Saito Taiju, Goto Rie, Rivers Nicola, Yamaha Etsuro
South Ehime Fisheries Research Center, Ehime University, Ehime, Japan.
Education Program in Reproduction and Development, Department of Obstetrics and Gynaecology, Monash University, Clayton, Australia.
Methods Mol Biol. 2019;1920:327-341. doi: 10.1007/978-1-4939-9009-2_20.
The induction of germ-line chimerism in fish is a strategy for the reproduction of endangered or genetically valuable fish species. Chimeras can be created by transplanting a single primordial germ cell or multiple blastomeres from a donor into a sterile host embryo. When the host reaches sexual maturity, it will produce donor-originating gametes throughout its reproductive life span. This technique provides unique experimental conditions for basic biology research in model fish species like zebrafish. The success of cell transplantation relies on the effective sterilization of host embryos, the correct identification of developing germ cells, and the synchronization of migratory cues between the host and the transplanted cells. Developments in non-transgenic methods of germ cell ablation and identification have made germ cell transplantation more applicable to use in conservation and aquaculture. In this chapter, we provide a protocol for germ cell labeling by injection of chimeric RNA or FITC-dextran, the sterilization of host embryos using an antisense morpholino oligonucleotide, and two methods for producing germ-line chimeras in zebrafish: single primordial germ cell transplant and blastomere transplant.
在鱼类中诱导生殖系嵌合体是繁殖濒危或具有遗传价值鱼类物种的一种策略。嵌合体可以通过将单个原始生殖细胞或多个卵裂球从供体移植到无菌宿主胚胎中产生。当宿主达到性成熟时，它将在整个生殖寿命中产生源自供体的配子。这项技术为斑马鱼等模式鱼类的基础生物学研究提供了独特的实验条件。细胞移植的成功依赖于宿主胚胎的有效绝育、发育中生殖细胞的正确识别以及宿主与移植细胞之间迁移信号的同步。生殖细胞消融和识别的非转基因方法的发展使生殖细胞移植更适用于保护和水产养殖。在本章中，我们提供了通过注射嵌合RNA或FITC-葡聚糖进行生殖细胞标记的方案、使用反义吗啉代寡核苷酸对宿主胚胎进行绝育的方案，以及在斑马鱼中产生生殖系嵌合体的两种方法：单个原始生殖细胞移植和卵裂球移植。
