植物细胞基因工程研究的现况和问题

现代生物学无论在理论上和方法上所取得的突飞猛进发展正给植物科学以巨大的推动。由于植物细胞培养、突变体选择、原生质体和细胞融合等方面的研究迅速进展,以及DNA分子体外重组和基因无性繁殖等技术的引入,已使利用培养的原生质体或细胞作为实验系统,来进行植物细胞的遗传操作成为     

书讯:水稻生物技术及遗传工程

农业生物技术领域的迅猛发展导致了对分子和细胞生物学的深入研究,并开发了新的基因转移方法。新出版的《粮食作物的生物技术:水稻生物技术及遗传工程》一书提供了一份详细的综述和近期水稻遗传工程的研究进展。本书的内容包括组织培养方面、细胞生物学和原生质体技术,还包括关于水稻基因转移的详细资料、转基因植物的特性鉴定和转基因植物的大田实验结果。本书分为以下各章:作为全球性作物     

酿酒酵母原生质体的融合

原生质体融合技术开始于动植物细胞,特别是Willin等报道了PEG有助于高等植物细胞原生质体融合以后,这一技术广泛地应用于植物、微生物原生质体的融合和动物细胞的融合。原生质体的制备、培养、融合和发育为研究细胞分化、膜结构与功能、定向育种等生物学     

植物原生质体的细胞壁再生

细胞壁作为植物细胞重要的组成部分,在决定细胞形状、维持机械支撑、吸收养分等方面发挥重要功能。因此,揭示植物细胞壁合成的调控机制具有重大的生物学意义。基于植物组织水平研究细胞壁的生物合成具有难以控制时间尺度、观察空间狭小等局限性。原生质体作为去除细胞壁的单个细胞是研究细胞壁再生的理想系统。在过去的几十年里报道了大量关于植物原生质体再生细胞壁的研究,但是关于细胞壁再生的机制尚不清楚。该综述介绍了目前应用于植物原生质体再生细胞壁研究的主要技术和取得的研究进展,并且对该领域的后续发展进行了展望,为进一步阐明植物细胞壁生物合成的机制提供理论参考。     

拟南芥六个新的small RNAs 的克隆和功能预测

以模式植物拟南芥为例, 建立了一种克隆small RNA 分子的技术平台, 为今后开展small RNA 分子的生物学功能研究提供技术支撑。通过用抗病信号分子水杨酸(SA) 处理拟南芥叶片后, 进行small RNA 分子群体的分离与接头连接、PCR 扩增、T - 载体克隆与检测、测序分析和生物信息学分析等一系列实验, 成功地克隆了一些small RNAs, 并对其表达和功能进行了分析。     

植物细胞的转化系统

本文系统地论述了目前植物细胞各种转化系统的技术特点、基本原理和研究进展,对植物细胞转化概念的广泛含义进行了初步的讨论,文中指出了转化系统对植物细胞分子遗传学及遗传工程的重要意义,最后指出了转化系除了上述各种转化系统外,用已转化的冠瘿瘤细胞原生质体与正常的原生质体融合也能实现 T-DNA 的转移,并得到属间体细胞杂种。这是一种借助细胞融合技术的转化系统。     

拟南芥六个新的small RNAs的克隆和功能预测

以模式植物拟南芥为例,建立了一种克隆small RNA分子的技术平台,为今后开展small RNA分子的生物学功能研究提供技术支撑.通过用抗病信号分子水杨酸(SA)处理拟南芥叶片后,进行small RNA分子群体的分离与接头连接、PCR扩增、T-载体克隆与检测、测序分析和生物信息学分析等一系列实验,成功地克隆了一些small RNAs.并对其表达和功能进行了分析.     

用TUNEL法检测植物原生质体的凋亡

TUNEL是近年来发展的一种对凋亡细胞进行原位检测的方法,可以特异性地标记完整的凋亡细胞核或凋亡小体的染色体3'_OH断裂末端,但在植物细胞中的应用还不多。本文报道应用TUNEL法检测胡萝卜原生质体的凋亡,并与DNA电泳、彗星电泳等方法进行了比较,结果表明它是一种适用于植物原生质体凋亡检测的灵敏度较高的方法。     

“检测生物组织中糖类、脂肪和蛋白质”教学组织

“检测生物组织中糖类、脂肪和蛋白质”是高中生物学必修1《分子和细胞》的一个重要实验。从实验原理、教学组织方式、检测方法以及学习评价等方面对这一实验的教学进行了分析,为高中生物学教学提供参考材料。     

单分子检测在生物学和药理学研究中的应用

单分子检测可以在单分子水平上原位、实时的获取各种生物化学信息,研究传统分析方法及生物学方法难以解决的问题,在生物学和药理学领域的研究中具有重要的意义.该文综述了荧光单分子检测在细胞生物学、分子生物学、药理学等领域的应用,并对其发展前景进行了展望.     

