• Asian-Australasian Journal of Animal Sciences
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• v.30 no.9
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• pp.1245-1252
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• 2017

Objective: Phellodendron amurense (P. amurense) and Humulus japonicus (H. japonicus) are closely involved in anti-oxidative response and increasing antioxidant enzymes activities. However, the effects of their extracts on development of preimplantation bovine embryos have not been investigated. Therefore, we investigated the effects of P. amurense and H. japonicus extracts on developmental competence and quality of preimplantation bovine embryos. Methods: After in vitro fertilization, bovine embryos were cultured for 7 days in Charles Rosenkrans amino acid medium supplemented with P. amurense ($0.01{\mu}g/mL$) and H. japonicus ($0.01{\mu}g/mL$). The effect of this supplementation during in vitro culture on development competence and antioxidant was investigated. Results: We observed that the blastocysts rate was significantly increased (p<0.05) in P. amurense ($28.9%{\pm}2.9%$), H. japonicus ($30.9%{\pm}1.5%$), and a mixture of P. amurense and H. japonicus ($34.8%{\pm}2.1%$) treated groups compared with the control group ($25.4%{\pm}1.6%$). We next confirmed that the intracellular levels of reactive oxygen species (ROS) were significantly decreased (p<0.01) in P. amurense and/or H. japonicus extract treated groups when compared with the control group. Our results also showed that expression of cleaved caspase-3 and apoptotic cells of blastocysts were significantly decreased (p<0.05) in bovine blastocysts derived from both P. amurense and H. japonicus extract treated embryos. Conclusion: These results suggest that proper treatment with P. amurense and H. japonicus extracts in the development of preimplantation bovine embryos improves the quality of blastocysts, which may be related to the reduction of ROS level and apoptosis.

• Journal of Life Science
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• v.14 no.3
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• pp.375-384
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• 2004

Fibroblast growth factors (FGFs) are known to induce multiple functions in early development, including mesoderm formation, gastrulation movement and antero-posterior patterning. The induction of mesoderm from Xenopus presumptive ectoderm and the combination effect on inducing organs of bFGF(basic FGF) and HGF (Hepatocyte Growth Factor) were studied. Explants were cultured in the combined solution for 3 days to normal embryo arrive at St. 43. These effects on combined dose were examined by histological experiment and by immunohistochemical method. The concentrations of growth factors were tested in 0, 0.5, 1, 10 and also tested in 50 ng/ml of bFGF, and 0, 1, 10, 50 and 100ng/ml of HGF respectively. The synergistic effects were seen in the combined-dose of bFGF and HGF rather than in single dose. Various organs were differentiated and highest inducing effects were seen at the combined concentration of 1 ng/ml of bFGF and 10ng/ml of HGF, and at the concentration 10ng/ml of bFGF and 1 ng/ml of HGF. The bFGF induces various organs from cultured animal cap explants and the effects are time and dose-dependent. HGF is also a potent mitogen for renal tubular cells and for mature hepatocytes in primary culture. Eyes were developed in high percentage at the combined concentration of 1 and 10ng/ml of bFGF, and 1 and 10 ng/ml of HGF. From the induced eye and normal embryonic eye, RPE65 was commonly detected by monoclonal antibodies 40All and 25F5 and the localization of RPE65 was seen by AP reaction.

• Korean Journal of Medicinal Crop Science
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• v.2 no.3
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• pp.241-245
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• 1994

Effects of media and plant growth regulators on the germination of somatic embryos of Angelica tree(Aralia elata Seem.) was studied for the mass production of Angelica tree through tissue culture. MS medium was found to be the most effective for the germination of somatic embryos(65% germination rate), Among the MS medium, the medium containing 25% less inorganic salts and 1% less sucrose was found to be the most effective. Gelling agent with 0.2-0.3% gelrite promoted the germination of somatic embryo$(65{\sim}70%)$ and caused good growth of shoots and roots. 0.1 mg/l of BA and kinetin treatment caused $65{\sim}70%$ germination rate of somatic embryos and good growth of shoots and roots, and resulted in high percentage of dry matter. 1mg/l or 5 mg/l treatment of putrescine, and 10 mg/l treatment of spermidine caused 90% germination rate of somatic embryos and good growth of plant organs, and inhibited vitrification of regenerated plants.

• Korean Journal of Plant Tissue Culture
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• v.25 no.1
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• pp.69-74
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• 1998

