Bovine imprinted gene analysis in assisted reproduction techniques

辅助生殖技术中的牛印记基因分析

• 批准号: 524915-2018
• 负责人: Sirard, MarcAndré
• 金额: $ 2.29万
• 依托单位: Université Laval
• 依托单位国家: 加拿大
• 项目类别: Collaborative Research and Development Grants
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=684150
• 关键词: Bovine; imprinted; gene; analysis; assisted

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.

通过体外成熟和受精生产牛胚胎正在成为奶牛基因组革命的重要工具。 根据父亲和母亲的基因型选择胚胎的能力，以及在怀孕开始前在胚泡水平上这样做的可能性，正在改变动物的生产和选择方式。对这些胚胎的分子分析表明，根据父母的年龄以及文化或激素环境，这些胚胎存在一些差异。 这种影响可能是暂时的，但在某些情况下，它可能会影响出生后的表型，这表明需要更多的研究来了解观察到的变化。 该项目基于这样的假设：一些甲基化模式是在卵泡发生的最后部分获得的，并且可能受到为产生这些卵母细胞而创造的环境的影响。这种差异在培养的第一周内可能不会被消除，或者在这段时间内对条件不敏感。为了检验这一假设，将从青春期前供体（8）获得的32个胚胎（第12天）与从成人获得的胚胎进行比较，这些胚胎在体外用（8）或不用血清（8）或在体内（对照）（8）产生。 将从这些胚胎中提取DNA并进行亚硫酸氢盐处理以揭示甲基化标记。然后将对10个印迹基因的分析进行比较，以评估模式是否被修改，以及是否所有胚胎都受到影响或仅受一个子集的影响。第二部分将评估48个胚泡和活检中4种最佳印迹指示剂的甲基化水平，以开发快速第7天测试，允许监测培养条件或卵巢刺激的任何修改。这项工作将产生标记物，以评估所需的激素和培养环境，以尽量减少ART产生的牛胚胎中的表观遗传干扰。