Study On The Fertilization In Vitro Of Laboratory Rats

Rats are one of the most important laboratory animals in the research of life sciences, the technique of in vitro fertilization in rats not only provides an economical way to preserve a large number of rats strains, but aslo can promote the research of procreation field, such as the mechanism of molecule, the fetation and so on. It was significant either in practical use or in theory. The contents and results of experiments as follows:1. The effects of PMSG/hCG dosages on superovulation in immature rats were investigated. Results showed that rats at 4-5 weeks of age are more sensitive to superovulation, which can obtain the highest numbers of oocytes with 20 IU PMSG and hCG, and the average number was 54.6. It's much more higher than other combinations.2. Vaginal smear method was used to check different stage of estrus cycle in rats, and the changes of vaginal epithelium cells were analysed. Results showed that vaginal epithelium cells changed regularly in different estrus cycle. Most of the cells were white cells in diestrus, stoned epithelium cells were the main cells in proestrus, keratinized epithelium cells in estrus, and three kinds of cells in metestrus.3. The technique of embryo transfer in rats was studied, and the quality of oocytes obtained by different methods were compared. Results showed that oocytes obtained from superovulation have similar development abiliy compared with oocytes obtained from natural ovulation, and the rate of pregnancy showing no significant difference after embryo transfer(63.3% VS 60%).4. Three mediums, which were mKRB, IVF30 and mR1ECM-BSA were used in the IVF in rats. Results showed that the fertilization rate was 73.8% when used IVF30 as capacitation medium, showing significant difference compared with mR1ECM-BSA and mKRB(13.5% VS 21.1%).5. Some pivotal factors on the results of IVF were observed. Results showed that SD could also obtain cleavage without capacitation in the medium IVF30, and the fertilization rate was 33.8%. Fertilization rates became higher and higer along with prolongs of capacitation time in certain scope, and obtained the highest rate after capacitating 7 hours(83.2%), showing significant difference compared with other time; Collected oocytes at different time after given hCG, then fertilization in vitro, the results showed the best collecting time of IVF in SD was 13 h whose fertilization rate was 88.6%, the fertilization rate descended along with the time prolonging. The fetilization rate was 70.9%, 78.6% and 73.8% respectively after oocytes encountered sperm 6, 9 and 12 hours, showing no significant difference.6. The effects of BSA, PVA and Glocose on the in vitro development of zygotes in rats were studyed. Results showed BSA and PVA have no significnat difference on the zygotes development before 4-cells, but PVA could support 21.4% zygotes developed to blastula, while there was none zygote developed to blastula using the medium contained BSA; While glocose was essential to the development of zygotes in vitro, and the rate of blastula was higher in the medium containing 5.5 mM or 7.5 mM glocose than containing 0.2 mM glocose(23.5%, 21.4% and 4.1% respectively).7. The IVF on more rat strains using IVF30 medium were studied, and the 2-cell embryos obtained from IVF were transferred to recipients to observe the development ability. Results showed that IVF30 medium also suitable for the IVF of Wistar, GK and F344 rats, and the fertilization rate was 72.6%, 87.8% and 71.6% respectively, but they showed no siginificant difference. After embryo transfer, the rate of pregnancy was 12.5%, it's lower than contrast(60%).