Characterization and germ line differentiation of non-rodent embryonic stem cells

转基因奶牛的奶是复杂重组生物制药蛋白质的宝贵来源，比传统的生物反应器更安全，需要更少的资本投资。目前，培育转基因农场动物的技术是体细胞核移植（SCNT），这种方法效率低且成本高。最近，我们开发了生殖细胞移植（GCT）作为一种新的，潜在的更有效的替代转基因大型动物的生产。在用于SCNT或GCT之前可以修饰和表征的细胞系的可用性将大大提高用于重组蛋白生产的转基因动物的效率和商业可行性。本项目的目的是从山羊胚胎中获得干细胞，并研究这些细胞向雄性生殖细胞的分化。山羊对重组蛋白的生产具有重要的商业价值。山羊也是研究生殖系分化的理想的大型动物模型，因为胚胎发生、SCNT和胚胎移植已经被很好地理解，我们已经开发了用于该物种生殖系分化的移植试验。我们最近获得了山羊胚胎干细胞系。在这里，我们将验证来自山羊胚胎的细胞系可以分化成身体的所有组织类型（胚胎干细胞的一个标志性特征）的假设。然后，我们将使用我们开发的动物模型来测量胚胎来源的干细胞向产生精子的功能性男性生殖细胞的分化。这些研究将验证暴露于睾丸环境会诱导生殖细胞分化的假设。最后，我们将验证一个假设，即存在于睾丸中的干细胞可以恢复为未分化的干细胞，类似于来自胚胎的干细胞。本项目的预期影响是建立从胚胎干细胞中获得功能性雄性生殖细胞的大型动物模型。胚胎或生殖细胞衍生干细胞的可用性有望转化为更有效的方式来产生转基因大型动物，用于大规模生产重组蛋白。重要的是，该项目将为学术界和生物技术行业的高素质人才提供理想的培训环境。

The milk of transgenic dairy animals is a valuable source of complex recombinant, biopharmaceutical proteins that is safer and requires lower capital investment than traditional bioreactors. The current state of the art for generating transgenic farm animals is somatic cell nuclear transfer (SCNT), which is inefficient and expensive. Recently, we developed germ cell transplantation (GCT) as a novel, potentially more efficient alternative for the production of transgenic large animals. Availability of cell lines that can be modified and characterized before use for SCNT or GCT would substantially improve efficiency and commercial viability of transgenic animals for recombinant protein production. Objectives of this project are to derive stem cells from goat embryos and to study differentiation of these cells into male germ cells. The goat is of significant commercial interest for recombinant protein production. The goat also is an ideal large-animal model for studying germ lineage differentiation because embryogenesis, SCNT and embryo transfer are well understood, and we have developed transplantation assays for germ line differentiation in this species. We recently derived a goat embryonic stem cell line. Here, we will test the hypothesis that cell lines derived from goat embryos can differentiate into all tissue types of the body (a hallmark characteristic of embryonic stem cells). We will then use animal models developed by us to measure differentiation of embryo-derived stem cells to functional male germ cells that produce sperm. These studies will test the hypothesis that exposure to the testis environment will induce germ cell differentiation. Finally, we will test the hypothesis that stem cells present in the testis can revert to undifferentiated stem cells resembling those derived from the embryo. The expected impact of this project is to establish a large animal model for deriving functional male germ cells from embryonic stem cells. Availability of embryo- or germ cell derived stem cells is expected to translate into more efficient ways to generate transgenic large animals for large scale production of recombinant proteins. Importantly, this program will provide an ideal training environment for highly qualified individuals for academia and biotech industry.