. Plant Nutr,1979,25:437-448.
Okamoto K, Murai T, Eguchi G, et al. Enzymic echanism of starch breakdown in germinating rice seeds. Plant Physiol,1982,70:905-911.
Olsen O A, Brow R C, Lemmon B E. Pattern and process of wall formation in developing endosperm. Bioassay,1995,17:448-456.
Olsen O A, Linnestad C, Niehols.S E. Developmental biology of the cereal endosperm. Trende in plant seienee,1999,4(7):253-257.
Olsen O A. Endosperm development:cellularization and cell fate specification. Annu Rev Plant Physiol Plant Mol Bio,2001,52:233-267.
Oparka K. J, Harris N. Rice protein-body formation:all types are initiated by dilation of the endoplasmic retieulum. Planta,1982,154:184-188.
Otegui M, Staehelin L A. Syneytial-type cell plates:a novel kind of cell plate involved in endosperm cllularization of Arabidopsis. The Plant Cell.2000.12:933-947.
Otegui M S.. Mastronarde D N, Kang B-H. Bednarek S Y, and Staehelin L. A. Three-dimensional analysis of syncytial-type cell plates during endosperm cellularization visualized by high resolution electron tomography. The Plant Cell,2001,3:2033-2051.
Parker M L, Hawes C R. The Golgi apparatus in developing endosperm of wheat (Triticum aestivum. L.). Planta,1982,154:277-283.
Parker M L. The relationship between A-type and B-type starch granules in the developing endosperm of wheat. Journal Cereal Science,1985,3:271-278.
Ramaehandran C, Raghavan V. Changes in nuelear DNA content of endosperm cells during grain development in rice (Oryza sativa). Ann Bot,1989,64(4):459-468.
Richard G M, Turner P F, Napier J A. Transport and deposition of cereal prolamins. Plant Physiology and Biochemistry,1996,34:237-243.
Rost T L. The ultrastructure and physiology of proteinbodies and lipids from hydrated dormant and nondormant embryos of Setaria Lutescens (Gramineae). Amer J Bot,1972,59: 607-616.
Rubin R, Levanony H, Galili G. Evidence for the presence of two different types of protein bodies in wheat endosperm. Plant Physiology,1992,99:718-724.
Sanderfoot A A, Raikhel N V. The specificity of vesicle trafficking:coat proteins and snares. Plant Cell,1999,11:629-641.
Sato T.and Shamoto M. Stain Technology,1973,48(5):223-227.
Shewry P R, Napier J A, Tatham A S. Grain storage proteins:structures and biosynthesis. Plant Cell,1995,7:945-956.
Shewry P R, Halford N G. Cereal grain storage proteins:structures, properties and role in grain utilization. Journal of Experimental Botany,2002,53:947-958.
Sodhi N S, Singh N. Morphological, thermal and rheological properties of starches separated from rice cultivars grown in India. Food Chem,2003,80:99-108.
S(?)rensen M B, Mayer U, Lukowitz W. Cellularisation in the endosperm of Arabidopsis tha lianais coupled to mitosis and shares multiple components with eytokinesis. Development, 2002,129:5567-5576.
Stewart A. Nield H, Lott J N A. An investigation of the mineral content of barley grains and seedlings. Plant Physiol,1998,86:93-97.
Taiz L, Zeiger E. Plant Physiology(3rd edition). Sunderland:Sinauer Associates, Inc., Publishers.2002:321.
Tanaka K, Ogawa M, Kasai Z. The rice scutellum. Ⅱ. A comparison of scutellar and aleurone electron-dense particles by transmission electron microscopy including energy-dispersive X-ray analysis. Cereal. Chem,1977,54:684-689.
Tashiro T, Ebata M, Ishikawa M. Studies on white belly rice kernel. Japan J Crop Sci,1980, 49(3):482-488.
Thomson W W, Whatley J M. Development of nongreen plastids. Ann Reu Plant physiol,1980, 31:375-394.
Tian G-W, You R-L, Guo F-L, Wang X-C. Mierotubular cytoskeleton X-C. Microtubular cytoskeleton in wheat.Cytol. Tokyo,1998,63:427.
Webb M C, Gunning B E S. The microtubular cytoskeleton during development of the zygote, proembryo and free-nuclear endosperm in Arabidopsis thaliana(L.) Heynh. Palanta,1991, 184(2):187-195.
Wang H, Li J, Bostock R M, Gilechrist D G. Apoptosis:A functional paradigm for programmed plant cell death induced by a host-selective phytotoxin and invoked during development. Plant Cell 1996a,8:375-391.
Wang M, Oppeijk B J, Lu X, Van V D, Schilperoort R A. Apoptosis in barley aleurone during germination and its inhibition by abscisic acid. Plant Mol Biol,1996b,32:1125-1134.
Whatley J M. Chloroplast development in Primary leaves of Phaseolus vulgaris. New Phyiol, 1974,73:1097-1110.
Whatley J M. Variation in the basic pathway of chloroplast development. New Phytol,1977, 78; 407-420.
Whatley J M. A suggested cycle of plastid developmental interrelationships. New Phytol,1978, 80:489-502.
Wei C X, Lan S Y, Xu Z X. Ultrastructural features of nucleus degradation during programmed cell death of starchy endosperm cells in rice. Acta Botanica Sinica,2002, 44(12):1396-1402.
Yamagata H, Tanaka K. The site of synthesis and accumulation of rice storage proteins. Plant Cell Physiol,1986,27:135-145.
Young T E, Gallie D R. Analysis of Programmed cell death in wheat endosperm reveals differences in endosperm development between cereals. Plant Molecular Biology.1999, 39:915-926.
Young T E, Gallie D R. Programmed cell death during endosperm development. Plant Mol Biol,2000,44:283-301.
