Abstract: The present study was undertaken to establish the in vitro production system of sexed embryos of dairy cattle by optimizing culture system of the bovine embryo production, such as fertilization and embryo culture in vitro. The results showed that oocytes maintained at least three layers of cumulus were suitable of the karyoplasms maturation in matured medium added 10 ng/ml EGF during the maturation in vitro; the survival time of the sperm was decreased by the addition of caffeine. Embryos cultured with CRlaa added of BSA, followed in SOFaa with 5%FBS could develop high rate of blastocyst, and the embryo developmental percentage could be improved by the addition of 50ng/ml IGF-I. There was no direct influence on the developmental quality of the embryos fertilized in vitro with sexed semen although the fertilization rate had a slight drop (P<0.05), and the pregnant rate was about 40% after the embryos transferred.