• Korean Journal of Plant Resources
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• v.17 no.2
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• pp.161-168
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• 2004

Optimal regeneration conditions of ginseng transformants were studied. It has been known that Ferritin Light Heavy Chain (FLHC) gene remove the several heavy metal by combination, store and transport. To obtain the ginseng tolerant to heavy metal, binary vector was introduced in Agrobacterium by tri-parental mating and then Agrobacterium tumefaciens MP90/FLHC was selected on the AB media and MS media containing kanamycin. Explants were co-cultured with Agrobacterium tumefaciens MP90/FLHC, which contained NPT II as a selectable marker, tadpole ferritin heavy chain (FLHC) gene and human ferritin light chain gene and then a number of embryos were induced. The induced embryo transferred to shooting media consisting of MS medium supplemented with GA 10 mg/L. As a result of examination that induced the normal growth of transfomants, transformants showed the equivalent growth in both root and shoot on the media containing the 1/3 MS.

• Korean Journal of Plant Tissue Culture
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• v.26 no.2
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• pp.71-76
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• 1999

Anthers of two hot pepper cultivars, Milyang-jare and Geryongsan-jare, were cultured on MS medium containing 0.1 mg/L 2,4-D and 0.1 mg/L kinetin. The influence of pretreatment at 4$^{\circ}C$ and 32$^{\circ}C$ on induction of microspore embryo was investigated. Milyang-jare was superior to the Geryongsan-jare in microspore embryo induction. The 32$^{\circ}C$ pretreatment increased embryo induction compared to the 4$^{\circ}C$ pretreatment while the 4$^{\circ}C$ pretreatment stimulated callus induction. Microspore embryos were regenerated to plantlets in the same medium or hormone free medium at 32$^{\circ}C$ treatment but most embryos failed to develop directly into plantlets at 4$^{\circ}C$ treatment. The optimal period of the 32$^{\circ}C$ pretreatment was 3 days in Milyang-jare and 6 days in Geryongsan-jare. The 32$^{\circ}C$ pretreatment was essential for induction and growth of microspore embryo in pepper.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.1
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• pp.50-56
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• 2019

These studies were conducted to evaluate developmental competence of follicular oocyte collected from the ovaries of Hanwoo cows with the high offspring meat quality (1++ and 1+ grade). Cumulus oocyte complexes from individual cows were matured, fertilized and cultured using protocols of in-vitro maturation (IVM), invitro fertilization (IVF) and in-vitro culture (IVC). The rates of blastocyst development from Hanwoo cows with the offspring meat quality grades of 1++ and 1+ were 18.6 and 21.2%, respectively. The rates of blastocyst development were 26.3, 20.7, 20.7, 17.2 and 31.2% from Hanwoo cows with the meat quality grades of 1++, 1+, 1, 2 and 3, respectively. Fiftyseven transferable embryos were recovered from 11 Hanwoo donor cows (5.2/head) with the high offspring meat quality grades of 1++ and 1+ in vivo, and the pregnancy rate after embryo transfer was 61.1%. In conclusion, these results suggest that in vitro embryo production from the ovaries of cows with the high meat quality grades using individual culture system can be used an efficient method for livestock improvement. In addition, for the successful industrialization of embryo transfer, conception rate should be improved.

• Clinical and Experimental Reproductive Medicine
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• v.47 no.4
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• pp.284-292
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• 2020

Objective: This study investigated whether adding outer-well medium to inhibit osmotic changes in culture media in a dry-type incubator improved the clinical outcomes of in vitro fertilization-embryo transfer (IVF-ET) cycles. Methods: In culture dishes, the osmotic changes in media (20 µL)-covered oil with or without outer-well medium (humid or dry culture conditions, respectively) were compared after 3 days of incubation in a dry-type incubator. One-step (Origio) and G1/G2 (Vitrolife) media were used. Results: The osmotic changes in the dry culture condition (308 mOsm) were higher than in the humid culture conditions (285-290 mOsm) after 3 days of incubation. In day 3 IVF-ET cycles, although the pregnancy rate did not significantly differ between the dry (46.2%) and humid culture (51.0%) groups, the rates of abortion and ongoing pregnancy were significantly better in the humid culture group (1.5% and 49.5%, respectively) than in the dry culture group (8.3% and 37.8%, respectively, p<0.05). In day 5 IVF-ET cycles, the abortion rate was significantly lower in the humid culture group (2.2%) than in the dry culture group (25.0%, p<0.01), but no statistically significant difference was observed in the rates of clinical and ongoing pregnancy between the dry (50.0% and 25.0%, respectively) and humid culture groups (59.5% and 57.3%, respectively) because of the small number of cycles. Conclusion: Hyperosmotic changes in media occurred in a dry-type incubator by evaporation, although the medium was covered with oil. These osmotic changes were efficiently inhibited by supplementation of outer-well medium, which resulted in improved pregnancy outcomes.