陈建敏孙德兰.淀粉体遗传研究的现状与展望,植物遗传资源学报,2008,92：258-262.
陈娟,王忠,莫亿伟等.水稻颖果的充实与呼吸活性的关系.扬州大学学报(农业与生命科学版),2005,26(2)：61-65.
陈娟,王忠等.不同施氮处理对水稻颖果灌浆和呼吸活性的影响.中国水稻科学.2006,20(4)：396-400.
程方民,胡东维,丁元树.人工控温条件下稻米垩白形成变化及胚乳扫描结构观察.中国水稻科学,2000,14(2)：83-87.
崔克明.植物细胞程序死亡的机理极其与发育的关系.植物学通报,2000,17(2)：97-107.
付洪兰.实用电子显微镜技术.高等教育出版社,2004：12-13.
符文英,向远鸿等.食用优质稻水胚乳显微结构研究.湖南农业大学报,1997,23(5)： 411-418.
高松洁,郭天财,吴雪峰,阎凌云.小麦淀粉合成关键酶与淀粉主要理化特性研究进展.河南农业大学学报.2002,36(4)：313-318.
顾蕴洁,熊飞,王忠等.水稻和小麦胚乳发育的比较.南京师范大学学报：自然科学版,2001,24(3)：65-74.
顾蕴洁,王忠,陈义芳,陈刚,黄海.高粱胚乳发育的观察与研究.南京师范大学学报(自然科学版),2002,25(3)：99-108.
顾蕴洁,王忠,陈娟,赵国勇.水稻果皮的结构与功能.作物学报,2002,28(4)：439-444.
郭文善.小麦籽粒胚乳细胞发育规律及其调控制.扬州：扬州大学博士论文,1997.
胡适宜.被子植物胚胎学.北京：人民教育出版社,1982,10-164.
胡适宜,徐丽云.显示环氧树脂厚切片中多糖、蛋白质和脂类的细胞化学方法.植物学报,1990,32(11)：841-846.
华振基,王俊.春小麦籽粒发育的形态学及解剖学的研究Ⅲ：胚乳初期发育的发生及解剖学观察.宁夏农学院报,1982,1：1-9.
黄百渠.拟南芥菜子叶中蛋白体形成的超微结构和免疫电镜定位研究.实验生物学报,1989,22：247-257.
黄景华,王广金,孙岩,刁艳玲.小麦花粉管通道形成时期的研究.黑龙江农业科学,2004,2：20-22.
季清娥,郑恒清,徐珍秀等.品质不同的稻米胚乳淀粉粒的微观结构观察.福建农业大学学报,1998,27(2)：241-244.
康海岐,常红叶,许育彬等.水稻籽粒胚乳的石蜡切片方法改良及其结构发育观察.西北植物学报,2008,28(5)：1069-1074.
蓝盛银,徐珍秀.水稻胚乳淀粉粒形态发育的亚显微结构研究.武汉大学学报(自然科学版),1997,2：73-74.
蓝盛银,钟方旭,杨泽敏,靳德明,徐珍秀.水稻淀粉胚乳程序性细胞死亡中的去核化.实验生物学报,2004,37(1)：34-44.
李师翁,屠骊珠.白刺胚和胚乳发育的研究.植物学报.1991,33(7)：500-506.
李师翁.被子植物核型胚乳细胞化机理研究的现状.植物学通报,1995,12(4)：13-16.
刘冬娟,祝素文,李艳杰.硼砂甲苯胺蓝染液在半薄切片染色上的应用.中国医科大学学报,2004,33(1)：33.
刘向东,徐是雄,卢永根.水稻胚囊壁的形成与发育观察.植物学报,1997,39(11)：985-990.
刘智,王玲玲,周卫东,陈义芳,王忠.固定和染色方法对水稻胚乳细胞结构观察的影响.中国水稻科学,2011,25(6)：667-671.
罗玉英.玉竹花粉中造粉质体形成及增殖的超微结构研究.电子显微学报,1995,14(3)：166-169.
李睿,蓝盛银,徐珍秀.水稻胚乳发育中线粒体向淀粉质体转变及琥珀酸脱氢酶的定位. 电子显微学报,2002,21(2)：118-121
毛同林,吴晓东,杨世杰.洋葱根冠外缘细胞编程性死亡过程中的超微结构研究.电子显微学报,2001,20：21-26.
宋今丹.高锰酸钾固定的培养细胞的膜性结构.电子显微学报,1990,9(3)：1.
松田智明,川原治之助,长南信雄.日本作物学会纪事,1979,48：155-162.
松田智明.登熟期の水稻子房にぉける淀粉贮藏细胞の微细构造と转流、蓄积机构につぃて.日本作物学会纪事,1984,53(别号1)：144-147.
沈波.早籼稻垩白形成中胚乳淀粉粒发育的电镜观察.中国水稻科学,2000,14(4)：225-228.
盛婧,郭文善,朱新开,封超年,彭永欣.不同专用类型小麦籽粒淀粉粒形成过程.作物学报,2004,30(9)：953-954.
汤圣祥,江云珠,李双盛等.早籼稻胚乳淀粉体的扫描电镜观察.作物学报,1999,25(2)：269-273.
田代亨,河野恭广.水稻胚乳组织の发育生理学研究第1报胚乳组织の细胞学的不均一性仁”にっぃて.日本作物学会纪事,1972,41：521-530.
王爱国,罗广华,邵从文等.大豆种子超氧化物歧化酶的研究.植物生理学报,1983,9：77-84.
王长发,张篙午.冷型小麦旗叶衰老和活性氧代谢特性研究.西北植物学报,2000,20(5)：727-732.
