• Korean Journal of Animal Reproduction
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• v.23 no.4
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• pp.313-321
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• 1999

The purpose of these study was to investigate the use of a glass micropipette (GMP) as a vessel for vitrification of bovine IVP blastocysts, to compare the post-thaw survival rates of bovine blastocysts frozen in GMP with those frozen in OPS that have been previously investigated, and to improve the hatching rate following vitrification with GMP method. The GMP vessel permits higher freezing and warming rate than the OPS due to the higher heat conductivity of the glass and lower mass of the solution that contains the embryos. Groups of three bovine IVP blastocysts were sequentially placed into vitrification solution before being loaded into either the OPS or GMP vessels and immersed into L$N_2$within 20 to 25 sec. Post-thaw blastocysts were serially washed in 0.25 and 0.15 M sucrose in HM and TCM-199 for each 5 min, respectively, and then cultured in TCM 199 supplemented with 10% FCS for 24 h. The rate of blastocyst re-expanding did not significantly different for OPS (75.9%) and GMP (80.0%) methods (P＞0.05). The hatching rates in OPS (34.1%) and GMP (37.5%) methods were significantly lower than that in control group (54.3%) (P＞0.05). In addition, the rate of blastocyst re-expanding was significantly lower if blastocysts were vitrified in the wide portion of the micropipette rather than the narrow portion of the micropipette (83.3 vs 56.7%) (P＞0.05), even though three blastocysts were loaded per vessel. The hatching rate in 0.05% pronase solution treatment for 30, 60 and 90 see (45.9, 54.7 and 57.5%) were significantly higher than that in control (35.0%), even though there was not significantly different between 30 see and control. These results indicate that both vitrification vessels can provide high survival rates of bovine IVP blastocysts. However, the GMP vessel has the advantage over the OPS, in that the former does not need a cap to protect the vessel from floating after immersion in L$N_2$. The location of the embryos (narrow or wide portion of immersion) were considered to be limiting factors to the viability of bovine IVP embryos. The exposing in 0.05% pronase solution for 60 or 90 see can increase hatching rates of post-thaw bovine IVP blastocysts.

• Korean Journal of Animal Reproduction
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• v.24 no.2
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• pp.199-208
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• 2000

This study was carried out to investigate of factors concerning to ultrasound-guided follicular aspiration; level of vacuum pressure, diameter of use needle, effect of FSH hormone and conception rate after embryos transfer. 1. Oocytes collection number were 4.2$\pm$2.9 e.a to luteuml phase and follicular phase were 4.4$\pm$3.5 e.a to ovaries of Hanwoo. 2. We taked proper level of aspiration vacuum pressure was 40~120 mmHg to oocytes collection. Oocytes collected number were 4.2$\pm$3.2, 4.3$\pm$3.4, 4.5$\pm$3.4 e.a. to 40, 80, 120mmHg, respectively, follicles aspiration rate were 49, 47, 45%. 3. Effect of collection needle diameter was not difference significantly(P＜0.05), oocytes collected number were 4.4$\pm$3.5 e.a to 170 and 3.0$\pm$1.8 e.a to 18G needle, collected oocytes quality were no difference significantly (P＜0.05). 4. Follicles increase number to FSH hormone injection were 6.2$\pm$2.3 e.a to intramuscle and 1.1$\pm$2.7 e.a. to epithelial injection method. 5. Conception rate derived from E.T. was 11.1% to freezing embryos and 46.2% to fresh E.T., difference significantly(P＜0.05).

• Korean Journal of Animal Reproduction
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• v.23 no.3
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• pp.247-256
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• 1999

