Roberts J M, Axel R
Cell. 1982 May;29(1):109-19. doi: 10.1016/0092-8674(82)90095-2.
We used gene transfer to identify frequent genetic rearrangements responsible for activating mutant genes in mammalian cells. We transformed an aprt- tk- cell with a plasmid containing a wild-type aprt gene and a truncated, promoterless tk gene. Transformants that integrate a single copy of this plasmid exhibit the aprt+ phenotype but remain tk-. Tk+ variants result from 20 to 50 fold amplification of the linked plasmid along with significant lengths of flanking DNA. They produce aberrant transcripts such that multiple genes are required to generate sufficient enzyme to convert the cell to the tk+ phenotype. One striking feature of the amplified aprt+ tk+ clones is the frequency (10(-4) ) at which aprt- tk+ mutants appear. These phenotypic requirements are such that the tk gene must remain amplified while all the amplified aprt genes become inactivated. The structure of the amplified DNA indicates that within aprt- cells, all amplified units bear identical mutations. These data suggest that these cells possess an efficient correction mechanism that maintains sequence homogeneity among repeated genetic elements.
我们利用基因转移技术来鉴定导致哺乳动物细胞中突变基因激活的常见基因重排。我们用一个含有野生型 aprt 基因和一个截短的、无启动子的 tk 基因的质粒转化 aprt- tk- 细胞。整合了该质粒单拷贝的转化体表现出 aprt+ 表型，但仍为 tk-。Tk+ 变体是由连接的质粒 20 至 50 倍的扩增以及相当长度的侧翼 DNA 产生的。它们产生异常转录本，以至于需要多个基因来产生足够的酶将细胞转化为 tk+ 表型。扩增的 aprt+ tk+ 克隆的一个显著特征是 aprt- tk+ 突变体出现的频率（10(-4)）。这些表型要求使得 tk 基因必须保持扩增状态，而所有扩增的 aprt 基因都失活。扩增 DNA 的结构表明，在 aprt- 细胞内，所有扩增单元都带有相同的突变。这些数据表明这些细胞拥有一种有效的校正机制，可维持重复遗传元件之间的序列同质性。
