• 한국수정란이식학회지
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• 제23권3호
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• pp.223-227
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• 2008

The cryopreservation of Hanwoo embryos has become an integral part of assisted reproduction in animal. The objective of this study was to assess the effect of The objectives of this study were: (1) to evaluate the influence of bovine embryo developmental stage on in vitro embryo development after freezing, (2) to study the efficiency compared with conventional freezed embryos at different embryo source. For conventional slow-freezing, day 7 or 8 expanded blastocysts were collected. The standard freezing medium was 1.8 M ethylene glycol (EG). Embryos were equilibrated in 1.8 Methylene glycol(EG) with 0.1 M sucrose in Dulbecco's phosphate-buffered saline (D-PBS) supplemented with 0.5% bovine serum albumin. Embryos were then loaded individually into 0.25 ml-straw and placed directly into cooling chamber of programmable freezer precooled to $-7^{\circ}C$, after 2 min, the straw was seeded, maintained at $-7^{\circ}C$ for 8 min, and then cooled to $-35^{\circ}C$ at $0.3^{\circ}C$/min, plunged and stored in liquid nitrogen for at least 3 days. For thawing, the straw containing embryos were warmed in air for 10 see and exposed to $37^{\circ}C$ water for 20 sec. Straws were then removed from $37^{\circ}C$ water. Rates of blastocyst survive and hatched were evaluated at 12 to 48h post-warming. The re-expansion and hatched rates of morula embryos were significantly lower than those obtained for blastocysts and expansion blastocysts (31.6%, 10.5% vs, 68.9%, 22.2% vs, 73.7%, 53.6%, respectively). No differences in re-expansion rates were found between in vivo and in vitro blastocysts. whereas hatched rates was significantly higher (51.2%) in vivo compared with in vitro embryos (18.6%). in conclusion, demonstrate that conventional freezing can be used successfully in cryopreservation of in vitro and in vivo bovine embryos, and that it might be considered for use in commercial programs and embryo preservation.

• 한국수정란이식학회지
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• 제10권2호
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• pp.163-169
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• 1995

In the experiment I for maas production of bovine early embryos, 18~20hpi fertilized eggs (756 eggs) and parthenogenic eggs (618 eggs) which were treated by 10% ethanol were cultured in both TGM and CZB. In the experiment II, suppiment effects each in CZB and CRlaa were tested by matured and fertilized oocytes which were after 18~20hpi. In the case of experiment I after 48hr, the cleavage rates of normally fertilized eggs were 66.6% in TCM treatment and 77.7% in CZB treatment, and after 240h the blastocysts were 7.5% in TCM and 14.1% in CZB. In the parthenogenic eggs, the deavage rates at 48hr were 39.6% in TCM and 57.5% in CZB, and at 240h, the blastocysts were 0.9% in TCM and 4.4% in CZB. These results showed that the effects of CZB on developmental ability to parthenogenic eggs as well as nomally fertilized eggs are relatively high. In experiment W, the effect of exposing the cleaved embryos to CZB for 30h on the blastocyst formation was examined. Similar rates of blastocyst formation were obtained both in TCM and CZB, suggesting that CZB exposure. during ealry development is critical. In experiments III ~ V, the effects of supplements were examined. The cleavage rates of CZB treatments at 48h were 83.8% in control, 78.1% in BSA+A.A+SIT, 75% in 5% FCS+A. A+SIT, 88.6% in BSA+A.A+SIT and not co-cultured BSA+A.A+SIT had 85.7% and in the case of 240h blastocysts showed 22.6, 0.0, fl.1, 6.5 and 0%, respectively. As a result, this study showed that CZB was effective culture system for in vitro development, and that CZB and CR$_1$aa had no significant differences and effects between them. It may be concluded that in the simple media containing supplements could replace the co-culture systems of bovine early embryo development.