王兰,刘向东等.同源四倍体水稻胚乳发育：极核融合和胚乳细胞化.中国水稻科学,2004,18(4)：281-289.
王兰,刘向东等 同源四倍体水稻胚乳发育：糊粉层细胞壁纤维素物质发育、胚乳淀粉积累胼胝质“套”.中国水稻科学,2004,18(6)：507-514.
王萌,周舟,兰盛银.不同大麦淀粉胚乳细胞中淀粉和贮藏蛋白的积累.中国农业科学,2004,37(10)：1464-1467
王萍,任力.树脂包埋半薄切片染色的改进及体会.大连医科大学学报,2007,29(5)：509-510.
王全喜.植物学.北京：科学出版社,2004：21.
王爽,王春梅,陈丹,黄晓峰.环氧树脂半薄切片的多色性染色方法.第四军医大学学报,2002,23(9)：775.
王晓曦,杨献辉,程凤明等.小麦胚乳结构中蛋白质分布及其对面团流变学特性影响的研究.郑州工程学院学报,2001,22(1)：63-66.
王余龙,蔡建中.水稻籽粒受容活性及其控制途径——Ⅱ.籽粒含水率与～(14)C光合产物分配及转化的关系.江苏农学院学报,1990,11(4)：27-31.
王忠,李卫芳,顾蕴洁,陈刚,石火英,高煌珠.水稻胚乳发育及其养分输入的途径.作物学报,1995,21：520-527.
王忠,陈刚,李克武,桂玉清,顾蕴洁.稻麦胚乳游离核的分裂.江苏农学院学报,1996, 17(1)：11-20.
王忠,顾蕴洁,李卫芳,练兴明,陈刚.关于大麦胚乳发育的研究.中国农业科学,1996,29：38-45.
王忠,顾蕴洁,李卫芳,陈刚,石火英,陈秀花.小麦胚乳发育及其养分输入的途径.作物学报,1998,24：536-543.
王忠,顾蕴洁,李卫芳,黄山,李克武.水稻糊粉层的形成及其在萌发过程中的变化.扬州大学学报,1998,1(1)：32-34.
王银改,王清改,翟素平.Image J软件辅助分析在网织红细胞计数中的应用.临床检验杂志,2005,23(3)：210.
翁寿松.日立的场发射扫描电镜.电子工业专用设备,1993,22(1)：30-33.
星川清亲.小麦胚乳形成初期的发生学研究,上海市科学技术编译馆出版,1960.
星川清亲.米の胚乳发育に关する组织形态的研究,第4报,糊粉层の分化と发育にっぃて.日作纪,1967,36：216-220.
星川清亲.日本作物学会纪事,1967,36(1)：151-161.
星川清亲.日本作物学会纪事,1967,36：203-227.
星川清亲.日本作物学会纪事,1967,36：398-407.
星川清亲.日本作物学会纪事,1968,37：87-106.
星川清亲.日本作物学会纪事,1968,37：207-216.
星川清亲.日本作物学会纪事,1984,53：153-162.
星川清亲著,蒋彭炎,许德海译.解剖图说稻的生长.上海：上海科学技术出版社,1980,273.
席湘媛.大葱胚和胚乳的发育.植物学报.1991,29(5)：459-464.
熊飞,王忠,陈刚等.不同籼稻颖果发育及其结构与品质性状.中国水稻科学,2006,20(1)：43-46.
徐是雄,徐雪宾.稻的形态与解剖.北京：农业出版社,1984：45-57.
徐是雄,刘向东,冯九焕,卢永根.水稻IR36及光温敏核雄性不育系培矮64S小孢子母细胞减数分裂期间微管骨架的变化.植物学报,2001,43(3)：221-226.
徐是雄,刘向东,朱洪亮,卢永根.水稻雌配子体发生过程中胚囊内微管骨架变化的进步研究.植物学报,2001,43(9)：910-917.
韦存虚.水稻胚乳发育中细胞核与细胞器的结构消长与功能研究.武汉：华中农业大学,2002.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞发育中的蛋白体的形成.作物学报,2002,28(5)：591-594.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞淀粉质体被膜与其增殖的关系.电子显微学报,2002,21(2)：124-128.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞发育中的蛋白体的形成.作物学报,2002,28(5)：591-594.
韦存虚,蓝盛银,徐珍秀.水稻胚乳发育中ATP酶的超微细胞化学定位和功能分析.中国农业科学,2003,36(3)：259-262.
韦存虚,张军,周卫东等.小麦胚乳发育过程中贮藏蛋白的积累和蛋白体的形成.麦类作物学报,2009,29(1)：73-78.
韦存虚,张军,周卫东,陈义芳,许如根.大麦胚乳发育过程中贮藏蛋白的积累和蛋白体的形成.作物学报,2009,35(2)：334-340.
杨弘远.用整体染色与透明技术观察胚囊、胚、胚乳和胚状体.植物学报,1986,28(6)：575-581.
杨貌仙.小麦胚乳发育的研究.植物学报,1957,6(1)：39-52.
杨貌仙.小麦胚乳游离核繁殖的研究.植物学报,1962,11(1)：7-9.
杨学举,屈平,张彩英等.小麦A型淀粉粒大小对淀粉特性及面包品质的影响.河北农业大学学报,2004,27(5)：1-5.
叶庆华.植物生物学.厦门：厦门大学出版社,2002：32.
银永安,齐军仓,李卫华,曹连莆,王自布.小麦胚乳A、B型淀粉粒理化特性研究.中国农业科学,2010,43(11)：2372-2379.
尤瑞麟.小麦珠心细胞衰退过程的超微结构研究.植物学报,1985,27：345-353.
