Name: surgical size reduction of zebrafish for the study of embryonic pattern scaling
Uploaded: 2019-05-03T14:30:00
Description: Watch this Scientific Journal Video about Surgical Size Reduction of Zebrafish for the Study of Embryonic Pattern Scaling at JoVE.com

这项工作得到了日本科学技术署 PRETO 方案 (JJPR11AA) 和国家卫生研究院赠款 (R01GM107733) 的支持。

所有与鱼类有关的程序都是在哈佛医学院动物护理和使用机构委员会 (IACUC) 的批准下进行的。
1. 工具和试剂制备
<ol><li>
做一个钢丝圈来切割胚胎
	<ol>
<li>取20厘米的不锈钢丝, 它坚硬且无腐蚀性, 直径为40μm。将电线循环到玻璃毛细管中 (外径 1.0 mm, 内径 0.5 mm, 无长丝), 使顶部有一个小循环 (环路长度为 1.0 mm)</li>
		<li>在钢丝圈之间的玻璃毛细管尖端放...

<ol>
	<li>Cooke, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scale+of+body+pattern+adjusts+to+available+cell+number+in+amphibian+embryos.">Scale of body pattern adjusts to available cell number in amphibian embryos.</a> Nature. 290, 775-778 (1981).</li><li>Driesch, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Entwicklungsmechanische+Studien:+I.+Der+Werthe+der+beiden+ersten+Furchungszellen+in+der+Echinogdermenentwicklung.+Experimentelle+Erzeugung+von+Theil-+und+Doppelbildungen.">Entwicklungsmechanische Studien: I. Der Werthe der beiden ersten Furchungszellen in der Echinogdermenentwicklung. Experimentelle Erzeugung von Theil- und Doppelbildungen.</a> Zeitschrift fur wissenschaftliche Zoologie. , (1892).</li><li>Morgan, T. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Half+embryos+and+whole+embryos+from+one+of+the+first+two+blastomeres.">Half embryos and whole embryos from one of the first two blastomeres.</a> Anatomischer Anzeiger. 10, 623-638 (1895).</li><li>Ishimatsu, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Size-reduced+embryos+reveal+a+gradient+scaling-based+mechanism+for+zebrafish+somite+formation.">Size-reduced embryos reveal a gradient scaling-based mechanism for zebrafish somite formation.</a> Development. 145, (2018).</li><li>Megason, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+toto+imaging+of+embryogenesis+with+confocal+time-lapse+microscopy.">In toto imaging of embryogenesis with confocal time-lapse microscopy.</a> Methods in Molecular Biology. 546, 317-332 (2009).</li><li>Graeden, E., Sive, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Live+imaging+of+the+zebrafish+embryonic+brain+by+confocal+microscopy.">Live imaging of the zebrafish embryonic brain by confocal microscopy.</a> Journal of Visualized Experiments. (26), e1217 (2009).</li><li>Rosen, J. N., Sweeney, M. F., Mably, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microinjection+of+zebrafish+embryos+to+analyze+gene+function.">Microinjection of zebrafish embryos to analyze gene function.</a> Journal of Visualized Experiments. (25), e1115 (2009).</li><li>Yuan, S., Sun, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microinjection+of+mRNA+and+morpholino+antisense+oligonucleotides+in+zebrafish+embryos.">Microinjection of mRNA and morpholino antisense oligonucleotides in zebrafish embryos.</a> Journal of Visualized Experiments. (27), e1113 (2009).</li><li>Sorlien, E. L., Witucki, M. A., Ogas, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficient+Production+and+Identification+of+CRISPR/Cas9-generated+Gene+Knockouts+in+the+Model+System+Danio+rerio.">Efficient Production and Identification of CRISPR/Cas9-generated Gene Knockouts in the Model System Danio rerio.</a> Journal of Visualized Experiments. (138), e56969 (2018).</li><li>Kemp, H. A., Carmany-Rampey, A., Moens, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generating+chimeric+zebrafish+embryos+by+transplantation.">Generating chimeric zebrafish embryos by transplantation.</a> Journal of Visualized Experiments. (29), e1394 (2009).</li><li>Mizuno, T., Shinya, M., Takeda, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cell+and+tissue+transplantation+in+zebrafish+embryos.">Cell and tissue transplantation in zebrafish embryos.</a> Methods in Molecular Biology. 127, 15-28 (1999).</li><li>Cooke, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Control+of+somite+number+during+morphogenesis+of+a+vertebrate,+Xenopus+laevis.">Control of somite number during morphogenesis of a vertebrate, Xenopus laevis.</a> Nature. 254, 196-199 (1975).</li><li>Inomata, H., Shibata, T., Haraguchi, T., Sasai, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scaling+of+dorsal-ventral+patterning+by+embryo+size-dependent+degradation+of+Spemann's+organizer+signals.">Scaling of dorsal-ventral patterning by embryo size-dependent degradation of Spemann's organizer signals.</a> Cell. 153, 1296-1311 (2013).</li><li>Gomez, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Control+of+segment+number+in+vertebrate+embryos.">Control of segment number in vertebrate embryos.</a> Nature. 454, 335-339 (2008).</li><li>Lauschke, V. M., Tsiairis, C. D., Francois, P., Aulehla, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scaling+of+embryonic+patterning+based+on+phase-gradient+encoding.">Scaling of embryonic patterning based on phase-gradient encoding.</a> Nature. 493, 101-105 (2013).</li><li>Schindelin, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fiji:+an+open-source+platform+for+biological-image+analysis.">Fiji: an open-source platform for biological-image analysis.</a> Nature Methods. 9, 676-682 (2012).</li><li>Almuedo-Castillo, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scale-invariant+patterning+by+size-dependent+inhibition+of+Nodal+signalling.">Scale-invariant patterning by size-dependent inhibition of Nodal signalling.</a> Nature Cell Biology. 20, 1032-1042 (2018).</li><li>Koos, D. S., Ho, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+nieuwkoid+gene+characterizes+and+mediates+a+Nieuwkoop-center-like+activity+in+the+zebrafish.">The nieuwkoid gene characterizes and mediates a Nieuwkoop-center-like activity in the zebrafish.</a> Current Biology. 8, 1199-1206 (1998).</li><li>Yamanaka, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+homeobox+gene,+dharma,+can+induce+the+organizer+in+a+non-cell-autonomous+manner.">A novel homeobox gene, dharma, can induce the organizer in a non-cell-autonomous manner.</a> Genes and Development. 12, 2345-2353 (1998).</li><li>Jesuthasan, S., Stahle, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+microtubules+and+specification+of+the+zebrafish+embryonic+axis.">Dynamic microtubules and specification of the zebrafish embryonic axis.</a> Current Biology. 7, 31-42 (1997).</li><li>Schier, A. F., Talbot, W. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+zebrafish+organizer.">The zebrafish organizer.</a> Current Opinion in Genetics and Development. 8, 464-471 (1998).</li></ol>