单分子荧光检测技术的发展及其在植物生物学研究中的应用

单分子荧光检测技术是利用荧光基团对目的分子标记后,在单分子水平成像并追踪分子的构象变化、动力学特征以及分子之间相互作用的研究方法.相较于传统分子生物学和遗传学的研究手段,单分子检测技术可以对单个分子的动态和特性进行分析,特别是瞬时或偶发性的事件,从而更加深入地挖掘在群体测量中被掩盖的信息.该技术已广泛应用于动物细胞生物...     

Responses of Chrysanthemum Cells to Mechanical Stimulation Require Intact Microtubules and Plasma Membrane–Cell Wall Adhesion

Plant cells are highly susceptible and receptive to physical factors, both in nature and under experimental conditions. Exposure to mechanical forces dramatically results in morphological and microstructural alterations in their growth. In the present study, cells from chrysanthemum (Dendranthema morifolium) were subjected to constant pressure from an agarose matrix, which surrounded and immobilized the cells to form a cell-gel block. Cells in the mechanically loaded blocks elongated and divided, with an axis preferentially perpendicular to the direction of principal stress vectors. After a sucrose-induced plasmolysis, application of peptides containing an RGD motif, which interferes with plasma membrane-cell wall adhesion, reduced the oriented growth under stress conditions. Moreover, colchicines, but not cytochalasin B, abolished the effects of mechanical stress on cell morphology. Cellulose staining revealed that mechanical force reinforces the architecture of cell walls and application of mechanical force, and RGD peptides caused aggregative staining on the surface of plasmolyzed protoplasts. These results provide evidence that the oriented cell growth in response to compressive stress requires the maintenance of plasmalemma-cell wall adhesion and intact microtubules. Stress-triggered wall development in individual plant cells was also demonstrated.     

植物实验生殖生物学与生殖工程：现在与未来

植物实验生殖生物学是植物实验胚胎学发展的新阶段,其主要特征为操作技术水平的提高与多学科综合性研究的加强。花粉原生质体、生殖细胞、精子、胚囊、卵细胞的操作、雌雄配子体外融合、配子-体细胞杂交等,代表了当前的主要研究趋势。实验生殖生物学与基因工程相结合,开辟了植柏物生殖工程新技术领域的前景。对生殖工程的意义与内容提出了轮廓设想。     

植物实验生殖生物学与生殖细胞工程:现在与未来

植物实验生殖生物学是植物实验胚胎学发展的新阶段,其主要特征为操作技术水平的提高与多学科综合性研究的加强。花粉原生质体、生殖细胞、精子,胚囊、卵细胞的操作、雌雄配子体外融合、配子-体细胞杂交等,代表了当前的主要研究趋势。实验生殖生物学与基因工程相结合,开辟了植物生殖工程新技术领域的前景。对生殖工程的意义与内容提出了轮廓设想。     

Interaction of purified DNA with plant protoplasts of different cell cycle stage: The concept of a competent phase for plant cell transformation

Summary The polycation mediated attachment of purified tritiated DNA to plant protoplasts has been measured by quantitative microautoradiography. The automated grain counting technique used, also provides information on the cell cycle stage of individual protoplasts, which circumvents the need to synchronize the plant cell population before preparation of protoplasts. With protoplasts from asynchronously dividing suspension cultures of Nicotiana syhestris (NS-1), S-phase protoplasts appear to be inefficient binders of ³H-DNA, as compared with G₁ or G₂ protoplasts. Protoplasts derived from a tumour line of Crepis capillaris (CAPT) exhibit ³H-DNA binding at all cell cycle phases, but Sphase protoplasts appear to be preferential binders. These differences are discussed with reference to cell cycle kinetics, membrane charge variation and the possibility of increasing the efficiency of genetic transformation of higher plant cells in culture.     

