Developmental analyzes of primordial-germ cells and induction of germ-line chimera in fish

1)采用全载原位杂交技术分析金鱼胚胎发生过程中输精管mRNA的分布。金鱼和斑马鱼的vas mRNA分布有一定的差异。2)在金鱼中胚层缺陷胚中分化出功能性PGCs。3)金鱼在原肠形成过程中，PGCs通过收敛和伸展运动进行被动迁移。4)将含PGCs的移植物移植到囊胚期的宿主胚胎中，成功地诱导了种系嵌合体。5)供体精子仅在生殖系嵌合体中以不育杂交种作为寄主胚胎产生。6)建立了gfp标记PGCs的转基因鳟鱼品系。7)建立了一种流式细胞仪纯化活PGCs的技术。8)采用cDNA减法分离PGCs中特异性表达的基因。9)利用分离的gfp标记的PGCs进行PGCs移植技术是为了建立一种将供体PGCs重新引入种系到受体鱼的程序。

1) Distribution of vas mRNA were analyzed by whole mount in situ hybridization during embryogenesis in goldfish. There are some difference of distribution of vas mRNA between goldfish and zebrafish.2) In goldfish, functional PGCs were differentiated from mesoderm deficient embryo.3) In goldfish, PGCs migrated passively by convergence and extension movement during gastrulation.4) Germ-line chimera was successfully induced by transplantation of graft, containing PGCs, to host embryo at the blastula stage.5) Sperm derived from donor was only generated in germ-line chimera by using sterile hybrid as host embryo.6) Transgenic trout strains having GFP-labeled PGCs were established.7) A technique to purify live PGCs using a flow cytometer was developed.8) Genes expressed specifically in PGCs were isolated by the cDNA subtraction method.9) A technique for PGCs transplantation using isolated GFP-labeled PGCs was developed in order to establish a procedure for re-introducing donor PGCs in to the germline to recipient fish.