Generation of a Non-Leaky Heat Shock-Inducible Cre Line for Conditional Cre/Lox Strategies in Zebrafish
Generation of a Non-Leaky Heat Shock-Inducible Cre Line for Conditional Cre/Lox Strategies in Zebrafish
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DOI：
10.1002/dvdy.22497
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发表时间：
2011-01-01
影响因子：
2.5
通讯作者：
Brand, Michael
中科院分区：
文献类型：
--
作者：
Hans, Stefan;Freudenreich, Dorian;Brand, Michael
Cre介导的定点重组已成为精确操纵哺乳动物基因组不可缺少的工具。最近，我们发现Cre在斑马鱼中也是高效的，并且可以通过配体诱导的Creer(T2)实现重组的时间控制。以前已经尝试通过使用温度可诱导的hsp701启动子来分别有条件地驱动Cre或EGFP-Cre的表达来控制重组。然而，在本研究中，我们证明即使在允许的温度下，hsp70l启动子也具有基础泄漏，从而导致Cre介导的重组。为了防止非条件重组，我们将hsp70l启动子与mCherry标记的配体诱导的Creer(T2)结合起来。在允许的温度下，在没有配体三苯氧胺()的情况下，没有观察到无条件的重组，这表明Creer(T2)受到严格的调控。取而代之的是，全面的部位特异性重组是在热诱导和给药后介导的。发展动力学240：108-115,2011年。(C)2010年Wiley-Liss公司
Cre-mediated site-specific recombination has emerged as an indispensable tool for the precise manipulation of the mammalian genome. Recently, we showed that Cre is also highly efficient in zebrafish and temporal control of recombination can be achieved by using the ligand-inducible CreER(T2). Previous attempts have been made to control recombination by using the temperature inducible hsp70l promoter to conditionally drive the expression of Cre or EGFP-Cre, respectively. However, in this study we demonstrate that the hsp70l promoter possesses a basal leakiness resulting in Cre-mediated recombination even at permissive temperatures. In order to prevent non-conditional recombination, we combined the hsp70l promoter with a mCherry-tagged ligand-inducible CreER(T2). At permissive temperatures and in the absence of the ligand tamoxifen (TAM), no non-conditional recombination is observed indicating tight regulation of CreER(T2). Instead, comprehensive site-specific recombination is mediated following heat induction and administration of TAM. Developmental Dynamics 240: 108-115, 2011. (C) 2010 Wiley-Liss, Inc.