CAJ | 학술논문

The objective of this study was to establish an efifcient system of producing early monozygotic twin bovine embryosin vitro using the blastomere separation and coculture technique.In this study, early eight-cel embryos were chosen to op-timize the separation method, and multi-coculture tactics were applied to improve the efifciency of this production system. Bovine embryo blastomeres (groups of at least 30 at the eight-cel stage) were separated into eight segments (to regard an eight-cel embryo as a tangerine, a blastomere as one segment) and one, two and four segments (blastomeres) were cultured respectively in microwels on the bottom of the four-wel dish (Nunc, Denmark) with 400 μL of culture medium under parafifn oil.Four different types of coculture tactics (cocultured with nothing, intact embryos, bovine cumulus cels (bCCs), intact embryos & bCCs) were applied to the group of four segments (blastomeres).Finaly, diameter and inner cel mass (ICM):trophectoderm (TE) cel ratio was measured as a criterion to assess the quality of the twin embryos which were derived from bovine separated blastomeres.Our results showed that rate of blastocyst formation of the four segments group was signiifcantly greater than one or two group (P<0.05).In addition, rate of blastocyst formation was signiifcantly increased when the four segments were cocultured with intact embryo & bCCs (P<0.05).Although the ICM, TE and total cels of blastocysts derived from separated blastomeres was less than the control group from intact embryo (P<0.05), more important quality indicator of the blastocyst diameter and ICM:TE cel ratio was similar between our experimental group and the control group (P>0.05).Thus, these results suggest that combined with intact embryos & bCCs coculture system, culturing four isolated segments (blastomeres) per microwel is an efifcient system of producing early monozygotic twin bovine embryos.Furthermore, our results also indicate that the quality of blastocysts derived from separated blastomere may be similar to those derived from intact eight-cel embryos.