• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.76-76
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• 2001

The objective of this study was to optimize the vitrification method of in vitro produced bovine embryos. Thus, in vitro produced embryos at 8 cell, morula and blastocyst stages were vitrified on electron microscope grids (EM grids) or in open pulled straws (OPS) with EG5.5 (5.5 M ethylene glycol, 1.0 M sucrose and 10% FBS in m-DPBS medium) freezing solution and their survival rates after thawing were compared. The embryos on EM grids or in OPS were briefly exposed to EG5.5 freezing solution and plunged directly into liquid nitrogen within 30 to 35 sec. Post-thawed embryos were serially diluted in 0.5, 0.25 and 0.125 M sucrose in m-DPBS, each for 1 min, and then cultured in CRI aa medium supplemented with 10% FBS. Embryonic survival rate was assessed as re-expanded and hatched rates of those embryos after warming. The rates of re-expansion embryos did not significantly different between EM grid (8 cell: 42.10%, morula: 66.66% and blastocyst: 77.08%) and OPS (8 cell: 47.36%, morula: 61.90% and blastocyst: 83.33%) methods. In addition, the hatched rates in EM grid (8 cell: 31.57%, morula: 57.14% and blastocyst: 72.91%) were similar to those in OPS (8 cell. 34.21%, morula: 50.00% and Blastocyst: 77.08%). Interestingly, even at the same blastocyst stage, the in vitro survival of day 7 embryos (EM grid: 79.48 and OPS: 87.18%) was higher than those of day 8 embryos (EM grid: 72.10 and OPS: 82.06%). The total cell number of blastocyst developed in vitro after vitrification was examined with Hoechst 33342 staining to compare the embryo quality among different treatment groups. The total cell number of blastocyst was not significantly different between vitrified groups (EM grid: 162.4$\pm$8.0 and OPS: 158.4$\pm$7.1) and unvitrified control (168.0$\pm$5.6). These results indicate that both vitrification containers can provide the high rate of embryo survival. Moreover, the OPS container may not need a cap to protect the container from floating after immersion in L$N_2$. Therefore, this study suggest that bovine embryos can be cryopreserved easily, effectively and successfully by vitrification method using EM grid or OPS with EG5.5 freezing solution. In the future, the Pregnancy rate would be investigated after transfer of our vitrified embryos into the appropriated recipients.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.78-78
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• 2002

The role of heat shock proteins in shielding organism from environmental stress is illustrated by the large-scale synthesis of these protein by the organism studied to date. However, recent evidence also suggests an important role for heat shock protein in fertilization and early development of mammalian embryos. Effects of elevated in vitro temperature on in vitro produced bovine embryos were analysed in order to determine its impact on the expression of heat shock protein 70 (HSP70) by control and frozen-thawed after in vitro fertilization (IVF) or nuclear transfer (NT). The objective of this study was to assess the developmental potential in vitro produced embryos with using of the various containers and examined expression and localization of heat shock protein 70 after it's frozen -thawed. For the vitrification, in vitro produced embryos at 2 cell, 8 cell and blastocysts stage after IVF and NT were exposed the ethylene glycol 5.5 M freezing solution (EG 5.5) for 30 sec, loaded on each containers such EM grid, straw and cryo-loop and then immediately plunged into liquid nitrogen. Thawed embryos were serially diluted in sucrose solution, each for 1 min, and cultured in CRI-aa medium. Survival rates of the vitrification production were assessed by re-expanded, hatched blastocysts. There were no differences in the survival rates of IVF using EM grid, cryo-loop. However, survival rates by straw were relatively lower than other containers. Only, nuclear transferred embryos survived by using cryo-loop. After IVF or NT, in vitro matured bovine embryos 2 cell, 8 cell and blastocysts subjected to control and thawed conditions were analysed by semiquantitive reverse transcription polymerase chain reaction methods for hsp 70 mRNA expression. Results revealed the expression of hsp 70 mRNA were higher thawed embryos than control embryos. Immunocytochemistry used to localization the hsp70 protein in embryos. Two, 8-cell embryos derived under control condition was evenly distributed in the cytoplasm but appeared as aggregates in some embryos exposed frozen-thawed. However, under control condition, blastocysts displayed aggregate signal while Hsp70 in frozen-thawed blastocysts appeared to be more uniform in distribution.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.92-92
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• 2002

