Studies on the differentiation of pigment cells with use of transgenic medaka fish bearing the gene for mouse tyrosinase

携带小鼠酪氨酸酶基因的转基因鱼是通过将其微注射到一种橙色的米达卡（Oryzias latipes）的卵母细胞核中或通过电穿孔将其引入受精卵中来生产的。我们成功地获得了41条棕色皮肤色素沉着的转基因鱼，前者获得了3条，后者获得了38条。研究发现，这些转基因鱼的棕色皮肤色素沉着源于“无色素黑色素细胞”的黑色素生成，这种色素分布在该物种的橙色变体中。利用抗小鼠酪氨酸酶抗体的免疫荧光揭示了小鼠型酪氨酸酶在这些黑素细胞中的明显分布。转基因鱼皮肤上的电子显微镜显示，黑素细胞安装了许多大小不均匀的黑素小体，具有晶格状的内部结构，这与鱼类黑色素细胞常见的“多泡”结构不同，并且在黄色细胞和白色细胞中检测到少量黑色素沉积。这些结果强烈提示转染的基因在medaka中以细胞类型特异性的方式表达。转基因鱼和来自橙色变异的鱼杂交产生的后代具有野生型棕色和橙色的色素，比例大约为1:1，这表明根据孟德尔定律，被转染的基因进行了传递。

Transgenic fish bearing the gene for mouse tyrosinase are produced by microinjecting it into the oocyte nucleus of or by introducing it by electroporation into the fertilized egg of an orange-colored variant of medaka (Oryzias latipes). We successfully obtain 41 transgenic fish exhibiting brown skin pigmentation, 3 fish by the former method and 38 by the latter. Brown skin pigmentation in these transgenic fish is found to result from melanogenesis in "amelanotic melanophores", which are distributing in an orange-colored variant of this species. immunofluorescence using the anti-mouse tyrosinase antibody discloses and apparent distribution of mouse-type tyrosinase in these melanophores. Electron microscopy on the skin of transgenic fish discloses that the melanophores are installed with numerous, unevenly sized melanosomes having a lattice-like internal structure, which is different from "multivesicular" ones common to fish melanophores, and that little melanin deposition is detectable in xanthophores and leucophores. All these findings strongly suggest that the gene transfected is expressed under a cell-type specific manner in medaka. Crossing between a transgenic fish and a fish from an orange-colored variant yields offsprings having wild-type brown and orange-colored pigmentation at an approximate rate of 1 : 1, suggesting transmission of the transfected gene according to the Mendelian law.