• 한국수정란이식학회지
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• 제10권3호
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• pp.251-256
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• 1995

This study was experimented that developmental effects of bovine in vitro fertilized embryos by coculture system and supplementation of energy materials into simple media. With the ovaries from slaughter house in vitro maturation by 24h, in vitro fertilization was performed with sperms collected by Percoll gradient method. Fertilized embryos were cocultured in 15% FCS+CZB medium with BOEC(bovine oviductal epithelial cell), GCM (granulosa cell monolayer) and MEFC(mouse embryonic fihrohlast cell). And also in this study, there was trying to improve the early developmental rate of embryos by addition of concentration-controlled Na-pyruvate, D-glucose which were used as energy sources into CZB medium. In vitro developmental rate was confirmed by the cleavage rate of 48h post-IVF and the embryo development rate at 240h culture. In the coculture system BOEC had 20.0% of blastocysts rate, which was higher than that of other coculture systems. To determine the optimum concentration for early embryo developmental rate rapidly, through the gradient of concentrations of Na-pyruvate and D-glucose, we focused on the cleavage rate at 48h and blastocysts rate at 240h. In case of Na-pyruvate, cleavage rate and developmental rate over 3-cell were lower at the concentration of 1.OOrnM than the other treatment concentrations, otherwise the blastocysts rate was higher as 23.2% than the others. That result showed that as like reported group which had higher develop-mental rate over 3-cell was also higher to the blastocysts rate. In case of D-glucose, there was no effects through the concentration changes. It was the result of this study for which the use of BOEC coculture system and 1.OOmM Na-pyruvate as an energy source had an effect upon embryo development.

• 한국수정란이식학회지
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• 제14권1호
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• pp.9-15
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• 1999

This study was conducted to investigate the effect of $\alpha$-tocopherol and cysteamine with Whitten's medium in supporting the development on in vitro maturation(IVM), in vitro fertilization (IVF) and in culture(IVC) on porcine oocytes. When the immature oocytes were cultured of $\alpha$-tocopherol for 40h, the nuclear maturation rates were 39, 4, 52.5 and 54.1%, respectivley. The nuclear maturation rates of treat groups were signficantly (P<0.05) higher than those of non-treat groups. After matureation, the oocytes were inseminated in vitro in medium 199 with ejaculated spermatoza for examination of sperm penetration, polyspermy, male pronuclear(MPN) formation, and cleavage rate. Sperm penetration rates of treat higher than the control groups(P<0.05), and MPN formation rates were significantly(P<0.05) higher on treated groups (24.3~53.1%) than control groups(14.2~21.4%). After insemination, the cleavage rates at 120hr were groups higher than control groups(P<0.05).

• Clinical and Experimental Reproductive Medicine
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• 제11권2호
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• pp.59-67
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• 1984

In vitro fertilization have been performed to know whether the frozen semen has fertilizing ability and can be used clinically. The results of cultured and developed embryos obtained are as follows: 1. The semen was frozen in three media for the good viability. The viability was more than 50% and the motility was also moderate (grade III), 2. As the 33 oocytes were collected from 45 follicles, the oocyte recovery rate was 73.3%. Among them, mature and immature ova were 5% each, and premature ova were 69.7%, When the first polar body was appeared, above ova were inseminated after adequate incubation with activated sperms. 3. The main components of three freezing medium containing egg yolk, glycerol and pyruvate respectively were the best for sperm viability, and Ham's F-10 medium was used for the fertilization and culture of eggs. 4. The results of in vitro fertilization of 33 ova, showed the second polar body developed in 12%, polyspermia in 24%, 1-cell embryo in 21% and 2-cell embryo in 9%. One mature ova developed to blastocyst via 16-cell to 32-cell embryo. The fertilization rate was 66%. 5. Above mentioned results represent that the frozen semen has fertilizing ability and can be used practically in the clinic.

