Salinger Andrew P, Justice Monica J
Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA.
CSH Protoc. 2008 Apr 1;2008:pdb.prot4985. doi: 10.1101/pdb.prot4985.
INTRODUCTIONThis protocol describes chemical mutagenesis of male mice using N-ethyl-N-nitrosourea (ENU), which is the most efficient method for obtaining mouse mutations in phenotype-driven screens. A fractionated dose of ENU, an alkylating agent, can produce a mutation rate as high as 1.5 × 10(-3) in male mouse spermatogonial stem cells. Treatment with ENU produces point mutations that provide a unique mutant resource: They reflect the consequences of single gene changes independent of position effects, provide a fine structure dissection of protein function, display a range of mutant effects from complete or partial loss of function to exaggerated function, and discover gene functions in an unbiased manner. After treatment with ENU, mice are mated in genetic screens designed to uncover mutations of interest. Screens for dominant, recessive, and modifying mutations can be performed.
引言
本方案描述了使用N-乙基-N-亚硝基脲（ENU）对雄性小鼠进行化学诱变，这是在表型驱动筛选中获得小鼠突变的最有效方法。烷基化剂ENU的分次剂量可在雄性小鼠精原干细胞中产生高达1.5×10⁻³的突变率。用ENU处理会产生点突变，从而提供独特的突变资源：它们反映了单基因变化的后果，与位置效应无关，对蛋白质功能进行精细结构剖析，显示从功能完全或部分丧失到功能亢进的一系列突变效应，并以无偏见的方式发现基因功能。用ENU处理后，将小鼠用于旨在发现感兴趣突变的遗传筛选。可以进行显性、隐性和修饰突变的筛选。
