• Journal of Life Science
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• v.27 no.11
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• pp.1369-1375
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• 2017

The purpose of this study was to investigate the cellular characterization of phospholipase C-${\delta}1$ in olive flounders (Paralichthys olivaceus). In general, phospholipase C signaling pathways are distributed in nuclei at plasma membranes and in cytoplasms, although the pathways' nuclear localization mechanisms are unclear. P. olivaceus duplicates type-A PoPLC-${\delta}1$ (PoPLC-${\delta}1A$), which has a high similarity to the human isoform PLC-${\delta}$; type-B PoPLC-${\delta}1$ (PoPLC-${\delta}1B$ [Sf]), which has a low similarity to the human isoform PLC-${\delta}$ and the alternative splice variant PoPLC-${\delta}1B$ (Lf), which has a nuclear localization signal (NLS) and a nuclear export signal (NES) for nuclear imports and exports, respectively. This study confirmed the effects of the cellular localization and translocation of GFP-tagged PoPLC-${\delta}1A$, PoPLC-${\delta}1B$ (Sf) and PoPLC-${\delta}1B$ (Lf). It administered treatments of $Ca^{2+}$ ionophore ionomycin and endoplasmic reticulum (ER)-$Ca^{2+}$ pump inhibitor thapsigargin to hirame natural-embryo (HINAE) cells. A laser-scanning confocal microscope was used. GFP-tagged PoPLC-${\delta}1A$ was distributed to the cellular organelles, rather than to the cytoplasms and cytomembranes, when PoPLC-${\delta}1B$ (Lf) and PoPLC-${\delta}1B$ (Sf) were localized at the plasma membranes. The treatments of ionomycin and thapsigargin showed the accumulation of PoPLC-${\delta}1A$ in the nuclei when PoPLC-${\delta}1B$ (Lf) nucleocytoplasmic shuttling and PoPLC-${\delta}1B$ (Sf) nucleocytoplasmic shuttling were not observed. The results were the first evidence that PoPLC-${\delta}1A$, which contains functional, intact NES sequences, has a main role in nucleocytoplasmic shuttling and translocation in fish.

• Journal of Bio-Environment Control
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• v.29 no.2
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• pp.141-152
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• 2020

The rapid on-site measurement of hydroponic nutrients allows for the more efficient use of crop fertilizers. This paper reports on the development of an embedded on-site system consisting of multiple ion-selective electrodes (ISEs) for the real-time measurement of the concentrations of macronutrients in hydroponic solutions. The system included a combination of PVC ISEs for the detection of NO₃, K, and Ca ions, a cobalt-electrode for the detection of H₂PO₄, a double-junction reference electrode, a solution container, and a sampling system consisting of pumps and valves. An Arduino Due board was used to collect data and to control the volume of the sample. Prior to the measurement of each sample, a two-point normalization method was employed to adjust the sensitivity followed by an offset to minimize potential drift that might occur during continuous measurement. The predictive capabilities of the NO₃ and K ISEs based on PVC membranes were satisfactory, producing results that were in close agreement with the results of standard analyzers (R² = 0.99). Though the Ca ISE fabricated with Ca ionophore II underestimated the Ca concentration by an average of 55%, the strong linear relationship (R² > 0.84) makes it possible for the embedded system to be used in hydroponic NO₃, K, and Ca sensing. The cobalt-rod-based phosphate electrodes exhibited a relatively high error of 24.7±9.26% in the phosphate concentration range of 45 to 155 mg/L compared to standard methods due to inconsistent signal readings between replicates, illustrating the need for further research on the signal conditioning of cobalt electrodes to improve their predictive ability in hydroponic P sensing.

• Journal of Embryo Transfer
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• v.18 no.3
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• pp.171-178
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• 2003

This study was conducted to investigate the effect of electric fusion methods on cell fusion rate and embryo development by somatic cell nuclear transfer in Korean Native Cattle. The KNC ear cell was cultured in vitro for confluence in serum starvation condition(DMEM＋0.05％ FBS) for cell confluence. The zona pellucida of IVM oocytes were partially dissection using micro pipette. Ear cells were transferred into an enucleated oocyte. The reconstructed embryos were electrically fused with Zimmermann Cell Fusion Medium(ZCFM). Nuclear transfer embryos were activated with a combination of 10${\mu}{\textrm}{m}$ calcium ionophore(5 min) and 2.0mM 6-DMAP(3 hr). The activated embryos were cultured in CR1 -aa medium contains 0.3％ BSA or 10％ FBS at 37$^{\circ}C$, 90％ $N_2$, and 5％ $CO_2$in incubator for 6 days. The fusion rates were 51.6％(chamber) and 68.9％(needle), respectively and there were significantly difference between the fusion method(P<0.05). But, lysis rates were not significantly different(10.7％, 11.5％), respectively. The cleavage rates were significantly different between the chamber method(73.2％) and needle method(80.3％), respectively(P<0.05). The rates of early embryos(2∼4cells) and blastocysts of chamber and needle methods were 54.1％, 61.1％ and 18.4％, 26.3％ respectively, and needle method was significantly higher than chamber method(P<0.05). But, morulae formation rate were not significantly differences between the chamber(6.7％) and needle(6.2) method(P <0.05). These result suggest that electric fusion of needle method was to be profitable for nuclear transfer embryo fusion rate, blastocyst formation rate and reduce of oocyte lysis.

