Name: protocol for rnai assays in adult mosquitoes (a. gambiae)
Uploaded: 2007-07-04T00:00:00
Description: Watch this Scientific Journal Video about Protocol for RNAi Assays in Adult Mosquitoes A. gambiae at JoVE.com

<p class="jove_content"> RNAi介导的基因沉默在蚊子需要事前准备和提纯感兴趣的基因的具体dsRNAs目标。双链RNA的合成，我们建议Ambion公司Megascript套件，这使得使用一个T7 RNA聚合酶介导<em>的体外</em>转录反应。对于纯化的双链RNA，我们建议Qiagen公司的RNeasy试剂​​盒。双链RNA的样品应量化，并调整为3微克/μL在水中。其他材料，您需要包括：细尖镊子，载玻片，玻璃microcapillary针，微量（我们使用Drummond的Nanoject第二），?...

<ol>
	<li>Boisson, ., 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> , (2006).</li><li>Blandin, ., 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> , (2002).</li></ol>

<p>基因沉默的效率是高度可变的，取决于多项因素，包括成绩单和蛋白质的转，超过利率，并通过细胞和器官的双链RNA吸收效率。我们发现，沉默开始早在1天注射后，可长达6天注射后。使用成绩单和蛋白质的检测方法，如定量PCR和Western印迹，以验证沉默。 </p>

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Megascript</td>
<td>kit</td>
<td>Ambion</td>
<td>&nbsp;</td>
<td>for dsRNA synthesis; T7 RNA polymerase-mediated in vitro transcription reaction</td>
</tr>
<tr>
<td>RNeasy</td>
<td>&nbsp;</td>
<td>Qiagen</td>
<td>&nbsp;</td>
<td>For dsRNA purification</td>
</tr>
<tr>
<td>dsRNA</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>should be quantified and adjusted to 3 &#x03BC;g/ &#x03BC;L in water</td>
</tr>
<tr>
<td>fine-tipped forceps</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>glass slide</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>glass microcapillary needles</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>microinjector</td>
<td>&nbsp;</td>
<td>Drummond</td>
<td>Nanoject II</td>
<td>&nbsp;</td>
<tr>
<td>light microscope</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>mounted above a cold block</td>
</tr>
<tr>
<td>Petri dish</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>covered, glass</td>
</tr>
<tr>
<td>paper towels</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>10% sucrose</td>
<td>food</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>A. gambiae</td>
<td>Animal</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>mosquitos</td>
</tr>		</tbody>
		</table>
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protocol for rnai assays in adult mosquitoes (a. gambiae)

<p class="jove_content">反向遗传方法已被证明非常有用，确定哪些基因底层电阻载体病原体在蚊子。该视频协议说明Dimopoulos实验室使用的冈比亚按蚊的蚊子，其中海港的疟疾寄生虫注入dsRNA的一个方法。操纵注射的设置和注入胸廓的dsRNA的技术说明。</p>

Watch this Scientific Journal Video about Protocol for RNAi Assays in Adult Mosquitoes A. gambiae at JoVE.com

Protocol for RNAi Assays in Adult Mosquitoes A. gambiae

成蚊（冈比亚按蚊的RNAi实验议定书）

Reverse genetic approaches have proven extremely useful for determining  which genes underly resistance to vector pathogens in mosquitoes.  This video ...

Research

JoVE Journal

Biology

Protocol for RNAi Assays in Adult Mosquitoes (A. gambiae)

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