Grunwald D J, Streisinger G
Institute of Molecular Biology, University of Oregon, Eugene 97403.
Genet Res. 1992 Apr;59(2):103-16. doi: 10.1017/s0016672300030317.
Recessive lethal mutations and mutations at the gol-1 locus were induced in the zebrafish by exposure of mature sperm to the alkylating agent ethyl nitrosourea (ENU). Embryonic lethal phenotypes were recognized among the parthenogenetic progeny of mutagenized animals or among the progeny of daughters of mutagenized animals. Novel specific locus mutations were identified by the failure of mutagenized chromosomes to complement pre-existing mutant alleles at the gol-1 locus. Each mutagenized individual harboured approximately 10 embryonic lethal mutations in its germ line and about 1 in 500 mutagenized animals harboured a new mutation at the gol-1 locus. Three lines of evidence indicate that the majority of mutations that were recovered following treatment of mature sperm with ENU were probably point mutations. First, the soma and germ lines of mutagenized animals were mosaic, as expected following simple alkylation of sperm DNA. Second, mutations induced by ENU at the gol-1 locus affected pigmentation but not viability, unlike the majority of mutations induced at this locus with gamma-irradiation. Third, the ratio of specific locus:recessive lethal mutations induced by ENU was approximately 50-fold lower than the ratio observed following mutagenesis with gamma-rays. Comparison of the incidence with which embryonic recessive lethal mutations were induced with the incidence with which specific locus mutations arose indicates that there are greater than 5000 genes essential to the development and viability of the zebrafish embryo.
通过将成熟精子暴露于烷化剂甲基亚硝基脲（ENU），在斑马鱼中诱导出隐性致死突变和gol-1位点的突变。在诱变动物的孤雌生殖后代或诱变动物女儿的后代中识别出胚胎致死表型。通过诱变染色体不能与gol-1位点预先存在的突变等位基因互补来鉴定新的特异性位点突变。每个诱变个体在其生殖系中大约携带10个胚胎致死突变，并且大约每500个诱变动物中有1个在gol-1位点携带新突变。三条证据表明，用ENU处理成熟精子后恢复的大多数突变可能是点突变。第一，诱变动物的体细胞和生殖系是嵌合体，这与精子DNA简单烷基化后的预期一致。第二，与用γ射线在该位点诱导的大多数突变不同，ENU在gol-1位点诱导的突变影响色素沉着但不影响活力。第三，ENU诱导的特异性位点：隐性致死突变的比率比用γ射线诱变后观察到的比率低约50倍。将诱导胚胎隐性致死突变的发生率与出现特异性位点突变的发生率进行比较表明，斑马鱼胚胎的发育和活力有超过5000个必需基因。