The present study was performed to improve the reproductive disturbance as well as the elimination of microbiological contamination for animals bred under conventional conditions followed by in vitro fertilization and embryo transfer techniques including embryo and sperm freezing, using a mouse strain(M. m. molossinus-tt＠Kist) showing the abnormal behavior disorder derived from Korean wild mice (Mus musculus molossinus). Moreover, hematological and serum biochemical analyses were also carried out to obtain the basic data of this mouse strain The results are summarized as follows: 1. In comparison with hematological data, the numbers of RBC and platelet of this mouse strain were appeared as the higher value those that of the same aged inbred strains such as BALB/c, DBA/2, C57BL/6 and C3H /Hen. However, no differences were found in values of WBC, Hb and Ht. Moreover, total cholesterol of this strain showed a low value but triglyceride, total protein and albumin values were similar as in inbred strains. 2. The average numbers of superovulated oocytes treated with 2.5/2.5 IU and 5.0/5.0 IU of PMSG/hCG were 11.6 and 12.7, respectively. The fertilization rates of 2.5/2.5 IU PMSG /hCG treatment(87.9%) was higher than 5.0/5.0 IU treatment(52.0%) (p＜0.05) and the developmental rate of 2 cell stage embryos were 외 so appeared as higher value 99.0% and 90.6%, respectively. 3. The rates of in vitro fertilization treated with frozen sperm(24.8%) was significantly lower than of that fresh sperm(87.9%), (p＜0.05). 4. The five, six and ten heads of offspring were obtained from frozen-thawed 2 cell embryos by in vitro fertilized, 2 cell embryos from in vitro fertilized by frozen-thawed spermatozoa. and 2 cell embryos by in vitro fertilization, respectively. These offspring developed the expected disease about 2 weeks after birth, which was confirmed that the disease character of this mutant mouse strain was reliably reproduced. 5. MHV(Mouse hepatitis virus) and Staphylococcus aureus were successfully eliminated from conventional animals by in vitro fertilization-embryo transfer and the use of SPF recipient animals.

• Korean Journal of Poultry Science
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• v.40 no.3
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• pp.171-178
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• 2013

The purpose of this study was to evaluate the sperm viability, normal acrosome and mitochondrial activity in the frozen-thawed fowl semen by different cryoprotectants. The experiment was carried out on 10 sexually adult roosters of Ogye. The semen was collected twice a week and pooled semen was diluted 1:1 EK extender containing no cryoprotectant at $5^{\circ}C$. After equilibration for 30 minutes, diluted chicken semen was diluted 1:1 extender containing either 7% dimethylacetamide (DMA), 7% dimethylformamide (DMF) or 7.5% methylacetamide (MA) at final concentration and was put in 0.5 mL plastic straws and frozen for 30 minutes by exposure to liquid nitrogen vapor 4 cm above the surface of liquid nitrogen, followed by plunging into liquid nitrogen. Frozen semen was thawed in water bath at $5^{\circ}C$ for 2 minutes. For cytometric analysis, the frozen-thawed semen was diluted with EK extender to a final concentration of 90 million spermatozoa per mL. Sperm membrane integrity was evaluated as SYBR-14 and propidium iodide (PI). Acrosome integrity was assessed with fluorescein isothiocyanate-labeled PSA and PI. The percentage of mitochondrial function was estimated by using Rhodamine123 (R123) and PI. In conclusion, freezing rooster semen by using 7% DMF as cryoprotectant was significantly highest in rates of survival and mitochondrial function while its rate of damage of acrosome was significantly lowest. As a result, DMF is the cryoprotectant that has the lowest influences on sperm membranes and acrosome integrity. Therefore it could be used for freezing method of animal genetic conservation method for poultry diversity.

• Korean Journal of Agricultural and Forest Meteorology
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• v.14 no.3
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• pp.124-131
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• 2012

This study was carried out to evaluate a possible change in freeze risk for 'Changhowon Hwangdo' peach buds in three major peach growing areas under the future climate projected by RCP8.5 emission scenario. Mean values of the monthly temperature data for the present decade (2000s) and the future decades (2020s, 2050s, 2080s) were extracted for farm lands in Icheon, Chungju, and Yeongcheon-Gyeongsan region at 1km resolution and 30 sets of daily temperature data were generated randomly by a stochastic process for each decade. The daily data were used to calculate a thermal time-based dormancy depth index which is closely related to the cold tolerance of peach buds. Combined with daily minimum temperature, dormancy depth can be used to estimate the potential risk of freezing damage on peach buds. When the freeze risk was calculated daily for the winter period (from 1 November to 15 March) in the present decade, Icheon and Chungju regions had high values across the whole period, but Yeongcheon-Gyeongsan regions had low values from mid-December to the end of January. In the future decades, the frequency of freezing damage would be reduced in all 3 regions and the reduction rate could be as high as 75 to 90% by 2080's. However, the severe class risk (over 80% damage) will not disappear in the future and most occurrences will be limited to December to early January according to the calculation. This phenomenon might be explained by shortened cold hardiness period caused by winter warming as well as sudden cold waves resulting from the higher inter-annual climate variability projected by the RCP8.5 scenario.