• Journal of Embryo Transfer
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• v.18 no.1
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• pp.35-41
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• 2003

Three experiments were performed to develop an optimal culture system of in-vitro derived porcine embryos. In experiment 1, embryos were cultured in two different basic culture media (NCSU-23 and PZH) supplemented with BSA (4mg/ml), FBS (10%), PVA (3mg/ml). Although no difference on the percentage of embryos developed to blastocysts was found (P>0.05), more 2-cell embryos (59.5% vs. 44.7 to 52.0%) were obtained when embryos were cultured in NCSU-23 added with BSA. In experiment 2, identical treatment was conducted using PZM medium. More (P<0.05) 2-cell embryos (54.4% vs. 40.7 to 44.5%) and blastocysts (12.5% vs. 5.8 to 9.0) were produced when embryos were cultured in PZM added with BSA (4mg/ml). In experiment 3, embryos were cultured in media as follows; 1) NCSU-23 without BSA for 192 hours; 2) NCSU-23 with BSA for 192 hours; 3) NCSU-23 with BSA for 96 h and then subsequently injected with FBS into culture drops. Although no difference on the percentage of embryos developed to blastocysts was found (P>0.05), more 2-cell embryos (63.8 to 69.2% vs. 52.0%) were obtained when embryos were cultured both in NCSU-23 added with BSA throughout culture duration and in FBS supplemented medium. In conclusion, no difference was found when porcine preimplantation embryos were cultured in NCSU-23 added with various macromolecules. However, when PZM medium was used, PZM medium supplemented with BSA showed more potential to support embryo development. Finally, FBS supplemented into culture drop 96h post-insemination did not show any benefits on embryo development.

• Clinical and Experimental Reproductive Medicine
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• v.24 no.3
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• pp.377-383
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• 1997

The effects of embryo number and incubation volume on the development of mouse embryos were evaluated. The growth rate of two-cell mouse embryos to attached blastocyst stage and the growth rate of blastocysts to early somite stage were assessed after culture in different incubation volumes and embryo densities. Embryos were collected from ICR female mice superovulated with pregnant mare serum gonadotropin and human chorionic gonadotropin and mated by ICR males. In experiment 1, groups of one, five, ten, twenty 2-cell embryos were cultured in 10-, 50-, 500-, 1000-${\mu}l$ drops of BWW media under mineral oil at $37^{\circ}C$ in a humidified atmosphere of 5% $CO_{2}$ and 95% air. As the incubation volume decreased, significantly (p<0.05) higher rates of embryos reached morular and blastocyst stage on day 3 and 4 culture, respectively. In experiment 2, groups of one, five, ten, twenty blastocysts were cultured in 1- and 2-ml volumes of CMRL 1066 media under same condition as in experiment 1. However the reverse was the result. Decreasing the number of embryos incubated per volume from 1 to 20 significantly (p<0.05) increased the number of blastocysts reaching the late egg cylinder (LEC) and early somite (ES) stage on day 6 and 8 culture, respectively, regardless of incubation volume. Blastocysts cultured in 2ml had higher (p<0.05) development rates to LEC and ES stage on day 6 and 8 culture, respectively, than embryos cultured in 1ml. Our results suggest that the effects of embryo number and incubation volume on the development of mouse embryos are stage specific and the shifting point was between hatching and EEC stage.

• Korean Journal of Plant Tissue Culture
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• v.24 no.3
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• pp.129-135
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• 1997

Morphogenetic responses were investigated by culturing embryo and leaf explants of Korean wild type tea plant, Camellia sinensis (L.) O. Kuntze. Induction of direct somatic embryogenesis as well as adventitious and/or axillary shoots was obtained from mature zygotic embryo cultures on Murashige and Skoog (MS) basal medium having 5 to $20\mu\textrm{M}$cytokinin a lone. Morphogenetic response was decreased dramatically by the addition of auxins tested. One hundred percent of induced and isolated shoots formed roots after four weeks of culture on half-strength MS or quarter-strength Schenk and Hildebrandt (SH) media supplemented with $10\mu\textrm{M}$indole-3-butyric acid (IBA). Immature zygotic embryos were shown to be a suitable explant for embryogenic callus formation in the presence of 2, 4-dichlorophenoxyacetic acid(2, 4-D) in basal medium. Mature zygotic embryo originated leaves were used to test their ability for mophogenesis by incorporating plant growth regulators such as IBA, naphthyl-1-acetic acid (NAA), and 6-benzylaminopurine (BAP). Apparently, the morphogenetic responses of the cultured explant sources on the types and/or levels of plant growth regulators tested were observed visually.

