본 연구에서는 수정란 이식을 위해 선발된 대리모에 이식 전 백신의 투여 시기가 임신과 유산에 미치는 효과와, 임신한 대리모의 분만 예정일 전에 백신의 투여가 송아지의 질병과 생존에 미치는 효과를 검토하였다. 실험 1에서는 한우 체외 수정란의 이식에 선발된 대리모의 백신 투여 시기가 임신과 유산에 미치는 영향을 검토하였다. 이식 전 주째 백신 투여시 임신율이 31.5%로서 대조군 및 주 투여군의 42.4 및 45.9%에 비하여 유의하게 낮았다(p<0.05

본 연구는 체외수정란에서 유래한 한우 송아지의 생존에 미치는 각종 요인들에 대하여 분석하여 체외수정란을 이용한 수정란 이식과 송아지의 효율성을 향상시키고자 실시하였다. 분만된 송아지의 기형율은 전 시험군에서 비슷한 경향이었다. 질병 발생율은 임신 기간, 분만 유도, 분만처치 방법과 형태 및 난산 처리 방법에 따른 차이가 없었다. 그러나 경산우가 미경산우, 봄과 겨울(20.4 및 )이 여름과 가을(4.3 및 ), 단태가 쌍태 및 정상 분만이 난산에 비하여

Cloned calves derived from somatic cell nuclear transfer (SCNT) have been frequently lost by sudden death at 1 to 3 month following healthy birth. To address whether placental anomalies are responsible for the sudden death of cloned calves, we compared protein patterns of 2 placentae derived from SCNT of Korean Native calves died suddenly at two months after birth and those of 2 normal placentae obtained from AI fetuses. Placental proteins were separated using 2-Dimensional gel electrophoresis. Approximately 800 spots were detected in placental 2-D gel stained with coomassie-blue. Then, image analysis of Malanie III (Swiss Institute for Bioinformatics) was performed to detect variations in protein spots between normal and SCNT placentae. In the comparison of normal and SCNT samples, 8 spots were identified to be up-regulated proteins and 24 spots to be down-regulated proteins in SCNT placentae, among which proteins were high mobility group protein HMG1, apolipoprotein A-1 precursor, bactenecin 1, tropomyosin beta chain, H+-transporting ATPase, carbonic anhydrase II, peroxiredoxin 2, tyrosine-rich acidic matrix protein, serum albumin precursor and cathepsin D. These results suggested that the sudden death of cloned calves might be related to abnormal protein expression in placenta.

본 실험은 한우 종빈우의 임신말기에 Se과 Vit. E 투여가 종빈우에서 태어난 송아지의 발육능력을 조사하고 혈액화학치 및 혈중 Vit. E 농도에 미치는 영향을 조사하여 다음과 같은 결과를 얻었다. 1. Se과 Vit. E를 분만 1개월전에 투여 한 한우 종빈우에서 태어난 송아지의 혈액성분에 미치는 영향을 검토한 결과, 평균 혈중 albumin은 3.36, 3.51, 3.43, 3.45, 3.83 및 3.31g/d1로서 Vit. E 2000IU 투여구가 여타구보다 높은 성적을 나타냈다(P<0.05). 평균 혈중 cholesterol함량은 Vit. E 1500IU 투여구가 여타구보다 높은 함량을 나타냈으나, 혈중 BUN, creatiine, glucose, inorganic phosphorous, calcium, total protein 및 triglycerides 함량은 처리구간 유의적인 차이는 없었다(P>0.05). 2. Se과 Vit. E를 분만 2개월 전에 투여한 한우 종빈우에서 태어난 송아지의 혈액성분에 미치는 영향을 검토한 결과, 평균 혈중 albumin, cho-lesterol, BUN, creatiine, glucose, inorganic phos-phorous, calcium, total protein 및 triglycerides 함량은 처리구간 유의적인 차이는 없었다(P>0.05). 3. 분만 1개월 전에 Se과 Vit. E 투여한 한우 종빈우에서 태어난 송아지의 혈중 vitamin E 농도에 미치는 영향을 검토한 결과, 한우 종빈우에서 태어난 송아지의 생후 15일째 혈중 Vit. E 농도는 처리구간 커다란 차이가 없었으나, 생후 45일째 혈중 Vit. E 농도는 투여구가 대조구보다 높은 농도를 나타냈다. 모든 처리구에서 송아지의 생후 일령이 증가함에 따라 혈중 Vit. E 농도가 증가하는 경향을 보였다. 4. 분만 전 2개월 전에 Se과 Vit. E를 투여한 한우 종빈우에서 태어난 송아지의 생후 15일과 45일째 혈중 Vit. E의 농도는 처리구간 커다란 차이는 없었다(P>0.05).