The somatic embryos of Daucus carota L. cv. Hongshim developed in MS basal medium far 4 weeks had varied number of cotyledons. Palisade and spongy parenchyma of cotyledon were not clearly discriminated in all seedlings developed from the embryos. No independent existence of collateral vascular bundle was observed in all seedlings with various types of cotyledon ; instead, vascular bundles were either interconnected or partially connected with one another. Most of the cotyledonary bases on hypocotyl showed short cylinder structure which encircle plumule. The vascular tissues of cotyledonary bases and nodes of seedlings with jar-shaped or 1 cotyledon were connected in ring forms, showing the pattern of ectophloic shiphonostele, and similar ring form structure was also found in the vascular arrangement of 5 cotyledon seedlings. The vascular bundles of seedlings with 2, 3 and 4 cotyledons in many cases had independently arranged within the cotyledonary bases and nodes, showing the pattern of eustele. In hypocotyl, tetrarch or hexarch xylems prevailed in seedlings with jar-shaped cotyledon or 1 and 5 cotyledon; tetrarch xylems prevailed in 2 cotyledon seedlings; and triarch xylems prevailed in 3 cotyledon seedlings. In most of seedlings, cortex vascular bundles were dispensed in the region from cotyledonary node to hypocotyl, but double vascular bundles were also observed occasionally. In roots, diarch xylems were observed in most of seedlings with 2 cotyledons, triarch xylems in half of seedlings with 3 cotyledons, and diarch xylems in most of the remaining seedlings with the occasional occurrences of tetrarch xylems.

• Korean Journal of Poultry Science
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• v.35 no.3
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• pp.241-245
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• 2008

Lentiviral vector system is efficient vehicles for the delivery of exogenous genes, and it is generally used in the generation of transgenic chickens. In this study, we used recombinant lentiviral vectors to generate transgenic chicks that express the human superoxide dismutase-3 gene driven by the chicken ovalbumin promoter. It is well known that superoxide dismutases(SODs) are believed to play a crucial role in protecting cells against oxygen toxicity. There are three forms of SOD proteins: cytosolic Cu-Zn SOD, mitochondrial Mn SOD, and extracellular SOD(SOD-3). The recombinant lentivirus containing the human SOD-3 gene was injected into the subgerminal cavity of freshly laid eggs. Subsequently, the embryos were incubated to hatch using phases II and III of the surrogate shell ex vivo culture system. From 341 injected embryos, the 78 chicks hatched after 21 days incubation. The hatched chicks were screened for the human SOD-3 gene by using PCR. Two of 47 male chickens that survived to sexual maturity contained the human SOD-3 gene in their semen. These results showed that our transgenic chicken generation system was completely established.

• Journal of Plant Biotechnology
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• v.32 no.1
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• pp.31-35
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• 2005

To obtain regeneration system of onion, we analyzed the effects of 2,4-D and BA concentration on the embryogenic callus induction from mature embryos. The highest embryogenic callus induction ratio was shown on MS medium (Murashie and Skoog 1962) containing $2.5\;\cal{mg/L}\;or\;5\;\cal{mg/L}$ picloram after mature embryos were placed on medium. When induced callus were cultured on half strength of MS medium containing $1\;\cal{mg/L}$ Kinetin, the highest shoot formation ratio was observed on MS medium containing $1\;{mg/L}$ 2,4-D and $1\;{mg/L}$ BA. Embryogenic callus were cultured in MS liquid medium containing $1\;\ccal{mg/L}$ of 2,4-D and $1\;\cal{mg/L}$ BA. The suspension cultured cell clumps could be mass propagated. Embryogenic callus were friable, but non-embryogenic callus included a lot of moisture, hence the identification between embryogenic and non-embryogenic callus as easily achieved. When embryogenic callus as cultured on half strength of MS medium containing $1\;\cal{mg/L}$ Kinetin, shoots were induced. The whole plantlet was obtained on rooting medium containing $0.5\;\cal{mg/}$ of NAA.

• Reproductive and Developmental Biology
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• v.36 no.3
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• pp.173-181
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• 2012

Sphingosine-1-phosphate (S1P) has a many function involved proliferation, differentiation and survival of many cells. In this study, to investigate whether S1P improve the developmental competence of porcine embryos, 50 nM of S1P were supplemented during in vitro maturation (with EGF or without EGF) medium and/or in vitro culture (IVC) medium. Addition of S1P was significantly increased the rate of oocytes reaching metaphase II (MII) compared to the control (83.5 vs. 64.1%) in without EGF medium, but not with EGF medium (89.5 vs. 84.6%). When treated with $1{\mu}M$ of N1N-dimethylsphingosine (DMS), a sphingosine kinase inhibitor which is blocked endogenous generation of S1P, the meiotic progression rates to MII stage (without EGF: 45.2 and with EGF: 66.7%) were significantly decreased and degeneration rates (without EGF: 51.2 and with EGF: 30.1%) were increased in both medium compared to control group during IVM periods. Also, the rates of blastocyst formation was significantly increased in the S1P treated group compared to control group (29.0 vs. 19.2%) of EGF supplemented medium, whereas there were no effect in the EGF free medium (9.0 vs. 10.5%). After 12 h IVM, the phosphorylation of ERK1 and ERK2, which is major signaling pathway of MAP kinase, were increased in the S1P group than that of control or DMS group. When supplemented of S1P during IVC, the rates of blastocyst formation and total cell number (30.2% and 40.6) were significantly increased in S1P-treated group compared with control (20.1% and 32.5), DMS (12.3% and 25.1), and S1P plus DMS group (24.7% and 33.6). The percentage of apoptosis nuclei in the S1P group was significantly decreased than other groups. Also, the rates of blastocyst formation (26.7 vs. 14%) and total cell number (42.8 vs. 32.5) were significantly increased in the S1P group than those of control group when S1P added during the entire IVM and IVC periods. Taken together, our results indicate that S1P supplementation in IVM and/or IVC medium affects beneficial effect of meiotic maturation and subsequent developmental competence of porcine embryos.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.2
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• pp.147-153
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• 2001