• 한국수정란이식학회지
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• 제7권1호
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• pp.41-47
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• 1992

1. In vitro system, LR and FSR accelerated and facilitated meiotic progression, and LH selectively improved cytoplasmic maturation which is required to promote the formation of a male pronucleus. 2. Caffeine (2mM) in the fetilization medium was required not only for inducing zona penetrating ability of boar also for developing to the male pronucleus of the penetrat- ing spermatozoa in vitro. 3. The germinal vesicle (GV)stage was observed for the first 17.6 hr;germinal vesicle break-down (GVBD)stage between 17.6~26.4 hr ;metaphase I (M-I)from 26.4 - 30. 9hr;anaphase I(A-I)ranged from 30. 9~33.4hr;telophase I(T-I) at 33.4~34.4hr; and metaphase II(M-II) at 34.4-48hr. 4. The addition of 10%(v /v) pig follicular fluid (pFF) to maturation media significantly increased the rate of nuclear maturation of pig oocytes (p<0.01), whereas the rate of nuclear maturation of pig oocytes among three different media did not differ. 5. The presence of a primary culture of POEC promotes in vitro development of early cleavage stage pig embryos.

• 한국수정란이식학회:학술대회논문집
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• 한국수정란이식학회 2005년도 제5회 발생공학 국제심포지엄 및 학술대회
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• pp.59-60
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• 2005

• 한국수정란이식학회지
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• 제9권1호
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• pp.73-83
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• 1994

The objective of this study was to develop an effective in vitro production system capable of obtaining more porcine embryos from immature oocytes. These experiments were thus conducted to examine the effect of oocytes type and maturation time on the in vitro maturation(IVM) and fertilization(IVF) of oocytes and the in vitro development (IVD)of IVF embryos. 1. The degree of oocyte maturation based on cumulus expansion index(GEI) did not differ for A- and B-typed oocytes but the index of oocyte type C was lower(P<0.05) than that of other oocyte types. 2. When the oocytes of type A and B were matured for 36, 42 and 48hrs, the GEl was not different between the 36- and 42-h maturation but the GEl after 48hrs was greatly lower(P<0.05) than that of other maturation times. 3. The highest cleavage rate(48.6%) of IVF oocytes was obtained from A typed oocytes and 42-h maturation but the developmental potential based on cleavage index was the highest when B-typed oocytes were matured for 42hrs.

• 한국수정란이식학회지
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• 제12권1호
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• pp.27-36
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• 1997

The studies were carried out to investigate the effects of co-culture with cumulus cells and oviduct epithelial cells on the in vitro fertilization and cleavage rate of bovine follicular cocytes and to determine the optimum thawing temperature and equilibration time on in vitro developmental rate of frozen bovine embryos. The ovaries were obtained from slaughtered Korean native cows. The follicular oocytes were cultured in TGM-199 medium containing 10 IU /ml의 PM SG, 10 IU /ml의 hCG, ip g/ml의 $\beta$-estradiol and 10% FCS for 24~48 hrs in incubator with 5% $CO_2$ in air at 38.5$^{\circ}C$. The bovine embryos following dehydration by cryoprotective agents and a various concentration of sucrose were directly plunged into liquld nitrogen and thawed in 3$0^{\circ}C$ water. Survival rate was defined as developmental rate on in vitro culture or FDA-test. The results are sunanarized as followes :1. The in vitro fertilization and in vitro developmental rates of bovine oocytes co-cultured with cumulus cells in TCM499 medium were 75.0~76.8% and 17.3~27.6%, respect-ively. And in-vitro fertilization rates of cumulus-enclosed oocytes(55.4%)were significantly(p<0.05) higher than cumulus-denuded oocytes (23.1%). 2. The in vitro fertilization and in vitro developmental rates of bovine oocytes co-cultured with l$\times$ l04cells /ml, 1 x l06cells /ml, lx l08cells /ml and 1 x l015cells /ml oviduct epithelial cells in TCM-199 medium were 74.5~77.8% and 15.7~21.20 respectively.3. The in-vitro fertilization and in vitro developmental rates of bovine oocytes cocultured in '1CM-199 media containing PMSG, hCG, PMSG+hCG. PMSG+$\beta$-estradiol, hCG+$\beta$-estradiol 0 to 40 hrs after insemination were 74.0~77.4% and l8.9~23.l%, re-spectiv ely.4.The survival rates of bovine embryos thawed after rapid freezing in the freezing medium containing a various concentration of sucrose added 1.5M and 2.OM glycerol,DMSO and propanediol were 23.5~31.4% and 20.6~34.l%, respectively. 5. The temperature thawed at 3$0^{\circ}C$ after rapid freezing of bovine embryos resulted in a significantly higher embryos survival rate than did at 2$0^{\circ}C$ and 35$^{\circ}C$.6. The equilibration time on the survival rates of bovine embryos was attained after short period of time(2.5~5 min.) in the freezing medium higher than long period of time (10~20min.). (Key words : bovine embryos, co-culture, freezing, in vitro development)