Considerable attention has been focused on the cryopreservation of semen and estrus induction in dog, as consequence of poor productivity caused by long anestrus period, in order to enhance the productivity of youngs and to preserve the breeds. The objectives of this study were to evaluate semen quality after cryopreservation and to evaluate the Pregnancy rate after insemination (AI). Fifty infertilie dogs (age 2∼3 years) were selected for the study and divided into three different estrus induction treatment groups. Group 1: dogs (n=15) were given clomifene (0.1 mg/kg) orally for five days at 12 hr intervals. Group 2: dogs (n=15) were given bromocriptine (50 $\mu\textrm{g}$/kg) orally for five days at 12 hr intervals, followed by single injection intravenously of 500 IU GnRH (Group 3, n=20) when pro-estrus occurred. The rates of pregnancy in estrus inducted dogs mated naturally compared to those inseminated artificially with ejaculated fresh semen and frozen-thawed semen. Estrus detection was performed using the method of vaginal smear and confirmed by the plasma progesterone assay. The ejaculated semen to freeze was exposed to a mixture of Tris extender with cryoprotectant (Trisma, 81 mM: TES, 209 mM: citric acid, 6 mM; glucose, 5 mM; glycerol, 8%) and cryopreserved gradually by slow-cooling at 17 cm above the surface of liquid nitrogen (LN$_2$) for 23 min. The motility of frozen-thawed spermatozoa was assessed by phase-contrast microscopy. To assess their viability and acrosome content, spermatozoa were stained with a vital stain and Fluorescence conjugated lectin Pisum Savitum Agglutinin (FITC/PAS), respectively. Pregnancy was confirmed by ultrasonograpy on day 25, 35 and 55 post insemination. The use of fresh semen, the pregnancy rates were observed 66.6, 66.6, 75.0 and 83.3% in natural estrus, clomifene induced, bromocriptine induced and a combination of GnRH and bromocriptine, respectively. The use of frozen-thawed semen, the pregnancy rates were observed 66.6, 33.3, 50.0 and 60.0% in natural estrus, clomifene induced, bromocriptine induced and a combination of GnRH and bromocriptine, respectively. No difference was observed in the number of offspring produced among natural estrus and treated groups inseminated with fresh or frozen-thawed semen. In conclusion, the pregnancy rate of dogs treated with a combination of GnRH and bromocriptine was more effective than use of clomifene or bromocriptine only. In addition, frozen-thawed semen can be used successfully far artificial insemination in dog.

• Analytical Science and Technology
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• v.20 no.4
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• pp.347-354
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• 2007

A simple and rapid high-performance liquid chromatography assay for the determination of residual novobiocin levels in bovine, porcine, chicken, flatfish and japanese eel muscle has been developed and validated. The separation condition for HPLC/UV was optimized with phenyl hexyl ($4.6{\times}150mm$, $5{\mu}m$) column with 10 mM monobasic sodium phosphate buffer (pH 2.5)/acetonitrile (50/50, v/v) as the mobile phase at a flow rate of 1.0 mL/min and detection wavelength was set at 254 nm. Residues were extracted from tissue by blending with methanol and lipid materials were removed with n-hexane. Then, the methanol extract was evaporated to dryness under a nitrogen stream, reconstituted in the mobile phase. Aliquot of the organic extract was decanted and filtered through $0.45{\mu}m$ syringe filter. The $20{\mu}L$ of the resulting solution was injected into the HPLC system. The calibration ranges were $0.5{\sim}5{\mu}g/g$ and calibration curves were linear with coefficients of correlation better than 0.95. The limits of quantification were $0.5{\mu}g/g$ for all muscles. The recoveries of bovine, porcine, chicken, flatfish and japaneseel muscles were 99.8%, 102.4%, 91.0%, 104.0% and 93.0%, respectively. The procedures were validated according to the CODEX guideline, determining specificity, linearity, accuracy, precision, quantitation limit and recovery.

• Journal of The Korean Society of Grassland and Forage Science
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• v.27 no.4
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• pp.287-296
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• 2007