张松林,金芝兰.谷子胚和胚乳的发育.西北植物学报,1994,14(4)：249-254,
张伟成,严文海,高小彦.小麦胚乳细胞化初期胞壁形成特征的观察.植物学报,1990,32(1)：76-78.
郑晓鹰,顾增辉,徐本美,郑光华.几种蔬菜种子的活力试验.中国农业科学,1982,4：58-64.
中华医学会.临床技术操作规范(病理学分册).北京：人民军医出版社,2004.
钟瑛.改良半薄切片多色性染色方法.临床与实验病理学杂志,1992,8(3)：228.
朱徵.高碘酸—锡夫试剂反应作为一种染色方法在植物组织学上的应用.植物学报,1963,11(2)：155-166.
朱美君,王学臣.植物的编程性细胞死亡.植物生理学通讯,1999,5：353-357.
Badenhuize N P. Encyclopaedia of plant physiology, Vol VI(Ruhland ed.). Berlin:Springer Verlag,1958:137-153.
Barlow K K, Lee J W, Vesk M. Morphological development of storage protein bodies in wheat. Mechanisms of Regulations of Plant Growth, Wellington:Royal Society of New Zealand, 1974:793.
Bechtel D B, Gaines R L, Pomeranz Y. Protein secretion in wheat endosperm:formation of the matrix protein. Cereal Chemistry,1982,59(5):336-343.
Bechtel D B, Gains R L, Pomeranz Y. Early stages in wheat endosperm formation and protein body initiation. Annu Bot,1982,50:507-518.
Bechtel D B, Gaines R L, Pomeranz Y. Early stages in wheat endosperm formation and protein body initiation. Ann Bot,1982,73:949-955.
Bechtel D B, Zayas I Y, Kaleikau L, et al. Size distribution of wheat starch granules during endosperm development. Cereal Chemistry,1990,67:59-63.
Bechtel D B, Wilson J D, Shewry R P. Immunocytochemical localization of the wheat storage protein triticin in developing endosperm tissue. Cereal Chemistry,199l.68(6):573-577.
Becraft P W, Asuncion-Crabb Y. Positional cues specify and maintain aleurone cell fate in maize endosperm development. Development,2000,127(18):4039-4048.
Beehtel D R, Juliano B O. Formation of protein bodies in the starchy endosperm of rice:a rein vestigation. Ann. Bot.1980,45:503-509.
Bell P R. Megaspore abortion:a consequence of selective apoptosis. Int J Plant Sci,1996,157: 1-7.
Briarty L G, Hughes C E, Evers A D. The developing endosperm of wheat-a stereological analysis. Ann Bot,1979,44:641-658.
Brown R C, Lemon B E, Olsen O A. Endosperm development in barley:microtubule involvement in the morphogenetic pathway. Plant cell,1994,6:1241-1252.
Brown R C, Lemmon B E, Olsen O A. Polarization prediets the pattern of cellularization in cereal endosperm. Protoplasma,1996,192:168-177.
Brown R C, Lemmon B E, Stone B A, Olsen O A. Cell wall (1→3)-and(1-3,1-4)-β-glueans during early grian development in rice(Oryza sativa L.). Planta,1997,202:414-426.
Brown R C., Lernrnon B E., Nguyen H, Olsen O-A. Development of endosperm in Arabidopsis thaliana. Sex Plant Reprod.1999,12:32-42.
Buchanan B B, Gruissem W, Jones R L. Biochemistry&molecular biology of plants.北京：科 学出版社,2002,18-19.
Buttrose M S. Submicroscopic development and structure of starch granules in cereal endosperm. J Ultrastructure Research,1960,4:231-257.
Buttrose M S. Ultrastructure of the developing wheat endosperm. Austrlian Journal of Biology Science,1963,16:305-317.
Chamberlin M A, Homer H T. Nutrition of early embryo development in Glycine max (L.)Merr. Alller J Bot,1990,77:20.
Chamberlin M A, Horner H T. An ultrastruetural study of endosperm wall formation and degeneration in Glycjne max(L.) Merr.AlnerJBot,1990,77:83.
Chamberlin M A, Horner H T, Palmer R G. Early endosperm, embryo, and ovule development in Glycine max(L.) Merr. Int J Plant Sci,1994,155(4):421-436.
Chamberlin M A, Homer H T. Palmer R G. Nuelear size and DNA content of the embryo and endosperm during the irinitial stages of development in Glycine max (Fabaceae). Amer J Bot,1993,80,(10):1209-1215.
Chitra lekha P. Bhandari N N. Cellularization of endosperm in AsPhodelus trenuifolius Cav.. Phytomorphology,1992,42(3-4):185-193.
Chung M C. Development of rice caryopsis. National Science Council Monthly,1983,11(10): 959-976.
Dang J M, Copeland L. Studies of the fracture surface of rice grains using environmental scanning electron microscopy. J Sci Food Agric,2004,84:707-713.
Dellaporta S L, Calderon-Urrea A. The sex determination proeess in maize. Science,1994,266: 1501-1505.
Dominique Cledat, Serge. Battu, Redouane. Mokrini, Philippe. J. P. Cardot. Rice starch granule charact-erization by flow cytometry scattering techniques hyphenated with sedimentation field-flow fractionation. Journal of chromatography A.2004,1049: 131-138.
Dute R R, Peterson C M. Early endosperm development in ovules of soybean, Glycine max(L.) Merr.AnnBot,1992,69:263-271.
Fineran B A, Wild D J, Ingerfeld M. Initial wall formation in the endosperm of wheat. Triticum aestivum:a reevaluation.CAN J BOT,1982,60:1776-1795.
Folsom M W, Cass D D. Embryo sac development in soybean:the central cell and aspects of fertilization. Amer J Bot,1992,79(12):1407-1017.