• Journal of Life Science
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• v.23 no.2
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• pp.213-221
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• 2013

Bibimbab (mixed rice) is a traditional Korean one-dish meal. This study was carried out to investigate the anti-inflammatory and antioxidative effects of raw and seasoned ingredients used in Bibimbab (Cucurbita moschata P., Platycodon grandiflorum A., Vigna radiata L., Porphyra yezonensis udea, Allium ampeloprasum L., Pterdium aguilinum, Raphanus sativus). Human mast cells (HMC-1) were pretreated with 70% ethanol extracts of Bibimbab and further cultured for an appropriate time after the addition of phorbol 12-myristate 13-acetate (PMA) and calcium ionophore A23187. Cell viability was determined by an MTT assay. None of the ingredients showed cytotoxic effects at a concentration of 1.0 ${\mu}g/ml$. Anti-inflammatory effects were analyzed at 0.01, 0.1, and 1.0 mg/ml concentrations of the raw, seasoned ingredients of PMA. A23187 stimulated HMC-1. Among the various ingredients, seasoned A. ampeloprasum L. extract showed the highest inhibition of TNF-${\alpha}$ and IL-6 secretion (90% and 93%, respectively) at a concentration of 1.0 mg/ml. The R. sativus extract showed the highest inhibition (85%) of IL-8 secretion. DPPH analysis of the antioxidation properties of the ingredients showed that raw and seasoned A. ampeloprasum extracts exhibited the highest DPPH free radical scavenging activity (67.50 and 73.65%, respectively). These results suggest that seasoned ingredients used in Bibimbab have lower anti-inflammatory effects in relation to TNF-${\alpha}$ and IL-6 secretion than raw ingredients in PMA- and A23187-treated HMC-1. In addition, the seasoned ingredients showed a tendency to increase antioxidative activity. Therefore, the ingredients used in Bibimbab have potential as anti-inflammatory and antioxidation agents.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.365-373
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• 2000

The purpose of this study was to investigate the effects of the fusion pulses and fusion media on fusion rate and the development of embryos produced by somatic cell nuclear transfer in Hanwoo (Korean cattle). Nuclear donor cumulus and fetal fibroblast cells were cultured in Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum at 38.5$^{\circ}C$ in a humidified atmosphere of 5% $CO_2$in air. The in vitro matured oocytes were enucleated and then the isolated donor cells were introduced. The cumulus cell and cytoplast were fused using one pulse of 70 volts for 40$mutextrm{s}$, two pulses of 70 volts for 40$mutextrm{s}$ and one pulse of 180 volts for 15$mutextrm{s}$. The fetal fibroblast cell and cytoplast were fused using one pulse of 180 volts for 15$mutextrm{s}$ or 30$mutextrm{s}$. The cumulus cell and cytoplast were fused using mannitol and Zimmerman cell fusion medium (ZCFM) as a fusion medium. The fused embryos were activated after the fusion with 10 $\mu$M calcium ionophore for 5 min and 2 mM 6-dimethyl- aminopurine for 3 h. The nuclear transfer embryos were cultured in 500 ${mu}ell$ well of modified CR1aa supplemented with 3 mg/$m\ell$ BSA in th $\varepsilon$ four well dish cove red with mineral oil. After 3 days culture, culture medium was changed into modified CRlaa medium containing 1.5 mg/$m\ell$ BSA and 5% FBS for 4 days. The incubation environment was 5% $CO_2$, 5% $O_2$, 90% $N_2$ at 38.5$^{\circ}C$. When the cumulus cells were fused with enucleated oocytes by three different fusion pulses, one pulse of 180 volts for 15 $mutextrm{s}$ yielded the highest fusion rate and developmental rate to blastocyst among the pulses (P＜0.05). When the fetal fibroblast cells were fused with enucleated oocytes, one pulse of 180 volts for 30$mutextrm{s}$ yielded significantly higher fusion rate compared with that for 15 $mutextrm{s}$(P＜0.05). The present result indicates that the fusion rate between karyoplast and cytoplast was affected by the cell type and the optimal fusion condition was different according to cell type or size. When the fusion was conducted by the use of mannitol and ZCFM, the fusion rate was 71.2% and 65.8%, respectively. The developmental rates to blastocyst were 37.8% and 39.8%, respectively. There was no significant difference between two fusion media in the developmental rate of cumulus cell nuclear transfer embryos. These results indicate that optimal electric current should be selected according to cell type.

• Korean Journal of Animal Reproduction
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• v.24 no.1
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• pp.49-57
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• 2000

This study was conducted to investigate the developmental potential of cloned embryos derived from bovine fetal fibroblast cells, and the effect of quiescent treatment, passage number and origin of donor cells on in vitro development of cloned embryos. Fetal skin and liver-derived fibroblast cells were transferred to enucleated oocytes after serum starvation or nontreatment (cycling). After electrofusion. reconstituted embryos were activated with $Ca^{＋＋}$-ionophore and cycloheximide, and cocultured for 7~9 days with BRL cells. Some blastocysts were transferred to recipient cows 7~8 days post estrus. The development rate to the blastocyst stage of serum starved cell-derived embryos was higher (25.3%) than that of actively dividing cells-derived embryos (15.9%), The rates of blastocyst formation were 23.1~25.0% after transfer of cell passaged 4 to 6 times, and 23.8 and 25.2% after transfer of fetal skin and liver cells, respectively. After embryo transfer, 34.4% and 15.6% of recipient cows were pregnant on Day 60 and 120, respectively, and one male calf was produced from skin-derived vitrified blastocyst. The result of this study showed that the development of cloned embryos. was enhanced by quiescent treatment, but did not different among the cells passaged 4 to 6 times, and between skin and liver cells. This result also confirms that offspring can be obtained from the vitrified clone embryo derived from fetal skin cell.