EXPERIMENTAL PLANT REPRODUCTIVE BIOLOGY AND REPRODUCTIVE CELL MANIPULATION IN HIGHER PLANTS: NOW AND THE FUTURE

Experimental plant reproductive biology has recently emerged as a new form of experimental embryology characterized by the direct experimental isolation and manipulation of reproductive cells and protoplasts. This work is fostered by current multidisciplinary trends in the sciences and serves to deepen our knowledge about the control of reproductive processes by providing novel in vitro material. The techniques of experimental plant reproductive biology also show great potential in providing new means for biotechnology, leading to significant refinements in plant breeding in higher plants that may eventually permit direct reproductive cell engineering. Recent achievements by our research group in experimental manipulation and biological studies of pollen protoplasts, generative cells, sperm cells, and embryo sacs are reviewed, and some ideas about future developments in this new area are presented.     

植物受精工程

植物受精工程是正在发展之中的高技术领域。近年来,雌雄配子的人工分离取得了较大进展,两者的体外模拟受精在玉米中已首次成功。本文对该领域的理论体系形成和实验系统创建作了评述,对目前存在的问题和今后的发展趋势进行了讨论。     

脂多糖诱导植物细胞产生NO的光学分子成像检测

脂多糖（lipopolysaccharides, LPS）是革兰氏阴性细菌细胞壁的主要成分,被植物感知后,启动植物防御反应。利用荧光探针分子成像及激光共聚焦扫描显微镜技术,在位、直观检测了LPS诱导下,拟南芥细胞产生重要信号分子一氧化氮（ NO）的时空特征。LPS诱导细胞产生大量NO,这些NO主要定位在细胞膜周围,且是在LPS处理90 min后出现。NO合成酶抑制剂L-单甲基精氨酸能明显抑制LPS诱导的NO生成,说明LPS诱导NO产生是NO合成酶途径依赖的。该研究结果有助于深入理解LPS作用机制以及NO信号传导通路的全貌,并为生物物理技术在相关植物生理研究中的应用提供一定的借鉴作用。     

PCR相关技术在植物病毒检疫检测中的应用

PCR和建立在PCR基础上的分子生物学技术以其灵敏、快速、简便等优点广泛应用于植物病毒的检测。阐述反转录聚合酶链式反应、免疫捕捉反转录PCR、PCR-单链构型多态性、实时荧光定量PCR、差异显示PCR和巢式PCR等相关技术的原理及其在植物病毒检测中的应用现状,以期为我国植物病毒的检疫检测提供有益参考。     

Trafficking of the human transferrin receptor in plant cells: effects of tyrphostin A23 and brefeldin A

Plant cells possess much of the molecular machinery necessary for receptor-mediated endocytosis (RME), but this process still awaits detailed characterization. In order to identify a reliable and well-characterized marker to investigate RME in plant cells, we have expressed the human transferrin receptor (hTfR) in Arabidopsis protoplasts. We have found that hTfR is mainly found in endosomal (Ara7- and FM4-64-positive) compartments, but also at the plasma membrane, where it mediates binding and internalization of its natural ligand transferrin (Tfn). Cell surface expression of hTfR increases upon treatment with tyrphostin A23, which inhibits the interaction between the YTRF endocytosis signal in the hTfR cytosolic tail and the mu2-subunit of the AP2 complex. Indeed, tyrphostin A23 inhibits Tfn internalization and redistributes most of hTfR to the plasma membrane, suggesting that the endocytosis signal of hTfR is functional in Arabidopsis protoplasts. Co-immunoprecipitation experiments show that hTfR is able to interact with a mu-adaptin subunit from Arabidopsis cytosol, a process that is blocked by tyrphostin A23. In contrast, treatment with brefeldin A, which inhibits recycling from endosomes back to the plasma membrane in plant cells, leads to the accumulation of Tfn and hTfR in larger patches inside the cell, reminiscent of BFA compartments. Therefore, hTfR has the same trafficking properties in Arabidopsis protoplasts as in animal cells, and cycles between the plasma membrane and endosomal compartments. The specific inhibition of Tfn/hTfR internalization and recycling by tyrphostin A23 and BFA, respectively, thus provide valuable molecular tools to characterize RME and the recycling pathway in plant cells.