Development and gene expression of nuclear transplants generated by transplantation of cultured cell nuclei into non‐enucleated eggs in the medaka Oryzias latipes

• Published in: Development, growth & differentiation growth & differentiation, 2003-04, Vol.45 (2), p.167-174
• Main Authors: Ju, Bensheng, Pristyazhnyuk, Inna, Ladygina, Tatiana, Kinoshita, Masato, Ozato, Kenjiro, Wakamatsu, Yuko
• Format: Article
• Language: English
• Subjects: Animals; Animals, Genetically Modified; Animals, Newborn; Blastula - physiology; Cell Culture Techniques - methods; Cell Nucleus - physiology; Chromosome Mapping; clone; cultured cells; Female; Freshwater; Gastrula - physiology; Gene Expression Regulation, Developmental - genetics; Genes, Reporter; Green Fluorescent Proteins; Luminescent Proteins - genetics; medaka fish; non‐enucleated eggs; Nuclear Transfer Techniques; nuclear transplantation; Oryzias - embryology; Oryzias - genetics; Oryzias latipes; Ovum - physiology
• ISSN: 0012-1592; 1440-169X
• DOI: 10.1034/j.1600-0854.2004.00687.x

To develop nuclear transplantation techniques for the medaka Oryzias latipes, nuclei of cultured cells from transgenic fish were transplanted into unfertilized eggs of the orange–red variety of O. latipes, without enucleation, in two experimental series. In the first experimental series, fibroblast cells cultured from the adult caudal fin were used as donors, which carried the green fluorescent protein (GFP) gene driven by the promoter of the medaka elongation factor 1α‐A gene. Wild‐type body color was another donor genetic marker used in this experimental series. In the second experimental series, cells cultured from 6‐day‐old embryos were used as donors, which carried the GFP genetic marker driven by the promoter of the medaka β‐actin gene. From more than 1000 eggs transplanted in each experiment, a considerable number of nuclear transplants developed to various embryonic stages showing stage‐ and tissue‐specific expression of the donor genetic markers, although the expression was mosaic in many cases. Three and six of the transplanted eggs in the first and second experimental series (0.3 and 0.5%, respectively) hatched, and the hatchlings expressing the genetic markers survived for up to 3 weeks. The chromosome number varied among cells in a single transplant embryo. The results obtained in these experiments may help future cloning efforts in fish.