• Korean Journal of Food Science and Technology
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• v.30 no.4
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• pp.862-870
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• 1998

This study shows the application of Ci-ELISA method for monitoring the denaturation of myosin by the frozen treatment in order to differentiate thawed beef from chilled. Hanwoo M.Semitendinosus (n=25) was treated under the two different frozen process as follows; simple frozen treatment (Exp-1) at 4 different temperatures, -10, -20, -50 and $-80^{\circ}C$, respectively, and repeated thawing-refreezing treatment (Exp-2) stored at 4 different temperatures, -10, -20, -50 and $-80^{\circ}C$, respectively. Antibodies (Abs) were produced from rabbits immunized with myosin whole molecule (MWM) isolated from beef round, heavy meromyosin S-1 (S-1) and light meromyosin (LMM) prepared by digestion of MWM. Each immunoglobulin G (IgG) was separated from antiserum. At 6 month storage, IA of anti-MWM IgG for myosin was decreased to 32.67, 32. 23, 51.52 and 34.27% in Exp-1 and to 14.82, 15.61, 25.3 and 23.7% in Exp-2 at -10, -20, -50 and $-80^{\circ}C$, respectively (P<0.05). In Exp-1, the reactivities of anti-LMM IgG were decreased to 25.12, 21.42, 49.05 and 28.96%, and those of Exp-2 were to 11.88, 9.56, 20.63 and 12.64% at -10, -20, -50 and $-80^{\circ}C$, respectively, at 6 times thawing (P<0.05). Conclusively, myosin was denaturated by freezing treatment and LMM or myosin rod part might have suffered from more extreme demage than HMM S-1, and samples at $-50^{\circ}C$ were slightly injured less than others by freezing treatment.

• Korean Journal of Animal Reproduction
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• v.23 no.2
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• pp.141-148
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• 1999

This study was to test whether the viability of bovine hatched blastocysts (HBs) can be maintained after vitrification and thawing. The HBs were produced in vitro at Day 9 and Day 10 after IVF, and they were classified to small (S-HBs; ø$\leq$300 ${\mu}{\textrm}{m}$) and large(L-HBs; ø＞300 ${\mu}{\textrm}{m}$) on the basis of embryo diameter using eyepiece micrometer. As freezing solution, we used EFS35 which consisted of 35% ethylene glycol (EG), 18% ficoll, 0.3 M sucrose and 10% FBS added in mDPBS. Vitrification was taken by two-step method, the HBs were equilibrated in 10% EG for 5 minutes and then shortly exposed in EFS35 and plunged into L$N_2$for 30~45 sec. After thawing, the survival rates were assessed by the re-expansion of the blastocoel during 2 h and 16 h of culture. The results obtained in these experiments were summarized as follows; 1) When the blastocysts(40.8%) recovered at Day 8 after IVF were further cultured for 24 h(Day 9 after IVF) and 48 h(Day 10 after IVF), the rates of HBs were 20.5% and 6.7%, respectively. Also, the total cell number of HBs on Day 9 was significantly higher than that of HBs on Day 10 (p＜0.01). 2) When the effects of freezing solution to the survival of Day 9 L-HBs were examined, the rate of vitrified group (75.7%) was significantly lower than 100% of control and exposed group(p＜0.05). 3) When the survival rates of vitrified HBs according to size and developmental age were examined, the data of L-HBs (75.5%) and S-HBs(63.6%) on Day 9 were slightly higher than those of L-HBs(64.3%) and S-HBs(60.7%) on Day 10. 4) Also, when the in vitro survival of Day 9 HBs was evaluated under different culture condition after thawing, the result in culture medium only (79.3%) was significantly higher than 43.2% in co-culture group (p＜0.05). These results demonstrated that bovine HBs can be successfully cryopreserved by two-step vitrification method using EFS35.

