The applicant's long-term objectives are to develop technologies that improve or modify reproductive performance of mammals. This application deals with the development of the technology for making cloned and transgenic animals by nuclear transfer. The procedure of cloning by nuclear transfer involves removal of the recipient egg chromosomes, transfer of the donor nucleus, and artificial activation of the recipient egg. The limiting factor in cloning pigs by nuclear transfer has been the inability to artificially activate the unfertilized egg to resume meiosis. Recently, this hurdle has been overcome by modifying the culture conditions and the treatments used to activate the eggs. The applicants propose to: 1) Evaluate methods of cell cycle synchronization and egg activation for nuclear transplantation; 2) Produce piglets derived from nuclear transplantation; and 3) Genetically modify the cell lines and use them as donors for nuclear transplantation. The pig has become an important research model for many human diseases ranging from cardiovascular disease, and toxicology research, to intestinal physiology. Development of this technology in the pig will facilitate genetic knock-outs and knock-ins. Development of the nuclear transfer technology may have wide utility.
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