COMPOSITIONS OF SEQUENTIAL SYNTHETIC CULTURE MEDIA FOR IN VITRO HUMAN EMBRYONIC DEVELOPMENT AND METHODS USING THEREOF

摘要

A sequential synthetic culture media composition for in vitro embryo development is provided to supply more physiological nutrition to embryo growing at each stage of early embryo, morula, and late embryo and be able to maintain high pregnancy rate and implantation rate when the embryo is transplanted on the womb. The composition consists of a synthetic culture buffer solution(SCB) and a synthetic adding solution for early embryo growth(SAG13) in a volumetric ratio of 85:15. The SCB is prepared by dissolving 91.53mM of NaCl, 5mM of KCl, 0.8mM of MgSO4À7H2O, 1.80mM of CaCl2À2H2O, 0.17mM of Na2HPO4, 0.08mM of KH2PO4, 5mM of KHCO3, 20mM of NaHCO3, 0.1 mM of sodium pyruvate, 11.74mM of D,L-sodium lactate, 0.58mM of D-glucose, 15ml of SNEasS(x66.67), 15ml of SEaaS(x66.67), 0.611mM of L-alanyl-L-glutamine, 0.3594mM of glycyl-L-glutamineÀH2O, 0.003mM of D-Ca-pantothenate, 0.000716mM of choline chloride, 0.00027mM of folic acid, 0.0011mM of i-inositol, 0.000819mM of nicotinamide, 0.000491mM of pyridoxal HCl, 0.0000266mM of riboflavin, 0.000296mM of thiamine HCl, 0.1052mM of penicillin-G, 0.0172mM of streptomycin-S, and 0.003mM of phenol red in 1 liter of ultra-pure water. The SAG13 is prepared by dissolving 14.44mM of NaCl, 5mM of KCl, 3.5mM of MgSO4À7H2O, 1.80mM of CaCl2À2H2O, 0.17mM of Na2HPO4, 0.08mM of KH2PO4, 5mM of KHCO3, 20mM of NaHCO3, 1.57mM of sodium pyruvate, 73.47mM of D,L-sodium lactate, 0.05mM of D-glucose, 0.1mM of Na2EDTAÀ2H2O, 15ml of SNEasS(x66.67), 15ml of SEaaS(x66.67), 0.611mM of L-alanyl-L-glutamine 0.3594mM of glycyl-L-glutamineÀH2O, 1.18mM of L-taurine, 0.003mM of D-Ca-pantothenate, 0.000716mM of choline chloride, 0.00027mM of folic acid, 0.0011mM of i-inositol, 0.000819mM of nicotinamide, 0.000491mM of pyridoxal HCl, 0.0000266mM of riboflavin, 0.000296mM of thiamine HCl, 0.1052mM of penicillin-G, 0.0172mM of streptomycin-S, 0.003mM of phenol red, 42g/L of albumin, 8g/L of globulin 1g/L of hyaluronic acid, 0.000033g/L of insulin, 0.0000333g/L of transferrin, 0.0000000333g/L of sodium selenite, 0.00003g/L of epidermal growth factor, 0.00000278g/L of insulin-like growth factor-II, 0.76U/L, 5.24U/L of FSH, and 1g/L of gelatine.