P-86: Production of Cloned Mice by Somaticm Cell Nuclear Transfer

Background: For several years, mammalian cloning by splitting an early embryo or nuclear transfer into oocytes method has been successfully performed. Cloning is now also possible using adult somatic cells. Although it has now been 15 years since the first cloned mammals were generated from somatic cells using nuclear transfer (NT), the success rate for producing live offspring by cloning is low, regardless of the cell type and animal species used. Nevertheless, the techniques have potential as important tools for future research in basic biology. Materials and Methods: In this study we have been used a stable NT method in the mouse, in which donor nuclei are directly injected into the oocyte using a piezoactuated micromanipulator. Female B6D2F1 induced to superovulate by consecutive injection of PMSG and HCG. 14-15 hours after HCG injection, cumulus-oocyte complexes were collected from oviducts and treated in M2+Hy medium. Then After enucleation of metaphase II oocytes, nucleus donor cells (Cumulus cells) were injected into separate enucleated oocytes. Following somatic-cell nucleus injection, groups of oocytes were placed in activation medium and incubation was continued at 37.5°C under 5% CO2. Where appropriate, 2-cell embryos were respectively transferred into oviducts of Pseudo-pregnant mice. Finally at 19 day after transfer embryos, live Pups were raised. The embryos for better nutrition were transferred in mouse cage, which coincided with the birth of cloned embryos. Results: Since the origin of isolated cumulus cells was black mice B6D2F1, the cloned mice were also black, confirming that the process of cloning was performed correctly. Conclusion: A somatic cell nuclear transfer technique could have applications in of cloning therapy, regenerative medicine and creation individual specific embryonic stem cells.