Department of Genetics, Max-Planck-Institute for Developmental Biology, Tübingen, Germany.
Zebrafish. 2011 Sep;8(3):119-23. doi: 10.1089/zeb.2011.0703. Epub 2011 Aug 23.
N-Ethyl-N-nitrosourea (ENU) treatment is the standard experimental method used for chemical mutagenesis of many vertebrate organisms commonly used in the laboratory. In zebrafish and medaka, the use of a repetitive, sublethal dose of 3-3.5 mM ENU has been shown to give the best balance between mutagenicity and toxicity. However, even at this concentration, a significant proportion of fish die during the treatment. Therefore, large numbers of fish are required to obtain a sufficient number of mutagenized founders at the end of the procedure. Additionally, it is quite common to have high levels of mortality in any particular dosing cycle. This may cause a mutagenesis experiment to suddenly fail after several weeks of work. Here we provide a very simple method for ENU mutagenesis of zebrafish using a subparalytic dose of clove oil as a sedative that drastically reduces the lethality of ENU treatment in fish. This facilitates ENU mutagenesis protocols considerably, facilitates higher dosing, and allows for sensitive strains of fish such as homozygous mutants to be mutagenized for use in genetic studies.
N-乙基-N-亚硝脲（ENU）处理是一种标准的实验方法，用于对许多在实验室中常用的脊椎动物进行化学诱变。在斑马鱼和青鳉中，使用重复的、亚致死剂量的 3-3.5mM ENU 已被证明在诱变率和毒性之间取得了最佳平衡。然而，即使在这个浓度下，仍有相当一部分鱼在处理过程中死亡。因此，为了在实验结束时获得足够数量的诱变创始者，需要大量的鱼。此外，在任何特定的给药周期中，死亡率都相当高。这可能导致经过数周的工作后，诱变实验突然失败。在这里，我们提供了一种非常简单的方法，使用丁香油作为镇静剂对斑马鱼进行 ENU 诱变，这可以大大降低 ENU 处理对鱼的致死率。这极大地简化了 ENU 诱变方案，允许更高剂量的处理，并使诸如纯合突变体等敏感品系的鱼能够进行诱变，用于遗传研究。
