Establishment of cryo-preservation method for ovary and testis and storage of wild mouse genes.

本研究旨在从基因资源保存的角度出发，通过冷冻卵巢和睾丸建立野生小鼠基因组存储系统。野生小鼠卵巢冷冻保存方法的建立及基因序列分析。在一系列实验小鼠中检测了卵巢成功冷冻的条件。将该方法应用于日本野生小鼠的MSM近交系卵巢时，获得了冷冻卵巢的小鼠，但产仔量很小。对新生小鼠睾丸冷冻损伤的低温保存方法的建立，在冷冻过程的不同阶段进行了组织学检查。在浸泡LN2之前，将生殖细胞缓慢冷却至-40ºC，许多生殖细胞仍保持正常形态。为了证实生殖细胞的生长和精子的产生，将冷冻的新生睾丸移植到组织相容的成年睾丸中。在上述冷冻方案的情况下，在移植后14天观察到减数分裂生殖细胞，在6周时在冷冻睾丸的精管中检测到精子，尽管尚未证实它们是否具有与卵母细胞受精的能力。目前正在研究将解冻的睾丸重建到宿主生殖系统。

This study aimed at an establishment of storage system for wild mouse genome by freezing of ovary and testis from a viewpoint of preservation of gene resources.1. Establishment of cryo-preservation method for ovary and strage of wild mouse genes.Successful freezing conditions of ovaries were detected in a series of experiments using laboratory mice. When the freesing method was applied to ovaries of MSM inbred strain derived from Japanese wild mice, mice derived from frozen ovaries were obtained, though litter sizes were very small.2. Establishment of cryo-preservation method for testisfreezing injuries of newborn mouse testes were examined histologically at various points of a freezing course. A lot of germ cells retained a normal morphology in the program of slow cooling til -40ﾟC before immersion into LN2.In order to confirm the growth of germ cells and the production of spermatozoa, the frozen newborn testis were transplanted into histocompatible adult testis. In the case of a freezing program described above, at 14 days after transplantation meiotic germ cells were observed and at six weeks spermatozoa were detected in the seminiferous tubules derived from the frozen testes, though it has not been confirmed whether they have a capacity of fertilization to oocytes.A reconstitution of thawed testis to a host reproductive system is under investigation.