• Asian-Australasian Journal of Animal Sciences
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• v.22 no.4
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• pp.483-491
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• 2009

In vitro produced porcine embryos have potential application in reproductive biotechnology. However, their development potential has been very low. This study evaluated the in vitro developmental ability and quality of cloned and parthenogenetic porcine embryos having 2-4 cells or 5-8 cells on Day 2 of in vitro culture. Analysis of results showed that 2 to 4 cell embryos had higher ability to form blastocysts than 5 to 8 cell embryos (p<0.05). Blastocysts produced from culture of 2 to 4 cell embryos also contained higher cell numbers and had lower BAX:BCLxL transcript ratio than those produced from 5 to 8 cell embryos (p<0.05), thereby suggesting 2 to 4 cell embryos have higher development potential. Further investigation revealed that 5 to 8 cell embryos had higher incidence (100${\pm}$0.0%) of blastomeric fragmentation than 2 to 4 cell embryos (15.2${\pm}$5.5% for parthenogenetic and 27.7${\pm}$7.1% for cloned embryos). This suggests that low development potential of 5 to 8 cell embryos was associated with blastomeric fragmentation. In conclusion, we have shown that morphological selection of embryos based on cell number on Day 2 of in vitro culture could offer a practical and valuable non-invasive means to select good quality porcine embryos.

• Journal of Embryo Transfer
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• v.11 no.3
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• pp.217-223
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• 1996

This study was conducted to investigate the effects of co-culture for the development rate to morula /blastocyst stages of early porcine embryos, derived from oocytes matured and fertilized in vitro, with porcine endometrial cell monolayers(PEM) in the two different media, respectively. The rates of embryos developed to 2-, 4-, 8~16-cell and morula /blastocyst stage were 49.6, 40.5, 28.2 and 15.3% in Ham's F-10 with PEM, and 55.3, 45.9, 32.7, and 17.6% in TCM-HEPES with PEM, respectively. The above development rates to morula /blastocyst stages were significantly higher than those of the embryos cultured in the Ham's F-10 and TGM-HEPES without PEM(P<0.05). The in vitro development rates to the morula /blastocyst stage of 1-cell embryos cultured in Ham's F-10 and TCM-HEPES without PEM were 0~1.2%. Especially, most of embryos were observed to arrest the development beyond 4-cell stages. As shown in the above results, the co-culture of in vitro produced porcine embryos with PEM in the two different media enhanced the development of fertilized eggs to morula /blastocyst stages in vitro. However, we didn't find out any differences for the in vitro development to morula /blastocyst stages between Ham's F-10 and TcM-HEPES media.

• Korean Journal of Animal Reproduction
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• v.20 no.3
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• pp.299-305
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• 1996

This study was conducted to investigate the effects of co-culture for the development rate to morula/blastocyst stages of early porcine embryos, derived from oocytes matured and fertilized in vitro, with porcine oviductal epithelial cell monolayers(POEC) in the two different media, respectively. The rates of embryos developed to 2-, 4-, 8∼16-cell and morula/blastocyst stage were 57.2, 48.2, 37.2 and 19.3% in Ham's F-10 with POEC, and 51.4, 41.2, 31.1, and 15.5% in TCM-HEPES with POEC, respectively. The above development rates to morula/blastocyst stages were significantly higher than those of the embryos cultured in the Ham's F-10 and TCM-HEPES with out POEC(P<0.05). The in vitro development rates to the morula/blastocyst stage of 1-cell embryos cultured in Ham's F-10 and TCM-HEPES without POEC were 1.1∼1.2%. Especially, most of embryos were observed to arrest the development beyond 4-cell stages. As shown in the above results, the co-culture of in vitro produced porcine embryos with POEC in the two different media enhanced the development of fertilized eggs to morula/blastocyst stages in vitro. However, we didn't find out any difference for the in vitro development to morula/blastocyst stages between Ham's F-10 and TCM-HEPES media.