• 한국가축번식학회지
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• 제25권2호
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• pp.181-189
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• 2001

본 연구에서는 ICSI에 의한 돼지 체외수정란의 생산효율을 높이고자, ICSI에 사용되는 돼지 체외성숙 난포란의 원심분리에 의한 효과, 세포질내에 주입하는 정자의 DTT전처리 효과, 체외성숙 난포란의 체외수정시 주로 사용되는 IVF방법과 ICSI방법 사용에 따른 수정율과 후기배로의 배발달율 조사 그리고 IVF방법과 ICSI방법에 의해 발달한 배반포기배의 할구수를 조사하여 다음과 같은 결과를 얻었다. ICSI에 사용되는 돼지 체외성숙 난포란을 이용하여 원심분리를 실시하지 않은 군과 실시한 군에 있어서 수정율과 후기배로의 발달율에 있어서는 두 처리군간에 유의적인 차이는 나타나지 않았으며, 또한 ICSI에 사용되는 정자를 주입전에 DTT를 처리한 군과 DTT를 처리하지 않은 군에서의 수정율과 배반포기배로의 발달율에 있어서도 유의적인 차이는 나타나지 않았다 그리고 체외성숙 후 IVF 또는 ISCI에 의한 체외수정 후 수정율과 후기배로의 발달율에 있어서도 두 처리군간의 유의성은 나타나지 않았다. IVF와 ICSI에 의해 체외수정된 수정란에 있어서 8일째까지 발달한 배반포기배의 할구수를 조사한 결과 각각 46.7$\pm$2.9개와 41.9$\pm$4.6개로 나타나 두 처리군간에 유의적인 차이를 나타내지 않았다. 이상의 실험 결과들을 종합해 보면, ICSI에 의한 체외수정란의 생산은 체외성숙후 난구세포가 제거된 난포란을 원심분리시키면 ICSI에 있어서의 편리를 도모할 수 있을 뿐만 아니라 주입된 정자의 관찰을 용이하게 하여 정확성을 높일 수 있고, 주입시 사용하는 배양액의 난포란내 진입을 최소화 할 수 있을 것으로 사료된다. 그러나 ICSI전 정자의 첨체반응 유도와 난포란의 활성화, 그리고 후기배로의 발달율 향상에 따른 할구수의 증가를 위한 연구가 진행되어야 할 것으로 사료된다. 지금까지의 이러한 결과들을 바탕으로 하여 ICSI에 의한 체외수정란의 생산을 위한 연구가 지속적으로 수행해 나가야 할 것이며, 또한 계속적인 연구로 ICSI를 이용하여 외부유전자의 도입에 의한 형질전환동물의 생산이 가능 할 수 있을 것으로 사료된다.

• Clinical and Experimental Reproductive Medicine
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• 제37권4호
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• pp.307-319
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• 2010

목적: 유리화 동결액의 조성조절과 냉각속도 증진을 통한 동결보호제의 농도를 낮추는 전략을 통해 세포에 미치는 독성을 감소시켜 유리화 동결 및 융해 후 생쥐 배아의 생존율 및 발생률을 증진시키고, 궁극적으로 배아의 유리화 동결법을 개선하고자 하였다. 연구방법: 생쥐 배아와 포배기를 그리드를 이용한 유리화 동결법을 이용하여 동결/융해하였다. 동결액 내 ethylene glycol와 dimethylsulphoxide (DMSO)의 농도와 당의 농도를 조절하여 생쥐 배아의 융해 후 생존율과 발생률을 관찰하였고, 냉각속도의 증가와 동결억제제의 농도와의 상관관계를 포배기의 융해 후 생존율과 발생률에 따라 비교하였다. 또한 융해 후 배아를 대리모에 이식하여 산자를 생산함으로 냉각속도의 효율성을 알아보았다. 결과: EG를 단독으로 사용한 동결보존액 보다는 DMSO와 혼합된 동결보존액의 사용이 보다 유리하다는 결과를 얻을 수 있었다. 슬러시 질소에 의한 냉각속도의 증가가 동결보존의 대상의 상해를 줄여 유리화 동결의 효율성을 증진시키는 것으로 생각된다. 결론: 혼합된 동결보호제를 사용하였을 때 생쥐 배아의 유리화 동결 후 생존과 발생률이 증진되었다. 슬러시 질소를 이용한 유리화 동결의 도입은 냉각속도의 증가를 통해 기존 유리화 동결방법의 효율을 증진시켜 생존율과 융해후 발생률, 임신율 증진에 기여하였다. 또한 냉각속도의 증진은 유리화 동결의 필수요건인 고농도의 동결억제제에 대한 노출을 감소시킬 수 있었다. 이러한 노력은 생식력의 보전을 위한 유리화 동결법의 효율 향상에 기여할 것이다.

