Name: transplantation of gfp-expressing blastomeres for live imaging of retinal and brain development in chimeric zebrafish embryos
Uploaded: 2010-07-19T17:00:00
Description: Watch this Scientific Journal Video about Transplantation of GFP-expressing Blastomeres for Live Imaging of Retinal and Brain Development in Chimeric Zebrafish Embryos at JoVE.com

这项研究是由一个国家卫生研究院（NIH）的核心格兰特（5P30EY008098）及以下的资金XW：美国国立卫生研究院批准R01EY016099和研究，以防盲职业发展奖的支持。我们感谢协助共焦成像基拉Lathrop。

第1天。制备 1。设置鱼杂交在晚上设置成对杂交斑马鱼。为了防止随机交配前所需的时间，使用分频器分开，在交配磁带的雌性和雄性的鱼。绿色荧光蛋白表达的转基因鱼，用于捐助卵裂球的来源。 pt104转基因株系（邹等 ，2008），用在这里，表达GFP的无所不在EF1启动子的控制下。 GFP的表达，使供体细胞的可视化实时成像。其他表达合适的荧光蛋白的?...

<ol>
	<li>Zou, J., Lathrop, K., Sun, M., Wei, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intact+RPE+maintained+by+Nok+is+essential+for+retinal+epithelial+polarity+and+cellular+patterning+in+zebrafish.">Intact RPE maintained by Nok is essential for retinal epithelial polarity and cellular patterning in zebrafish.</a> Journal of Neuroscience. 28 (50), 13684-13695 (2008).</li><li>Westerfield, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+guide+for+the+laboratory+use+of+zebrafish+(Danio+rerio).">A guide for the laboratory use of zebrafish (Danio rerio).</a> THE ZEBRAFISH BOOK. , (2007).</li><li>Ho, R. K., Kane, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell-autonomous+action+of+zebrafish+spt-1+mutation+in+specific+mesodermal+precursors.">Cell-autonomous action of zebrafish spt-1 mutation in specific mesodermal precursors.</a> Nature. 348, 728-730 (1990).</li><li>Catalano, A., Raymond, P., Goldman, D., Wei, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zebrafish+dou+yan+Mutation+Causes+Patterning+Defects+and+Extensive+Cell+Death+in+the+Retina.">Zebrafish dou yan Mutation Causes Patterning Defects and Extensive Cell Death in the Retina.</a> Developmental Dynamics. 236 (5), 1295-1236 (2007).</li></ol>

在这段视频中，我们展示了一个卵裂球移植的方法来分析在斑马鱼的视网膜和大脑发育。此协议也可以用来分析其他组织和器官的发展，在主机的胚胎捐赠者卵裂球，只需释放到相应的位置。与绿色荧光蛋白分子的标签产量比用荧光染料标记的葡聚糖（何和凯恩1990年）标签的背景清洁。这是因为右旋糖酐染料从死亡的供体细胞的碎片持续更长的时间比绿色荧光蛋白，从而推断成像（卡塔拉...

E3的蛋水： <ul><li> 5毫米氯化钠</li><li> 0.17 mM的氯化钾</li><li> 0.33毫米氯化钙2 </li><li> 0.33毫米硫酸镁4 </li><li> 0.1％亚甲蓝</li></ul>

transplantation of gfp-expressing blastomeres for live imaging of retinal and brain development in chimeric zebrafish embryos

在自己的位置和财产广泛胚胎发育过程中细胞的变化。这些变化是受细胞和它们的环境之间的相互作用。嵌合胚胎的遗传背景不同的细胞组成的，是伟大的工具来研究基因介导的细胞 - 细胞相互作用。斑马鱼胚胎早期发育阶段的透明度，允许在细胞水平上的组织和器官的形态发生的直接可视。在这里，我们展示了一个协议，以产生在斑马鱼胚胎的嵌合体视网膜和大脑的供体细胞进行实时成像。该协议包括移植针，准备移植绿色荧光蛋白GFP阴性主机，表达捐助卵裂球和捐助现场共聚焦显微镜下的细胞行为的审查。稍作修改，这个协议也可以被用来研究的其他组织和机关在斑马鱼的胚胎发育。使用GFP标签的供体细胞的优点进行了讨论。