• Journal of Embryo Transfer
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• v.15 no.2
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• pp.121-128
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• 2000

This study was conducted to enhance the efficiency of embryo transfer and embryos produced in vitro according to select good quality blastocyst among in vivo or in vitro produced embryos. After middle blastocys(MB) and late blastocyst(LB) stages embryos that developed in vitro for 7∼8 days were treated hypertonic solution(380 mOsm or 600 mOsm) and the rate of development to hatched blastocyst(HB) was examined. The results obtained were summarized as follows: The proportion of embryos developed to HB was higher than that of control regardless of hypertonic solution treatment. However, it was related to culture time in hypertonic solution The propotrion of embryos developed to HB of morphologically recovered embryos in 380 nOsm solution ws higher than that of morphologically shrinked embryos. However, treatment time was not significantly different the rates of HB development. Especially, the proportion of embryos developed to HB of morphologically recovered embryos of the 30-min-treated group was significantly higher than that of control (p<0.05). The results suggest that hypertonic solution treatment should enhance the efficiency for criterion of transferable blastocyst quality.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.1
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• pp.33-38
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• 2004

This study was aimed at testing the gene expression of antioxidant enzymes and apoptosis genes for in vitro culture in porcine embryos produced by in vitro maturation/in vitro fertilization (IVM/IVF). Pocine preimplantation embryos obtainted from IVM/IVF can be successfully culture in vitro, but they are delayed or stop to develop at specific developmental stage. Many factors such as reactive oxygen species and apoptosis in an IVM/IVF system followed by in vitro culture influence the rate of production of viable blastocysts. Porcine embryos derived from IVM/IVF were cultured in the atmosphere of 5% $CO_2$ and 20% $O_2$ at $38.5^{\circ}C$ in NCSU23 medium. The patterns of gene expression for antioxidant enzymes and apoptosis genes during in vitro culture in pocine IVM/IVF embryos were examined by the modified semi-quantitative single cell reverse transcriptase-polymerase chain reaction (RT-PCR). Porcine embryos produced by in vitro procedures were expressed mRNAs for CuZn-SOD, GAPDH and GPX, whereas transcripts for Mn-SOD and catalase were not detected at any developmental stages. Expression of caspase-3 mRNA was detected at 2 cell, 8 cell 16 cell and blastocyst, but p53 mRNA was not detected at any stages. The fas transcripts was only detected in blastocyst stage. These results suggest that various antioxidant enzymes and apoptosis genes play crucial roles in vitro culture of porcine IVM/IVF embryos.

• Korean Journal of Animal Reproduction
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• v.18 no.4
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• pp.251-256
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• 1995

Two experiments were conducted to study the production of in vitro fertilized bovine embryos and the viability of blastocysts cryopreserved by vitrification. In experiment 1, production rate of in vitro matured bovine oocytes after fertilization in medium containing bovine oviduct epithelial cells (BOEC), cumulus cells and granulosa cells to blastocysts were 18.4, 14.6 and 13.1%, respectively. Developmental percentages of blastocysts produced at day 6, 7 and 8 were 8.5, 10.6 and 15.2% respectively. Hatching rate of bovine embryos produced was 60.0%. In experiment 2, post-thawed surviving embryos in a vitrification solution consisting of 7.15M ethylene glycol, 2.5 mM ficoll and 0.3 M sucrose were 36.4% (56/154). Also, survival rate of bovine embryos after exposed to vitrification solution at 1, 2, 3, 4 and 5 min were 84.0, 88.0, 71.0, 48.0 and 24.0% respectively.

• Korean Journal of Veterinary Research
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• v.36 no.4
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• pp.919-927
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• 1996

In the last few years, methods for in vitro culture of early embryo stages from oocytes matured and fertilized in vitro using suitable cell culture systems have been established. But the factors affecting pregnancy rates following transfer of bovine embryos produced in vitro were not evaluated enough. So this study was performed to investigate the effects of quality and stage of embryos, parity and Corpus Luteum quality of recipients on pregnancy rates following non-surgical transfer of bovine embryos produced in vitro. Oocytes aspirated from small antral follicles of ovaries obtained at a local slaughter house were matured, fertilized with frozen-thawed semen and co-cultured for 6-7 days by utilizing co-culture system with bovine oviduct epithelial cell in vitro. After co-culture, embryos were transfered to recipients on day 7 (estrus=day 0). Recipients were monitored by ultrasonic scanning method or observation for estrus and rectal palpation after 50 days from transfer. The results of this study are follows. 1. Of the 70 recipients, 70%(49 of 70) had not showed estrus sign between day 0 and day 50, but 22.9%(16 of 70) was diagnosed not pregnant. Therefore the overall pregnancy rate of this study was 47.1%(33 of 70). 2. The pregnancy rate of recipients transfered with excellent(66.7%) and good(54.5%) embryos were higher than that of recipients transfered with fair embryos(15.8%) (p<0.05). 3. The pregnancy rate of recipients transfered with morula, compacted morula, blastocyst and expanded blastocysts were 46.2, 55.0, 62.5 and 50.0%, respectively. 4. The pregnancy rates of recipients transfered to heifer and cow were 54.5 and 55.2%, respectively. 5. The pregnancy rates of recipients with CL score I, II(66.7, 63.6%) were higher than those of recipients with CL score III (10%), (p<0.05). Success of transfer of embryos produced in vitro depends on many variables. The important factors identified in this study were the quality of embryos and the CL score of recipient animals after non-surgical transfer of embryos matured, fertilized and cultured in vitro.