The experimental work was conducted to determine the growth characteristics and yield of whole crop rice (cv. Suwon 468 and cv. Chuchungbeo) and soil properties using various type of livestock manure application on rice paddy land for 3 years ($2003{\sim}3005$). Compared Suwon 468 and Chuchungbeo, Suwon 468 has longer plant height and more DM yield than that of Chuchungbeo. Among livestock manure type, plant height was longer in order of liquid swine manure (LSM) > composted swine manure (CSM) > chemical fertilizer (CF) > composted cattle manure (CCM). Number of branch on Chuchungbeo had more than that of Suwon 468. Among livestock manure type, number of branch had more in order of LSM > CSM > CF > CCM. DM yield of whole crop rice (WCR) was affected by various types of livestock manure application and increased in order CSM > CCM = LSM. DM yield on the effects of application level of LSM was highest in LSM 75% + CF 25%. Plant diseases such as rice blast, damage by insect, smut, sheath blight occurred in LSM and CSM and there was not significantly different among application level of LSM. The nitrogen content of WCR by CSM was the highest of all treatments and the ripened ratio by CSM was in contrary order. Moreover the feed value of WCR was not significantly different among treatments. Soil pH, organic matter and total nitrogen was not different by LSM application whereas phosphorus content increased by LSM application. Cu and Zn content increased by LSM and CSM application and were not different by CCM as compared to control plots.

• Journal of The Korean Society of Grassland and Forage Science
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• v.14 no.1
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• pp.50-56
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• 1994

This study was investigated for the purposes of securing forage resource using idled rice paddy recently increased in accordance to a current trend of farm products' liberalization and also of presevation of environment by using cattle sluny as liquid manure, which is seriously increasing these days. In this study, mean annual dry matter yield and its seasonal variation with reed canarygrass, and a optimum rate of cattle sluny application were investigated. The results are as follows: 1. According to the conditions of cutting frequencies(3, 4 and 5 cutting per year), mean annual dry matter yield was recorded from 8.9 tons to 10.9 tons per hectare and was the highest at 3 cutting frequency. 2. The use of cattle sluny with the levels of between 300 and 360 kg N per hectare showed a significantly higher mean annual dry matter yield than that of the control (non-fertilization). 3. The treatments with 3 and 4 cutting frequencies(90 kg Nhdyear, 120 kg Nhdyear) recorded higher dry matter yields than the control of the former level by 1.23 tons and 2.34 tons respectively and in the treatment of 5 cutting frequency, the second level with cattle sluny of 300 kg Nhdyear showed an increased dry matter yield of 2.11 tons compared to the former level(l50 kg Nhdyear). With regards to nitrogen efficiency, one kg of nitrogen is applied to 13.7, 19.4 and 14.1 kg of dry matter yields in the conditions of 3, 4 and 5 cutting frequencies respectively. 4. In view of seasonal variance of annual dry matter yield, the second cut in 3 cutting frequency, the third cut in 4 cutting frequency and the third in 5 cutting frequency showed the highest ratio as 42, 37 and 32% respectively compared to the total. 5. Under the conditions of this study, the 'Input-Output curve' from 5 cutting frequency was the closest to sigmaformed process(i=0.9993) of various cutting frequencies, and the maximum marginal yield in the treatment was obtained at the level of 250 kg Nha with cattle sluny. The economic level of cattle sluny was between 371.0 and 402.2 kg N and the highest dry matter yield was obtained at 489.3 kg Mdyear in the same treatment

• Horticultural Science & Technology
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• v.33 no.2
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• pp.268-275
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• 2015

We evaluated the effect of mixed liquid fertilizer (MLF) on growth responses of plants and soil chemical properties. To do so, a pot experiment with cherry tomatoes (Lycopersicon esculentum var. cerasiforme) using loam soil was conducted for 81 days in a temperature-controlled glasshouse, and four N fertilization treatments were laid out in a completely randomized design with three replicates: control (C), chemical fertilizer treatment (CF), and two rates (MLF-0.5 and MLF-1.0) of MLF treatment. Soils were periodically sampled and analyzed for pH, EC, total N, inorganic N and total C, and growth characteristics of cherry tomatoes were measured. During the experimental periods, the pH of MLF soils was higher than that of CF soils. Soil total-N content increased right after CF-application and ultimately decreased to the level of the control (C) soil, while MLF-application slightly increased the level of soil total-N and this level remained unchanged throughout the experiment. The levels of soil inorganic N content increased after application of CF or MLF, but the initial increase disappeared in 56 days after transplanting (DAT). The dry weight of shoots and roots increased in CF or MLF plants, while the number of fruit increased only in MLF plants. Whereas soluble solid contents were higher in MLF plants than in the other (C and CF) plants, the titratable acidity was not different among treatments. However, no consistent effect of N treatments on major elements of the organs of cherry tomatoes was found. The amounts of N taken up by plants were 0.91 g for CF, 0.61 g for MLF-1.0, 0.43 g for MLF-0.5, and 0.25 g for control treatments, resulting in greater N efficiency for CF than for MLF.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.34 no.4
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• pp.389-393
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• 2001