본 실험은 한우 종빈우의 임신말기에 Se과 Vit. E 투여가 종빈우의 번식기능에 미치는 요인과 종빈우에서 태어난 송아지의 발육능력을 조사하였다. 1 분만 1개월전 Se과 Vit. E을 투여한 구에서 송아지의 발육성적을 조사한 결과, 생시체중은 각각 23.33, 24.00, 24.00, 24.50, 24.00 및 25.60㎏으로서 처리구가 대조구보다 다소 높은 성적을 나타냈다. 또한 이유시 체중과 일당증체량도 투여구가 대조구보다 다소 높은 성적을 나타냈으며, 이유시 일령은 투여구가 대조구보다 이유시 일령이 다소 적었으나 처리구간 유의적인 차이는 없었다(P>0.05). 2. 종빈우의 번식능력을 조사한 결과, 분만 후 초발정과 인공수정 휫수는 투여구가 대조구보다 초발정이 빨리 도래하였고, 수태당 인공수정 휫수도 적었으나 처리구간 유의적인 차이는 없었다(P>0.05). 3. Se과 Vit. E를 분만 2개월전에 4회 투여한 한우 종빈우에서 태어난 송아지의 생시체중, 이유시 체중 및 일당증체량은 투여구가 대조구보다 높은 성적을 나타냈으며, 이유시 일령은 투여구가 대조구보다 이유시 일령이 짧았으나 처리구간 유의적인 차이는 없었다(P>0.05). 4. 한우 종빈우의 번식능력을 조사한 결과, 분만 후 초발정은 유의적인 차이는 없었으며(P> 0.05), 수태 당 인공수정 횟수는 각각 2.00, 1.63, 1.25, 1.50, 1.33 및 1.46회로서 투여구가 대조구보다 인공수정 횟수가 적었으나 처리구간 커다란 차이가 없었다(P>0.05).

The objective of this study was to improve the efficiency of bovine embryo transfer by transferring of Hanwoo embryos into Hanwoo or Holstein recipients. The cryopreserved or fresh in vitro produced(IVP) embryos were transferred into uterine horn contralaterally or ipsilaterally to the corpus luteum. The recipients were inseminated by artificially on the next day of estrus. The pregnancy was diagnosed by rectal palpation at 60∼90 days after transfer of the embryos. The pregnancy rate by transfer of one or two embryos was 78%(7/9) and 74%(31/42), respectively. The pregnancy rates according to the grade of corpus lutea of recipients was 75% (20/27) and 82.0%(18/22) at the grade of A and B, respectively. Ten(67.0%) of 15 Holstein recipients transferred with IVP Hanwoo embryos and 5(42.0%) of 12 Holstein recipients transferred with frozen IVP Hanwoo embryos were pregnant. The single and twin calving ratio in Hanwoos was 77.0%(10/13) and 23.0%(3.13) in the recipients transferred with IVP embryos and 64.0%(7/10) and 27.0%(3/10) in the recipients transferred with frozen IVP embryos, respectively. Twenty-four pregnant cows following transfer of IVP embryos, 21(88.0%) calved the normal calves, and 2(8.3%) aborted. When the frozen IVP embryos were transferred, 16 pregnant cows calved 14(88.0%) normal calves and 2(13.0%) aborted. In conclusion, when one or two IVP bovine embryos were transferred into recipients, the A and B grade of corpus luteum resulted in high pregnancy rates. For the production of twin calves, transfer of the IVP or frozen IVP embryos could be suitable.

