Rat sperm cryobanking for genetic resources

描述（申请人提供）：种质冷冻库是无限期保存科学、医学、经济和环境重要遗传资源的有效方法。虽然大鼠是模拟人类疾病的重要系统，但没有建立的种质冷冻保存方案，这将允许具有成本效益的管理大鼠遗传资源。收集附睾精子是保存遗传学以备将来使用的最简单和最具成本效益的方法。然而，迄今为止，只有一个方案显示解冻后活力仅为10%。因此，迫切需要了解影响大鼠精子冷冻存活的生理和低温生物学因素，以提高大鼠精子的冷冻存活率。本申请的目的是开发用于附睾大鼠精子的冷冻方案，其将允许通过使用标准人工授精和体外受精方法重建遗传学。在过去的几年里，我们的实验室已经产生了大量的数据，与最佳的精子处理，Percoll分离，冷冻延长剂和冷却和加热条件的大鼠精子。这将有助于减少物理、低温、渗透胁迫和致死冰形成造成的伤害，并最终获得可接受的解冻后恢复率。我们实验室收集的数据与本申请中概述的冷冻生物学研究相结合，将使我们能够开发有效和可靠的大鼠附睾精子冷冻保存方案。 公共卫生相关性（由申请人提供）：本提案旨在开发有效的大鼠精子冷冻保存方法，该方法将用于通过常用的辅助生殖技术（如人工授精或体外受精（如需要））重建具有遗传重要性的大鼠品系。因此，拟议的研究将通过提供可靠的大鼠精子冷冻保存方案来解决生物医学界的问题，以避免连续的活体动物饲养，这对美国国立卫生研究院及其全国各地的研究人员来说是一个重大负担。

DESCRIPTION (provided by applicant): Germplasm cryobanking is an effective way of preserving scientifically, medically, economically and environmentally important genetic stocks indefinitely. Although rats are important system for modeling human disease, there is no established germplasm cryopreservation protocol which would allow cost-effective management of rat genetic resources. Collection of epididymal sperm is the simplest and most cost-effective method of archiving genetics for future use. However, to date, there is only one protocol showing only 10% post-thaw motility. Thus, there is an urgent need for understanding the physiological and cryobiological factors affecting rat sperm in order to improve the rat sperm cryosurvival. The goal of this application is to develop freezing protocols for epididymal rat sperm which would allow reconstitution of genetics by using standard artificial insemination and in-vitro fertilization methods. Over the last few years, our laboratory has generated ample amount of data related with optimal sperm handling, Percoll separation, freezing extender and cooling and warming conditions for the rat sperm. These would help reduce the injuries caused by physical, hypothermic, osmotic stress and lethal ice formation, and ultimately yield acceptable post-thaw recovery rate. The data which have been collected in our laboratory in combination with proposed cryobiologic studies outlined in this application will allow us to develop efficient and reliable cryopreservation protocol for epididymal rat sperm. PUBLIC HEALTH RELEVANCE (provided by applicant): This proposal aims at developing efficient cryopreservation method for rat sperm which would be used to reconstitute genetically important rat strains through commonly used assisted reproductive technologies such as artificial insemination or in-vitro fertilization if needed. The proposed studies will therefore address to the biomedical community by providing a reliable protocol for rat sperm cryopreservation in order to avoid continuous live animal housing which is a significant burden for National Institutes of Health and its investigators around the nation.