Fujita N, Yoshida M, Kondo T. et al. Characterization of SSIIla-Deficient Mutants of Rice: The Function of SSIlIa and Pleiotropic Effects by SSIIIa Deficiency in the Rice Endosperm. Plant Physiology.2007,144:2009-2023.
Gerhard P. Schwall, Richard Safford, Roger J. Westcott, et al. Production of very-high-amylose potato starch by inhibition of SBE A and B. Nature Biotechnology.2000,18: 551-554.
G. E. Vandeputte, J. A. Delcour. From sucrose to starch granule to starch physical behaviour:a focus on rice starch. Carbohydrate Polymers.2004,58:245-266.
Goldberg R B, Beals T P, Sanders P M. Anther development:basic principles and practical applications, Plant Cell,1993,5:1217-1228.
Gray J, Johal G S. Programmed cell death in plants. In:Anderson M, Roberts J A eds. Arabidopsis. Annual reviews, Boca Raton:Sheffield Academic Press,1998,360-394.
Greenberg J T. Programmed cell death:a way of lifef or plants. Proc Natl Acad Sci USA,1996, 93:12094-12097.
Groover A, Jones A M. Tracheary element differentiation uses a novel mechanism coordinating programmed cell death and secondary cell wall synthesis. Plant Physiol, 1999,119:375-384.
Hepler P K, Palevitz B A, Lancelle S A, et al. Cortical endoplasmic reticulum in plants J Cell Sci,1990,96:355-373.
Hiffins T J V. synthesis and regulation of major protein in seeds. Ann Rev plant Physiol,1984, 35:191-221.
Hughes C E. The developing endosperm of Triticum aestivum (L.):An ultrastructural and morphometric study. PhD Dissertation of University of Nottingham,1976.
Jensen W A., Shlz P. and Aston M.E. An ultrastruetoral study of early endosperm development and synergid chandges inunfertilized cotton ovuies. Planta,1977,133:179-189.
Jensen C G. Dynamic of spindle microtubule organization:kinetoehore fiber mierotubules of plant endosperm. J Cell Bio,1982,92:540-557.
Johnson K L, Degnan K A. Ross Walker J. Ingram GC AtDEKl is essential for specification of embryonic epidermal cell fate. Planta,1971,101:189-209.
Juliano, B.O. Rice in human nutrition/prepared in collaboration with FAO. Rome Published with the Cooperation of the International Rice Research Institute, Food and Agriculture Organization of the United Nations,1993.
Kim W T, Franceschi V R, Krishnan H B, et al. Formation of wheat protein bodies: involvement of the Golgi apparatus in gliadin transport. Plants,1988,173:173-182.
Kluth A, Sprunck S, Becker D. et al.5'deletion of a gbssl promoter region from wheat leads to changes in tissue and developmental specificities. Plant Molecular Biology.2002,49: 669-682.
Krishnan H B, White J A, Pueppke S G. Immunocytochemical localization of wheat prolamins in the lumen of the rough endoplasmic reticulum. Cananian Journal of Botany,1991:39: 2574-2577.
Kumamaru T, Ogawa M, Satoh H, et al. Protein body biogenesis in cereal endosperms//In: Olsen O A. Plant cell Monogr(8):Endosperm. Heidelberg:Springer-Verlag Berlin,2007: 141-158.
Lammeren V, M A A. Structure and function of the microtubular cytoskeleton during endosperm development in wheat:an immunofluorescence study. Protoplasm,1988, 14(5):613-622.
Langeveld S M J, van Wijk R, Stuurman N,et al. B-type granule containing protrusions and interconnection between amyloplasts in developing wheat endosperm revealed by transmission electron microscopy and GFP expression. Journal of Experimental Botany, 2000,51:1357-1361.
Levanony H, Rubin R, Altsehuler, Y. Evidence for a novel roate of wheat storage protein to vacuoles. J Cell Biology,1992,119:117-128.
Loussert C, Popineau Y, Mangavel C. Protein bodies ontogeny and localization of prolamin components in the developing endosperm of wheat caryopses. J Cereal Sci,2008,47: 445-456.
Lyon H O, Schulte E K, Prento P, et al. Standardized staining methods:feulgen-rossenbeck reaction for desoxyribonucleic acid and periodic acid-Schiff (PAS) procedure. Biotech Histochem,2002,77 (3):121-125.
Mares D.J, Seone B.A., Jeffery C. Early stages in the development of wheat endosperm Ⅱ. Ultrastructural observationon cell wall formation. Aust.J.Bot.1977,25:599-613.
M.S. Buttrose. Ultrastructure of the developing wheat endosperm. Aust J Biol Sci,1963.16: 305-317.
Niedeu U Z, Weber E, Neurnann D. Dictyosomes partieipate in the intracellular pathway of storage protein in developing vicia faba cotyledons.European J Cell Biol,1984,34:9-17.
Ogawa M,Tanaka K.Kasai Z. Energy-dispersive X-ray analysis of phytin globoids in aleurone particle of developing rice grains. Soil Sci. Plant Nutr,1979,25:437-448.
Okamoto K, Murai T, Eguchi G, et al. Enzymic echanism of starch breakdown in germinating rice seeds. Plant Physiol,1982,70:905-911.
Olsen O A, Brow R C, Lemmon B E. Pattern and process of wall formation in developing endosperm. Bioassay,1995,17:448-456.
Olsen O A, Linnestad C, Niehols.S E. Developmental biology of the cereal endosperm. Trende in plant seienee,1999,4(7):253-257.
Olsen O A. Endosperm development:cellularization and cell fate specification. Annu Rev Plant Physiol Plant Mol Bio,2001,52:233-267.
Oparka K. J, Harris N. Rice protein-body formation:all types are initiated by dilation of the endoplasmic retieulum. Planta,1982,154:184-188.
