• 한국수정란이식학회지
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• 제15권1호
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• pp.47-56
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• 2000

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.

• 한국수정란이식학회지
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• 제24권1호
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• pp.15-19
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• 2009

This study was carried out to collect the basic data about gestation lengths and offspring's birth weights and sex ratio of the dairy recipients transferred with Hanwoo IVF embryos. Blastocysts cultured for $7{\sim}8$ days were transferred to 96 and 167 heads for the basic data about gestation length and offspring's birth weight and sex ratio of the dairy recipients, respectively. The gestation lengths of the dairy recipients transferred with Hanwoo IVF embryos were $287.3{\pm}11.2$ ($257{\sim}311$) and $284.1{\sim}7.7$ ($266{\sim}295$) days in male and female of the offspring, respectively. The gestation lengths of the recipients were $290.7{\pm}10.4$, $285.8{\pm}3.4$, $280.0{\pm}9.3$ and $286.1{\pm}7.9$ days in spring, summer, autumn and winter of the calving season, respectively, and were significantly different among the calving season (p<0.05). The birth weights of male and female calves were $25.5{\pm}2.4$ ($21{\sim}30$) and $26.3{\pm}3.9$ ($21{\sim}40$ kg in offsprings of the dairy recipients transferred with Hanwoo IVF embryos, respectively. The sex ratio was 90.7 in the offsprings of the dairy recipients transferred with Hanwoo IVF embryos.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2002년도 춘계학술발표대회 발표논문초록집
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• pp.69-69
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• 2002

To investigate the effects of two antioxidants, aesculeitin and taurine, expression of apoptosis related genes and antioxidant enzyme gene in preimplantation Hanwoo embryos was determined by modified semi-quantitative single cell RT-PCR. Hanwoo embryos derived from in vitro maturation /in vitro fertilization were cultured in 5% CO₂ and 5% O₂ in CR₁aa medium at 37℃. Aesculeitin and taurine were added to medium at concentration of l㎍/㎖ and 2.5mM, respectively. (omitted)

• 한국수정란이식학회지
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• 제19권3호
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• pp.209-218
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• 2004

가축에서 생산성을 높이기 위해 사용되는 수정란이식술의 실용성을 알아보기 위하여 한우 및 젖소 36두로부터 과배란 처리를 하여 신선수정란은 25두에, 동결수정란은 17두에, 쌍태 생산을 위한 수정란 두 개 이식은 13두 수란우에 이식되어 그 생산 결과를 조사하였다. 1 과배란을 위한 호르몬 처리후 반응을 나타낸 공란우는 75.0％이었다. 2. 황체수와 비교하여 채란된 수정란 율은 91.8％ 이었다. 3. 비외과적으로 채란된 전체 난자 중 생존성이 있는 배반포는 66.4％이었고, 이식 가능한 배반포는 48.6％이었다. 4. 간이채란법으로 시도된 one-catheter method에 의한 양 자궁 관류성공률은 75.0％이었다. 5. 신선 수정란을 이식후 분만결과에 따른 수태율은 60.0％이었다. 6. 동결수정란을 융해하여 이식후 분만결과에 따른 수태율은 35.3％이었다. 7. 쌍태 생산을 위한 수정란 두 개 이식결과에서 신선수정란 사용시 28.6％, 동결수정란 사용시 16.7％의 쌍태분만을을 나타내었다. 8.수정란 이식 후 생산된 산자에서 수컷의 비율은 56.3％이었다. 9. 수정란 이식 계절에 따른 분만 결과에서 여름에 이식시 가장 낮은 수태율을 나타내었다.

• Reproductive and Developmental Biology
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• 제32권3호
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• pp.175-181
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• 2008

This study was conducted to investigate an effective recipient oocyte and culture system for producing of Hanwoo (Korean native cattle) somatic cell nuclear transfer (SCNT) embryos. Hanwoo ear skin fibroblasts were used as donor cells. In vitro matured Hanwoo or Holstein oocytes were enucleated, and single donor cells were transferred into the perivitelline space of the enucleated oocytes. The couplets were subsequently fused and activated. The reconstructed embryos were cultured in a conventional or sequential culture system. In the former, embryos were cultured in CR2aa medium for eight days; in the latter, embryos were cultured in modified CR2aa-A (mCR2-A) for three days and then further cultured in modified CR2aa-B (mCR2-B) for five days. In the experiment with the recipient oocyte, the rate of embryo development to the blastocyst stage was significantly (p<0.05) higher in Hanwoo recipient oocytes than in Holstein ones (48.8% vs 38.9%). BIastocysts derived from Hanwoo recipient oocytes contained significantly (p<0.05) higher numbers of total cells than those derived from Holstein recipient oocytes ($156.0{\pm}68.2$ vs $134.7{\pm}54.8$). There was no difference in the mean proportion of apoptotic cells in blastocysts between the sources of recipient oocytes. In the experiment with the embryo culture system, the blastocyst rate was somewhat higher in sequential system than in conventional system (50.0% vs 43.5%), though there was no significant difference. The numbers of total ($160.0{\pm}69.0$ vs $156.7{\pm}68.4$) and apoptotic cells ($14.0{\pm}10.4$ vs $11.8{\pm}6.4$) were not different between the culture systems. In conclusion, the present study demonstrated that Hanwoo recipient oocytes and the sequential culture system were more effective in supporting the production of Hanwoo SCNT embryos.

