• Clinical and Experimental Reproductive Medicine
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• v.24 no.2
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• pp.233-239
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• 1997

Embryos of most mammalian species grown in vitro would undergo developmental arrest at the approximate time of genomic activation. Stage-specific cell block and the resulting rapid loss of embryo viability in conventional culture media have limited the duration for which embryos may be cultured prior to transfer. As a result, embryos are usually transferred to the uterus at the 4-to 8-cell stage to avoid the loss of viability associated with long-term in vitro culture. Early transfer has led to asynchrony of the endometrium-trophectoderm interaction at the time of implantation and a resultant reduction in the rate of implantation. To overcome these problems, a variety of co-culture systems has been devised in which embryos can develop for a longer period prior to embryo transfer. Vero cells, derived from African green monkey kidney, share a common embryologic origin with cells from the genital tract. In addition, they are potentially safe to use, since they are highly controlled for viruses and other contaminants. Therefore, co-culture using Vero cells has been widely utilized to enhance embryo viability and development, although not without controversies. We thus designed a series of experiments to demonstrate whether Vero cells do indeed enhance mouse embryo development as well as to compare the efficacy of co-culturing mouse 1-cell embryos on Vero cell monolayer in both Ham's F-10 and human tubal fluid (HTF) culture media. 1-cell stage ICR mouse embryos were cultured either in the presence of Vero cells (Group A) or in conventional culture medium alone (Group B). In Ham's F-10 significantly more 3-to-8cell embryos developed in group A than group B (59.8 versus 10.0%; p<0.01). In contrast, there was no significant difference in embryonic development both group A and group B in HTF. However, significant differences were noted only in later embryonic stage (13 and 0%; p<0.05 of group A and B respectively, hatching or hatched). In Ham's F-10, we also could observe the beneficial effect of Vero cell on hatching process (70.7 and 42.1%; p<0.05 of group A and group B respectively).

• Clinical and Experimental Reproductive Medicine
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• v.47 no.4
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• pp.312-318
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• 2020

Objective: The objective of the study was to compare the effects of long-term and short-term embryo culture to assess whether there is a correlation between culture duration and clinical outcomes. Methods: Embryos were divided into two study groups depending on whether their post-warming culture period was long-term (20-24 hours) or short-term (2-4 hours). Embryo morphology was analyzed with a time-lapse monitoring device to estimate the appropriate timing and parameters for evaluating embryos with high implantation potency in both groups. Propensity score matching was performed to adjust the confounding factors across groups. The grades of embryos and blastocoels, morphokinetic parameters, implantation rate, and ongoing pregnancy rate were compared. Results: No significant differences were observed in the implantation rate or ongoing pregnancy rate between the two groups (long-term culture group vs. short-term culture group: 56.3% vs. 67.9%, p=0.182; 47.3% vs. 53.6%, p=0.513). After warming, there were more expanded and hatching/hatched blastocysts in the long-term culture group than in the short-term culture group, but there was no significant between-group difference in embryo grade. Regarding pregnancy outcomes, the time to complete blastocyst re-expansion after warming is shorter in women who became pregnant than in those who did not in both culture groups (long-term: 2.19±0.63 vs. 4.11±0.81 hours, p=0.003; short-term: 1.17±0.29 vs. 1.94±0.76 hours, p=0.018, respectively). Conclusion: The outcomes of short-term culture and long-term culture were not significantly different in vitrified-warmed blastocyst transfer. Regardless of the post-warming culture time, the degree of blastocyst re-expansion 3-4 hours after warming is an important marker for embryo selection.

• Korean Journal of Animal Reproduction
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• v.15 no.2
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• pp.97-101
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• 1991

This study was carried out to investigate the survival rate of in vitro culture after frozenthawed, to used DMSO(dimethyl sulfoxide), glycerol and ethylene glycol of cryorpotective agents at the zona pellucida removed and intact on the morulae and blastocysts. The results obtained from this study were as follows : 1. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the morulae was 86.0%, 87.1% and 83.3%, total or mean were 85.5%, respectively. 2. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed morulae was 53.2%, 42.3% and 37.5%, total or mean were 44.3%, respectively. 3. The survival rate of in vitro cultrue after frozen-thawed to used cryoprotective agents of three kinds at the blastocysts was 89.4%, 86.2%, total or mean were 86.7%, respectively. 4. The survival rate of in vitro culture after frozen-thawed to used cryoprotective agents of three kinds at the zona pellucida removed blastocysts was 55.8%, 51.6% and 40.6%, total or mean were 49.3%, respectively.

