Bovine imprinted gene analysis in assisted reproduction techniques

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The production of bovine embryos through in vitro maturation and fertilization is becoming an important tool of the genomic revolution in dairy cows. The capacity to select embryos based on the father and the mother genotypes and the possibility to do that at the blastocyst level before a pregnancy is initiated is changing the way animals are produced and selected. The molecular analysis of these embryos indicate some differences according to the age of the parent and according to the culture or hormonal environment. Such influences may be temporary but in some case, it may influence the post-natal phenotype suggesting that more research is required to understand the changes observed. This project is based on the hypothesis that some of the methylation patterns are acquired in the last portion of folliculogenesis and could be influenced by the environment created to generate these oocytes. Such differences may not be erased during the first week of culture or sensitive to the conditions during this time. To test this hypothesis, 32 embryos (day 12) obtained from pre-puber donors (8)will be compared to embryos obtained from adults and produced in vitro with (8) or without serum (8) or in vivo (control)(8). The DNA from these embryos will be extracted and bisulfited to reveal methylation marks. The analysis of 10 imprinted genes will then be compared across treatments to assess if the pattern is modified and if all embryos are affected or only a subset. The second part will assess the methylation level of the 4 best imprinted indicator in 48 blastocysts and biopsies to develop a rapid day 7 test permitting the monitoring of any modification to the culture conditions or ovarian stimulation. This work will generate markers to assess the required hormonal and culture environment to minimize epigenetic perturbation in bovine embryos generated by ART.