Nuclear transplantation by microinjection of inner cell mass and granulosa cell nuclei

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Abstract
The developmental potential of bovine inner cell mass (ICM) and somatic differentiated (granulosa cell) nuclei was investigated using nuclear transplantation. ICM blastomeres were isolated after immunosurgery of day 7 in vitro produced blastocysts and cumulus granulosa cells recovered from in vitro matured oocytes. Nuclear transplantation was carried out by microinjection of the lysed donor cells into enucleated mature oocytes. Oocytes were activated by three 0.2 kVcm−1/20 μs pulses in mannitol containing 100 μM Ca2+, with each pulse 22 min apart. Embryos were cultured in vitro for 7 days and blastocysts were transferred into recipients. ICM and granulosa cell donor nuclei directed 7% (20/304) and 9% (19/213) development to blastocysts, respectively. Fifteen blastocysts from ICM donors resulted in four pregnancies (27%) and two births. No pregnancy was detected with granulosa cell donors. The results illustrate the totipotency of ICM nuclei and indicate that granulosa cell nuclei promote preimplantation development of nuclear transplant embryos.