琼脂糖印迹法：观察植物表皮细胞的一种简易方法

本文介绍一种获得完整植物器官表皮细胞大小和数目的简易方法：琼脂糖印迹法。该方法根据琼脂糖凝固时具有可塑性的原理,通过对材料固定、包埋、切胶和显微观察等步骤从而获得材料表皮细胞的轮廓。该方法具有简单迅速、图像清晰、观测结果准确且应用广泛等优点,可使统计植物发育过程中细胞数目及大小的工作变得简单易行。     

介绍一种新的表皮细胞传代方法

我们设计了一种新的传代方法,在传代之前,先用 DispaseⅡ酶,将培养的表皮细胞片整个消化下来,再用0.25—0.3%胰酶冷消化将表皮细胞驱散,进行1:2传代获得成功,传代59次成功率为71.2%。若在培养后7—16天传代,成功率可达80%。本文讨论了此种传代方法比单纯用胰蛋白酶或用胰蛋白酶和EDTA 混合传代方法优越。     

琼脂糖凝胶直接杂交法

DNA印迹(Southern blot)杂交法是研究DNA分子结构,变异及其组成的一种分子生物学技术。自1975年闻世以来,在分子生物学,遗传学及分子病毒学等研究领域得到广泛应用,对这些学科的发展起了重要作用。由于该技术均需要首先将电泳后已变性的DNA从琼脂糖凝胶转移至支持膜上,因此,实验结果的好坏很大程度取决于吸印的效果,同时也受支持物     

一种简易RNA甲醛—琼脂糖凝胶变性电泳

构建ｃＤＮＡ文库 ,进行Ｎｏｒｔｈｅｒｎ杂交分析以及ＲＴ ＰＣＲ等分子生物学研究 ,都需要纯度高、完整性好的ＲＮＡ。鉴定ＲＮＡ的完整性通常需要进行甲醛 琼脂糖凝胶电泳。然而 ,传统的甲醛 琼脂糖凝胶电泳操作步骤繁杂[1 ] ,本文提出一种改进的甲醛 琼脂糖凝胶电泳 ,不仅简化了操作 ,而且效果很好。我们以向日葵总ＲＮＡ和ｍＲＮＡ甲醛 琼脂糖凝胶电泳为例 ,对这一方法进行介绍。1　材料与方法1.1　总ＲＮＡ的提取及ｍＲＮＡ的分离取向日葵花蕾 1ｇ ,按照改进的异硫氰酸胍法提取向日葵总ＲＮＡ[2 ] 。利用磁珠法分离ｍＲＮＡ[3] 。…     

利用洋葱鳞叶表皮细胞观察植物细胞结构实验的改进

以洋葱表皮细胞为研究材料,通过中性红液泡染色和质壁分离实验相结合,将能更清晰地观察到细胞壁,细胞质膜,细胞质,液泡膜,液泡,细胞核,核仁等结构,达到光学显微镜下观察植物细胞基本结构的教学目的。改进后的实验大大提高了利用洋葱表皮细胞观察植物细胞结构实验的教学效果,同时可以通过观察不同部位液泡体积变化,了解植物细胞的动态发育,使学生掌握了更多的知识点。     

一种用透明胶带粘取叶片表皮观察气孔的方法

用普通塑料透明胶带粘取叶片下表皮观察植物的气孔,比印迹法和撕取下表皮观察气孔的方法具有操作简单、速度快、真实性强的优点,尤其适宜于试管苗等幼嫩材料的气孔变化动态和形态学指标的研究.此法可以防止因保卫细胞失水而导致气孔开张度的变化,能真实地观察植物不同阶段和时期气孔的变化动态.     

一种从琼脂糖凝胶中同步回收DNA和琼脂糖的方法

介绍一种从琼脂糖凝胶同步回收DNA和琼脂糖的方法。利用0.25 mol/L异硫氰酸胍溶液(p H 8.0)溶解含有目的 DNA片段的的凝胶条,胶条溶解后,静置冰上10 min再加入预冷的异丙醇,琼脂糖呈颗粒状析出,通过离心即可初步分离DNA和琼脂糖。上清液用异丙醇沉淀回收DNA片段,利用50%PEG溶液沉淀琼脂糖。分别对0.2 kb、1 kb和10 kb长度的DNA片段进行回收,回收率分别为19.44%、36.40%、13.49%,回收的DNA纯度高,电泳条带清晰。琼脂糖均回收率为62.52%,回收琼脂糖脱水后的状态为白色颗粒。该方法切实可行,回收成本低廉,回收的DNA和琼脂糖可用于后续实验。     

植物高度木质化叶片表皮细胞制备方法

以凤凰茶(Camellia sinensis)叶片为例,介绍一种获取高度木质化植物叶片表皮细胞的简易方法——叶片燃烧法.该方法根据高度木质化的叶片在燃烧、过水之后,其上下表皮自行分开、细胞透明化,从而获得材料上下表皮细胞的轮廓.该方法具有简单迅速、制片效率高、上下表皮易区分及可靠稳定等优点,非常适用于木质化叶片表皮特征的研究,也可为植物学及生态学的相关研究方法提供参考.     

