Zimmer A, Gruss P
Max-Planck Institut für biophysikalische Chemie, Abt. Molekulare Zellbiologie, Göttingen, FRG.
Nature. 1989 Mar 9;338(6211):150-3. doi: 10.1038/338150a0.
Several mouse gene families related to Drosophila developmental control genes and containing a homoeobox, a paired box or a finger domain, have been cloned and structurally analysed. On the basis of structural similarities to the Drosophila genes and of their spatially and temporally restricted expression patterns during mouse embryogenesis, it has been proposed that these mammalian genes also are involved in the control of development. To elucidate the function of homoeobox genes by genetic means, mouse mutants must be generated. We have developed a technique for mutagenesis in vivo and have used it to mutate the homoeobox Hox 1.1 gene. In vivo mutagenesis was achieved through homologous recombination between an endogenous Hox 1.1 allele and a microinjected mutated gene in pluripotent embryonic stem (ES) cells. Mutant cells were identified by means of the polymerase chain reaction (PCR) and mutant clones were used to generate chimaeric mice. Because the homologous recombination event is formally a gene conversion event and no selection is required to screen for cells carrying the mutated allele, in vivo mutagenesis allows specific alterations in the target sequence to be made without the introduction of any other sequences.
已经克隆并对几个与果蝇发育控制基因相关且包含同源异型框、成对盒或指状结构域的小鼠基因家族进行了结构分析。基于与果蝇基因的结构相似性以及它们在小鼠胚胎发生过程中在空间和时间上受到限制的表达模式，有人提出这些哺乳动物基因也参与发育控制。为了通过遗传手段阐明同源异型框基因的功能，必须产生小鼠突变体。我们开发了一种体内诱变技术，并将其用于使同源异型框Hox 1.1基因发生突变。体内诱变是通过多能胚胎干细胞（ES细胞）中内源性Hox 1.1等位基因与显微注射的突变基因之间的同源重组实现的。通过聚合酶链反应（PCR）鉴定突变细胞，并使用突变克隆来产生嵌合小鼠。由于同源重组事件实际上是一个基因转换事件，并且不需要选择来筛选携带突变等位基因的细胞，因此体内诱变允许在不引入任何其他序列的情况下对靶序列进行特异性改变。
