Targeted genome editing in Aedes aegypti using TALENs
Targeted genome editing in Aedes aegypti using TALENs
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DOI：
10.1016/j.ymeth.2014.02.008
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发表时间：
2014-08-15
期刊：
影响因子：
4.8
通讯作者：
Adelman, Zach N.
中科院分区：
文献类型：
--
作者：
Aryan, Azadeh;Myles, Kevin M.;Adelman, Zach N.
库蚊，埃及伊蚊，既是节肢动物传播病毒（虫媒病毒）的主要媒介，也是虫媒病毒传播的遗传模式生物。TALE核酸酶（TALEN）是一组能够产生位点特异性DNA损伤的人工酶，由融合到对靶位点特异性的工程化DNA结合结构域的非特异性FokI内切核酸酶切割结构域组成。虽然TALEN已成为多种生物体中靶向基因破坏的重要工具，但应用于蚊子基因组是一种新方法。我们最近描述了使用TALEN来执行A.埃及人。在这里，我们提供了详细的方法，将允许其他研究实验室利用这项技术的潜力，了解蚊子的基因功能。我们描述了靶位点选择，瞬时胚胎为基础的测定，以快速评估TALEN活性，胚胎显微注射和下游筛选步骤，以确定靶位点突变。(C)2014 Elsevier Inc. All rights reserved.
The Culicine mosquito, Aedes aegypti, is both a major vector of arthropod-borne viruses (arboviruses) and a genetic model organism for arbovirus transmission. TALE nucleases (TALENs), a group of artificial enzymes capable of generating site-specific DNA lesions, consist of a non-specific FokI endonuclease cleavage domain fused to an engineered DNA binding domain specific to a target site. While TALENs have become an important tool for targeted gene disruption in a variety of organisms, application to the mosquito genome is a new approach. We recently described the use of TALENs to perform heritable genetic disruptions in A. aegypti. Here, we provide detailed methods that will allow other research laboratories to capitalize on the potential of this technology for understanding mosquito gene function. We describe target site selection, transient embryo-based assays to rapidly assess TALEN activity, embryonic microinjection and downstream screening steps to identify target site mutations. (C) 2014 Elsevier Inc. All rights reserved.