眼结膜印迹细胞法及其影响因素

本文摸索眼结膜印迹细胞法及其影响因素,证明该方法是一项简便,快速,有效地评价机体维生素Ａ营养状况的方法;取样材料,染色时间,温度,透明方法均可影响制片结果。该方法适合基层推广应用。     

观察洋葱表皮细胞实验的改进

在实验过程中,教师可以想办法培养学生的竞争意识与创新意识。竞争意识的培养可以通过组间或个人间进行比赛的形式获得,如比比谁先观察到玻片标本上的东西。创新意识的培养主要体现在实验的设计方面,不拘泥于书本上的方案,根据现有条件重新设计方案。在此过程中,通过评选最佳方案又可以培养学生的竞争意识。诸如此类的情感教育,完全可以让学生全面体会到实验教学的功效,在情感的指引下唤起学生内部的学习动机,巩固实验的浓厚兴趣。     

一种用于PCR模板制备的电泳产物简易回收方法

为了探索一种简便、有效而且能从琼脂糖凝胶中大量回收用于第2次PCR扩增的DNA电泳条带的方法,采用刀片切胶法和牙签插胶法从琼脂糖中回收DNA,并进行了两种方法的比较．结果显示牙签插胶法回收的DNA用作第2次PCR的模板,获得了清晰、稳定的PCR产物电泳条带,用该法成功地制备了一批DNA微阵列探针．由此可见牙签插胶法是一种简便、快速、有效的用于PCR模板的DNA琼脂糖凝胶回收法．     

A New Way to Investigate Living Flagellated/Ciliated Cells in the Light Microscope: Immobilization of Cells in Agarose

A method for cell immobilization of living flagellated/ciliated cells in agarose has been developed that allows single cells to be viewed for prolonged periods of time using high resolution light microscopy. Embedding in ultralow gelling, soft agarose preserves cellular functions of various flagellated/ciliated protists including delicate species, marine dinoflagellates, cryptomonads, contractile ciliates, etc. for days. Cell division, morphogenesis of cell organelles and intracellular movements can thus be studied for the first time in great detail. The method may also be useful for the isolation of flagellated/ciliated protists from nature and for the establishment of axenic clonal cultures in a single step.     

A Theoretical Model of Jigsaw-Puzzle Pattern Formation by Plant Leaf Epidermal Cells

Plant leaf epidermal cells exhibit a jigsaw puzzle–like pattern that is generated by interdigitation of the cell wall during leaf development. The contribution of two ROP GTPases, ROP2 and ROP6, to the cytoskeletal dynamics that regulate epidermal cell wall interdigitation has already been examined; however, how interactions between these molecules result in pattern formation remains to be elucidated. Here, we propose a simple interface equation model that incorporates both the cell wall remodeling activity of ROP GTPases and the diffusible signaling molecules by which they are regulated. This model successfully reproduces pattern formation observed in vivo, and explains the counterintuitive experimental results of decreased cellulose production and increased thickness. Our model also reproduces the dynamics of three-way cell wall junctions. Therefore, this model provides a possible mechanism for cell wall interdigitation formation in vivo.     

Western Blotting: Sample Preparation to Detection

Western blotting is an analytical technique used to detect specific proteins in a given sample of tissue homogenate or extract. It uses gel electrophoresis to separate native or denatured proteins by the length of the polypeptide (denaturing conditions) or by the 3-D structure of the protein (native/ non-denaturing conditions). The proteins are then transferred to a membrane (typically nitrocellulose or PVDF), where they are probed (detected) using antibodies specific to the target protein.Download video file.^{(47M, mov)}     

The Use of a Simple Electron Microscope Serology Procedure to Observe Relationships of Seven Potyviruses

The relationships between bean yellow mosaic (BYMV), bean common mosaic (BCMV), clover yellow vein (CYVV), lettuce mosaic (LMV), potato virus Y (PVY), turnip mosaic (TuMV) and celery mosaic (CeMV) viruses were studied in homologous and heterologous reactions, using simple and relatively rapid electron microscope serology decoration tests. The degree of relationship between these viruses was assessed by the intensity of antibody coating when the viruses were decorated by heterologous antibodies. A close relationship was observed between BYMV and CYVV, and between BYMV and LMV but not between CYVV and LMV. CeMV was quite closely related to BYMV and CYVV. Antibodies to BCMV and BYMV intensely decorated different strains of their own virus, but decoration was negligible in cross reactions.     

Involvement of Suppressor-Glucans and Plant Epidermal Cells in Host-Selective Pathogenesis of Phytophthora capsici

Water-soluble glucans (WSG) from a virulent isolate of Phytophthora capsici (PCAP-3) which were released during germination of cystospore markedly suppressed the elicitor-induced death of suspension-cultured cells of susceptible sweet pepper and tomato but not that of resistant pepper and tobacco. PCAP-3, its polygalacturonase (PGase)-deficient mutant (PCAP3-M16), and galacturonic acid non-utilizable mutant carrying the PGase (PCAP-1) activity could penetrate in epidermal cells of host leaves, but similarly caused a hypersensitive response (HR) on non-injured leaves of resistant host (sweet pepper). In the case of inoculation on press-injured leaves, however, both of the resistant and nonhost plant leaves became quite susceptible to PCAP-3similar to susceptible hosts, but not to PCAP3-M16 and PCAP-1. The results suggested that host-selectivity of P. capsici may be determined in the leaf epidermal cells where the suppressor glucans released during infection effectively suppressed the occurrence of hypersensitive reaction. Furthermore, during growth of the fungus in intercellular spaces of leaf tissues, PGase may contribute not only to the virulence experession but also the supply of initial nutrition for fungal growth in the intercellular space of host tissues.     

介绍一种高效、简单的DNA沉淀法

毫摩尔级的Ｚｎ２＋可以在适宜条件下导致核酸沉淀物的形成。优化后的ＺｎＣｌ２沉淀法的基本步骤是：５０ｋｂ和２０ｂｐ的ＤＮＡ样品分别置于５０ｍＭＴｒｉｓ（ｐＨ７．０）稀释,ＥＤ－ＴＡ到终浓度小于０．１ｍＭ中,加入０．０１Ｍ磷酸钠缓冲液（ｐＨ７．０）到终浓...