Parthenogenesis is maternally uniparental reproduction through the embryonic development of oocytes without fertilization. Artificial activation of mature oocytes could generate homozygous haploid embryos with the extrusion of the second polar body. However, the haploid embryos showed low embryo development in preimplantation embryos. In this study, we investigated whether the electronic fusion of the haploid embryos could enhance embryo development and ESC establishment in mice. Haploid embryos showed the developmental delay from 4-cell to the blastocyst stage. The haploid blastomeres of the 2-cell stage were fused electronically, resulting in that the fused embryos showed a significantly higher rate of blastocysts compared to non-fused haploid embryos (55% vs. 37%). Further, the embryonic stem cells (ESCs) derived from the fused embryos were confirmed to be diploid. The rate of ESC establishment in fused embryos was significantly higher compared to non-fused ones. Based on the results, we concluded that the electronic fusion of haploid embryos could be efficient to generate homozygous ESCs.

Somatic cell nuclear transfer derived embryonic stem cells (NT-ESCs) have significant advantages in various fields such as genetics, embryology, stem cell science, and regenerative medicine. However, the poor establishment of NT-ESCs hinders various research. Here, we applied fasudil, a Rho-associated kinase (ROCK) inhibitor, to develop somatic cell nuclear transfer (SCNT) embryos and establish NT-ESCs. In the study, MII oocytes were isolated from female B6D2F1 mice and performed SCNT with mouse embryonic fibroblasts (MEFs). The reconstructed NT-oocytes were activated artificially, and cultured to blastocysts in KSOM supplemented with 10 μM fasudil. Further, the blastocysts were seeded on inactivated MEFs in embryonic stem cell medium supplemented with 10 μM fasudil. A total of 26% of embryos formed into blastocysts in the fasudil treated group, while this ratio was 44% in the fasudil free control group. On the other hand, 30% of blastocysts were established NT-ESCs after exposure of fasudil, which was significantly higher than the control group (10%). The results suggest that fasudil reduced blastocyst development after SCNT due to inhibition of 2 cell cleavage while improved the establishment of NT-ESCs through the anti-apoptotic pathway.