• Asian-Australasian Journal of Animal Sciences
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• 제23권4호
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• pp.456-464
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• 2010

RNA interference (RNAi) is an acknowledged useful and effective tool to study gene function in various cells. Here, we suppressed the Connexin 43 (Cx 43) gene expression during in vitro development of ovine pre-implantation embryos using the RNAi method. The 353 bp Cx 43 double-stranded RNA was microinjected into in vitro fertilized ovine zygotes, and the levels of target mRNA and protein were investigated. Control groups included uninjected zygotes or those injected with RNase-free water. The dsRNA injection resulted in the specific reduction of Cx 43 transcripts as analyzed by quantitative real-time RT-PCR and decreased protein levels as shown by Western blot analysis at the blastocyst stage. Microinjection of Cx 43 dsRNA led to 20.3%, 21.7% and 34.5% blastocyst rates and 19.2%, 37.5% and 41.3% hatched blastocyst rates in Cx 43 dsRNA-injected, water-injected and uninjected groups, respectively. Then the RNAi could not significantly affect cell number and cell death rates of blastocysts. Therefore, suppression of Cx 43 dsRNA and proteins did not apparently affect the development potential of ovine pre-implantation embryos but may play a role in embryo quality. RNAi technology is a promising approach to study gene function in early ovine embryogenesis.

• 한국동물생명공학회지
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• 제37권4호
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• pp.226-230
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• 2022

The sheep can be reproduced by natural mating as well as applied reproductive biotechnology, embryo transfer (ET). However, this method in sheep is influenced by several factors such as season, photoperiod, latitude, temperature, nutrition, and breed. In addition, there is still less research on assisted reproductive technologies in small ruminants, compared to other livestock species such as cattle and pigs. Because there has been a need for an optimization and a continuous improvement of ET techniques in small ruminants. the main objective of this study was to evaluate the conception rate obtained after ET in Mongolian sheep (Dorper breed). After embryo recover, code 1 and 2 embryos (morula or blastocyst stage) for ET in the present study were 63% (63/100) and 24% (24/100), respectively. Then Each single embryo was transferred to a synchronized recipient who prepared by estrous synchronization protocol with fluorogestone acetate-cloprostenol sodium. The results demonstrated that an average conception rate and lambing rate was 35.6% (31/87) and 33.3% (29/87), respectively. Further study is still necessary, but these results indicated that single embryo of Mongolian sheep with the present protocol was enough to conducting ET when the genetically superior sheep were necessary to be expanded.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2001년도 발생공학 국세심포지엄
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• pp.25-34
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• 2001

The Committee has, for the 9th consecutive year, compiled statistics of embryos collected and transferred worldwide. Geographically, all regions have participated in the survey; however, there are still some places where it has been impossible to retrieve such data (particularly in Asia). The statistics, therefore, are partially underestimated. By contrast, in other areas, the system has proven to be more efficient than before, particularly in North and South America. This has resulted in the present report, which gives a somewhat more satisfactory picture of the current situation of the ET industry. For the second year, it has also been possible to collect data for several species other than cattle. In cattle, the number of in vivo-derived embryos collected and transferred has once again increased with more than half a million embryos trans-ferred in 1999(520,712), a new record. The number of bovine in vitro-produced embryos has remained stable this year as compared with the previous year(approximately 30,000 embryos transferred). However, there are still teams that have not Yet reported their data. More than 10,000 embryos in each of the ovine and caprine species and close to 2,000 cervid embryos were reported transferred in 1999. Some 500 embryos of the equine species and a few thousand in the porcine and rabbit species have also been transferred in 1999. It is concluded that the ET industry continues to be very active, and, in many species, it encompasses a larger segment of the livestock population overall, which is to the farmer s benefit.

• Journal of Animal Science and Technology
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• 제57권8호
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• pp.27.1-27.6
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• 2015

This study was designed to investigate the effects of ${\alpha}$-tocopherol and granulosa cells monolayer on nuclear maturation and cleavage rates of ovine cumulus-oocyte complexes (COCs). The COCs (n = 2814) were matured in maturation medium supplemented with various concentration of ${\alpha}$-tocopherol (0, 5, 10, $15{\mu}g/ml$), oocytes were incubated at $39^{\circ}C$ with 5 % $CO_2$ for 24 h in three culture systems: (a) maturation medium (MM; n = 884), (b) co-cultured with granulosa cells (CG; n = 982) and (c) co-cultured with granulosa cells and cells were further cultured in MM for 12 h (CG + 12hMM; n = 948). Our results showed that ${\alpha}$-tocopherol had no effect on GVBD and MII as compared to control group, but when ${\alpha}$-tocopherol added to maturation medium the rate of cleavage decreased. This indicates interaction of above mentioned factors in any of the treatments showed no significant differences on the rate of maturation and cleavage stages (MII, GVBD and cleavage) (p > 0.05). The oocytes co-cultured with granulosa cells for 24 h had beneficial effects on cleavage rate. The maximum MII and cleavage rates were achieved when oocytes had extra 12 h culture in the maturation medium without granulosa cells. Results also showed our modified co-culture system (CG + 12hMM), improved rates of MII and the cleavage in comparison with other studied maturation systems.