从历史上看, 在脊椎动物中, 大小缩小主要是使用两栖动物胚胎, 在第12条阶段沿动物-植物轴将胚胎一分为二.然而, 当我们将青蛙和斑马鱼胚胎一分为二时, 主要有两个区别。首先, 在斑马鱼胚胎耐受分分割 (囊胚阶段) 的阶段, 组织者位于脂肪边缘 18、19、20、21 的...

科学家们早就知道, 胚胎有一个显着的能力, 形成恒定的身体比例, 虽然胚胎大小可以很大的差异, 在自然和实验条件下1, 2,3。尽管几十年的理论和实验研究, 这种鲁棒性的大小变化, 称为缩放, 其潜在的机制仍然是未知的许多组织和器官。为了直接捕捉开发系统的动力学, 我们在斑马鱼4中建立了一种可重复、简单的尺寸缩小技术, 在活体成像5中具有很大的优势。
斑马鱼曾作为脊椎动物的模型, 研究生物学的多个学科, 包括发育生物学。特别是, 斑马鱼是体内活体成像的理想选择,因为 1) 发育可以在母亲和蛋壳之外正常进行, 2) 胚胎是透明的。此外, 胚胎还能承受一定的温度和环境波动, 从而能够在实验室条件下进行研究。此外, 除了常规基因表达摄动的吗啡和 mrna 注射7,8, 最近的进展 crispr/cas9 技术使反向遗传学在斑马鱼高效9。此外, 胚胎学中的许多经典技术, 如细胞移植或组织手术, 可以应用4,10,11。
减少大小的技术最初是在两栖动物和其他非脊椎动物中开发的.例如, 在另一种流行的脊椎动物模型月子劳维斯, 在囊胚阶段沿着动物-植物轴进行混合, 可以产生大小减少的胚胎 12,13.然而, 在我们手中, 这种一步的方法会导致斑马鱼的背化或腹形胚胎, 大概是因为背侧决定因素分布不均, 人们无法从胚胎的形态中知道它们的定位。在这里, 我们演示了另一种两步切碎技术的斑马鱼, 产生正常发育, 但较小的胚胎。有了这一技术, 细胞首先从动物极点中取出, 这是一个缺乏组织者活动的天真细胞区域。为了平衡蛋黄和细胞的数量, 蛋黄被去除, 这对上皮和随后的形态发生很重要。在这里, 我们详细介绍了这个协议, 并给出了模式形成中大小不变性的两个例子;体细胞形成和腹侧神经管模式。结合定量成像, 我们利用尺寸缩小技术, 考察了体细胞和神经管的大小在减少胚胎大小方面的影响。

<table border="0" cellpadding="0" cellspacing="0" style="width:608px;" width="609">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">60 mm PYREX Petri dish</td>
			<td style="width:107px;">CORNING</td>
			<td style="width:119px;">&#160;3160-60</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Agarose</td>
			<td style="width:107px;">affymetrix</td>
			<td align="right" style="width:119px;">75817</td>
			<td>For making a mount for live imaging</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Agarose, low gelling temperature Type VII-A</td>
			<td style="width:107px;">SIGMA-ALDRICH</td>
			<td style="width:119px;">A0701-25G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CaCl2</td>
			<td style="width:107px;">EMD</td>
			<td style="width:119px;">CX0130-1</td>
			<td>For 1/3 Ringer&#39;s solution</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CaSO4</td>
			<td style="width:107px;"></td>
			<td style="width:119px;"></td>
			<td>For egg water</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Cover slip (25 mm x 25 mm, Thickness 1)</td>
			<td style="width:107px;">CORNING</td>
			<td style="width:119px;">2845-25</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Disposable Spatula</td>
			<td style="width:107px;">VWR&#160;</td>
			<td style="width:119px;">80081-188</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Foam board</td>
			<td style="width:107px;">ELMER&#39;S</td>
			<td align="right" style="width:119px;">951300</td>
			<td>For microscope incubator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Forcept (No 55)</td>
			<td style="width:107px;">FST</td>
			<td style="width:119px;">11255-20</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Glass pipette</td>
			<td style="width:107px;">VWR</td>
			<td style="width:119px;">14673-043</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">HEPES</td>
			<td style="width:107px;">SIGMA Life Science</td>
			<td style="width:119px;">H4034</td>
			<td>For 1/3 Ringer&#39;s solution</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">INCUKIT XL for Cabinet Incubators</td>
			<td style="width:107px;">INCUBATOR Warehouse.com</td>
			<td style="width:119px;"></td>
			<td>For microscope incubator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Instant sea salt</td>
			<td style="width:107px;">Instant Ocean</td>
			<td align="right" style="width:119px;">138510</td>
			<td>For egg water</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">KCl</td>
			<td style="width:107px;">SIGMA-ALDRICH</td>
			<td style="width:119px;">P4504</td>
			<td>For 1/3 Ringer&#39;s solution</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Methyl cellulose</td>
			<td style="width:107px;">SIGMA-ALDRICH</td>
			<td style="width:119px;">M0387-100G</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">NaCl</td>
			<td style="width:107px;">SIGMA-ALDRICH</td>
			<td style="width:119px;">S7653</td>
			<td>For 1/3 Ringer&#39;s solution</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Petri dish</td>
			<td style="width:107px;">Falcon</td>
			<td align="right" style="width:119px;">351029</td>
			<td>For making a mount for live imaging</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Phenol red</td>
			<td style="width:107px;">SIGMA Life Science</td>
			<td style="width:119px;">P0290</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Pipette pump</td>
			<td style="width:107px;">BEL-ART PRODUCTS</td>
			<td style="width:119px;">F37898</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Pronase</td>
			<td style="width:107px;">EMD Millipore Corp</td>
			<td style="width:119px;">53702-250KU</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Tricaine-S (MS222)</td>
			<td style="width:107px;">WESTERN CHEMICAL INC</td>
			<td style="width:119px;">NC0135573</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ultra thin bright annealed 316L dia. 0.035 mm Stainless Steel Weaving Wires</td>
			<td style="width:107px;">Sandra</td>
			<td style="width:119px;"></td>
			<td>The wire we used was obtained ~20 years ago and we could not find exactly the same one. This product has the same material and diameter as the one we use.</td>
		</tr>
	</tbody>
</table>

surgical size reduction of zebrafish for the study of embryonic pattern scaling

在发育过程中, 胚胎表现出显著的能力, 使他们的身体模式与他们的身体大小相匹配;即使在较大或较小的胚胎中, 它们的身体比例也保持在一定范围内。虽然这种规模现象在一个多世纪以来引起了人们的注意, 但对其基本机制的了解有限, 部分原因是缺乏对不同大小胚胎发育动态的定量描述。为了克服这一限制, 我们开发了一种新的技术, 通过手术缩小斑马鱼胚胎的大小, 这对活体成像有很大的优势。我们证明, 在囊胚阶段分别平衡切除细胞和蛋黄后, 胚胎可以在合适的条件下迅速恢复, 发育成较小但原本正常的胚胎。由于这种技术不需要特殊的设备, 它很容易适应, 并可用于研究广泛的缩放问题, 包括形态原介导的模式的鲁棒性。

蛋黄体积减少对正常形态很重要 正如最近在 Almuedo-castylo 等人 17中所描述的, 在不减少蛋黄体积的情况下, 可以减少胚胎的大小。为了与蛋黄体积减少和不减少, 我们执行了两步切碎 (包括囊胚和蛋黄) 和只切碎 (图 2和补充电影 1)。与对照 (仅 dechorionation) 相比, 两步切碎的胚胎在整个发育阶段显示出看似正常的整体形态 (大小差异?...

Watch this Scientific Journal Video about Surgical Size Reduction of Zebrafish for the Study of Embryonic Pattern Scaling at JoVE.com

Surgical Size Reduction of Zebrafish for the Study of Embryonic Pattern Scaling

在这里, 我们描述了一种在不破坏正常发育过程的情况下减小斑马鱼胚胎大小的方法。这种技术可以研究模式缩放和针对大小变化的开发鲁棒性。

斑马鱼手术尺寸缩小术中胚胎形态缩放的研究

In the developmental process, embryos exhibit a remarkable ability to match their body pattern to their body size; their body proportion is maintained ...

虽然规模是发育系统的普遍和基本属性之一，但许多器官和组织尚未了解其基本机制。本视频中介绍的斑马鱼的尺寸缩小技术将促进您对动态缩放过程基础机制的理解。与斑马鱼合作的优点是易于进行活成像、遗传学和定量分析。我们的技术也非常简单，易于应用，没有任何专门的设备或强化培训。胚胎的健康是这项技术成功的关键。使用幼鱼作为父母，并小心通过去除胆小鱼尽量减少损害。也玩在蛋黄的伤口大小，和多少细胞被移除。由于我们使用的胚胎非常年轻和脆弱，在移植和切菜时需要非常小心地处理。同时，它需要快速，以便程序可以在正确的时间窗口完成开发。视觉演示可以让人们了解如何有效地产生更小的胚胎。为了制作用于胚胎切的钢丝环，通过玻璃毛细管将20厘米长的硬质和非腐蚀性不锈钢丝喂给，在顶部形成一个1毫米长的小环。将一小滴透明指甲油放在等级毛细管的尖端，在毛细管和钢丝环之间，以将指甲油放在到位位置，确保不将任何指甲油放在环上，因为它可能会损坏胚胎。然后让它干。使用实验室胶带将带环的玻璃毛细管连接到木筷子上，使大约两厘米半的玻璃毛细管延伸到筷子之外，使工具的筷子部分以后不会浸入水中。在100毫米玻璃培养皿中，用塑料铲将约0.5毫升的2%甲基纤维素分散在底部中心附近，均匀地摊开约0.5毫米的厚度。将约30毫升的三分之一林格溶液倒入菜面，并允许其扩散到盘子的其余部分，覆盖甲基纤维素。将先前脱卵的胚胎放在256个细胞的K细胞阶段，放在2%甲基纤维素上，确保胚胎被定向到它们的一侧。要切割炸管，请使用电线环在动物杆附近切割，垂直于动物植物轴，并切掉大约 30 到 40% 的爆炸细胞。然后轻轻敲击两端，帮助剩余的细胞粘住。对于蛋黄，使用安装的电线在植物杆附近通过划破蛋膜，使蛋黄渗出几分钟，然后愈合。当蛋黄停止渗出时，使用移液器将胚胎移出同一盘中的甲基纤维素以进行恢复。对所有胚胎重复炸管和蛋黄切，然后在胚胎恢复时让盘子不受干扰30分钟。然后，使用玻璃移液器将胚胎转移到先前准备的35毫米玻璃盘中，其中含有新鲜三分之一的林格溶液，并把它们放在28.5摄氏度的培养箱中，以便完全恢复，然后再继续成像。在用胚管和蛋黄进行两步切后，胚胎在发育阶段与控制胚胎相比，除了大小差异，似乎是正常的整体形态。另一方面，仅被切碎的胚胎则表现出一种特殊的形态，尤其是在早期阶段。在表皮期间，胚胎有收缩和缩进的外观。在以下索米特阶段，中线结构被发现在许多轴向层被夷为平地。在后期阶段，与蛋黄相邻的体体结构，如中脑和后脑仍然表现出相对扁平的形状，可能是因为相对较大的蛋黄的张力增加。索米特形成的时间推移成像表明，在对照胚胎和切碎的胚胎中，后期形成的卵石大小与早期形成的胚胎大小相比较小。此外，在整个苏米特形成阶段，切碎的胚胎比控制胚胎的胚胎小。当两步切克技术在膜mCherry注射胚胎上进行时，受精后20小时脊髓的成像显示，由于尺寸缩小，神经管高度降低。两步斩波对于获得更小但正常的胚胎具有高效率非常重要。重要的是要记住，不要切蛋黄，同时从爆炸中切掉细胞。对于蛋黄，做一个小伤害，它的内容渗出。蛋黄膜将在适当的缓冲液中自然愈合。由于切碎的胚胎可以快速恢复，因此任何方法或分析都可以应用于正常胚胎，可以按照缩小尺寸的方法进行。这种技术的简单性和多功能性是非常强大的，因为任何人都可以引入这种技术来研究斑马鱼中任何组织或器官的结垢问题。我们相信，通过这项技术，研究人员可以解决各种系统以前未触及的缩放问题。我们已经在苏米特和神经管模式的缩放方面取得了重要的发现。