• 한국동물생명공학회지
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• 제38권2호
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• pp.84-88
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• 2023

Because multiple ovulation embryo transfer (MOET) in cattle includes several benefits such as wide spreading of genetically superior offspring for long distance, this biotechnological method has been widely applied to Hanwoo. When the recipients are not stayed close after embryo recovery from donor, the embryos are moved to other farms via several vehicles (car, train, and airplane). However, air travel induces lesser oxygen level, increased vibration, lower air pressure, higher noise, and increased exposure of cosmic radiation to living things than ground level. It was still unknown that fresh embryos obtained from multiple ovulation of Hanwoo could maintain their fertility after being transported via air plane, the present case report introduced a clinical case of MOET in Hanwoo after shipping fresh embryos via air transportation. The donor was multi-ovulated via follicle-stimulating hormone series of injection, which was followed by a gonadotrophin-releasing hormone injection and artificial insemination twice. The embryos were recovered by the uterine flushing, packed in ministraws, transported to recipients for 6 h including 1 h air flight, and then transferred to the synchronized recipients. During pregnancy diagnosis of early gestation period, 5 of 7 recipients (71.4%) presented no heat signs and showed fetal sacs with fluid under transrectal ultrasonography. After normal gestation period, all recipients naturally delivered healthy calves (male n = 2 and female n = 3) without abortion, stillbirth, and premature birth. The present case report indicated that transportation of fresh embryos for MOET via domestic flight in Korea did not affect to their fertility.

• 한국수정란이식학회지
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• 제26권3호
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• pp.153-158
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• 2011

This study was performed in order to simplify the operation and minimize stress of donor and be readily available in the field with low cost and high quality embryos using the Direct Embryo Collection (DEC). Donors, at random stages of the estrous cycle, received a CIDR. 7 days later, 200 mg FSH was treated with 40, 30, 20, 10 mg FSH levels in declining doses twice daily by intramuscular injection for 4 days. On the 3rd day administration of FSH, 25 mg $PGF_2{\alpha}$ was administered and CIDR was withdrawn. After FSH injections were complete, donors were artificially inseminated twice at 12 hr intervals. The donor cattle received 250 ${\mu}g$ GnRH at time of 1 st insemination and embryos were recovered 8 days after the 1st insemination. Embryo collection from superovulated donors was performed to flushing by non-surgical methods of 3-way, 2-way and DEC (l-way). The average number of recovered embryos were 11.25${\pm}$0.63, 12.5${\pm}$0.65 and 11.75${\pm}$0.48 from operations of 3-way, 2-way and DEC methods, respectively. There were no significant differences among the embryo collection methods. Also, The average number of transferable embryos were 6.25${\pm}$0.48, 7.25${\pm}$0.48 and 7.25${\pm}$0.63 from each embryo collection procedures. The number of transferable embryos was no differences among the 3-way, 2-way and DEC methods, respectively. Meanwhile, the ratio of transferable embryos for all recovered embryos from DEC methods was higher as 61.7 % than 55.6 %, 58 % from methods of 3-way, 2-way. And the flushing solution required for recovering embryos by DEC method was significantly lower as 0.28${\pm}$0.32 1 than 1.8${\pm}$0.12 1, 1.75${\pm}$0.10 1 from 3-way, 2-way methods (p<0.05). Also, the time required for recovering embryos by DEC methods was significantly lower as 27${\pm}$2 min than 51${\pm}$3, 45${\pm}$2 min, respectively (p<0.05). In conclusion, these results suggest that DEC method for embryo collection may be effectively used for production of in vivo embryos using less flushing solution and, it might be effectively available in the field compared to conventional embryo recovery methods using 3-way or 2-way balloon catheter.

• Journal of Animal Science and Technology
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• 제62권4호
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• pp.449-459
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• 2020

