Factors affecting in vitro embryo production and cryopreservation in cattle

In this dissertation, fundamental biological questions were investigated with the aim to identify improved culture conditions for in vitro produced cattle embryos. Such culture conditions include defined/semi-defined and sequential media such as KSOM, SOF and KSOM-SOF which might ultimately affect embryo development, sex ratio, and viability following cryopreservation or transfer into recipients. The effect of modified IVF media on post-fertilization embryo development, duration of sperm-oocyte co-incubation, and on blastocysts\u27 viability following vitrification was also investigated. Although in vitro production of mammalian embryos has led to the use of several culture media, combinations of culture media such as KSOM and SOF without co-culture have not been systematically tested previously. The rate of embryo development in KSOM or KSOM-SOF was investigated to determine if they affect blastocyst survival rate post-vitrification, and whether male and female embryos survive vitrification differently. This research has clearly demonstrated that IVF media affects post-fertilization embryo development and blastocyst survival post-vitrification. The modified M 199 (m-M199) compares favorably to Brackett and Oliphant (BO) medium and may offer an advantage for 18 h extended IVF period. Embryos produced from cell-free sequential culture media (KSOM-SOF) survived vitrification without compromising their quality. However, male embryos reach blastocyst stage earlier, and survive vitrification better, than their female counterparts. This potential skewing of sex ratio ought to be taken into account when preserving beef or dairy blastocysts, where the sex of the progeny is of major economic importance. Embryo sexing, prior to cryopreservation or transfer, might provide one solution by offering selection based upon sex.