In Vitro Fertilization Of Kongming Mouse

In vitro fertilization and the culture of fertilized eggs have been widely used in the field of medicine and biology, but still need further optimizing. Kunming mouse(KM), bred in China by near inbred, has been an ideal laboratory animal model because of the well-known inheriting background, short reproduction cycle and the strong environmental adapt ability. The purpose of this study is to establish optimal, effective system for in vitro fertilization and embryo culture and improve the quality of embryo. In these experiments, we studied the in vitro fertilization of KM mouse from five aspects of screening the optimum embryos culture medium, effecting of heparin on sperm, the effect of different capacitation solution on rate of vitro fertilization, the effect of different culture media on the development of Kunming mouse embryos, the effect of different culture systems on the development of Kunming mouse embryos. And the result showed that:1. G1/G2 and KSOM medium containing 10%NBS could support 2-cell embryos to development blastocyst (87.08% and 89.52%).The development rates of blastocyst in KSOM containing 10%FBS was significantly higher than that in the media containing BSA, the difference were extreme(P<0.01).2. To screen the optimum heparin concentrations in the capacitation solution HTF of mouse. The rate of fertilization in vitro in the media without heparin (41.75%)was significantly higher(P<0.01) than those with heparin at 1μg/mL (31.00%),3μg/mL(18.80%),5μg/mL(22.43%).3. The rate of vitro fertilization of HTF (MR-070,85.71%) which was foreign imported reagent was significant higher than those of HTF,T6 and Whittingham (38.58%,10.22% and 24.64%) self-prepared. The fracture rate of fertilized eggs in HTF was lower (3.57%) than those three other self-made media (23.62%,20.44%,32.61%).4. The rate of blastocyst in G1/G2 was 77.50%, which was higher than imported KSOM(22.78%),and self-made KSOM (0).5. Granulosa cell co-cultured with mouse embryos could support IVF embryos to overcome 2-cell block, and the rates of 4-cell(37.97% vs 59.72%),morula(27.85% vs 48.61%),blastocyst (22.70%vs45.83%)could be all improved significantly than non-culture system.