칡소의 귀세포를 이용한 핵이식 시에 전기 융합조건이 핵이식란의 발생에 미치는 영향을 알아보고, 칡소의 귀세포와 한우의 태아섬유아세포를 이용한 핵이식란을 이식하여 수태율을 비교하였다. 전기자극 조건에 따른 핵이식 후 융합율은 51∼68%의 범위로 차이는 없었으나, lysis 율은 1.9kv/cm의 10us, 20us에서 각각 0.0과 38.7%, 2.0kv/cm의 10us, 20us에서 각각 2.9와 37.5%, 2.1kv/cm의 10us, 20us에서 각각 21.2와 51.8%을 보였다. 난할율은 1.9kv/cm의 10us, 20us에서 각각 75.8과 69.8%, 2.0kv/cm의 10us, 20us에서 각각 76.9와 68.8%, 2.1kv/cm의 10us, 20us에서 각각 70.5와 68.5로 큰 차이가 없었다. 그러나 배반포 발생율은 1.9kv/cm의 10us, 20us 각각 19.5와 48.6%, 2.0kv/cm의 10us, 20us에서 각각 20.0과 40.9%, 2.1kv/cm의 10us, 20us에서 각각 44.2와 27.0%로 각 조건에서 시간에 따른 차이를 보였다. 핵이식에서 공여세포와 세포질간의 융합율과 lysis율은 전압과 통전시간에 영향을 받으며 전압을 높이는 것이 통전시간을 길게 하는 경우보다 수핵난자의 세포질에 상해를 줄이고 배반포 발생에 유리 하였다. 한우의 태아섬유아세포의 핵이식후 생산된 배반포를 5마리의 수란우에 이식한 결과 두 마리가 임신되었고(40.0%), 칡소의 귀 세포를 핵이식한 후 생산된 배반포를 19마리에 이식한 결과 3마리가 임신되었다(15.8%).

본 연구는 체세포 및 체세포 핵이식란의 염색체 이상에 체세포의 계대의 영향성을 조사하고자 하였다. 한우 성축의 귀 조직으로 얻어진 세포를 공여세포를 체외성숙 후 제핵된 난자에 핵이식을 실시하였으며, 1.9kv/cm, 20/2times의 전기자극으로 융합후 5/ml의 ionomycin에서 4min, 1.9mM 6-DMAP에서 4h동안 배양함으로써 활성화를 유도하였다. 핵이식란은 CRlaa에서 4일간 배양 후 8 -cell단계에서 중기상의 유도를 위하여 상기 배양액 1ml당 0.05 colcemid에서 6-8시간 더 배양하였다. 이후, 6% Fetal bovine serum이 함유된 1% sodium citrate용액에 20분간 저장처리 후, methanol 5 : aceticacid 1 : distilled water 4로 1차, methanol 3: aceticacid 1 로 조성된 2차, methanol 4 : acetic acid 3 : distilled water 1의 3차고정액으로 1분간 재 침지시켰다. 고정 처리가 완료된 slide는 4% Giemsa용액으로 염색한 후 광학현미경 하에서 핵형 양상을 검경하였다. 체세포의 5계대에서는 684개의 spreads를 검경한 결과 염색체 수는 72%가 정상으로 60개이었고, 24%가 60개 이하였으며 4%가 60개 이상을 보였다. 10계대도 5계대와 비슷하여 71%가 정상, 26%가 60개 이하, 3%가 60개 이상이었고, 15계대에서는 55%가 정상이었고, 30%가 60개이하, 15%가 60개 이상을 보였다. 10계대 까지는 mixoploid의 비율의 변화가 없었으나 15계대에서 현저하게 늘어남을 볼 수 있었다. 또한 체외수정란과 핵이식란의 비교에서는 체외수정란은 250개의 spreads를 검경한 결과 염색체 수는 95.6%가 정상으로 60개이었고, 2.0%가 60개 이하, 2.4%가 60개 이상이었으나, 핵이식란은 204개를 검경하여 88%가 정상이었고, 4.9%가 60개이하, 7.1%가 60개 이상을 보임으로써, 핵이식란이 체외수정란에 비하여 염색체 이상의 비율이 높았다. 따라서 계대에 따라 체세포의 염색체이상의 비율이 상대적으로 증가하고, 체세포 핵이식에 따른 염색체 이상이 생길 수 있음을 알 수 있었다. (이 논문은 농림부 연구비에 의해 수행되었음)

1994년 6월부터 8월까지 금호강 본류의 유수변 초본식생을 식물사회학적으로 조사하였으며, 수질환경과 관련된 식생의 분포특성을 밝혔다. 금호강 본류의 하변식생은 여뀌-미꾸리낚시군락(Persicaria sieboldi-Persicaria hydropiper community), 고마리군락(Persicaria thunbergii community), 물봉선군락(Impatiens textori community), 줄군락(Zizania latifolia co

This study was carried out to investigate the effect of co-culture system(bovine oviduct epithelial cells; BOEC) and defined culture system (modified TALP ; mTALP) on the conception of embryos transferred, and pregnancy and twin birth rates after transfer of fresh or frozen-thawed bovine blastocysts produced in vitro were also evaluated. Oocytes from the slaughterhouse ovaries were matured and fertilized using general protocol. The results obtained were as the following. The pregnancy rate after transfer was higher in co-culture group than in mTALP group, but was not signficantly different, and there is no difference between fresh embryo group and frozen-thawed embryo group in conception rate. The conception rate was not different whether 3∼4 blastocysts or 2 blatocysts transferred into a recipient, but the production rate of twin calves was significantly higher (p<0.05) when 3∼4 embryos transferred. The average birth weight of twin calves(24.38kg) was numerically, but not significantly lighter than that of single calves(26.68kg).

