The biology of spermatogonial development in sheep

精原干细胞是精子的成体前体；它们的健康对物种的繁殖至关重要。精原干细胞移植是一种从供体睾丸中分离干细胞Sg并将其注入受者的生精小管中由受者产生供体精子的技术。小鼠的精原干细胞移植已被证明是研究精子发生的一种有价值的新工具，并为利用干细胞Sg保存和传播遗传学打开了可能性。尤其是羊业将受益于更多的方法来运输遗传学，而不需要运输动物，以及从成本更低的方式直接将基因引入生殖系。公羊是研究哺乳动物精原细胞出生后发育的很好的啮齿动物替代品。由于公羊相对较小和便宜，公羊羔羊没有被阉割，所以屠宰场有现成的所有年龄段的睾丸供研究。在研究干Sg时必须克服的一个主要问题是，它们是睾丸中的一个小种群。因此，为了寻找对它们的健康重要的分子，我们将用激光从公羊羔羊睾丸的冰冻切片中收集单个干细胞Sg，然后从细胞中提取RNA，通过PCR对重要的基因转录本进行量化。这些RNA还将用于利用微阵列对这些细胞表达的基因进行广泛的筛选。这些信息将用于提高公羊精原移植的效率。我们还将开发一种方法，利用逆转录病毒将基因永久整合到供体干细胞Sg中。最后，我们将使用精原移植和转导的供体干细胞Sg来测试筛选结果中确定的基因在公羊精原细胞发育过程中的功能。我们将为农场畜牧业和寻求大型动物疾病模型的研究人员展示精原移植的效率。

Spermatogonial stem cells (stem Sg) are the adult precursors of sperm; their health is critical to propagation of species.  Spermatogonial transplantation is a technique that isolates stem Sg from donor testes and injects them into the seminiferous tubules of a recipient for production of donor sperm by the recipient animal.  Spermatogonial transplantation in mice has proven to be a valuable new tool for studying spermatogenesis, and has opened the possibility of using stem Sg for preservation and dissemination of genetics.  In particular the sheep industry would benefit from additional ways to transport genetics without shipping animals, and from less costly ways to introduce genes directly into the germ line. Rams are a good alternative to rodents for studying post-natal development of spermatogonia in mammals.  They are comparatively small and cheap, and ram lambs are not castrated, so there is a ready supply of testes of all ages from the abattoir for study.  A major problem that must be overcome in studying stem Sg is that they are a minor population in the testis.  Therefore in order to look for molecules important to their health, we will collect individual stem Sg from frozen sections of ram lamb testes with a laser and then extract RNA from the cells to quantify important gene transcripts by PCR.  The RNA also will be used for a broad screening of genes expressed by these cells using microarrays.  The information will be used to increase the efficiency of spermatogonial transplantation in rams.  We also will develop a way to permanently incorporate genes into donor stem Sg using a retrovirus.  Finally we will use spermatogonial transplantation and transduced donor stem Sg to test the function of genes identified in the screens in development of ram spermatogonia.  In the process, we will demonstrate efficiency of spermatogonial transplantation for the farm animal industries and for researchers seeking large animal models of disease.