This study was carried out to improve a technique of embryo transfer for twin calves production in Hanwoo cattle. Blastocysts for the donor of embryo transfer were classified into three criteria by accessment of morphology; early blastocyst, blastocyst and expanded blastocyst. Tow embryos were introduced transcervically into utrerine horn either of Hanwoo or Holstein by ipsilaterally or contralaterally to the corpus luteum. Thiry-six out of 57 recipients cows were inseminated by artificially on the next day of estrus, and followed by transfer of embryos into contralaterally. The pregnance rates of recipients following transfer of bovine embryos of day 7, 8 and 9 was 43.5, 18.2 and 8.3%, respectively. These results appeared that these was a significant (P<0.05) difference between on day-7 embryos and day-9 embryos, but not between on day-8 and day-9 embryos. Although there was not significant(P<0.05) difference in the pregnancy rates between the blastocysts(11/25, 44%) and expanded blastocysts(2/19, 10.5%) and between the blastocysts and early blastocysts(2/13, 15.4%), the embryos at blastocyst stage are more suitable than others for obtaining higher rate of pregnancy. There was no significant difference on pregnancy of the embryos transferred prior to presence(6/21, 29%) or absence (9/36, 25%) of artificial insemination. On pregnancy of Holstein, 2(15.4%) out of 13 recipients were pregnant in heifer. Similar Pregnancy rates were obtained between 1∼2 parities and 3∼4 parities by 30% (6/20) and 27.3%(3/11), respectively. Taken together, there was not significant difference in pregnancy rate due to small number of recipients used for this experiment. Both of Hanwoo and Holstein introduced the embryos by contralsterally to the corpus luteum were slightly higher pregnancy rate compare to by ipsilaterally (12/41, 29.3% vs, 3/16, 18.8%). The ratio of production of twin and single calves in Holstein was 20% (9/45) and 2.2% (1/45), respectively. However, in Hanwoo cows both of production of twin and single were similar as 8%. This result suggests that Holstein as recipients was superior to Hanwoo cows for production of twin calves. Out of all 15 pregnant, 12(80%) were produced a total of 22 normal calves in which the others composed of abnormal, as judging as 2(13.3%) for abortion and 1(6.6%) for stillbirth during the pregnant period.

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 establish an effective system for embryo transfer of techniques by analyzing several factors affecting the gestation length and the weight changes of calves produced from embryo transfer in Korean cattle. The results obtained in study on factors affecting the gestation length and the weight changes of calves produced from embryo transfer were as fallow; 1) The gestation length and the birth weight did not differ between male and female, but the weight changes after birth were remarkablely different between sex(P<0.05). 2) The gestation length between heifers and cows was not different, and body weights at birth and 6 months were remarkabley heavy in cows(P<0.05). Weight changes after 6 months were not different. 3) The gestation length and the birth weight were significantly different between the single and twin calving (P<0.05). Weight of twin at 6 and 12 months were remarkabely light. 4) Calving seasons did not affect the gestation length the and the birth weight. Weaning weight was significantly heavy(P<0.05), but weight changes after weaning were no different among the calving seasons. Conclusivley, this results suggest that cows will be better when considering growth of calves and twin produced from embryo trnsfer in Korean cattle.

The objective of this study was to produce calves by transfer of embryos derived from slaughter house(SHD) and ultrasound-guided ovum pick-up (OPU). At 60 hrs after injection of 400 mg FSH dissolved in 25% polyvinylpyrrolidone(PVP) by single dose, ultrasound-guided follicular oocyte aspiration was ferformed. Day-7 and day-8 blastocysts produced by in vitro maturation (IVM), fertilization (IVF) and culture(IVC) of the oocytes derived from SHD and OPU were nonsurgically transferred into recipients. The results obtained were as follows. The cleavage rate and the development rate to blastocysts were not significantly (P<0.05) different between the oocytes obtained by SHD (72.9% vs. 34.1%) and OPU (75.9% vs. 38.4%). The oocyte recovery rate from the number of follicles by ultrasound-guided aspiration were not significantly (P<0.05) different between Holstein (61.7%) and Hanwoo (60.1%), but the rate of oocytes useful for IVF was significantly (P<0.05) higher in Hanwoo (69.3%) than Holstein (59.6%). The cleavage rate and the development rate to blastocysts was not significantly (P<0.05) different between Holstein (74.9% vs. 39.2%) and recipients on day 8 of estrus cycle resulted in 13 pregnancies (34.2%). One of them was sacrificed during gestation period due to mastitis and another was aborted spontaneous. The resulting 14 calves were morphologically normal at birth. Seventy eight fresh OPU-IVF embryos were transferred into 21 recipients on day 8 of estrus cycles, resulting in pregnancy of 12 recipients (41.4%). Two of them were sacrificed during gestation period due to mastitis and the other two were aborted. Nevertheless, the 11 OPU-calves have been born normally.

The development potential of bovine somatic cells was evaluated using nuclear transfer. A single donor cell derived from fetus of HanWoo(Korean Native Cattle) was selected and deposited into perivitelline space of each enucleated oocyte before electrical fusion and activation. Nuclei of donor cells starved for 7 days (37%) tended to support the development of reconstitute embryo the blastocyst stage better than those of donor cells starved 3, 14 and 30 days. The cleavage rate was significantly lower(P<0.05) in reconstitute embryos derived from large size donor cells(51.2%), than those from small medium size donor cells(76.6 and 73.5, respectively). The developmental rate to blastocyst of reconstructed embryos from medium size donor cells was higher than those from small and medium size donor cells. This study demonstrates that an appropriate culture period for induction into quiescent stage and the size of donor cells effect on the efficiency of nuclear transfer using cultured bovine cells.