• Journal of Embryo Transfer
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• v.24 no.1
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• pp.71-76
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• 2009

It is not easy for porcine embryos produced by in vitro systems to develop into blastocysts with high quality. To solve this problem, many researchers have developed novel culture methods. However, the formation of blastocysts with high quality is still low. In this study, we aimed to produce piglet following transfer of in vitro produced early embryos ($2{\sim}4$ cell stage embryos) or morula and blastocyst. The $2{\sim}4$ cell stage embryos were transferred to five estrus-synchronized recipients (200 embryos per recipient). One of the five sows farrowed three piglets, which contain two live piglets and one dead piglet, 114 days after embryo transfer. However, two recipients transferred with morula and blastocysts did not farrow. Microsatellite analysis confirmed that the genomic DNA of two live piglets were not genetically identical to that of the recipient. These results indicate that it is possible to obtain piglets by transfer of early embryos produced by in vitro production (IVP) systems.

• Animal Bioscience
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• v.35 no.2
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• pp.177-183
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• 2022

Objective: The present study evaluated the efficiency of embryo development and pregnancy of somatic cell nuclear transfer (SCNT) embryos using different source-matured oocytes in Camelus dromedarius. Methods: Camelus dromedarius embryos were produced by SCNT using in vivo- and in vitro- matured oocytes. In vitro embryo developmental capacity of reconstructed embryos was evaluated. To confirm the efficiency of pregnancy and live birth rates, a total of 72 blastocysts using in vitro- matured oocytes transferred into 45 surrogates and 95 blastocysts using in vivo- matured oocytes were transferred into 62 surrogates by transvaginal method. Results: The collected oocytes derived from ovum pick up showed higher maturation potential into metaphase II oocytes than oocytes from the slaughterhouse. The competence of cleavage, and blastocyst were also significantly higher in in vivo- matured oocytes than in vitro- matured oocytes. After embryo transfer, 11 pregnant and 10 live births were confirmed in in vivo- matured oocytes group, and 2 pregnant and 1 live birth were confirmed in in vitro- matured oocytes group. Furthermore, blastocysts produced by in vivo-matured oocytes resulted in significantly higher early pregnancy and live birth rates than in vitro-matured oocytes. Conclusion: In this study, SCNT embryos using in vivo- and in vitro-matured camel oocytes were successfully developed, and pregnancy was established in recipient camels. We also confirmed that in vivo-matured oocytes improved the development of embryos and the pregnancy capacity using the blastocyst embryo transfer method.

• Journal of Animal Reproduction and Biotechnology
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• v.38 no.3
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• pp.143-150
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• 2023

Background: The successful production of superior or transgenic offspring from in vitro produced embryos in cattle relies heavily on the quality of blastocyst stage embryos. In order to enhance the developmental competency of these embryos, a novel culture method was devised. Methods: This study utilized stem cell culture medium (SCM) from hESCs as a supplement within the culture medium for bovine in vitro produced embryos. To gauge the efficacy of this approach, in vitro fertilized embryos were subjected to culture in CR1aa medium enriched with one of three supplements: 0.3% BSA, 10% FBS, or 10% SCM. Results: The blastocyst development and hatching rates of one-cell zygotes cultured in CR1aa medium supplemented with SCM (23.9% and 10.2%) surpassed those cultured in CR1aa medium supplemented with BSA (9.3% and 0.0%) or FBS (3.1% and 0.0%) (p < 0.05). Furthermore, post-zygotic gene activation, cleaved embryos cultured in CR1aa medium supplemented with SCM (57.8% and 34.5%) exhibited notably higher rates (p < 0.05) compared to those cultured with BSA (12.9% and 0.0%) or FBS (45.7% and 22.5%) supplementation. Furthermore, the microinjection of SCM into the cytoplasm or pronucleus of fertilized zygotes resulted in elevated blastocyst development and hatching rates, particularly when the microinjected embryos were subsequently cultured in CR1aa medium supplemented with SCM from the 8-cell embryo stage onwards (p < 0.05), in contrast to those cultured with FBS supplementation. Conclusions: In conclusion, this study conclusively demonstrated that the incorporation of SCM into the culture medium significantly enhances the developmental progress of preimplantation embryos.