108. INTERSPECIES SOMATIC CELL NUCLEAR TRANSFER IS DEPENDENT ON COMPATIBLE CYTOPLASMIC FACTORS AND MITOCHONDRIAL DNA
Y. Jiang B , R. Kelly A B , A. Peters B , H. Fulka B , D. A. Mitchell A and J. C. St John A BA Centre for Reproduction and Development, Monash Institute of Medical Research, Clayton, VIC, Australia.
B Clinical Sciences Research Institute, University of Warwick, Coventry, UK.
Reproduction, Fertility and Development 22(9) 26-26 https://doi.org/10.1071/SRB10Abs108
Published: 6 September 2010
Abstract
Interspecies somatic cell nuclear transfer (iSCNT) offers significant opportunities to analyze and understand nuclear-cytoplasmic interactions. Using a murine-porcine interspecies model, we investigated the importance of nuclear-cytoplasmic compatibility, specifically mitochondrial DNA (mtDNA), on successful development. Transfer of somatic murine fetal fibroblasts into enucleated porcine oocytes resulted in extremely low blastocyst rates (0.4%); increased DNA strand breaks; deficient nuclear pore complex arrangements and increased aberrant karyokinesis than observed in porcine-porcine SCNT embryos. Using allele specific-PCR analysis, murine mtDNA was detected at ever-decreasing levels to the blastocyst stage, with peak levels being 0.14 ± 0.055% in 2-cell embryos. Furthermore, these embryos reduced total mtDNA copy number during preimplantation development in a manner similar to porcine embryos. Injecting mouse embryonic stem cell extract and mitochondria along with the murine donor cell into a mitochondria depleted porcine oocyte, increased blastocyst zona pellucida thinning and blastocyst rates significantly (0.4% vs 3.42%) compared to the non-supplemented iSCNT group. They also had significantly more murine mtDNA at the 2-cell stage than the non-supplemented embryos, which was maintained throughout preimplantation development. At later stages of preimplantation development, they possessed 48.00% ± 17.38% murine mtDNA and exhibited a mtDNA copy number profile similar to murine embryos. Overall, these data demonstrate that the addition of species compatible cytoplasmic factors and mitochondrial DNA improve developmental competence of iSCNT embryos.