• Journal of Embryo Transfer
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• v.22 no.2
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• pp.89-95
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• 2007

The present study was conducted to examine some factors affecting in vitro development and fecundity of embryos recloned with somatic cell nuclear transfer (SCNT). Fibroblast cells retrieved from the ear of a 3-week-old, cloned Korean goat (Jinsoonny) were used as karyoplast donors and serum-starvation was conducted in tissue culture medium (TCM)-199 supplemented with 0.5% FBS. Recipient oocytes were surgically collected by flushing the oviducts 35 h after hCG injection following FSH priming. The zonae pellucidae of the oocytes were partially perforated with a laser drill and a donor cell was transferred into an enucleated oocyte. The couplets were electrically fused and activated by ionomycin (5 min) and 6-DMAP (4 h). The reconstructed embryos were cultured in mSOF medium containing 0.8% BSA at $39^{\circ}C$ in an atmosphere of 5% $CO_2$, 5% $%O_2$, 90% $N_2$ for 12 to 15 h. Re-cloned embryos (2- to 4-cell stages) were surgically transferred into the oviducts of the recipients and pregnancy was subsequently diagnosed by progesterone assay and ultrasound on Days 21 and 63 of pregnancy. The fusion rate following 1st fusion pulse was higher (p<0.05) in 2nd cloning (65.9%) compared to 1st cloning (51.0%), but it was not different in the other groups. The rate of cleavage after fusion was significantly higher (p<0.05) in 1st (77.7%) than in 2nd cloning (56.0%). A total of 175 re-cloned embryos were transferred into 28 recipients. On day 21 and 60 after transfer, 11 (39.3%) and 4 recipients (17.4%) were pregnancy, respectively. In comparison of pregnancy rate by estrous synchronization, a total of 66 and 109 re-cloned embryos were transferred into 11 recipients in natural estrus and 17 recipients in induced estrus, respectively. Five (45.4%) and 2 recipients (18.2%) in natural estrus were pregnant on days 21 and 63 while 6 (35.3%) and 2 (11.8%) recipients in induced estrus were pregnant, respectively. These results show that recloning of goat can be achieved by SCNT and estrous synchronization between donor and recipient animals may be one of the major factors affecting success rate.

• Journal of Embryo Transfer
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• v.19 no.1
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• pp.43-51
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• 2004

The present study was conducted to investigate the effects of fusion and/or activation protocol on in vitro development of porcine somatic cell nuclear transfer (SCNT) embryos. Porcine fetal fibroblast cells were transferred into the perivitelline space of enucleated in vitro matured oocytes. Cell fusion and activation were induced simultaneous fusion/activation (SA) or delayed activation (DA) with or without cytochalasin B (CB) treatment with electic pulses in 0.28 M mannitol-based medium. The SCNT embryos were cultured in vitro for 7 days and stained with Hoechst 33342 to determine the number of nuclei. After 7 days culture, cleavage and blastocyst formation rates were 72.4% and 7.6% in SCNT and 76.3% and 20.4% in parthenotes. To examine the effect of electric field strengths on development of SCNT embryos, oocytes were fused two pulses of 110 V/mm, 130 V/mm or 150 V/mm for 30 sec post-injection. The fusion and cleavage rates in 130 V/mm group (70.2% and 72.6%) and 150 V/mm group (72.6% and 70.5%) were higher (P<0.05) than 110 V/mm group (47.1% and 48.6%), respectively. However, the rate of embryos developing to the blastocyst stage (8.1%, 9.7% and 10.7%) were not different among three groups. The cleavage rates and the blastcyst formation rates were not different among three treatment groups (SA group, 71.4% and 9.7%; SA+CB treatment group, 74.7% and 8.0%; DA+CB treatment group, 70.8% and 11.2%, respectively). And, no different in the number of cells in blastocysts was observed among the three groups (22.5$\pm$12.8, 23.3$\pm$11.2 and 21.6$\pm$10.4, respectively). These result suggest that two pulses of 130 V/mm or 150 V/mm for 30 sec with SA treatment or DA treatment are enough for fusion/activation of porcine somatic cell nuclear transfer (SCNT) embryos to develop to the blastocyst stage.