• Reproductive and Developmental Biology
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• 제32권4호
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• pp.245-248
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• 2008

In the present studies, we have intended to compare the EDS (20% EG + 20% DMSO + 0.4 M sucrose + 10% FCS) and EDT (20% EG + 20% DMSO + 0.3 M trehalose 10% FCS) methods for vitrification of canine oocytes, in order to improve the vitrification methods. The survival rate of vitrified-thawed oocytes using the EDS method was $15.1{\pm}1.8%$ (p<0.05), which was lower than that of the control group $(66.7{\pm}2.5%)$. About $45{\sim}55%$ of the vitrified-warmed oocytes showed normal morphology, as assessed by PI staining. However, the ratio of survival rate of oocytes showed lower than that of normal morphology in comparison between EDS method and control group. The survival and developmental rates of vitrified-warmed oocytes by the EDS and EDT methods were $16.7{\pm}1.4%\;and\;11.1{\pm}0.8%$ and $8.3{\pm}1.4%\;and\;4.4{\pm}1.8%$, respectively (p<0.05). The results were significantly lower than the control group $(66.7{\pm}2.5%\;and\;16.7{\pm}3.7%)$. However, the survival rate of vitrified-warmed oocytes using EDS method showed higher than that in the ETS group.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.125-125
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• 2003

The objective of this study was to evaluate the effect of GnRH or estradiol in a CIDR-based timed Al (TAI) protocol on follicular turnover, synchronized ovulation and pregnancy rates in Holstein cows. Cows were treated at random stages of the estrus cycle with an insertion of an intravigal progesterone (1.9 g) device (CIDR, Day 0) and either no other treatment (control group; n=10), injection of 100 ug fertirelin acetate (GnRH group; n=10) or 4 mg estradiol benzoate (estradiol group; n=10). Seven days later devices were removed and an injection of 25 mg $PGF_{2$\alpha$}$ was administered. On Day 9, 100 ug GnRH was administered. Cows received a fixed-time insemination 16 h after injection of the GnRH. (중략)

• 한국가축번식학회지
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• 제13권3호
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• pp.141-148
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• 1989

The effects of seeding method and optimum time for freezing embryos according to the developmental stages on embryo survival rates after rapid freezing were determined using the FDA-test. The summarized results are as follows : 1. In the rapid freezing of embryos, the sucrose added medium together with Co-seeding or non-seeding showed the FDA scores of 4.67 and 4.20, respectively, but, raffinose addition obtained FDA scores of 4.27 and 3.97. 2. The developmental stage of embryos at freezing was most critical on the survival of embryos after thawing. Higher FDA scores were obtained in the order of blastocyst stage(4.94), morula stage(3.82) and ealy stage(2.65) in sucrose added medium. The same trend was observed in the raffinose added medium with an order or 4.91, 4.47 and 2.32. 3. Microscopic study of embryo before freezing and post-thawing indicated that the embryo showed shrinkage within 5 minutes after the embryo was transfer to the freezing medium. When thawed embryo was tranfered to the dilution medium, swelling of the embryo was observed and there after it reshrank indicating the removal of cryoprotectant from the embryo. The size of the embryo recovered to the original state when it was moved into a PBS-solution.