Objective: The present study was performed to investigate the efficiency of partial laser assisted hatching (p-LAH; lased 1/2 ZP width from ZP edge) on hatching of mouse blastocysts. Methods: We used non-contact $1.48{\mu}m$ diode laser (MTM, Switzland) to create a precise hole on zona pellucida. 2-cell embryos were collected from the mouse (ICR) oviduct at 48 hours after hCG administration. Collected 2-cell embryos were cultured in the P-1 medium supplemented with 0.4% BSA. For experiments, embryos at 8-cell stage were used after $20{\sim}22$ hours in culture. After conventional (c-LAH) or partial laser assisted hatching, the embryos were further cultured in P-1 medium supplemented with 0.4% BSA for 3 days. To compare efficiency of complete and partial laser assisted hatching, hatching rate, hatching time and blastocyst diameter and zona pellucida thickness at hatching time were investigated. Embryos were examined every 12 hours. Blastocyst diameter and zona pellucida thickness at hatching time were measured with an ocular micrometer. Results: Hatching rates of p-LAH group (84.2%) was significantly higher than that of control group (39.3%), but there was no difference between the p-LAH (84.2%) and c-LAH (91.2%). p-LAH group was hatched 12 hours earlier than control group, but hatched 12 hours later than c-LAH group. The diameter of blastocyst at hatching time of p-LAH group ($113.1{\pm}6.4{\mu}m$) was smaller than that of control group ($122.2{\pm}5.0{\mu}m$), but larger than that of c-LAH group ($102.2{\pm}2.7{\mu}m$). Zona pellucida thickness at hatching time of p-LAH group ($6.4{\pm}0.9{\mu}m$) was thicker than that of control group ($4.5{\pm}1.5{\mu}m$), but thinner than that of c-LAH group ($10.0{\pm}0.8{\mu}m$). Conclusion: These results suggest that p-LAH may maintains the cell arrangement of early embryos to ensure successful development and prevent precocious hatching of blastocyst when compare to c-LAH and conventional (acidic tyrode) AH. Thus, p-LAH may provide a valuable and effective AH technique for human ART program.

• Korean Journal of Plant Resources
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• v.31 no.5
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• pp.515-521
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• 2018

This was carried out to develop a chromosome-doubled (12x) persimmon that will be used as a crossing parent to select seedless persimmon cultivars with the change of the consumption trend recently. To obtain a chromosome-doubled (12x) persimmon, colchicine was applied at the meristem of seedlings in vitro derived from cross among hexaploid persimmon (Diopyros kaki Thunb.). These were treated with 0.03%, 0.05% and 0.1% colchicine respectively for doubling chromosome, and it was most effective at the concentration of 0.05% colchicine. After colchicine treatment, we conducted tests to elucidate conditions for inducing shoot and root development. As the result, the shoots grew best when cultivated at 1/2MS media plus 10 and $30{\mu}M$ zeatin respectively, and the roots grew best when cultivated at 1/2MS media after dipping for 5 seconds at 10 mM NAA+5% DMSO. We also compared seedlings that have chromosome (6x) do not doubled and crossing parents (6x) and chromosome-doubled seedlings (12x). As the result, these chromosome-doubled seedlings (12x) showed lower stomatal density and larger stomatal size.

• Korean Journal of Poultry Science
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• v.41 no.2
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• pp.87-92
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• 2014

The avian embryos have been used as a good model to study embryonic development. Due to its unique development in the eggshell, avian embryos can be cultured and hatch in the surrogate eggshell system. In this study, we examined the viability, normal development and hatchability of Korean Oge (KO) chicken embryos in White Leghorn (WL) surrogate eggshells. Donor KO embryos at 3-day and 4-day-old were transferred into recipient WL eggshells, incubated for further 18 days at $37.5^{\circ}C$ with 70% of humidity until hatching. The viability of 3-day-old KO embryos at 7, 14 and 21 day in surrogate eggshell were 70.0%, 43.8% and 23.1%, respectively. In contrast, the viability of 4-day-old KO embryos at 7, 14 and 21 day in surrogate eggshells were 87.1%, 55.6% and 36.0%, respectively. The hatchability of KO embryos transferred into surrogate eggshells at 3-day-old was 23.1%, whereas embryos transferred at 4-day-old was 36.0%. Furthermore, the development of all viable embryos from 3-day group and 4-day group were normal. Our results suggested that culture of KO embryos in WL surrogate eggshells is highly possible, and transfer of donor embryos at 4-day-old may yield higher percentage of hatchability. This study may provide potential knowledge for the conservation of wild and endangered birds through surrogate system.