Otegui M, Staehelin L A. Syneytial-type cell plates:a novel kind of cell plate involved in endosperm cllularization of Arabidopsis. The Plant Cell.2000.12:933-947.
Otegui M S.. Mastronarde D N, Kang B-H. Bednarek S Y, and Staehelin L. A. Three-dimensional analysis of syncytial-type cell plates during endosperm cellularization visualized by high resolution electron tomography. The Plant Cell,2001,3:2033-2051.
Parker M L, Hawes C R. The Golgi apparatus in developing endosperm of wheat (Triticum aestivum. L.). Planta,1982,154:277-283.
Parker M L. The relationship between A-type and B-type starch granules in the developing endosperm of wheat. Journal Cereal Science,1985,3:271-278.
Ramaehandran C, Raghavan V. Changes in nuelear DNA content of endosperm cells during grain development in rice (Oryza sativa). Ann Bot,1989,64(4):459-468.
Richard G M, Turner P F, Napier J A. Transport and deposition of cereal prolamins. Plant Physiology and Biochemistry,1996,34:237-243.
Rost T L. The ultrastructure and physiology of proteinbodies and lipids from hydrated dormant and nondormant embryos of Setaria Lutescens (Gramineae). Amer J Bot,1972,59: 607-616.
Rubin R, Levanony H, Galili G. Evidence for the presence of two different types of protein bodies in wheat endosperm. Plant Physiology,1992,99:718-724.
Sanderfoot A A, Raikhel N V. The specificity of vesicle trafficking:coat proteins and snares. Plant Cell,1999,11:629-641.
Sato T.and Shamoto M. Stain Technology,1973,48(5):223-227.
Shewry P R, Napier J A, Tatham A S. Grain storage proteins:structures and biosynthesis. Plant Cell,1995,7:945-956.
Shewry P R, Halford N G. Cereal grain storage proteins:structures, properties and role in grain utilization. Journal of Experimental Botany,2002,53:947-958.
Sodhi N S, Singh N. Morphological, thermal and rheological properties of starches separated from rice cultivars grown in India. Food Chem,2003,80:99-108.
S(?)rensen M B, Mayer U, Lukowitz W. Cellularisation in the endosperm of Arabidopsis tha lianais coupled to mitosis and shares multiple components with eytokinesis. Development, 2002,129:5567-5576.
Stewart A. Nield H, Lott J N A. An investigation of the mineral content of barley grains and seedlings. Plant Physiol,1998,86:93-97.
Taiz L, Zeiger E. Plant Physiology(3rd edition). Sunderland:Sinauer Associates, Inc., Publishers.2002:321.
Tanaka K, Ogawa M, Kasai Z. The rice scutellum. Ⅱ. A comparison of scutellar and aleurone electron-dense particles by transmission electron microscopy including energy-dispersive X-ray analysis. Cereal. Chem,1977,54:684-689.
Tashiro T, Ebata M, Ishikawa M. Studies on white belly rice kernel. Japan J Crop Sci,1980, 49(3):482-488.
Thomson W W, Whatley J M. Development of nongreen plastids. Ann Reu Plant physiol,1980, 31:375-394.
Tian G-W, You R-L, Guo F-L, Wang X-C. Mierotubular cytoskeleton X-C. Microtubular cytoskeleton in wheat.Cytol. Tokyo,1998,63:427.
Webb M C, Gunning B E S. The microtubular cytoskeleton during development of the zygote, proembryo and free-nuclear endosperm in Arabidopsis thaliana(L.) Heynh. Palanta,1991, 184(2):187-195.
Wang H, Li J, Bostock R M, Gilechrist D G. Apoptosis:A functional paradigm for programmed plant cell death induced by a host-selective phytotoxin and invoked during development. Plant Cell 1996a,8:375-391.
Wang M, Oppeijk B J, Lu X, Van V D, Schilperoort R A. Apoptosis in barley aleurone during germination and its inhibition by abscisic acid. Plant Mol Biol,1996b,32:1125-1134.
Whatley J M. Chloroplast development in Primary leaves of Phaseolus vulgaris. New Phyiol, 1974,73:1097-1110.
Whatley J M. Variation in the basic pathway of chloroplast development. New Phytol,1977, 78; 407-420.
Whatley J M. A suggested cycle of plastid developmental interrelationships. New Phytol,1978, 80:489-502.
Wei C X, Lan S Y, Xu Z X. Ultrastructural features of nucleus degradation during programmed cell death of starchy endosperm cells in rice. Acta Botanica Sinica,2002, 44(12):1396-1402.
Yamagata H, Tanaka K. The site of synthesis and accumulation of rice storage proteins. Plant Cell Physiol,1986,27:135-145.
Young T E, Gallie D R. Analysis of Programmed cell death in wheat endosperm reveals differences in endosperm development between cereals. Plant Molecular Biology.1999, 39:915-926.
Young T E, Gallie D R. Programmed cell death during endosperm development. Plant Mol Biol,2000,44:283-301.
陈建敏孙德兰.淀粉体遗传研究的现状与展望,植物遗传资源学报,2008,92：258-262.
陈娟,王忠,莫亿伟等.水稻颖果的充实与呼吸活性的关系.扬州大学学报(农业与生命科学版),2005,26(2)：61-65.
陈娟,王忠等.不同施氮处理对水稻颖果灌浆和呼吸活性的影响.中国水稻科学.2006,20(4)：396-400.
程方民,胡东维,丁元树.人工控温条件下稻米垩白形成变化及胚乳扫描结构观察.中国水稻科学,2000,14(2)：83-87.
崔克明.植物细胞程序死亡的机理极其与发育的关系.植物学通报,2000,17(2)：97-107.