• Korean Journal of Environment and Ecology
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• v.28 no.5
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• pp.500-509
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• 2014

To prevent freezing of the road by fallen snow, Calcium chloride($CaCl_2$) as a deicer is used to very often and it can be harmful to roadside trees. This study was conducted to investigate the effects of Calcium chloride($CaCl_2$) as a deicer on growth and physiological traits of Acer triflorum according to different concentration of $CaCl_2$. We measured growth, chlorophyll contents, gas exchangement characteristics, chlorophyll fluorescence and mineral nutrition concentration in plant and soil. The experimental group was composed of four treatments including 0mM(control), 9mM(0.5 %), 18mM(1.0 %), 54mM(3.0 %). Before germinating new shoot, the dissolution of $CaCl_2$ was irrigated twice interval of a week. At 30 days after treatment, all treatments decreased total cholorophyll content, photosynthetic rate, transpiration rate, stomatal conductance and photochemical efficiency($F_v/F_m$) with increasing concentration of $CaCl_2$ and especially, they significantly reduced in 3.0 % treatment. In contrast, chlorophyll a/b ratio increased with an increase of $CaCl_2$ concentration and water use efficiency increased in 1.0 % and 3.0 % treatments. At 50 days after treatment, all treatments were decreased in chl a, chl b, total chlorophyll content, carotenoid content, photosynthetic capacity, photochemical efficiency($F_v/F_m$) and quantum yield of photosystem II(${\Phi}_{PSII}$) compared with control and 3.0 % treatments were withered. $Ca^{2+}$ and $Cl^-$ were accumulated in leaves and soil, which inhibited water absorption and electron transport and it caused the reduction of height growth rate more than 50 %. Although there was a little difference according to time and $CaCl_2$ concentration, all treatments decreased in growth rate and physiological activity slowed down. As time passed, these results got worse. Therefore we need to take a measure earlier in order to minimize damage of trees.

• Journal of the Korean GEO-environmental Society
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• v.13 no.12
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• pp.5-16
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• 2012

The second scientific antarctic station of South Korea is under construction at Terra Nova Bay located in eastern Antarctica. Ground condition in the Antarctica is frozen in general, but there are seasonal frozen grounds with active layers sporadically. When the active layer is frozen, frost heaving occurs that might cause the differential movement of frozen ground and the failure of structures. Therefore, it is necessary to determine the frost heaving susceptibility of soils at Terra Nova Bay before starting antarctic station construction. This study presents experimental investigation of the frost heaving susceptibility of soil samples with variation of particle sizes and unfrozen water contents. The soil samples were taken from five different locations at Terra Nova Bay and physical properties, unfrozen water content, and frost heaving tests were performed. For the frost heaving tests, soil specimens were frozen with constant freezing temperatures at the top and with drainage at the bottom in order to stimulate the frost heaving. The frost heaving tests provide volume expansion, volumetric strain, and heaving rate which can be used to analyze the relationship between the frost heaving vs. particle size and the frost heaving vs. unfrozen water content. Experimental results show that the more the fine contents exist in soils, the more frost heaving occurs. In addition, the frost heaving depends on unfrozen water content. Experimental data can be used to evaluate the frost heaving susceptibility of soils at the future construction site in the Antarctica.

• Journal of the Korean Geotechnical Society
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• v.27 no.10
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• pp.13-23
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• 2011

Bearing capacity of pile foundations in cold region is dominated by adfreeze bond strength between surrounding soil and pile perimeter. It denotes that adfreeze bond strength is the most important design parameter for foundations in cold region. Adfreeze bond strength is affected by various factors like 'soil type', 'frozen temperature', 'normal stress acting on soil/pile interface', 'loading rate', 'roughness of pile surface', etc. Several methods have already been proposed to estimate adfreeze bond strength during past 50 years. However, most methods have not considered the effect of normal stress for adfreeze bond strength. In this study, both freezing temperature and normal stress have been controlled as primary factors affecting adfreeze bond strength. A direct shear box was used to measure adfreeze bond strength between sand and aluminum under different temperature conditions. Based on the test results, the relation between shear strength of frozen sand and adfreeze bond strength have been investigated. The test results showed that both of shear strength and adfreeze bond strength tend to increase with decreasing frozen temperature or increasing confining pressure. The ratio of shear strength and adfreeze bond strength, expressed as $r_s$, decreased initially frozen section but increased at much lower frozen temperature and there were uniform intervals under the different normal stress conditions. A method for predicting adfreeze bond strength using $r_s$ has finally been proposed in this study.