Watch this Scientific Journal Video about Transplantation of GFP-expressing Blastomeres for Live Imaging of Retinal and Brain Development in Chimeric Zebrafish Embryos at JoVE.com

Transplantation of GFP-expressing Blastomeres for Live Imaging of Retinal and Brain Development in Chimeric Zebrafish Embryos

我们展示了协议，实时成像胚胎发育过程中的细胞行为产生的嵌合斑马鱼的胚胎。

移植绿色荧光蛋白表达对视网膜和大脑发育的实时成像嵌合斑马鱼胚胎的卵裂球

Cells change extensively in their locations and property during embryogenesis. These changes are regulated by the interactions between the cells and their ...

Research

JoVE Journal

Biology

Two-photon axotomy and time-lapse confocal imaging in live zebrafish embryos

双光子现场斑马鱼的胚胎干切断和时间推移共焦成像

Zebrafish have long been utilized to study the cellular and molecular mechanisms of development by time-lapse imaging of the living transparent embryo.  ...

科研

生物学

Live Imaging of the Zebrafish Embryonic Brain by Confocal Microscopy

现场共聚焦显微镜对斑马鱼的胚胎脑成像

In this video, we demonstrate the method our lab has developed to analyze the cell shape changes and rearrangements required to bend and fold the ...

Generating Chimeric Zebrafish Embryos by Transplantation

移植嵌合斑马鱼胚胎生成

One of the most powerful tools used to gain insight into complex developmental processes is the analysis of chimeric embryos. A chimera is defined as an ...

Live Imaging of Cell Motility and Actin Cytoskeleton of Individual Neurons and Neural Crest Cells in Zebrafish Embryos

在斑马鱼胚胎的单个神经元和神经嵴细胞的细胞运动和肌动蛋白细胞骨架的实时成像

The zebrafish is an ideal model for imaging cell behaviors during development in vivo. Zebrafish embryos are externally fertilized and thus easily ...

Development of automated imaging and analysis for zebrafish chemical screens.

斑马鱼化工屏幕自动成像和分析的发展。

We demonstrate the application of image-based high-content screening (HCS) methodology to identify small molecules that can modulate the FGF/RAS/MAPK ...

Two-Photon-Based Photoactivation in Live Zebrafish Embryos

在现场的斑马鱼胚胎的基于双光子Photoactivati​​on

Photoactivation of target compounds in a living organism has proven a valuable approach to investigate various biological processes such as embryonic ...

Dechorionation of Medaka Embryos and Cell Transplantation for the Generation of Chimeras

鳉胚胎和细胞移植嵌合体生成Dechorionation

Medaka is a small egg-laying freshwater fish that allows both genetic and embryological analyses and is one of the three vertebrate model organisms in ...

Imaging Glycans in Zebrafish Embryos by Metabolic Labeling and Bioorthogonal Click Chemistry

在斑马鱼胚胎的代谢标记和Bioorthogonal点击化学成像聚糖

Imaging glycans in vivo has recently been enabled using a bioorthogonal chemical reporter strategy by treating cells or organisms with azide- or ...

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

Live Imaging of Apoptotic Cell Clearance during Drosophila Embryogenesis

凋亡细胞清除过程中实时成像<em&gt;果蝇</em&gt;胚胎发育

The  proper elimination of unwanted or aberrant cells through apoptosis and  subsequent phagocytosis (apoptotic cell clearance) is crucial for normal  ...

Analysis of Oxidative Stress in Zebrafish Embryos

在斑马鱼胚胎氧化应激分析

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes ...