Name: mosaic analysis of gene function in postnatal mouse brain development by using virus-based cre recombination
Uploaded: 2011-08-01T12:00:00
Description: Watch this Scientific Journal Video about Mosaic Analysis of Gene Function in Postnatal Mouse Brain Development by Using Virus-based Cre Recombination at JoVE.com

这项工作是由来自美国国立卫生研究院研究所（NINDS）RO1的赠款支持。

1。准备用于注射的病毒<ol><li> rAAVs均购自商业供应商的建议，但他们也可以在自己的实验室中产生的（见下面的讨论）。 〜1 × 12，每毫升的基因组副本（GC /毫升）的滴度和病毒解决方案通常是生产，可在全滴度用来处理大量的细胞。此外，他们可能会被摊薄产生稀疏标签的理想水平。适当稀释，必须由用户决定，但建议开始1:10稀释。 </li><li>使用前在冰上解冻病毒。转移到一个小（〜...

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这里介绍的新生儿病毒注射方法提供了一个简单而快速的方式来产生体内马赛克产后大脑发育的研究。该方法需要利用floxed等位基因目前可用的，以及那些正在通过高通量基因靶向项目6。使用Cre转基因表达相比，这种方法提供了一种快速的方法来测试基因在不同细胞类型的功能，携带floxed等位基因的小鼠可直接用于实验，整个病毒注射过程从开始到结束，可在不到一小时完成。此?...

<table border="1"><tbody><tr><th>试剂名称</th><th>公司</th><th>目录编号</th><th>评论（可选） </th></tr><tr><td> rAAV8 - CRE，- GFP - DsRed的， </td><td>矢量Biolabs公司</td><td> ＃7060，＃7061，客户订单</td><td> http://www.vectorbiolabs.com </td></tr><tr><td>哈佛泵11加</td><td>哈佛器械</td><td> ＃702208 </td><td> （没有脚踏板端口） </td></tr><tr><td>视网膜色素上皮细胞注射套件</td><td>世界精密仪器</td><td> RPE - KIT </td><td>包含连通管，注射针（36G），持针器</td></tr><tr><td> NanoFil注射器，100微升</td><td>世界精密仪器</td><td> NANOFIL - 100 </td><td>包括可重复使用的装载针</td></tr><tr><td> D - PBS </td><td> Invitrogen公司</td><td> 14040-117 </td><td></td></tr><tr><td> GFP抗体</td><td>鸟类</td><td> GFP - 1020 </td><td></td></tr><tr><td> DsRed的抗体</td><td> Clontech公司</td><td> 632496 </td><td></td></tr><tr><td>加热块</td><td> VWR </td><td> 97042-610 </td><td></td></tr><tr><td> ROSA26R鼠标</td><td>杰克逊实验室</td><td> 003309 </td><td></td></tr></tbody></table>

mosaic analysis of gene function in postnatal mouse brain development by using virus-based cre recombination

不仅依赖于胚胎发育的主要神经通路建立正常的脑功能，但也发育时神经回路的成熟和完善。 Misregulation在这个阶段，可能会导致神经和精神疾病，如自闭症和精神分裂症1,2。许多基因已在胎儿的大脑研究，发现许多 3-5发育过程至关重要。然而，他们在出生后大脑功能在很大程度上是未知的，部分原因是因为他们在小鼠的缺失往往会导致新生儿发育期间，以杀伤力，部分原因是因为他们在早期发展的要求，阻碍了产后的分析。为了克服这些障碍，这些基因的floxed等位基因，目前正在生成在小鼠 6 。当结合表达Cre重组酶在特定的细胞类型，有条件的删除，可以实现在产后的大脑研究基因功能的转基因等位基因。然而，这种方法需要额外的等位基因和额外的时间（3-6个月），以产生适当的基因型的小鼠，从而限制了在鼠脑大规模扩展的遗传分析。 在这里，我们展示了一个互补的方法，使用病毒的表达Cre重组迅速，系统地研究这些floxed等位基因在出生后大脑发育。通过注入重组腺相关病毒（rAAVs）7,8编码的Cre新生儿的大脑，我们可以删除该基因在大脑的不同区域的利益。通过控制病毒滴度和共表达荧光蛋白标记，我们可以同时实现镶嵌基因的失活和稀疏神经元的标签。这种方法绕过了许多基因在早期发展的要求，使我们能够学习他们的细胞自主的功能在许多关键的过程，在出生后大脑发育，包括轴突和树突状增长，分支，和平铺，以及突触的形成和完善。该方法已成功地使用在我们自己的实验室（未公布结果）和其他8,9，并可以扩展到其他的病毒， 如 9，以及慢病毒的shRNA或主导活性蛋白 10的表达，。此外，通过与电相结合的技术，以及最近开发的光学成像工具11，这种方法提供了一个新的战略，以研究如何遗传途径影响神经电路的发展和在小鼠和大鼠的功能。

Watch this Scientific Journal Video about Mosaic Analysis of Gene Function in Postnatal Mouse Brain Development by Using Virus-based Cre Recombination at JoVE.com

Mosaic Analysis of Gene Function in Postnatal Mouse Brain Development by Using Virus-based Cre Recombination

一个<em&gt;在体内</em&gt;产后脑的方法来测试基因功能描述。表达CRE和/或荧光蛋白的重组自动增值服务注入新生小鼠大脑。马赛克基因的失活和稀疏的神经标签实现，使神经电路的发展过程的关键基因功能的快速分析。

利用基于病毒的Cre重组马赛克产后小鼠脑开发基因功能分析

Normal brain function relies not only on embryonic development when major neuronal pathways are established, but also on postnatal 
development when ...

Research

JoVE Journal

Neuroscience

Gene Delivery to Postnatal Rat Brain by Non-ventricular Plasmid Injection and Electroporation

基因传递到产后大鼠脑非心室质粒注射和电穿孔

Creation of transgenic animals is a standard approach in studying functions of a gene of interest in vivo. However, many knockout or transgenic animals ...

科研

神经科学

Mouse in Utero Electroporation: Controlled Spatiotemporal Gene Transfection

Mouse in Utero Electroporation: Controlled Spatiotemporal Gene Transfection

在子宫内电穿孔：鼠标控制时空基因Transefection

In order to understand the function of genes expressed in specific region of the developing brain, including signaling molecules and axon guidance ...

In ovo Electroporation in Chick Midbrain for Studying Gene Function in Dopaminergic Neuron Development

In ovo Electroporation in Chick Midbrain for Studying Gene Function in Dopaminergic Neuron Development

在OVO在鸡胚中脑多巴胺神经元发育中的基因功能研究电穿孔

Dopaminergic neurons located in the ventral midbrain control movement, emotional behavior, and reward mechanisms1-3. The dysfunction of ventral midbrain ...

Imaging Serotonergic Fibers in the Mouse Spinal Cord Using the CLARITY/CUBIC Technique

影像学检查HT能纤维在小鼠脊髓使用CLARITY / CUBIC技术

Long descending fibers to the spinal cord are essential for locomotion, pain perception, and other behaviors. The fiber termination pattern in the spinal ...

Inducing Cre-lox Recombination in Mouse Cerebral Cortex Through In Utero Electroporation

Inducing Cre-lox Recombination in Mouse Cerebral Cortex Through In Utero Electroporation

宫内电穿孔诱导小鼠大脑皮质中氧合酶的重组

Cell-autonomous neuronal functions of genes can be revealed by causing loss or gain of function of a gene in a small and sparse population of neurons. To ...

Homochronic Transplantation of Interneuron Precursors into Early Postnatal Mouse Brains

Interneuron 前体 Homochronic 移植早期小鼠脑的研究

Neuronal fate determination and maturation requires an intricate interplay between genetic programs and environmental signals. However, disentangling the ...

Studying Brain Function in Children Using Magnetoencephalography

脑磁图在儿童脑功能研究中的应用

Magnetoencephalography (MEG) is a non-invasive neuroimaging technique which directly measures magnetic fields produced by the electrical activity of the ...

Production, Purification, and Quality Control for Adeno-associated Virus-based Vectors

与腺相关病毒的向量的生产、纯化和质量控制

Gene delivery tools based on adeno-associated viruses (AAVs) are a popular choice for the delivery of transgenes to the central nervous system (CNS), ...

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset

常见马莫塞的全皮质电皮质阵列的慢性植入

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent ...

Transplantation of Chemogenetically Engineered Cortical Interneuron Progenitors into Early Postnatal Mouse Brains

将化学遗传工程皮质内神经内生前生移植到早期产后小鼠大脑

Neuronal development is regulated by a complex combination of environmental and genetic factors. Assessing the relative contribution of each component is ...