Research

JoVE Journal

Developmental Biology

Large-scale Zebrafish Embryonic Heart Dissection for Transcriptional Analysis

大规模的斑马鱼胚胎的心脏解剖的转录分析

The zebrafish embryonic heart is composed of only a few hundred cells, representing only a small fraction of the entire embryo. Therefore, to prevent the ...

科研

发育生物学

Use of the TetON System to Study Molecular Mechanisms of Zebrafish Regeneration

使用提顿体系的研究斑马鱼再生的分子机制

The zebrafish has become a very important model organism for studying vertebrate development, physiology, disease, and tissue regeneration. A thorough ...

Hepatocyte-specific Ablation in Zebrafish to Study Biliary-driven Liver Regeneration

肝细胞特异性消融在斑马鱼研究胆驱动肝再生

The liver has a great capacity to regenerate. Hepatocytes, the parenchymal cells of the liver, can regenerate in one of two ways: hepatocyte- or ...

Analysis of Zebrafish Larvae Skeletal Muscle Integrity with Evans Blue Dye

斑马鱼幼虫骨骼肌诚信与伊文思蓝分析

The zebrafish model is an emerging system for the study of neuromuscular disorders.  In the study of neuromuscular diseases, the integrity of the muscle ...

Generation of Parabiotic Zebrafish Embryos by Surgical Fusion of Developing Blastulae

联体斑马鱼胚胎一代发展囊胚的手术融合

Surgical parabiosis of two animals of different genetic backgrounds creates a unique scenario to study cell-intrinsic versus cell-extrinsic roles for ...

Culture of Murine Embryonic Metatarsals: A Physiological Model of Endochondral Ossification

小鼠胚胎跖骨文化：软骨内骨化的生理模型

The fundamental process of endochondral ossification is under tight regulation in the healthy individual so as to prevent disturbed development and/or ...

In Vivo Imaging of Transgenic Gene Expression in Individual Retinal Progenitors in Chimeric Zebrafish Embryos to Study Cell Nonautonomous Influences

In Vivo Imaging of Transgenic Gene Expression in Individual Retinal Progenitors in Chimeric Zebrafish Embryos to Study Cell Nonautonomous Influences

在嵌合斑马鱼胚胎的个别视网膜祖细胞转基因表达的体内成像来研究细胞非自治的影响

The genetic and technical strengths have made the zebrafish vertebrate a key model organism in which the consequences of gene manipulations can be traced ...

A Method for Characterizing Embryogenesis in Arabidopsis

A Method for Characterizing Embryogenesis in Arabidopsis

Given the highly predictable nature of their development, Arabidopsis embryos have been used as a model for studies of morphogenesis in plants. However, ...

Organotypic Culture Method to Study the Development Of Embryonic Chicken Tissues

脊髓培养法研究胚胎鸡组织的发展

The embryonic chicken is commonly used as a reliable model organism for vertebrate development. Its accessibility and short incubation period makes it ...

Adult Zebrafish Injury Models to Study the Effects of Prednisolone in Regenerating Bone Tissue

成人斑马鱼损伤模型研究泼尼松对再生骨组织的影响

Zebrafish are able to regenerate various organs, including appendages (fins) after amputation. This involves the regeneration of bone, which regrows ...