A MUTATION DETECTION SYSTEM FOR MAMMALS AT INDIVIDUAL LEVEL

在个体水平上定量检测哺乳动物的致突变作用，对于评价环境致突变剂、致癌物以及近年来在现代生活中使用的遗传毒性物质对人类的影响具有重要意义。然而，需要数十万只小鼠来进行突变频率的定量实验。因此，我们尝试建立一个只需要少量动物就能在个体水平上检测突变的检测系统，该系统是将噬菌体遗传学和转基因小鼠系统相结合构建的。将E. coli中导入λ噬菌体EMBL3。当与它们的蛋白质混合时，λ噬菌体的DNA可以在体外包装以产生λ噬菌体颗粒。用适当的E.将纯化的含supF的λ噬菌体DNA注射到小鼠受精卵中，建立转基因小鼠。以supF基因为探针，对从小鼠尾部提取的DNA进行Southern分析，检测发育小鼠中转基因的存在。到目前为止，我们已经获得了几个品系的转基因小鼠。我们现在正在通过限制性酶切分析来检查转基因小鼠中转基因的组织。一旦建立了具有适当特性的菌株，将来自小鼠精子的DNA置于体外包装系统中，以分析体内发生的supF基因突变。

Quantitative assay for mutation induction in mammals at individual level is of importance to evaluate the effect on humans of environmental mutageiis and carcidogens as well as genotoxic agents, which have been recently used in modern living. Over hundreds thousands of mice are required, however, to carry out experiments for quantitation of mutation frequencies. We therefore tried to develop an assay system in which only a small number of animals are required for the detection of mutation at the individual level.The system has been constructed by combination of phage genetics and transgenic mouse system. As a marker gene, supF gene of E. coli was introduced into lambda phage EMBL3. When mixed with their proteins, the DNA of lambda phage can be packaged in vitro to yield lambda phage particles. Mutation in supF gene in the lambda phages is easily detected by using a proper E. coli as an indicator cells.Purified DNA of the lambda phage containing supF was injected into fertilized eggs of mice to establish a transgenic mouse. Presence of the transgene in the developed mice was examined by southern analysis of the DNA extracted from tails using supF gene as a probe. To date, we have obtained several strains of transgenic mice. We are now examining the organization of the transgene in the transgenic mice by restriction analysis. Once a strain with appropriate properties has established, DNA from sperms from the mice is subjected to the in vitro packaging system to analyze the mutation in supF gene occurred in vivo.