• Journal of Applied Biological Chemistry
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• v.53 no.1
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• pp.1-7
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• 2010

xylA promoter is a major promoter in xylose operon of Escherichia coli. xylA promoter is sufficient as the promoter for the construction of new expression vector because this promoter was tightly controlled and induced by the addition of xylose. For the construction of xylose-inducible expression vector, 600 bp of xylA promoter was ligated between AatII and HindIII of pUC18, named pXA600. In order to investigate the effect of XylR protein encoded by xylR gene on the xylA promoter, 1,988 bp of xylR gene including its promoter was ligated into downstream of multiple cloning site to the opposite direction of xylA promoter in pXA600, named pXAR600. For the measurement of expression level, 3,048 bp of lacZ structural gene was fused into xylA promoter in both plasmids pXA600 and pXAR600 as a reporter gene, named pXA600-lacZ and pXAR600-lacZ, respectively. The $\beta$-galactosidase activity of pXA600-lacZ and pXAR600-lacZ in E. coli JM109 was determined to be 1,641 and 2,304 unit by the induction with xylose in LB medium, respectively. The $\beta$-galactosidase activity of pXAR600-lacZ/JM109 was about 1.4 times higher by the induction with xylose than that of pXA600-lacZ/JM109. The $\beta$-galactosidase activity of pXA600-lacZ and pXAR600-lacZ in E.coli JM109 showed 6,282 and 9,320 unit by the induction with xylose in DM minimal medium, respectively. A regulator, xylR protein works as an activator for the gene expression by the addition of xylose in the xylose-inducible vectors because the level of gene expression in pXA600 is increased by the insertion of xylR gene into the same vector. The xynA gene of Streptomyces thermocyaneoviolaceus cloned in pXA600 and pXAR600 was successfully expressed in E. coli BLR(DE3). As a result, plasmids pXA600 and pXAR600 using xylA promoter are sufficient as new expression system to produce a foreign protein in E. coli.

• The Journal of Korean Academy of Prosthodontics
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• v.49 no.3
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• pp.191-196
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• 2011

Purpose: In this study, we evaluated the bond-strength between a few metal ceramic-alloys and veneered-porcelain and found if the bond strength of metal-ceramic alloy with lower gold content than the presently used gold alloy was high enough. Materials and methods: For this study, after plasticizing porcelain only for gold alloy, metal specimen was fabricated using Ni-Cr alloy and gold alloy with different gold content. Three point flexural test were performed to measure their bond-strength. Results: The bonding strength of Group 1 to porcelain was $40.62{\pm}3.32$ MPa, which was the highest (P<.05). In sequence of decreasing value, Group 2 (75%) was $37.47{\pm}1.57$ MPa, Group 3 (52%) $35.85{\pm}1.48$ MPa, Group 4 (51.5%) $35.04{\pm}1.34$ MPa, Group 5 (32%) $33.17{\pm}1.62$ MPa, Group 6 (10%) $30.75{\pm}1.21$ MPa. Bonding strength of Group 3 and Group 4 to porcelain did not show statistically significant difference with comparison to that of Group 5 (Duncan's test, P>.05), while there was a significant difference between that of Group 2, Group 3 and Group 4 and that of Group 6 (Duncan's test, P<.05). The bonding strength between gold alloy and porcelain increased according to the content of gold. In all experimental groups showed higher value than 25 MPa, which is the least value recommended by ISO 9693. Conclusion: In all groups, bonding strength was higher than 25 MPa, which is the least value of ISO9693. Therefore, it is considered that metal gold alloy with low gold content is clinically usable.

• The Journal of Korean Academy of Prosthodontics
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• v.48 no.4
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• pp.273-279
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• 2010

Purpose: The purpose of this in vitro study was to compare the marginal adaptation of a ceramic-pressed-to-metal restoration with traditional metal-ceramic restoration. Materials and methods: Duplicating the prepared resin tooth, 20 metal dies were fabricated. Twenty metal copings of 2 groups which were metal ceramic restoration and pressed to metal restoration were fabricated. The marginal opening of each coping was measured with Microscope (BX 60M-36E $41D^{(R)}$: Olympus, Japan). After porcelain build-up, the marginal opening of metal ceramic restoration and pressed to metal restoration ($PoM^{(R)}$: Ivoclar vivadent., Liechtenstein) were also evaluated in the same method. The measurements were analyzed using Wilcoxon Signed Ranks test and Mann-Whitney U test. Results: Within the limits of this study, the results were as follows. 1. Metal-ceramic restorations in coping state ($64.93{\pm}12.48\;{\mu}m$) in compared with Metal ceramic restorations after porcelain build-up ($63.43{\pm}12.86\;{\mu}m$) had no significant difference in marginal adaptation. 2. Pressed-metal-ceramic restorations in coping state ($50.00{\pm}12.28\;{\mu}m$) in compared with Pressed metal ceramic restorations after porcelain build-up ($56.72{\pm}13.80\;{\mu}m$) had no significant difference in marginal adaptation. 3. Metal-ceramic restorations in compared Pressed-metal-ceramic restorations had no significant difference in marginal adaptation. Conclusion: Pressed-metal-ceramic restorations have the advantage of being technically less change through using of the lost-wax technique and this allows for the convenience of a full-contour ceramic wax-up as opposed to the more technique-sensitive layering method. Pressed-metal-ceramic restorations may be considered in clinic on the basis of the result of this study and the advantage of this system.