The objective of this study was to evaluate the embryonic development ability and the appearance of blastocysts of bovine in vitro fertilized oocytes cultured in different culture media, and also to evaluate survival rate after thawing of frozen embryos by using 1.5 or 1.8M ethylene glycol(EG) with sucrose or trehalose. Fertilized oocytes were divided into three groups; i ) monolayer of cumulus /granulosa cell prepared by TGM 199+5% calf serum(TGM199), ii)GRlaa+5% CS, iii)SOF+5% CS, and they were cultured after insemination for 9 days, at 39˚C, under 5% in air, but SOF+5% CS was cultured at 39˚C, under 5% 02, 5% GO2, 99% N2. Blastocysts derived from GRlaa + 5% CS on day 7~8 after insemination were frozen by using 1.5M EG or 1.8M EG with/without 0.2M sucrose or O.1M trehalose. The development rate of blastocysts on day 7 after insemination in SOF+5% CS was significant higher than in TCM199 or CR1aa(P<0.05). The appearance rate of blastocysts on day 7-8 after insemination was higher than in TCM199, when fertilized oocytes were cultured in GRlas or SOF. The survival rate of frozen blastocysts after thawing tended to increase, when blastocysts were frozen by using 1.8M EG with 0.2M sucrose or O.1M trehalose. These results indicated that SOF or CRlaa media with amino acids was superior to TCM199 with monolayer in terms of blastocyst development in culturing of in vitro fertilized bovine nocytes, and sucrose or trehalose was supposed to prevent embryos from the freezing shock.

A combined technology of transvaginal ovum pick-up(OPU) system with in vitro-oocyte manipulation technique can be used for improving reproductive efficiency in the cattle. The objective of this study was to establish a newly-conceived breeding program using OPU in the pregnant cows. The OPU trial was performed in pregnant cows every 10 days from 40 through 90 days of artificial insemination (Al), and number of follicles in ovary, number of retrieved oocytes and embryo development following in vitro-fertilization, were evaluated. Reduced number of follicles in the ovaries of pregnant cows was firstly detected from 70 days after A' and a significant (P<0.05) decrease in the follicle number (5.4 follicles /donor) was found at 90 days than at 40, 50, 60 and 80 days after Al (8.0~9.2). A similar pattern was also observed in the number of oocytes retrieved by OPU apparatus during experimental period. When retrieved oocytes were matured and inseminated in vitro with frozen bull semen, development of the oocytes to the blastocyst stage was not significantly affected by the retrieval time. Four embryos (morula or blastocyst stage) derived from oocytes retrieved from pregnant cows were nonsurgically transferred to four recipient cows on day 7 of estrus cycle. For the first time in Korea, three of four transferred embryos developed to live calves with normal physiological parameters. In conclusion, an effective breeding program employing pregnant cow can be developed by use of OPU trial and in vitro culture techniques of oocytes ; OPU system could be repeated in pregnant cows with no risk of abortion and viable offsprings were borne after transfer to the recipients.

The objectives of the present study were improvements in the efficiency of developmental rates to morula and blastocyst stages to produce a large number of genetically identical nuclear transplant embryos. The oocytes collected from slaughterhouse ovaries were matured for 24 h and then enucleated and cultured to allow cytoplasmic maturation and gain activation competence. And then the donor embryos were treated for 12 h with 10 g /ml nocodazole and 7.5 g /ml cytochalasin B to synchronize the cell cycle stage at 26 h after the onset of culture. The blastomeres were transferred into the perivitelline space of the enucleated nocytes and blastomeres and oocytes were fused by electrofusion. The cloned embryos were then cultured in various conditions to allow further development. The age of the recipient(30 vs 40 h) had no significant effect on the fusion rates(82.4 vs 82.1%) and the developmental rates to morula /blastocyst(9.8 vs 11.0%). Effect of Nocodazole treatment on the donor cell cyle synchronization to improve the developmental rates of bovine nuclear transplant embryos was significantly higher than control group(21.4 vs 10.1%, p<0.05). Significant differences were in the percentage of fusion rates(72.9,77.1vs 61.9%) in three types of fusion medium(PBS(+), mannitol and sucrose, p<0.01). The developmental rates of bovine nuclear transplant embryos appeared to be highest in mSOF medium under 5% 0 condition, but no significant differences were found when compared with TCM199-BOEC and mSOF under two different oxygen ratio(5 and 20%).