• Journal of Embryo Transfer
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• v.24 no.3
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• pp.145-150
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• 2009

This study was conducted to examine the effects of human follicular fluid and gonadotropin (FSH+HCG+rhEGF) on in vitro maturation, fertilization and development of human immature oocytes. Cumulus-oocyte complexes (COCs) were collected following for in vitro fertilization and embryo transfer (IVF-ET) cycles of the patients. At the time of oocytes collection, oocytes were classified into MII, MI and GV in accordance with their appearance (MII: Fully mature oocyte at metaphase II of meiosis; MI: Nearly mature oocytes at metaphase I of meiosis; GV: Immature oocytes at prophase I of meiosis). After controlled ovarian stimulation using gonadotropin(FSH) and human chorionic gonadotropin (HCG) in 70 ICSI cycles, 158 MI to MII matured oocytes were intracytoplasmic sperm injection (ICSI) ${\sim}4$ h after in vitro culture and 553 MII oocytes were ICSI after denudation. The aspirated MI and GV oocytes were cultured in culture medium containing 10% (v/v) serum protein substitute (SPS), 10% (v/v) human follicular fluid (hFF) and 10% (v/v) serum protein substitute (SPS)+1 IU/ml FSH+10 IU/ml HCG+10 ng/ml recombinant human epidermal growth factor (rhEGF). The maturation rate of immature oocytes was similar among the three group. When maturation medium was supplemented with 10% SPS, 10% hFF or gonadotropins, the fertilization rate of in vitro matured oocytes was higher in 10% SPS (80.0%), but there was no statistical significance (78.2%; hFF, 76.9%; gonadotropin, p>0.05). The development rate of human embryos developed to $6{\sim}8$ cells were not significant difference in the medium containing SPS, hFF and gonadotropins (65.6%, 65.9% and 66.7%). The results of these study suggest that human follicular fluid and gonadotropins supplemented in the culture medium was not effected on the in vitro maturation, fertilization and development of human immature oocytes.

• Toxicological Research
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• v.22 no.2
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• pp.127-133
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• 2006

Recently, we have reported that the environmental pollutant 2-bromopropane (2-BP) induces a significant embryo-fetal developmental toxicity in rats. However, the cause of developmental toxicity and the relationship between maternal and developmental toxicities could not be elucidated because the developmental toxicity of 2-BP was observed only in the presence of maternal toxicity The in vitro teratogenicity study using whole embryo culture was carried out to understand the teratogenic properties and the possible mechanism of teratogenicity induced by 2-BP in rats. Rat embryos aged 9.5 days were cultured in vitro for 48 hrs at medium concentrations of 0, 1, 3, or 10 mg/ml of 2-BP. Embryos were evaluated for growth, differentiation, and morphological alterations at the end of the culture period. At 10 mg/ml, 2-BP caused a delay in the growth and differentiation of embryos and an increase in the incidence of morphological alterations, including altered yolk sac circulation, abnormal axial rotation, craniofacial hypoplasia, open neuropore, absent optic vesicle and kinked somites. At 3 mg/ml, only a delay in the growth and differentiation of embryos was observed. There were no adverse effects on embryonic growth and development at the concentration of 1 mg/ml. The results showed that the exposure of 2-BP to rat embryos results in a developmental delay and morphological alterations at dose levels of 3 mg/ml culture media or higher and that 2-BP can induce a direct developmental toxicity in rat embryos.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.8
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• pp.1117-1123
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• 2009

This study investigated the effects of IGF-I and EGF on the development of blastocysts or hatched blastocysts during the in vitro culture of embryos from immature porcine oocytes. After the in vitro maturation and fertilization of cumulus-oocyte complexes (COCs) and their culture in vitro in PZM3 medium, we examined the embryo development rate for 168 h. When different concentrations of IGF-I (0, 1, 10, 20 ng/ml) were supplemented to fertilized porcine embryos in vitro, there were no significant differences in cleavage rate, blastocyst development rate or blastocyst hatching rate among the treated groups. On the other hand, when different concentrations of EGF (0, 1, 10, 20 ng/ml) were supplemented to the in vitro culture medium, blastocyst development rate was highest in the group in which EGF was not supplemented and, specifically, it was higher than in the 20 ng/ml treatment group (p<0.05). When 10 ng/ml IGF-I and 1 ng/ml EGF were supplemented separately or simultaneously, there were no significant differences among the treated groups in blastocyst hatching rate and the number of cells in each condition. This study demonstrated that the addition of IGF-I and EGF into PZM3 medium did not enhance development of the blastocyst stage and total cell number in blastocysts.