|Budget Amount *help
¥2,200,000 (Direct Cost : ¥2,200,000)
Fiscal Year 1993 : ¥1,000,000 (Direct Cost : ¥1,000,000)
Fiscal Year 1992 : ¥1,200,000 (Direct Cost : ¥1,200,000)
This study aimed at establishing a storage system for wild mouse genes or genomes by freezing of embryos and gametes from a view point of the preservation of gene resources, and a succesful freezing condition of an ovary, which included oocytes as female gametes, was developed with a series of experiments.
1. Histological examination : In various points of a frozen course, which was made of slow cooling of 0.1ﾟC/min to -30ﾟC after seeding at -7ﾟC and rapid cooling to liquid nitrogen temperature, freezing injuries of the ovary were examined histologically. Till -30ﾟC damage of the ovary was little and a lot of oocytes with normal morphology were observed. On the other hand in the ovary immersed into liquid nitrogen after that almost all oocytes were injured.
2. Transplantation of frozen ovary : New borns were obtained from the control transplantation and till the -30ﾟC of cooling, but not from that of liquid nitrogen. This result showed a good correlation to that of the histological observation of the frozen ovary.
3. Examination of cooling rate : New borns were obtained from a transplantation of frozen ovary at conditions of cooling rates of 0.1, 0.2, 0.5ﾟC/min to -30ﾟC.The shortest interval between the transfer and the birth of the first new borns derived from the frozen ovary was 43,40,141 days each(control : 41 days). This result suggested that the rate of 0.5ﾟC/min, which was usually used for embryo freezing, was not appropriate for the ovary freezing.
4. Establishment of freezing program : Successful freezing of the ovary was performed by using a program which consisted of slow cooling at 0.2ﾟC/min to -40ﾟC and following rapid cooling to -180ﾟC.As an average litter size was 2.0 and new borns were not always obtained from all frozen ovary, more improvements of freezing conditions seemed to be indispensable for the practical use of the cryo-preservation of the ovary.