Development of a detection system for germ-cell mutagenesis with high sensitivity and multiple genetic endpoints using the Japanese Medaka

本研究的目的是开发一种高灵敏度、多遗传终点的日本稻属植物生殖细胞诱变检测系统。大约十年前，我们开始开发一种非哺乳动物测试系统，用于使用日本Medaka检测环境生殖细胞诱变。1994年中期，我们通过20多代的兄弟姐妹重复交配，成功地建立了具有3个标记位点（b/b; gu/gu; lf/lf）的自交系试验品系AA2。到1995年2月8日，我们已经检查了872961个胚胎，对应了2299892个基因座。其中对照组有195,554个胚胎，对应于501,917个位点。自发性胚胎死亡率为5.6%，几乎与报道的小鼠（5.8%）相当。自发总突变率为3.8x10-5/座，自发活突变率为4.2x10-6/座。在目前的测试系统中，我们成功地将显性致死突变、总突变、活活突变和外部畸形这四个遗传终点整合到一个个体中，尽管外部畸形的定量不如其他三个终点成功。这些遗传终点对伽马射线和裂变中子辐射表现出良好的剂量-响应关系。此外，无论辐照时雄性生殖细胞成熟阶段的差异如何，总突变率降至可活突变率的程度约为90%。然而，男性生殖细胞暴露于ENU后，根据成熟阶段和ENU浓度的不同，会导致不同的减少。

The purpose of the present study is to develop a detection system for germ-cell mutagenesis with high sensitivity and multiple genetic endpoints using the Japanese Medaka Oryzias latipes. About ten years ago, we started developing a non-mammalian test system for detecting environmental germ-cell mutagenesis using the Japanese Medaka. In the middle of the FY 94, we succeeded in establishing a inbred tester strain with three marker loci (b/b ; gu/gu ; lf/lf) , named AA2 inbred strain, by repeating brother-sister mating for more that 20 filial generations. By February 8,1995, we have examined 872,961 embryos that correspond to 2,299,892 loci. Among these were the control with 195,554 embryos corresponding to 501,917 loci. The rate for spontaneous embryonic death was 5.6%, a value almost comparable to the reported one in the mouse (5.8%). The spontaneous total mutation rate and the spontaneous viable mutation rate were, respectively, 3.8x10-5/locus and 4.2x10-6/locus. In the present test system, we succeeded in integrating four genetic endpoints, i.e., dominant lethal mutation, total mutation, viable mutation and external malformation, into one individual, although the quantitation of external malformations was not as successful as the remaining three endpoints. These genetic endpoints showed good dose-response relationships for gamma-rays and fission neutron radiation. Furthermore, the extent of reduction of total mutation rate down to viable mutation rate was about 90%, irrespective of the difference in stages of maturation of male germ cells at the time of exposure to radiations.However, exposure of male germ cells to ENU resulted in different reduction depending on the maturation stages as well as ENU concentration.