• Korean Journal of Soil Science and Fertilizer
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• v.34 no.4
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• pp.255-264
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• 2001

This study was conducted to investigate changes in inorganic phosphorus fractions in phosphorus accumulated plastic film house soils under cropping condition. Pot experiment was conducted with surface soils taken from plastic film house fields cultivated for 3, 8 and 16 years. Phosphorus fertilizer was applied at the rates of 0 (P0), $100kg\;ha^{-1}$ (recommended application rate. P1) and $200kg\;ha^{-1}$ (P2) as fused phosphate. Crops were grown sequentially in the order of corn-Chinese cabbage-Chinese cabbage-corn for 3 years. The content of easily soluble-P fraction was decreased with cropping. There were significantly positive relationship not only between the contents of easily soluble-P and the amounts of soil available P ($r=0.839^{**}{\sim}0.952^{**}$ for Bray-1 P, $r=0.895^{**}{\sim}0.967^{**}$ for Lancaster P, and $r=0.491^{**}{\sim}0.821^{**}$ for Olsen P), but also between the amounts of P absorbed by plants and the amounts of easily soluble P decreased ($r=0.644^{**}{\sim}0.822^{**}$). The decrease of easily soluble-P during cropping period could be described by a first-order reaction. The number of cropping times needed to decrease the content of easily soluble-P to an index level of $0.2mg\;kg^{-1}$, which is commonly reported as the desired concentration for soil P, was predicted in the range of 26~33 cropping times by using the equations. Regardless of P fertilization, the proportion of Al-P to total P was little varied during cropping period, but the proportion of Fe-P to total P increased with cropping. Although the content of Ca-P was high before cropping, the proportion of Ca-P to total P was increased with cropping. The proportion of reductant soluble-P to total P was little varied for P fertilizer treatment but was increased for no P fertilizer treatment. The residual-P was decreased during cropping period due to the absorption by crops and the conversion to other inorganic P fractions.

• Reproductive and Developmental Biology
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• v.29 no.3
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• pp.175-180
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• 2005

This study was conducted to examine the effects of demecolcine-assisted enucleation and recipient cell cycle stage on the development of bovine somatic cell nuclear transfer (NT) embryos. In vitro cultured oocytes for $16\~20$ h were classified by first polar body (1st PB) extrusion and cell cycle stage (MI and MII) and treated $0.4\;{\mu}L/mL$ demecolcine for 40 min before enucleation. Enucleated oocytes were fused electrically with bovine ear skin cells, activated by Ca-ionophore+DMAP, and cultured in vitro. Most of eggs ($86.2\%$) treated with demecolcine protruded a chromosome mass and enucleated efficiently ($98.8\%$, (P<0.05). Demecolcine did not have a deteriorative effect on the development of NT embryos. Developmental rate of NT embryos reconstituted with oocytes extruded 1st PB significantly higher than that of NT embryos produced by oocytes without 1st PB ($18.2\%\;vs.\;4.6\%\cdot$, P<0.05). Cleavage and blastocyst formation rate of embryos reconstituted with MI oocytes ($69.4\%\;and\;5.9\%$, respectively) were significantly lower than those of embryos reconstituted with MII oocytes ($96.7\%\;and\;23.9\%$, respectively, P<0.05). From the present result, it is suggested that domecolcine is useful for the enucleation of recipient oocytes in bovine NT procedures, and MII oocytes rather than MI oocytes are more appropriate for recipient cytoplasm. Although, the potential to develop into blastocysts of NT embryos produced by 1st PB-nonextruded and MI oocytes was very low, these oorytes could be used for NT.

