Name: large scale zebrafish-based in vivo small molecule screen
Uploaded: 2010-12-30T17:00:00
Description: Watch this Scientific Journal Video about Large Scale Zebrafish-Based In vivo Small Molecule Screen at JoVE.com

1）斑马鱼的卵收集<ol><li>化学屏幕一天前的下午，成立10到20的斑马鱼的繁殖坦克。每个水箱装满水从水产养殖系统。使用渔网，将一个成年男性和一到两个成年女性在每个养殖池的内部容器。独立分频器从对方的男性和女性的鱼。标签的笼子，并把他们的盖子。 </li><li>在屏幕上午上，删除从养殖池的分隔，使斑马鱼进行交配。在未来1小时的课程，让受精卵通过电网秋天在每个内部容器的底部?...

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	<li>Bayliss, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+modulation+of+receptor+signaling+inhibits+regenerative+angiogenesis+in+adult+zebrafish.">Chemical modulation of receptor signaling inhibits regenerative angiogenesis in adult zebrafish.</a> Nature chemical biology. 2, 265-273 (2006).</li><li>Burns, C. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+assay+for+small+molecules+that+modulate+zebrafish+embryonic+heart+rate.">High-throughput assay for small molecules that modulate zebrafish embryonic heart rate.</a> Nature chemical biology. 1, 263-264 (2005).</li><li>Hao, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+structure-activity+relationship+study+of+dorsomorphin+analogues+identifies+selective+VEGF+and+BMP+inhibitors.">In vivo structure-activity relationship study of dorsomorphin analogues identifies selective VEGF and BMP inhibitors.</a> ACS chemical biology. 5, 245-253 (2010).</li><li>Hong, C. C., Peterson, Q. P., Hong, J. Y., Peterson, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Artery/vein+specification+is+governed+by+opposing+phosphatidylinositol-3+kinase+and+MAP+kinase/ERK+signaling.">Artery/vein specification is governed by opposing phosphatidylinositol-3 kinase and MAP kinase/ERK signaling.</a> Curr Biol. 16, 1366-1372 (2006).</li><li>Milan, D. J., Peterson, T. A., Ruskin, J. N., Peterson, R. T., MacRae, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drugs+that+induce+repolarization+abnormalities+cause+bradycardia+in+zebrafish.">Drugs that induce repolarization abnormalities cause bradycardia in zebrafish.</a> Circulation. 107, 1355-1358 (2003).</li><li>Peterson, R. T., Link, B. A., Dowling, J. E., Schreiber, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Small+molecule+developmental+screens+reveal+the+logic+and+timing+of+vertebrate+development.">Small molecule developmental screens reveal the logic and timing of vertebrate development.</a> Proceedings of the National Academy of Sciences of the United States of America. 97, 12965-12969 (2000).</li><li>Yu, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dorsomorphin+inhibits+BMP+signals+required+for+embryogenesis+and+iron+metabolism.">Dorsomorphin inhibits BMP signals required for embryogenesis and iron metabolism.</a> Nature chemical biology. 4, 33-41 (2008).</li><li>Zon, L. I., Peterson, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+drug+discovery+in+the+zebrafish.">In vivo drug discovery in the zebrafish.</a> Nature reviews. 4, 35-44 (2005).</li><li>Stern, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Small+molecules+that+delay+S+phase+suppress+a+zebrafish+bmyb+mutant.">Small molecules that delay S phase suppress a zebrafish bmyb mutant.</a> Nature chemical biology. 1, 366-370 (2005).</li><li>Peterson, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+suppression+of+a+genetic+mutation+in+a+zebrafish+model+of+aortic+coarctation.">Chemical suppression of a genetic mutation in a zebrafish model of aortic coarctation.</a> Nature biotechnology. 22, 595-599 (2004).</li></ol>

规划斑马鱼为基础的化学屏幕时，必须特别注意​​正在审议的表型，这种表型的背景率的稳健性。诱导表型的化学抑制器的屏幕，这一点尤为重要。例如，对于引起热休克诱导转基因表型，导致表型的条件下再现，必须精确地映射出启动屏幕前，以避免不可接受的高误报率。通过精心规划，斑马鱼化工屏幕，这就需要适度的资本投资，可能会导致的细胞信号转导和生理的新型调制器的发现，以?...