This study was carried out to investigate the effect of co-culture system(bovine oviduct epithelial cells; BOEC) and defined culture system(modified TALP ; mTALP) on the development of IVM-IVF embryos, and survival of in vitro produced blastocysts after freezing and thawing. Occytes from the slaugheterhous ovaries were matured and fertilized using general protocol. The results obtained were as the following: 1. Survival rates of frozen-thawed blastocysts using 10% glycerol as cryoprotectant was higher in day 7 blastocysts than in Day 8 and 9 blastocysts from co-cultrue system, but survival rate of frozen-thawed blastocysts was higher in Day 10 blastocysts than in day 8 and 9 blastocysts from defined culture system. Regardless of their age, survival rate of frozen-thawed blastocysts was significantly higher (p<0.05) in co-culture system than in defined culture system. 2. The cell number of blastocysts was significanlty higher (p<0.05) in Day 7 blasotcysts than in Day 8 and 9 blastocysts from co-cultures, but the cell number of blsstocysts was significantly higher (p<0.05) in Day 10 blastocysts than in Day 8 and 9 blastocysts from defined culture system. Regardless of the culture system, blastocysts with higher cell number showed higher survival rates after freezing and thawing.

The objectives of the study were to establish sperm separation method and duration of insemination for bovine IVF. Oocytes from slaughterhouse ovaries were matured and fertilized using general protocol. After 18 or 42 h of insemination, six to ten embryos were placed into a 30 drop of each medium, and the embryos were examined 7~10d post in semination without medium renewal. First, we compared Percoll gradient will swim-up technique for sperm separation. There was no difference in cleavage rates between them, but the development rates over morula stage of oocytes fertilized with sperm separated by Percoll gradient was significantly higher than that sperm selected by swim-up technique (p<0.05). Second, we evaluated development of bovine embryos derived from the IVF procedure with different durations(18 vs 42 h) of fertilization. There was also no difference in cleavage rates, but the development to blastocyst stage of oocytes exposed in cleavage rates, but the development to blastocyst stage of oocytes exposed to sperm for 42 h was significantly higher than that exposed for 18 h (p<0.05). In conclusion, Percoll gradient can be used for sperm selecton, improving of embryonic development. Also, 42h of IVF may improve the development of bovine embryos.

백운산의 삼림식생과 토양환경을 파악하기 위하여 식물사회학적 조사방법에 의하여 삼림식생을 분류하고, 각 군락의 구조적 특성을 밝혔으며, 토양환경을 조사하였다. 삼림식생은 신갈나무군락(전형 하위군락, 철쭉꽃하위군락), 굴참나무군락, 물푸레나무군락, 소나무군락 등 4개의 자연군락과 일본잎갈나무식재림, 잣나무식재림 등 2개의 식재림으로 분류되었다. 식생의 계층구조는 매우 발달한 편으로 신갈 나무군락은 백운산의 거의 전역에 걸쳐 표고 500m이상부터 분포하고,

Immature nocytes and in VitrO matured Oocytes collected from the slaughtered Korean cattle were frozen slowly with 10% ethylene glycol+5% polyvinyl pyrolidine+0.05M trehalose (l0EPT), 10% ethylene glycol+5% ficoll+0.05M sucrose (1OEFS), or 10% ethylene glycol+5% ficoll+0.05M trehalose (l0EFT) by cell freezer (experiment 1). And also,They were ultra-rapidly frozen with 30% ethylene glycol+10% polyvinyl pyrolidine+0.5M trehalose (3OEPT) or 30% ethylene glycol+18% ficoll+0.5M sucrose (3OEFS) using electron microscope grid (experiment 2). In experiment 1, the cleavage rate was 23.0% when immature oocytes were frozen slowly using various cryoprotectants descrihed above, and 5.1% of cleaved oocytes developed to over morula stage after in Vitro fertilization (IVF). There were no significant differences among these groups. When matured oocytes were frozen slowly, the total cleavage rate was 19.7%, and over morula stage was 3.2%. lOEPT (4.8%) and EFS (4.4%) were slightly more effective than l0EFT (0.0%) for development in vitro. Only in l0EFT treated group, immature oocytes have higher developmental capacity than matured ones, when they were frozen slowly and IVF after thawing. In experiment 2, oocytes were ultra-rapidly frozen using the electron microscope grid with two kind of cryoprotectants described above. In immature oocyte group, the cleavage rate was 13.9% and 5.8% of cleaved oocytes developed to over morula stage after IVF, and in matured group, 25.7 and 7.6%, respectively. There were no significant differences between two kind of cryoprotectants, but in ultra-rapid freezing using electron microscope grid, the efficiency is slightly higher in matured oocyte group.