Bovine And Mouse Oocytes Matured In Vitro

In vitro maturation is the process of gaining the oocytes from the mammal's ovary and culture in vitro into maturation.IVM is the fundamental technology of the trans-animal and the bacement of IVF. Although oocytes'nuclear can develop into maturation in vitro spontaneously, but the quality and the quantity of the matured oocytes in vitro is weaker than vivo and has a poor ability to develop into the blastula. The are lots of effects which caused the problem:the animals' age and health',the type of oocytes gained in vitro, the time and does of injection, the kinds of basic mediums, the hormone and growth factors added into the medium.To make a clear understanding of the IVM mechanism, we need to establish a system of the oocyte IVM.Bovine oocytes maturation technology is important on the agriculture and the IVM of mouse oocytes is also important as a mode animal.Objective:We try to find a proper system for the oocytes IVM of the mouse and bovine, and discuss the effects on the result of IVM,compare the maturation quality of DO and COCs at the same condition.Method and Results:1.Obtain bovine oocytes from the ovary with aspiration, rupture, aspiration and rupture, the result shows that, aspiration and rupture together is the best way to obtain bovine's oocytes.2.Use PMSG and hCG for stimulate and record the number of A,B,C oocytes, the result shows that the quantity of oocytes from the mouse without superovulation is low, it is necessary to inject PMSG.3.Use PMSG and hCG for stimulate, after 12h,24h,36h,48h collect oocytes and culture in vitro, the result shows that prolong the time of PMSG injection has no effects on the oocytes IVM,but peolong the time of hCG injection influence the maturation of oocytes.4.Use M199,M2 and DMEM as basic medium and added 10% FBS,culture B oocytes in vitro, the result shows that M199 is the best basic medium for the mouse oocyte maturation.5.Obtian A,B,C oocytes and culture in vitro, the result shows that the ability of maturation of the B oocytes is superior than A,C oocytes.6. Culture A,B oocytes and observe the number of oocytes on every level of cumulus expansion, the result shows that it is necessary that the cumulus expansion when the oocytes maturation, but the completely expansion is not necessary.7.Collect GV oocytes and get rid of the cumulus to obtain the DO and cuture DO and COC in vitro,the result shows that after oocytes IVM 4-10h, GVBD occurs and 8-14h PBI occurs.8.Use M199 as basic medium and add FSH at 0IU/mL 0.5IU/mL,1.0IU/mL,1.5IU/mL,2.0IU/mL, the result shows that cumulus can. stimulate the oocyte maturation when FSH added.9. Use M199 as basic medium and add LH at OIU/mL 0.5IU/mL,1.0IU/mL,1.5IU/mL,2.0IU/mL,the result shows that cumulus can stimulate the oocyte maturation when LH added.1.5IU/mLLH signamently enhace the mouse oocyte maturation. 10.Use M199 as basic medium and add E2 at.Ong/mL,5ng/mL,50ng/mL,500ng/mL,10000ng/mL the result shows that 50ng/mL E2 can enhance the mouse oocytes maturation and high does restrain maturation of oocytes.11.Use M199 as basic medium and add EGF at Ong/mL,20ng/mL,30ng/mL,40ng/mL,50ng/mL,100 ng/mL,40ng/mLEGF is the proper does to stimulate oocyte maturation, and the high does restrain maturation of oocytes.12.Collect COC, GC and DO after 24h in vivo and observe the expression of GDP-9 and BMP-15 with RT-PCR, the result shows that.the expression of BMP-15 in COC,DO and GC is equal, the result shows that expression of BMP-15 in COC are weaker than DO, while the expression of GDF-9 in DO and COC is equal,but in GC is weaker.