| Blastocysts of mouse and porcine were recovered and vitrified in this research, the experiment was carried out as the following: 1) study on vitrification of mouse blastocysts and expanded blastocysts; 2) study on post-vitrification mitochondrial injury of mouse blastocysts and expanded blastocysts; 3)study on super-ovulation of porcine;4)study on vitrification of porcine blastocysts and expanded blastocysts. Vitrification and post-vitrification mitochondrial injury of mouse embryos were studied to provide inference for subsequent study on vitrification of porcine embryos.1 Study on vitrification preservation of mouse blastocysts and expanded blastocystsCryoprotective solution containing 6M GLY+6.5%BSA(V1)and 8MEG+7%PVP(V2) and methods of straw and one-step OPS vitrification were used. Results showed that embryos which equilibrated for 20s has significantly higher rate of viability than those for 60s(P<0.05).Two cryopreservation solutions(Vl and V2) has no significant effect on developmental rate(P>0.05).Different thawing methods in straw vitrification, including two-step, three-step and four-step dilutions, were compared and results showed that four-step dilution as well as three-step dilution achieved significant higher developmental rate than two-step dilution(P<0.05).Freezing methods affected developmental rate. In this paper, three-step vitrification (V2)was found to be very significantly higher than straw vitrification and one-step OPS vitrification in developmental rate(P<0.01).Equilibrium time affected developmental rate in three-step OPS vitrification. During three-step OPS vitrification using V1 solution, 10-15 minutes equilibrium was positively compared to 20 minutes(P<0.05) , similarly, when using 2M V2 solution, 3-4minutes equilibrium was found significantly better than 5 minutes(P<0.05).Additionally, developmental stage was compared and no significant effect was found ( P>0.05 ) .2 Study on mitochondrial congregation of mouseIn this experiment, Rhodaminel23 molecular fluorescent staining was used to study mitochondrial congregation of mouse blastocysts in V1 solution and expanded blastocysts in V2 solution. Mitochondrial congregation was observed at oh, 4h, 8h, 12h, 18h and 24h and results indicated: regardless of using V1 or V2 solution, the same descent trend was found in rate of normal congregation as culture time went.3 Study on factors affecting super-ovulation of porcineEffect of breed, hormone dose, and super-ovulation times on super-ovulation of white-Large and Landrace was studied in this experiment. Results turned out: in case of number of CL, Beijing Black was significantly higher than White-Large and Landrace (P<0.01); while Landrace was significantly higher than White-Large (P<0.05).in case of viable embryos, Beijing Black significantly higher than Large-White (P<0.01) and Landrace (P<0.05).Two hormone doses were compared in number of CL and viable embryos, results showed that 1450 IU PMSG was significantly higher than 1025 IU PMAG(P<0.01).Additionally, super-ovulation times was compared and significantly less CL in repeated super-ovulation group than one-time super-ovulation(P<0.01).4 Study on cryopreservation of porcine embryos 'Porcine blastocysts and expanded blastocysts were used in this experiment. After treated with CB, embryos was frozen using conventional slow freezing, straw vitrification and OPS vitrification.In conventional slow freezing with 1.5M GLY as cryoprotectant, 25.38% and 20.41% developmental rate was achieved in blastocysts and expanded blastocysts respectively, and no significant difference was found ( P>0.05 ) .Expanded blastocysts were treated with CB in V1 and V2 solutions and subsequently vitrified using OPS method, results showed that no significant difference was found between two groups (P>0.05 ) ,but two groups were significantly worse than control ( P>0.01 )Effect of centrifugation on developmental rate of blastocysts using straw vitrification was studied and results showed that centrifugation significantly improved developmental rate ( P<0.01 ) .Furthermore, e... |
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