• Journal of The Korean Society of Grassland and Forage Science
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• v.19 no.2
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• pp.133-140
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• 1999

This pot experiment was conducted to find out the forage productivity and quality in a grass/clover sward as affected by the application of three different fertilizers; double superphosphate(DS), fused Mg-phosphate(FP), and complex fertilizer(CF), on newly reclaimed hilly soil. This 1st part was concerned with the forage yields, yield components and botanical composition. The results obtained are summarized as follows: 1. The significant differences in the yields(relative yields, %) of whole forages and legume were laid in the following increasing order; CF(relative yield: 150.4% in whole forages and 169.7% in ladino clover) > FP(125.4% in whole forages and 121.7% in ladino clover) > DS(100.0%). The yield of orchardgrass was significantly reduced in the DS plot, compared with the yields in the FP and CF plots, whereas these showed no significant differences between the FP and CF plots. 2. In the botanicl composition, it showed that the rates of grass : legume were 48:52% in the DS and FP, and 40:60% in the CF. It was recognized that the application of complex fertilizer contributed to the increasing of the yields of whole forages, and especially legume yield and rate in the mixed sward.

• Journal of Korean Ophthalmic Optics Society
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• v.17 no.2
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• pp.159-164
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• 2012

Purpose: Of the various methods of vision training, the essay aims to explore the effective ways of using the Fresnel prism lens in order to expand the positive fusional vergence for the patient having specific condition of convergence insufficiency or basic exophoria. Methods: 15 students of city of Daejeon university without an eye disease (average age $22.73{\pm}1.68$) were selected and underwent the subjective refraction test and binocular vision test, and recording their test results before vision training and replacing an identical frame with a lens of same quality after the full calibration, the lens was then adhered with the Fresnel prism lens and continued to train for thirty minutes daily during two weeks. Afterwards, the binocular vision test was reattempted. The observation of the change in the results of the binocular vision test in use of the fresnel prism lens in the vision training test was researched. Results: After training, the positive fusional vergence had increased to a number of $22.27{\pm}2.26$ $\Delta$, to 7.80 $\Delta$, at near, the fused cross cylinder test increased to an average of $0.55{\pm}0.09$ D, 0.40 D after training, showing a normal result. The value of negative relative accommodation after training had an average of $2.22{\pm}0.08$D, showing that 0.42 D had increased. The value of near point of convergence after training had an average of $6.13{\pm}0.53$ cm, showing that 2.80 cm had decreased. To patients who had convergence insufficiency or basic exophoria, the value of the near vision test that used the Fresnel prism lens which was able to expand BO positive fusional vergence had increased without phoria. Conclusions: The changes were tested and the effectiveness of the Fresnel prism lens, due to the nature of the lens itself, helped with both cosmetic effects and cost. It also allows good optical correction effects, in addition to these clinical effects indicated before. Therefore, it may be determined that the Fresnel prism lens binocular vision therapy for patients is more popular and highly recommended.

• Applied Biological Chemistry
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• v.33 no.1
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• pp.93-100
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• 1990

The optimum conditions of electric stimulation for electrofusion of protoplasts of petunia, carrot and soybean, and the effects of calcium, magnesium, protease, trypsin, triton X-100, concanavalin A, dimethyl sulfoxide(DMSO), glycerol monooleate and spermine on fusion frequency and/or viability of petunia protoplast were investigated. The optimum frequencies(Hz)-amplitudes(V/cm) of AC Pulse for protoplast pearl-chain formation were 10 kHz-20 V/cm and 1 MHz-60 V/cm for petunia, 100 kHz-40 V/cm and $1\;MHz-40{\sim}60\;V/cm$ for carrot, and $1\;MHz-40{\sim}80\;V/cm$ for soybean, respectively. The optimum condition of DC pulse treatment at the 1 MHz-60 V/cm-15sec treatment of AC for electrofusion of petunia protoplasts was 2.5 kV/cm-40 sec, and under this condition the fusion frequency and viability of protoplasts were 45 % and 10 %, respectively, Both of the protoplasts of carrot and soybean were not fused under the AC and DC conditions tested in this experiment. The electrofusion of petunia protoplasts was stimulated by calcium, and the fusion frequency and the viability of the protoplasts were 43 % and 11 % , respectively at the calcium concentration of 140 mM. Although fusion frequency was not affected by magnesium only, magnesium stimulated fusion frequency in the presence of calcium, and the viability and fusion frequency of petunia protoplasts were 45 % and 13 %, respectively, at 140 mM of magnesium-140 mM of calcium. The relative fusion frequencies of petunia protoplasts to the controls were increased by 2.4, 2.1, 1.6, 1.4, 1.8, 1.5 and 2.2 folds, respectively, by the treatments of protease, trypsin, triton X-100, concanavalin A, DMSO, glycerol monooleate, and spermine. The viabilities of petunia protoplasts were decreased by these substances.