• Applied Microscopy
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• 제44권4호
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• pp.111-116
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• 2014

This study investigates the developmental pattern of adhesive discs (ADs) to highlight the ontogeny and structural changes that occur during the growth of Boston ivy. Initiation to postmortem features of ADs were examined through light and scanning electron microscopy. The study also reveals a new finding of the dislocation of peripheral tissues of adaxial origin. Four phases of attachment are suggested with regards to its climbing behavior: 1) pre-attachment, 2) upon attachment, 3) after attachment, and 4) final attachment. During initiation, several ADs originate from tendril primordia without epidermal differentiation. However, different growth rates in the epidermis results in completely different ADs. ADs were discerned by size, shape, and color during expansion, but cells in the adaxial surface remained alive longer than the other side. Upon contact, the ADs demonstrate simultaneous growth and deterioration, but once attachment is established the latter process subdues to final stages. Epidermal transformation, adhesive secretion, cellular disruption, and mechanical stress were essential for the self-clinging nature of Boston ivy. The post-attachment sequence is also believed to be critical in achieving maximum mechanical strength to provide extensive support. The developmental process of ADs is prompted by tactile stimulation but in a highly organized and systematic manner.

• Proceedings of the National Institute of Ecology of the Republic of Korea
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• 제2권4호
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• pp.279-284
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• 2021

The dung beetle, Gymnopleurus mopsus (Coleoptera: Scarabaeidae), is one of endangered species in South Korea. It was last recorded in 1971. To restore this species, we introduced G. mopsus populations from eastern and southern regions of Mongolia in July 2019 and August 2019, respectively. One of the main tasks for the restoration of endangered insects is to develop breeding techniques to eventually incorporate these insects into the restoration system. In a series of laboratory experiments, we investigated effects of short-term hibernation periods on life-history traits of G. mopsus. Adult G. mopsus that had hibernated for 30, 60, and 90 days had lower survival rates than adults that had hibernated for 120 days. We also compared developmental time of these four experimental groups and found a significant difference in the egg - phase. However, the duration of hibernation did not affect the fecundity, brood-ball size, or body size of F1 adults. Follow-up studies are currently being conducted to further investigate the effect of a short-term hibernation period on population growth of G. mopsus under laboratory conditions.

• 농업과학연구
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• 제45권3호
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• pp.493-498
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• 2018

There has been widespread warming and a general increase in summer temperatures over the Korean peninsula ($0.5^{\circ}C$/10 years from 2001 to 2010). South Korea is transforming into a subtropical region, and the productivity of livestock is affected by the climatic changes. In this study, we investigated whether the summer heat waves affect the developmental competency of Korean native cattle (Hanwoo), a taurine type of cattle with a small portion of indicine varieties. We collected oocytes during the summer (heat stress, HS) and autumn (non-HS condition) and examined the developmental competencies including in vitro maturation and preimplantation embryo development. No significant differences were observed between the HS and non-HS oocytes in nuclear maturation (extrusion of the polar body); however, the cleavage and blastocyst rates were significantly lower in the HS group than those in the non-HS group. The lower developmental competence of the HS oocytes compared to the non-HS is, in part, due to insufficient cytoplasmic maturation because of a higher production of Reactive oxygen species (ROS) levels as well as peri/cortical distributed mitochondria in the HS oocytes after in vitro maturation. Next, we examined the ROS and mitochondria distribution and found a significant increase in the levels of ROS in the HS oocytes and a polarized distribution (pericortical cytoplasm) of mitochondria in the HS oocytes. In summary, impaired cytoplasmic maturation of oocytes from exposure to HS affects the preimplantation embryo development by dysfunction of mitochondria. To improve reproductive performance, embryo transfer using cryopreserved embryos/oocytes is recommended in the hot summer season of South Korea.