<ol><li>最小的20对成年斑马鱼所需的基因型。 </li><li>鱼网，养殖坦克，带有可拆卸的内胆和除法（水生境）。 </li><li>培养皿（10厘米）。 </li><li>塑料滤茶器。 </li><li>洗瓶containg胚胎水。 </li><li>一次性聚乙烯转移移液器。 </li><li>聚苯乙烯96圆底实验板（康宁COSTAR;洛厄尔，马）。 </li><li>玻璃巴斯德吸管（尔科技）。 </li><li>手动泵吸液管，10毫升（倍儿最先进的产品，Pequannock，新泽西州）。 </li><li> E3胚胎中期：5毫米氯化钠，0.17 mM的氯化钾， 氯化钙 0.33毫米，0.33毫米硫酸镁4，含有0.003％机动部队（苯，尿素，Sigma公司;圣路易斯，密苏里州）。溶解在1 L E3胚胎媒体0.3 - G机动部队，机动部队可以准备作为一个10倍的解决方案。解决方案包含机动部队应避光，用铝箔覆盖。 </li><li> 12通道移液器2-20μL（Eppendorf公司）。 </li><li> 12通道移液器，3-300μL（Eppendorf公司）。 </li><li>一次性聚苯乙烯吸管盆地，50毫升（尔科技）。 </li><li>小分子结构不同的化合物在96 - WLL格式排列在DMSO中的10毫米股票库。每个主板是分装成96孔聚丙烯存储板（康宁），并储存于-80 ° C，直到使用。 </li><li>铝箔封口磁带96孔板（NUNC，纽约州罗彻斯特）。 </li><li> DMSO（Sigma公司，圣路易斯，密苏里州）。 </li><li>基本孵化器，28.5℃（尔科技）。 </li><li>体视显微镜与透射光基地（徕卡显微系统，班诺克本，IL）。 </li></ol>

large scale zebrafish-based in vivo small molecule screen

鉴于他们的小胚胎的大小，快速发展，透明度，繁殖力，以及众多的分子，形态和生理上的相似性，对哺乳动物，已经成为一个强大的斑马鱼<em&gt;在体内</em&gt;表型为基础的药物屏幕和化学遗传分析平台。在这里，我们展示了一个简单，实用的方法大规模使用斑马鱼的胚胎小分子筛选。

Watch this Scientific Journal Video about Large Scale Zebrafish-Based In vivo Small Molecule Screen at JoVE.com

Large Scale Zebrafish-Based In vivo Small Molecule Screen

斑马鱼已成为一个强大的<em&gt;在体内</em&gt;表型为基础的药物屏幕和化学遗传分析平台。在这里，我们展示了一个简单，实用的方法大规模使用斑马鱼的胚胎小分子筛选。

斑马鱼为基础的大型在体内小分子屏幕

Given their small embryo size, rapid development, transparency, fecundity, and numerous molecular, morphological and physiological similarities to ...

Research

JoVE Journal

Biology

Large Scale Zebrafish-Based In vivo Small Molecule Screen

Large-Scale Screens of Metagenomic Libraries

宏基因组库的大型屏幕

Metagenomic libraries archive large fragments of contiguous genomic sequences from microorganisms without requiring prior cultivation.  Generating a ...

科研

生物学

Batch Immunostaining for Large-Scale Protein Detection in the Whole Monkey Brain

在整个猴脑进行大规模蛋白质检测的批次免疫染色

Immunohistochemistry (IHC) is one of the most widely used laboratory techniques for the detection of target proteins in situ. Questions concerning the ...

Isolation and Large Scale Expansion of Adult Human Endothelial Colony Forming Progenitor Cells

成人内皮细胞集落形成祖细胞的分离和大型规模扩张

This paper introduces a novel recovery strategy for endothelial colony forming progenitor cells (ECFCs) from heparinized but otherwise unmanipulated adult ...

Efficient Derivation of Human Cardiac Precursors and Cardiomyocytes from Pluripotent Human Embryonic Stem Cells with Small Molecule Induction

高效的推导人类心脏的前体和从多能干的小分子诱导人类胚胎干细胞的心肌细胞

To date, the lack of a suitable human cardiac cell source has been the major setback in regenerating the human myocardium, either by cell-based ...

Small-scale Nuclear Extracts for Functional Assays of Gene-expression Machineries

小规模的核提取物，基因表达机具功能分析

A great deal of progress in understanding gene expression has been made using in vitro systems.  For most studies, functional assays are carried out using ...

High-throughput Screening for Small-molecule Modulators of Inward Rectifier Potassium Channels

内向整流钾通道的小分子调节剂的高通量筛选

Specific members of the inward rectifier potassium (Kir) channel family are postulated drug targets for a variety of disorders, including hypertension, ...

Infinium Assay for Large-scale SNP Genotyping Applications

Infinium检测的大型SNP基因分型应用

Genotyping variants in the human genome has proven to be an efficient method to identify genetic associations with phenotypes. The distribution of ...

A Strategy for Sensitive, Large Scale Quantitative Metabolomics

敏感，大规模定量代谢组学的策略

Metabolite profiling has been a valuable asset in the study of metabolism in health and disease. However, current platforms have different limiting ...

A Manual Small Molecule Screen Approaching High-throughput Using Zebrafish Embryos

手册小分子屏幕接近高通量使用斑马鱼胚胎

Zebrafish have become a widely used model organism to investigate the mechanisms that underlie developmental biology and to study human disease pathology ...

Novel Production Protocol for Small-scale Manufacture of Probiotic Fermented Foods

对于益生菌发酵食品的小规模生产新生产协议

A novel dried bacterial consortium of Lactobacillus rhamnosus yoba 2012 and Streptococcus thermophilus C106 is cultured in 1 L of milk. This fresh starter ...