付洪兰.实用电子显微镜技术.高等教育出版社,2004：12-13.
符文英,向远鸿等.食用优质稻水胚乳显微结构研究.湖南农业大学报,1997,23(5)： 411-418.
高松洁,郭天财,吴雪峰,阎凌云.小麦淀粉合成关键酶与淀粉主要理化特性研究进展.河南农业大学学报.2002,36(4)：313-318.
顾蕴洁,熊飞,王忠等.水稻和小麦胚乳发育的比较.南京师范大学学报：自然科学版,2001,24(3)：65-74.
顾蕴洁,王忠,陈义芳,陈刚,黄海.高粱胚乳发育的观察与研究.南京师范大学学报(自然科学版),2002,25(3)：99-108.
顾蕴洁,王忠,陈娟,赵国勇.水稻果皮的结构与功能.作物学报,2002,28(4)：439-444.
郭文善.小麦籽粒胚乳细胞发育规律及其调控制.扬州：扬州大学博士论文,1997.
胡适宜.被子植物胚胎学.北京：人民教育出版社,1982,10-164.
胡适宜,徐丽云.显示环氧树脂厚切片中多糖、蛋白质和脂类的细胞化学方法.植物学报,1990,32(11)：841-846.
华振基,王俊.春小麦籽粒发育的形态学及解剖学的研究Ⅲ：胚乳初期发育的发生及解剖学观察.宁夏农学院报,1982,1：1-9.
黄百渠.拟南芥菜子叶中蛋白体形成的超微结构和免疫电镜定位研究.实验生物学报,1989,22：247-257.
黄景华,王广金,孙岩,刁艳玲.小麦花粉管通道形成时期的研究.黑龙江农业科学,2004,2：20-22.
季清娥,郑恒清,徐珍秀等.品质不同的稻米胚乳淀粉粒的微观结构观察.福建农业大学学报,1998,27(2)：241-244.
康海岐,常红叶,许育彬等.水稻籽粒胚乳的石蜡切片方法改良及其结构发育观察.西北植物学报,2008,28(5)：1069-1074.
蓝盛银,徐珍秀.水稻胚乳淀粉粒形态发育的亚显微结构研究.武汉大学学报(自然科学版),1997,2：73-74.
蓝盛银,钟方旭,杨泽敏,靳德明,徐珍秀.水稻淀粉胚乳程序性细胞死亡中的去核化.实验生物学报,2004,37(1)：34-44.
李师翁,屠骊珠.白刺胚和胚乳发育的研究.植物学报.1991,33(7)：500-506.
李师翁.被子植物核型胚乳细胞化机理研究的现状.植物学通报,1995,12(4)：13-16.
刘冬娟,祝素文,李艳杰.硼砂甲苯胺蓝染液在半薄切片染色上的应用.中国医科大学学报,2004,33(1)：33.
刘向东,徐是雄,卢永根.水稻胚囊壁的形成与发育观察.植物学报,1997,39(11)：985-990.
刘智,王玲玲,周卫东,陈义芳,王忠.固定和染色方法对水稻胚乳细胞结构观察的影响.中国水稻科学,2011,25(6)：667-671.
罗玉英.玉竹花粉中造粉质体形成及增殖的超微结构研究.电子显微学报,1995,14(3)：166-169.
李睿,蓝盛银,徐珍秀.水稻胚乳发育中线粒体向淀粉质体转变及琥珀酸脱氢酶的定位. 电子显微学报,2002,21(2)：118-121
毛同林,吴晓东,杨世杰.洋葱根冠外缘细胞编程性死亡过程中的超微结构研究.电子显微学报,2001,20：21-26.
宋今丹.高锰酸钾固定的培养细胞的膜性结构.电子显微学报,1990,9(3)：1.
松田智明,川原治之助,长南信雄.日本作物学会纪事,1979,48：155-162.
松田智明.登熟期の水稻子房にぉける淀粉贮藏细胞の微细构造と转流、蓄积机构につぃて.日本作物学会纪事,1984,53(别号1)：144-147.
沈波.早籼稻垩白形成中胚乳淀粉粒发育的电镜观察.中国水稻科学,2000,14(4)：225-228.
盛婧,郭文善,朱新开,封超年,彭永欣.不同专用类型小麦籽粒淀粉粒形成过程.作物学报,2004,30(9)：953-954.
汤圣祥,江云珠,李双盛等.早籼稻胚乳淀粉体的扫描电镜观察.作物学报,1999,25(2)：269-273.
田代亨,河野恭广.水稻胚乳组织の发育生理学研究第1报胚乳组织の细胞学的不均一性仁”にっぃて.日本作物学会纪事,1972,41：521-530.
王爱国,罗广华,邵从文等.大豆种子超氧化物歧化酶的研究.植物生理学报,1983,9：77-84.
王长发,张篙午.冷型小麦旗叶衰老和活性氧代谢特性研究.西北植物学报,2000,20(5)：727-732.
王兰,刘向东等.同源四倍体水稻胚乳发育：极核融合和胚乳细胞化.中国水稻科学,2004,18(4)：281-289.
王兰,刘向东等 同源四倍体水稻胚乳发育：糊粉层细胞壁纤维素物质发育、胚乳淀粉积累胼胝质“套”.中国水稻科学,2004,18(6)：507-514.
王萌,周舟,兰盛银.不同大麦淀粉胚乳细胞中淀粉和贮藏蛋白的积累.中国农业科学,2004,37(10)：1464-1467
王萍,任力.树脂包埋半薄切片染色的改进及体会.大连医科大学学报,2007,29(5)：509-510.
王全喜.植物学.北京：科学出版社,2004：21.
王爽,王春梅,陈丹,黄晓峰.环氧树脂半薄切片的多色性染色方法.第四军医大学学报,2002,23(9)：775.