• 한국수정란이식학회지
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• 제13권3호
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• pp.235-243
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• 1998

적정 배양액의 선정, ITS의 첨가와 BSA의 농도조절 및 NaCl 농도의 조절을 통해 소 수정란의 무혈청, 체세포배제 배양체계를 확립하기 위하여 수행한 실험에서 다음과 같은 결론을 얻었다. 1. 배양액으로 CRlaa, TALP 및 SOF를 사용하여 발육률을 검토한 결과, 발육률의 유의적인 차이는 나타나지 않았다. 2. 배양액내의 고분자물질원으로 BSA, FBS 및 PVA를 첨가하여 사용한 결과 BSA 및 FBS 첨가군이 PVA 첨가군보다 유의적으로 높은 발육률을 나타내었다(p〈0.01). 3. 배양액내의 BSA 농도를 달리 하면서(1, 3, 8 mg/ml) 1% ITS를 첨가하여 실험한 결과 BSA의 농도가 증가할수록 후기배로의 발육률이 높았으며 모든 군에서 ITS 첨가군이 후기배로의 발육률이 높았으나 BSA가 1 mg/ml로 첨가된 군에서만 ITS 첨가에 따른 유의적인 차이가 인정되었다(p〈0.05). 4. 배양액내의 N3Cl 농도를 114 mM과 90 mM 로 나누어 소 수정란을 배양한 결과 90 mM 군의 후기배로의 발육률이 114 mM 군에 비해 유의적으로 높게 나타났다(p〈0.05).

• Journal of Animal Science and Technology
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• 제63권2호
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• pp.272-280
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• 2021

Cumulus-oocyte complexes (COCs), which contain immature oocytes, are matured in vitro for in vitro embryo production. Oocyte and cumulus cells are then separated using hyaluronidase. To date, there have only been a few reported cases of the toxic effects of hyaluronidase on porcine oocytes. The aim of this study was to compare the effects of bovine testis-derived hyaluronidase and recombinant human hyaluronidase on oocyte denudation and quality. Porcine COCs were matured for 44 h and denuded using different hyaluronidase concentrations and exposure times. Then, oocytes were activated by electrical parthenogenesis. In experiment 1, COCs were denuded using bovine-derived, ovine-derived (Hirax), and human recombinant (ALT-BC4) hyaluronidases for 10 and 20 min. In experiment 2, bovine-derived and human recombinant (ALT-BC4 and ICSI Cumulase®) hyaluronidases were used to denude the COCs for 2 and 20 min. In both experiments the oocytes were all completely denuded, and there was no degeneration. Rate of embryo development was significantly increased in group treated ALT-BC4 for 2 min and not significantly different in other treatment groups. In general it slightly decreased with longer exposure times. These results have confirmed that different sources of hyaluronidase do not have detrimental effects on the quality of porcine oocytes and suggest that the human recombinant hyaluronidase ALT-BC4 is suitable for oocyte denudation with an increased blastocyst rate.

• Asian-Australasian Journal of Animal Sciences
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• 제16권5호
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• pp.650-654
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• 2003

The accuracy of Polymerase chain reaction (PCR) assay in sexing of sheep embryos was assessed in this study. A total of 174 ovine embryos produced in vitro at different stages of development (2, 4-8 cell stages, morula and blastocyst) were sexed. The universal primers (P1-5EZ and P2-3EZ) used in this assay amplified ZFY/ZFX-specific sequences and yielded a 445 bp fragment in both sexes. Restriction enzyme analysis of ZFY/ZFX-amplified fragments with Sac I exhibited polymorphism between sexes, three and two fragments in males and in females, respectively. For verification of accuracy, blood samples of known sex were utilized as positive controls in each test. The mean percentages of sex identification by this method at 2 cell, 4-8 cell, morula and blastocyst were $73.00{\pm}5.72$, $89.77{\pm}3.79$, $3.33{\pm}8.08$ and $79.6{\pm}9.09$, espectively with the over all male to female ratio of 1:0.87. It is concluded that the ZFY/ZFX based method is highly reliable for the sexing of sheep embryos.

• Journal of Animal Science and Technology
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• 제51권6호
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• pp.471-484
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• 2009

The peri-implantation period in mammals is critical with respect to survival of the conceptus (embryo/fetus and associated extraembryonic membranes) and establishment of pregnancy. During this period of pregnancy, reciprocal communication between ovary, conceptus, and endometrium is required for successful implantation and placentation. At this time, interferon tau (IFNT) is synthesized and secreted by the mononuclear trophectodermal cells of the conceptus between days 10 and 21~25. The actions of IFNT to signal pregnancy recognition and induce or increase expression of IFNT-stimulated genes (ISGs), such as ISG15 and OAS, are mediated by the Type I IFN signal transduction pathway. This article reviews the history, signaling pathways of IFNT and the uterine expression of several IFNT-stimulated genes during the peri-implantation period. Collectively, these newly identified genes are believed to be critical to unraveling the mechanism(s) of reciprocal fetal-maternal interactions required for successful implantation and pregnancy.