For an effective utilization, quality stability of instant powdered oyster soup made of canned oyster processing waste water (IPSW) was determined. Instant powdered soup from oyster hot-water extracts (IPSE) was also prepared by mixing hot-water extract powder (15 g) with table salt (5 g), cream powder (19 g), milk replacer (12 g), wheat flour (20 g), corn flour (15 g), starch (5 g), glucose (7.5 g) and onion powder (1.5 g). In preparing IPSW, mixed powder from wash water and boiling liquid waste, instead of powder from hot-water extracts and table salt, was added (powder from boiling liquid waste: powder from wash water= 12: 8) and other additives were added in proportion to those in the IPSE. The moisture content, water activity, peroxide value and fatty acid composition showed little changes during storage of the IPSW. The pH, volatile basic nitrogen content and brown pigment formation increased slightly, while white index decreased slightly during storage of IPSW. No significant difference was observed in the changes of food component during storage between IPSW and IPSE. According to a sensory evaluation, the change in quality of IPSW was negligible during 12 months of storage. from the results of the chemical experiment and sensory evaluation, IPSW packed with laminated film bag (OPP, $20{\mu}m/PE,\;20{\mu}m/paper,\;45g/m^3/PE,\;20{\mu}\;m/Al,\;7{\mu}\;m/PE,\;20{\mu}m$) was revealed to be preserved in good quality during 12 months of storage.

• Korean Journal of Poultry Science
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• v.41 no.4
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• pp.261-270
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• 2014

This study was conducted to establish the method for preserving chicken primordial germ cells (PGCs) that enables long-term storage in liquid nitrogen ($LN_2$) for developmental engineering or preservation of species. The purpose of this study is to clarify the effects of simple freeze-thaw treatment on viability of PGCs in chickens and to the optimal protocol for PGCs freezing. PGCs obtained from the germinal gonade of an early embryos of 5.5~6 day (stage 28) of Isa Brown, Korean Ogye (KO), White Leghorn and Commercial breeds, using the MACS method were suspended in a freezing medium containing a freezing and protecting agents (e.g. dimethyl sulfoxide (DMSO), ethylene glycol (EG) and propylene glycol (PG)). The gonadal cells, including PGCs, were then frozen in 1 of the following cryoprotectant treatments : 2.5%, 5%, 10%, 15%, and 0% cryoprotectant (DMSO, EG, PG) as a control. Effects of exposure to simple freezing, with different concentrations of the cryoprotectant solution, were examined. After simple freezing, the viability of PGCs after freeze-thawing was significantly higher for Commercial breeds ($88.7{\pm}2.4%$) than KO ($85.1{\pm}0.4%$), Isa Brown ($84.6{\pm}0.2%$) and White Leghorn ($85.9{\pm}0.1%$) (p<0.05) using 10% EG cryoprotectant. Therefore, these systems may contribute in the improvement of cryopreservation for a scarce species in birds preservation. This study established a method for preserving chicken PGCs that enables systematic storage and labeling of cryopreserved PGCs in liquid ($LN_2$) at a germplasm repository and ease of entry into a database.

• Journal of the Korean Society for Marine Environment & Energy
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• v.15 no.2
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• pp.67-75
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• 2012

Carbon-dioxide capture and storage (CCS) process is consisted by capturing carbon-dioxide from large point source such as power plant and steel works, transporting and sequestrating captured $CO_2$ in a stable geological structure. During CCS process, it is inevitable of introducing impurities from combustion, capture and purification process into $CO_2$ stream. Impurities such as $SO_2$, $H_2O$, CO, $N_2$, Ar, $O_2$, $H_2$, can influence on process efficiency, capital expenditure, operation expense of CCS process. In this study, experimental apparatus is built to simulate the behavior of $CO_2$ transport under various impurity composition and process pressure condition. With this apparatus, $N_2$ impurity effect on $CO_2$ mixture transportation was experimentally evaluated. The result showed that as $N_2$ ratio increased pressure drop per mass flow and specific volume of $CO_2-N_2$ mixture also increased. In 120 and 100 bar condition the mixture was in single phase supercritical condition, and as $N_2$ ratio increased gradient of specific volume change and pressure drop per mass flow did not change largely compared to low pressure condition. In 70 bar condition the mixture phase changed from single phase liquid to single phase vapor through liquid-vapor two phase region, and it showed that the gradient of specific volume change and pressure drop per mass flow varied in each phase.