王晓曦,杨献辉,程凤明等.小麦胚乳结构中蛋白质分布及其对面团流变学特性影响的研究.郑州工程学院学报,2001,22(1)：63-66.
王余龙,蔡建中.水稻籽粒受容活性及其控制途径——Ⅱ.籽粒含水率与～(14)C光合产物分配及转化的关系.江苏农学院学报,1990,11(4)：27-31.
王忠,李卫芳,顾蕴洁,陈刚,石火英,高煌珠.水稻胚乳发育及其养分输入的途径.作物学报,1995,21：520-527.
王忠,陈刚,李克武,桂玉清,顾蕴洁.稻麦胚乳游离核的分裂.江苏农学院学报,1996, 17(1)：11-20.
王忠,顾蕴洁,李卫芳,练兴明,陈刚.关于大麦胚乳发育的研究.中国农业科学,1996,29：38-45.
王忠,顾蕴洁,李卫芳,陈刚,石火英,陈秀花.小麦胚乳发育及其养分输入的途径.作物学报,1998,24：536-543.
王忠,顾蕴洁,李卫芳,黄山,李克武.水稻糊粉层的形成及其在萌发过程中的变化.扬州大学学报,1998,1(1)：32-34.
王银改,王清改,翟素平.Image J软件辅助分析在网织红细胞计数中的应用.临床检验杂志,2005,23(3)：210.
翁寿松.日立的场发射扫描电镜.电子工业专用设备,1993,22(1)：30-33.
星川清亲.小麦胚乳形成初期的发生学研究,上海市科学技术编译馆出版,1960.
星川清亲.米の胚乳发育に关する组织形态的研究,第4报,糊粉层の分化と发育にっぃて.日作纪,1967,36：216-220.
星川清亲.日本作物学会纪事,1967,36(1)：151-161.
星川清亲.日本作物学会纪事,1967,36：203-227.
星川清亲.日本作物学会纪事,1967,36：398-407.
星川清亲.日本作物学会纪事,1968,37：87-106.
星川清亲.日本作物学会纪事,1968,37：207-216.
星川清亲.日本作物学会纪事,1984,53：153-162.
星川清亲著,蒋彭炎,许德海译.解剖图说稻的生长.上海：上海科学技术出版社,1980,273.
席湘媛.大葱胚和胚乳的发育.植物学报.1991,29(5)：459-464.
熊飞,王忠,陈刚等.不同籼稻颖果发育及其结构与品质性状.中国水稻科学,2006,20(1)：43-46.
徐是雄,徐雪宾.稻的形态与解剖.北京：农业出版社,1984：45-57.
徐是雄,刘向东,冯九焕,卢永根.水稻IR36及光温敏核雄性不育系培矮64S小孢子母细胞减数分裂期间微管骨架的变化.植物学报,2001,43(3)：221-226.
徐是雄,刘向东,朱洪亮,卢永根.水稻雌配子体发生过程中胚囊内微管骨架变化的进步研究.植物学报,2001,43(9)：910-917.
韦存虚.水稻胚乳发育中细胞核与细胞器的结构消长与功能研究.武汉：华中农业大学,2002.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞发育中的蛋白体的形成.作物学报,2002,28(5)：591-594.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞淀粉质体被膜与其增殖的关系.电子显微学报,2002,21(2)：124-128.
韦存虚,蓝盛银,徐珍秀.水稻胚乳细胞发育中的蛋白体的形成.作物学报,2002,28(5)：591-594.
韦存虚,蓝盛银,徐珍秀.水稻胚乳发育中ATP酶的超微细胞化学定位和功能分析.中国农业科学,2003,36(3)：259-262.
韦存虚,张军,周卫东等.小麦胚乳发育过程中贮藏蛋白的积累和蛋白体的形成.麦类作物学报,2009,29(1)：73-78.
韦存虚,张军,周卫东,陈义芳,许如根.大麦胚乳发育过程中贮藏蛋白的积累和蛋白体的形成.作物学报,2009,35(2)：334-340.
杨弘远.用整体染色与透明技术观察胚囊、胚、胚乳和胚状体.植物学报,1986,28(6)：575-581.
杨貌仙.小麦胚乳发育的研究.植物学报,1957,6(1)：39-52.
杨貌仙.小麦胚乳游离核繁殖的研究.植物学报,1962,11(1)：7-9.
杨学举,屈平,张彩英等.小麦A型淀粉粒大小对淀粉特性及面包品质的影响.河北农业大学学报,2004,27(5)：1-5.
叶庆华.植物生物学.厦门：厦门大学出版社,2002：32.
银永安,齐军仓,李卫华,曹连莆,王自布.小麦胚乳A、B型淀粉粒理化特性研究.中国农业科学,2010,43(11)：2372-2379.
尤瑞麟.小麦珠心细胞衰退过程的超微结构研究.植物学报,1985,27：345-353.
张松林,金芝兰.谷子胚和胚乳的发育.西北植物学报,1994,14(4)：249-254,
张伟成,严文海,高小彦.小麦胚乳细胞化初期胞壁形成特征的观察.植物学报,1990,32(1)：76-78.
郑晓鹰,顾增辉,徐本美,郑光华.几种蔬菜种子的活力试验.中国农业科学,1982,4：58-64.
中华医学会.临床技术操作规范(病理学分册).北京：人民军医出版社,2004.
钟瑛.改良半薄切片多色性染色方法.临床与实验病理学杂志,1992,8(3)：228.
朱徵.高碘酸—锡夫试剂反应作为一种染色方法在植物组织学上的应用.植物学报,1963,11(2)：155-166.
朱美君,王学臣.植物的编程性细胞死亡.植物生理学通讯,1999,5：353-357.