Department of Animal Science

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Browsing Department of Animal Science by Subject "636.08245"

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    Effect of different culture media and incubation methods on culturing murine embryos in vitro using a semen straw as an alternative receptacle
    (2016-05) Madzhie, Lufuno Rosheen; Barry, D. M.; Nedambale, T. L.
    The aim of the study was to fertilize mouse oocytes and culture the resulting zygotes in bi-gas incubator and in a goat vagina and compare the in vitro embryo development in TCM-199 and Ham's F10 culture media until the blastocyst formation. The spermatozoa and oocytes of an F1 generation (Balb C x C57) were fertilized in Ham's F10 and TCM-199 in semen straws and in micro-drops in petri dishes. The embryos were cultured in TCM-199 and Ham's F10 in semen straws and micro-drops in petri dishes. The semen straws were incubated in a bi-gas incubator and goat vagina. The blastocyst-stage embryos were stained using Hoechst 33528 solution and blastomeres count. The results were analyzed by 2 X 3 factorial design and the student t-test was used to separate the mean and they showed that there was no statistica! difference (p > 0.05) between the media, receptacles and incubators. The overall fertilization rate was 94 % to 99 %. The semen straw with Ham's F10 incubated in the bi-gas incubator had the highest rate (80.5 %), reaching the blastocyst-stage of the embryos. The lowest rate of murine embryos reaching the blastocyst-stage were these cultured in semen straws incubated in the gaat vagina, with its highest rate of 60 %. The embryos blocking at the 8 - cell and morula stage recorded the lowest rate on bath culture media and incubation methods, compared to the ether embryo developmental stages. The overall mean number of blastomeres in the blastocyst-stage of the embryos ranged trom 85±9 - 90±9 cells in all receptacles and incubators. lt was concluded that the fertilization and culturing of murine embryos are possible in French semen straws incubated in a bi-gas incubator and in the gaat vagina as an alternative method of fertilizing oocytes and culturing murine embryos in addition TCM-199 and Ham's F10 can bath be used to fertilize oocytes and culture murine embryos until blastocyst formation embryo in vitro, incubated in a bi - gas incubator or in the vagina.
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    ItemOpen Access
    Effect of in vitro culture media and assisted hatching techniques on mice embryo survival rate following cryopreservation
    (2018-05-18) Serota, Nthabiseng Ruth; Nedambale, T. L.; Fushai, F.
    This study determined the effects of in vitro culture media (Ham’s F10 and TCM199) and assisted hatching techniques (laser or mechanical) on mice embryo survival following cryopreservation. Pure strain C57BL/6 (B6) female (50) and strain BALB /c (C) Male (25) mice were crossed to produce F1 generation of females which were injected for follicular growth and super ovulation at 6 weeks of age and from which embryos were produced 21 h later through in vivo fertilization. Embryos were randomly divided into Petri dishes with different culture media, and the development of embryos was assessed until the morula stage. At the morula stage, selected embryos were assisted to hatch using different techniques, and then cryopreserved in liquid nitrogen using the slow freezing method for a period of 1 week. After 1 week of cryopreservation, the embryos were thawed and cultured in the two different in vitro culture media for 72 hours. Thereafter, the numbers of embryos hatched or survived were recorded after 24 h, 48 h and 72 h. Data was analyzed using ANOVA in Minitab Software Version 16 (2010). Significant difference in embryo quality development was observed between in vitro culture media and stage of embryo development (P<0.05). In the TCM-199 in vitro culture medium, embryo quality development yielded 72, 69 and 69% from day 1 to day 3, while in Ham’s F10 embryo quality development yielded 68, 63 and 60% respectively. Relative to the control (18.1%) assisted hatching improved hatchability significantly (P<0.05) in the order laser (23.6%)>, mechanical (20.8%). There was significant (P<0.01) interaction between assisted hatching techniques and evaluation time, whereby laser assisted hatching was most successful at 48 h (42.0%) while mechanical assisted hatching was most successful at 72 h (36.8%). Cryopreservation reduced the embryo survival compared to fresh embryos. In conclusion laser was the best assisted hatching technique, while TCM-199 was the better medium for in vitro culture of embryo.