• Journal of Embryo Transfer
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• v.6 no.1
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• pp.25-32
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• 1991

The present study was performed to investigate the effects of co-culture with granulosa cells on in vitro fertilization and cleavage of early bovine embryo development. Bovine oocytes were matured for 20-24 hrs in vitro with granulosa cells or without and then fertilized in vitro using frozen-thawed spermatozoa treated with BO-caffeine, BO-BSA(2OmM heparin added). At l8hrs after insemination, oocytes were fixed and examined or further cultured in TCM 199 for 48hrs. The fertilization rates between the control(70.4%) and the groups of co-cultured with granulosa cell(2.5$\times$106 cells/ml; 71.6%, 5.0$\times$ 106/ml; 71.9%, l.0$\times$ 107/ml; 71.1%) did not differ significantly. The cleavage rates in the groups co-cultured with granulosa cell(2.5$\times$ 106 cells/mi; 43.6%, 5.0$\times$ 106/ml; 46.8%. l.0$\times$ 107/ml; 45.0%)were significantly higher than that of without granulosa cell, respectively(P<0.05). However there were no significant differences between the groups co-cultured with granulosa cells. The result indicated that co-culture with granulosa cell was effective means to cleavage of bovine follicular oocytes but did not affect the in vitro fertilization.

• 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.

• Journal of The Korean Society of Grassland and Forage Science
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• v.20 no.1
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• pp.7-12
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• 2000

This study was carried out to improve the ability of embryo formation and the efficiency of plant regeneration from suspension cultured cells of seed derived calli of orchardgrass (Dactylis glomerata L.). The frequency of formation of round cell and cell colonies was highest at 50 days after suspension culture in $N_6$ medium supplemented with $4\;g/{\ell}$ casein hydrolysate (CH), $20\;g/{\ell}$ sucrose and $30\;g/{\ell}$ sorbitol. The highest frequency of plant regeneration and somatic embryo formation was obtained from suspension cultured cells of 60 days. Addition of CH ($4\;g/{\ell}$) in suspension culture medium gave the highest frequency of embryo formation (39.6%) and plant regeneration (73.0%).

• Korean Journal of Plant Resources
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• v.17 no.3
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• pp.257-264
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• 2004

Selection of stress-tolerant ginseng lines in fields is very difficult because it is almost impossible to control properly the environmental conditions of soil. On the contrary, it can be studied with ease to search for stress-tolerant ginseng lines through in vitro culture because of easy manipulation of stress conditions. This study was conducted for the selection of ginseng pure lines tolerant to salt stress. Murashige ＆amp; Skoog(MS) media with 2.5 folds of KNO$_3$, NH$_4$NO$_3$, MgSO$_4$.7$H_2O$, KH$_2$PO$_4$, and CaC1$_2$.2$H_2O$ was established for the selection of ginseng pure lines tolerant to salt stress in vitro. Among 88 ginseng pure lines bred by Korea Ginseng and Tobacco Research Institute, Punggi Hwangsuk, 78093, 82886, 78135, 86024 and KG104 lines was tolerant to salt stress. For the stable production of quality Korean ginseng, genetic tolerance to salt stress is one of important factors since relatively high salt concentrations in the ginseng nursery soil environment of Korea. Ginseng inbred pure lines were tested for their tolerance to salt stress through in vitro culture technique.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2005.10a
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• pp.126-127
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• 2005

• Korean Journal of Plant Tissue Culture
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• v.27 no.4
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• pp.299-307
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• 2000

A large amount of information is currently available for somatic embryogenesis of plants. However, one common problem related to somatic embryos is that the conversion rate can be low for some species. Apical meristems are responsible for post-embryonic growth of the embryo. The low percentage observed is most likely a result of poor apical meristem development or defects in the meristem organization during somatic embryogenesis. In flowering plants, apical meristems are well developed by the late heart stage of zygotic embryo development. In conifers, such as white spruce, apical meristems are formed at the pre-cotyledon stage. Thus, apical meristem development occurs very early, prior to the maturation stage of embryo development. Once formed, meristems are stably determined. In the somatic embryo, as exemplified by white spruce, since embryo development is not synchronous, tissue differentiation including apical meristem formation occurs throughout the“maturation”stage. Different apical meristem organizations can be found among different individuals within a population. In contrast to their zygotic counterparts, the apical meristems appear not to be stably determined as their organization, as the shoot apical meristem especially, can be easily modified or disrupted. Precocious germination seldom results in functional plantlets. All these observations suggest that the conditions for somatic embryo maturation have not been optimized or are not suitable for meristem formation and development. The following strategies could improve meristem development and hence conversion: 1. Simulate in ouuio conditions to promote meristem development prior to the“maturation”treatment.2. Prevent deterioration of apical meristem organization during somatic embryo maturation.3. Promote further meristem development during embryo germination.

• Journal of Embryo Transfer
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• v.17 no.3
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• pp.195-201
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• 2002

This study was to assess the developmental capacity of oocytes matured in vitro after 20, 15, 10, 5 and 0 days of organ culture when ovaries were isolated from juvenile mice at 0-, 5-, 10-,15- and 20-day old, respectively, and to develop in vitro culture system that observed a view to morphology of ovaries and nucleus maturation of oocytes. The size of ovaries decreased 35.9%, 8.7%, 1.2% and 14.4% after 20, 15, 10, 5 days of organ culture when the ovaries were isolated from 0-, 5-, 10 and 15-day old mice, respectively. After organ culture, the recovery rates, diameters of oocytes and the number of oocytes progressed from GV to MII were increased as increasing age of mice.