• Journal of Plant Biotechnology
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• 제2권1호
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• pp.29-33
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• 2000

To develop an efficient propagation method for yew tree, zygotic embryos were cultured under various conditions. When dissected embryos were cultured on GA$_3$ containing media, the highest germination frequency was observed on WPM medium contaning 1.0 mg/L GA$_3$. For germination of the embryos, two different conditions were compared; culturing embryos with endosperm (Method I), and 2) culturing embryos only (Method II). Maximum germination was achieved in 0.5 mg/L GA$_3$ when embryos with endosperm were cultured on the media. Of the media tested, White and WPM medium were the most suitable on germination of embryos. The abnormality of yew embryos found was observed when it cultured on GA$_3$ or culture media. About 40% of the precociously germinated embryos could be developed into full seedlings. Seedlings contained taxol in high quantity (535 $\mu\textrm{g}$/g dry weight). In vitro techniques will be sewed as a useful tool for the development of transformed root cultures and biosynthesis studies.

• 한국가축번식학회지
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• 제16권3호
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• pp.217-224
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• 1992

These studies were carried out ot investigate on the transferred embryo development following ultrarapid frozen for 8-cell and morula of in vitro fertilization mouse embryos. The post-thaw embryo survival was evaluated and compared by cell stage of embryos and by equilibration time before ultrarapid freezing. The results obatined were summerized as follows: 1. The effects of equilibration time of 3 vs. 6 minutes before ultrarapid freezing and after thawing on the morphological survival and the viability of 8-cell and morulas embryos were not significant. 2. When the ultrarapid frozen-thawed 32 eight-cell and 33 morula embryos, and 30 fresh blastocysts were transferred to pseudopregnant recipient mice, the number of normal offsprings produced were 9(28.1%), 14(42.4%) and 18(60.0%), respectively. From the above resutls, it was concluded that the optimal conditions of pH osmolality of the media for mouse IVF and embryo culture, and the period of sperm preincubation might be 7.1, 310 mOsm and 120 min., respectively,a nd somewhat high conception rate might be resulted from transfer of frozen embryos of morula stage and fresh embryos of blastocyst stage.

• Reproductive and Developmental Biology
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• 제31권2호
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• pp.91-96
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• 2007

In vitro development of porcine embryo is affected by culture condition. One possible factor is osmolarity of culture medium. 1his study examined whether high osmolarity of culture medium at the early culture stage improves development of preimplantation porcine in vitro fertilization (IVF) and nuclear transfer (NT) embryos. NT and IVF embryos were divided into three groups and the basic medium was PZM-3 ($250{\sim}270$ mOsmol, control group). The control group of embryos was cultured in PZM-3 for whole culture period. Other two groups of embryos were cultured in a modified PZM-3 with 0.05 M sorbitol or 0.05 M sucrose ($300{\sim}320$ mOsmol, sorbitol or sucrose group) for the first 2 days, and then cultured in PZM-3 for further culture. NT embryos cultured in sucrose group showed a significantly higher developmental rate to the blastocyst stage with a decreased apoptosis rate compared to the sorbitol (p<0.05). For IVF, sucrose group showed a significantly increased the blastocyst formation rate with a decreased apoptosis rate compared to the control (p<0.05). This study represents that the high osmolarity in the early embryo culture stage can enhance the in vitro development of porcine NT and IVF embryos to the blastocyst stage with reduced apoptosis of cells.