Studies on Cryo-preservation of Registered Strains of Lentinula edodes

New strain needs to maintain desirable characteristics for long term when it was bred, but in lapse of time it degenerates into a bad condition. Therefore the influence of temperature on the viability and survival rates of Lentinula edodes strains were examined after cryopreservation. Also, liquid nitrogen preservation for L. edodes has been proved to be one of the most reliable method. However, a mechanical damage of strain is inevitable during cryopreservation of the fungus because the fungus is very sensitive to stress of cooling rate in the freezing process. So we tried to find out state change of L. edodes with a programmable freezer. L. edodes strains were preserved at $-20^{\circ}C$, $-80^{\circ}C$ and $-196^{\circ}C$ for 50 days. At $-20^{\circ}C$, its mycelial growth became extinct. When thawed, the growth of mycelia which were preserved at $-80^{\circ}C$ was fastest. Attempts were made to investigate viability of L. edodes strains after freezing at $-80^{\circ}C$ and $-196^{\circ}C$, respectively. As the result, more than 90% showed high survival rate of strains tested at $-80^{\circ}C$ and $-196^{\circ}C$. Mycelial growth between apical and basal parts of colony after freezing preservation for 50 days was compared. At apical and basal parts, the survival rates showed 100% at $-80^{\circ}C$, but 98% and 94% at $-196^{\circ}C$, respectively. We confirmed that the ice crystal formation temperatures of L. edodes strains were $-6.0^{\circ}C$ for Sanlim 1, $-5.5^{\circ}C$ for the Sanlim 2, $-4.0^{\circ}C$ for the Sanlim 3 and $-15.5^{\circ}C$ for the Sanzo 302. These results indicated that L. edodes strains showed completely different responses to the ice crystal formation. We knew the fact that even the same species, especially L. edodes, they displayed completely different responses to the same freezing condition. Also, this has nothing to do with the connection between temperature type and freezing point. And a protocol was tried to minimize state change of L. edodes strains using programmable freezer when they are frozen, but it was not effective on them.

표고 등록균주의 초저온 보존에 관한 연구

새로운 균주가 육종되었을 때 그 균주가 장기간 고유특성을 유지하기란 쉽지 않으며, 시간이 경과 할수록 균주는 퇴화를 거듭한다. 이에 따라 현재 많이 이용되고 있는 초저온에서의 균주보존 후 균사의 활력과 온도가 생존에 미치는 영향과 표고균주가 동결하는 동안 나타나는 상태변화를 이해하고자 하였다. 저온 처리별 균주보존결과 표고균주의 생존은 $-20^{\circ}C$에서 50일간 보존하면 사멸했으며, $-80^{\circ}C$와 $-196^{\circ}C$에서 보존한 균주들은 온도에 영향을 받지 않고 생존하였다. 저온 처리별 균사생장속도는 $-80^{\circ}C$에서 보존한 균주들의 재생속도가 가장 빨랐다. 균사의 정단부분과 말단부분에 대한 저온 처리별 균주보존 결과 부분별 위치는 균사의 활력과 생존에 영향을 끼치지 못했다. 프로그램이 가능한 동결장치를 이용하여 표고균주의 동결과정에서 나타나는 현상을 조사한 결과 균주별 각기 다른 어는점을 가지고 있었다. 또한 표고의 자실체 발생 온도형과 균주별 어는점과의 관계는 관련이 없었다. 프로그램이 가능한 동결장치를 이용하여 표고균주의 동결과정에서 나타나는 상태변화를 최소화하기 위해 일시적으로 낮은 온도로 동결을 시도하였지만 균주들은 온도에 영향을 받지 않았다.