Simple and Efficient Transgenesis with I-SceI Meganuclease in the Newt, Cynops pyrrhogaster
Simple and Efficient Transgenesis with I-SceI Meganuclease in the Newt, Cynops pyrrhogaster
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DOI：
10.1002/dvdy.22463
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发表时间：
2010-12-01
影响因子：
2.5
通讯作者：
Chiba, Chikafumi
中科院分区：
文献类型：
--
作者：
Casco-Robles, Martin Miguel;Yamada, Shouta;Chiba, Chikafumi
自 18 世纪以来，蝾螈就被认为是创伤后身体部位再生的理想模型。然而，由于技术限制，再生的分子机制仍然是个谜。在本研究中，为了打破这一障碍，我们通过采用I-SceI显微注射技术，建立了一个简单有效的蝾螈Cynopspyrrogaster转基因方案，以及一个两缸罐（TAT）系统，使我们能够在实验室中不断获得受精卵进行转基因。按照我们的方案，大约 20% 的注射胚胎将表现出非嵌合的广泛转基因表达，并在变态后存活。这一预期成功率比之前的协议高出约10倍，达到了实用水平。因此，我们的转基因方案与 TAT 系统相结合，可以提供一项关键技术，为揭开蝾螈身体部位再生的长期谜团开辟道路。发育动力学 239:3275-3284, 2010。(C) 2010 Wiley-Liss, Inc.
Newts have been recognized as an ideal model for body-parts regeneration after traumatic injury since the 18(th) century. However, molecular mechanisms underlying regeneration remain a mystery because of technical limitations. In the current study, to break this obstacle, we established a simple and efficient transgenic protocol for the newt Cynops pyrrhogaster by adapting an I-SceI microinjection technique, as well as a two-aquarium-tank (TAT) system that allows us to constantly obtain fertilized eggs in the laboratory for transgenesis. Following our protocol, similar to 20% of injected embryos would exhibit non-mosaic widespread transgene expression and survive beyond metamorphosis. This anticipated success rate is about 10 times higher than that obtained by previous protocols, reaching a practical level. Therefore, our transgenic protocol in conjunction with the TAT-system could provide a key technique to open the way to uncover the long mystery underlying body-parts regeneration of newts. Developmental Dynamics 239:3275-3284, 2010. (C) 2010 Wiley-Liss, Inc.