Here, we investigated the effects of different nutrient requirements (NR) on blood metabolites, transferable embryo number after multiple superovulations with artificial insemination (AI), body condition score (BCS), and estrus cycle in Hanwoo cow. Nineteen Hanwoo cows were randomly divided into three groups (80%, 100%, and 120% NR, containing 6, 8, and 5 individuals, respectively) and fed based on the NR. In experiment 1, glucose, total cholesterol, triglyceride, blood urea nitrogen (BUN), aspartate aminotransferase (AST), alanine aminotransferase (ALT), non-esterified fatty acids (NEFA), albumin (ALB), and total protein (TP) were analyzed. In experiment 2, total number of recovered embryos and transferable embryos was examined after embryo recovery and multiple superovulations with AI. In experiment 3, body weight, BCS, and estrus cycle were examined. In experiment 1, total cholesterol was significantly different among the 80%, 100%, and 120% NR groups (126.5 ± 3.3, 152.6 ± 2.4, and 177.4 ± 1.8 mg/dL, respectively, p < 0.05). The triglyceride and BUN levels in the 120% NR group were significantly higher than those in the 80% and 100% groups (p < 0.05). The NEFA levels were significantly different among the 80%, 100%, and 120% NR groups (440.5 ± 18.2, 318.5 ± 23.1, and 195.1 ± 8.5 ЧEq/L, respectively, p < 0.05). The AST and TP levels in the 80% NR group were significantly lower than those in the 100% and 120% NR groups (p < 0.05). In experiment 2, the 120% NR group showed a higher percentage of transferable embryos than the 80% and 100% groups (p < 0.01). The mean body weight and BCS among the 80%, 100%, and 120% NR groups were significantly different (p < 0.05). The estrus cycle in the 80% NR group was delayed compared with the 100% and 120% NR groups (20.8 ± 0.2 and 21.2 ± 0.5 days, respectively). In conclusion, the blood metabolic tests proved that Hanwoo cows with 120% NR can produce a large number of transferable embryos. Thus, 120% NR is the appropriate feeding level for this type of cows as it results in the production of a large number of transferable embryos by multiple superovulations with AI.

• 한국가축번식학회지
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• 제24권3호
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• pp.299-309
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• 2000

본 연구는 한우 체외수정란의 정확한 성판별을 위한 PCR의 적정조건을 검토하였고, Y 염색체 특이적 DNA primer(BOV97M : 141bp)와 소 특이적 DNA primer(216bp)를 이용한 2단계 PCR 방법으로 체외수정란의 발육속도에 따른 성판별율과 성비를 검토하였다. PCR 기법을 이용한 한우 체외수정란의 성판별은 Y 염색체 특이적 DNA primer와 소 특이적 DNA primer로 정확히 성을 판별할 수 있었으며, 웅성 수정란의 경우 141bp와 216bp에서 band가 나타났고, 자성 수정란의 경우 216bp에서 band가 나타났다. 한우 체외수정란의 경우 성판별 정확도를 높이기 위하여 투명대의 제거가 필요하며, 수정란의 DNA 추출방법에 따른 Y 염색체 특이적 band의 출현율은 Proteinase K와 반복 동결융해방법이 각각 45.2 및 53.3%로 나타났다. DNA의 양에 따른 성판별율은 zona free인 경우는 2$\mu$1와 10$\mu$1에서 각각 97.2%와 92.2% 였으며, zona intact의 경우는 12$\mu$1와 13$\mu$1에서는 각각 91.5%와 93.6%로서 zona free 수정란의 경우 2$\mu$1, zona intact의 경우 13$\mu$1의 DNA 양으로 성판별이 가능하였다. 한편, PCR 기법에 의한 한우 체외수정란의 성판별율은 96.0%였으며, 정확히 성판별을 할수 없는 비율은 4.0%였다. 성판별 수정란의 자성과 웅성의 비율은 46.3%와 49.7%로서 성비는 1.1:1이였으며, 발육단계간 자웅성비는 커다란 차이가 인정되지 않았으며, 발육단계가 진행될수록 성판별 정확도가 증가하였다.

• 한국수정란이식학회지
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• 제27권4호
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• pp.253-258
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• 2012

This study was carried out to effects of ethylene glycol concentration, sucrose and culture day of in vitro production embryo on slow-down freezing in Hanwoo. 6, 7, 8 and 9 day embryos produced in vitro were frozen using 1.8M EG+0.1M sucrose, 1.8M EG+0.5% BSA and 1.5M EG+0.1M sucrose media. Survivability was confirmed after frozen-thawed 24 and 48h and ICM, TE cell number were counted by Hoechst 33342 and PI staining after frozen-thawed 24h. As a result, 1.8M EG+0.1M sucrose group was most significantly (p<0.05) higher compared with the other treatment groups on survivability, TE and total cell number after frozen-thawed 24h ($94.2{\pm}2.6%$, $94.67{\pm}3.4$ and $129.67{\pm}5.5$). ICM number did not found significant (p<0.05) differences between the three treatment groups. in 6, 7, 8 and 9 day of embryos using three types of freezing media, frozen-thawed, 1.8M EG+0.1M sucrose groups with embryos cultured 8 day was significantly (p<0.05) highest survivability to $98.3{\pm}1.7%$ after frozen-thawed 24h. 1.5M EG+0.1 sucrose group with embryos cultured 9 day was significantly higher survivability than group of embryos cultured 8 day after frozen-thawed 24 and 48h. In conclusion, 1.8M EG+0.1M sucrose media is considered to be effective to cryopreservation of embryos cultured 8 and 9 day.