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Expression of $\beta$-Galactosidase Gene Microinjected into Xenopus Egg During Early Development (초기발생 동안 양서류 난에 미세주입된 $\beta$-galactosidase 유전자의 발현)

  • 차병직;정해문
    • The Korean Journal of Zoology
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    • v.33 no.3
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    • pp.365-372
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    • 1990
  • For the effort to produce transgenic amphibians, a plasmid DNA sequence (cytoplasmic actin promoter-linked bacterial $\beta$-galactosidase gene) was microinjected into fertilized Xenopus eggs. It appeared that the injection of 20 nl solution containing 1-2 ng of DNA was not toxic, but over 4 ng was toxic to embryonic development. The translational product of $\beta$-gal gene ($\beta$-galactosidase) had enzyme activity in all three germ layers of the embryo. Expression of the injected $\beta$-gal genes was first detected at mid-gastrula stage, and the activity persisted up to stage 43 (feeding tadpole) with decreased level of retention. However, the level of the expression was various among the injected individuals as well as each experiment. That is, $\beta$-galactosidase activities did not appear in all cells, instead a localized distribution pattern. Although other possibilities could not be omitted, this mosaic distribution of gene expression seemed to arise from unequal partition of the injected DNA into each blastomere during early cleavage.

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Effect of Red Ginseng Water Extract on Trypsin Activity (홍삼 물추출물이 Trypsin 활성에 미치는 영향)

  • Lee, Jong-Won;Kim, Na-Mi;Do, Jae-Ho
    • Journal of Ginseng Research
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    • v.28 no.3
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    • pp.127-131
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    • 2004
  • This study was carried out to investigate the effect of red ginseng water extract (RGWE) on trypsin activity. After extraction of fat soluble and saponin component from red ginseng powder by methyl alcohol, the residue was extracted with distilled water, and manufactured to water extract. The extract was dialyzed with different molecular cut off membrane. Trypsin activity demonstrated the highest level at the RGWE concentration of 9${\times}$10$\^$-2/% in reaction mixture, and also increased to 15% at 2.9${\times}$10$\^$-3/%. Km value was decreased and Vmax was increased in the present of red ginseng water extract. Red ginseng water extract was partially purified by dialysis, Bio-Gel P-I0 and DEAE-cellulose column chromatography. The active fraction demonstrated positive reaction to ninhydrin, DNS and folin reaction.

Immunological Studies on the Surface Antigens of Tumor Cells (종양세포 표면항원에 대한 분자면역학적 연구)

  • 김한도;김규원
    • The Korean Journal of Zoology
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    • v.32 no.2
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    • pp.142-152
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    • 1989
  • We have produced a new monoclonal antibody detecting common acute lymphoblastic leukemia antigen (CALLA) and designated as KP-22. CALIA detected by KP-22 is expressed on the all of the various cefl lines examined including common ALL. Burkitt's lymphoma, human fibroblasts and cultured normal human fibroblasts. However out of cell lines tested, a fraction of J-ALL and all of myelocytic leukemia and all other nonleukemia cell lines except for fibroblast are CALIA negative. Immunoprecipitation of solubilized 125 I-labeled membrane proteins from cultured human fibroblasts and leukemia cell lines with KP-22 revealed a major polypeptide chain with an apparent molecular weight of approximately 100 Kd and 95 Kd, respectively. Even though a microheterogeneity in terms of molecular weight between two CALLAs, the peptide mapping patterns of them &e identical indicating that such a microheterogeneity seems to be partly due to heterogeneous terminal sialic acid compositions added by a posttranslational modification process.

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Determination of Respiratory Activity of Mitochondria and Submitochondrial Particles by Using Dropping Mercury Electrode (적하수은전극을 이용한 미토콘드리아 및 Submitochondrial particles의 호흡활성측정)

  • Jung, Jin;Park, Sang-Gyu;Lee, Sang-Kee;Kim, Se-Ho
    • Applied Biological Chemistry
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    • v.28 no.4
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    • pp.271-277
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    • 1985
  • A polarograph with specially designed cell compartment usable in kinetic study of the mitochondrial respiration of a small sized sample was made, and its performance and experimental conditions were examined. An applied potential (ca-1.2V vs. SCE) which gives rise to the second step reduction of oxygen caused a considerable level of a residual current independent of oxygen, which is temporarily interpreted as the reduction current of the membrane-bound redox material(s) of mitochondria. A potential corresponding to the first slop reduction of oxygen (ca-0.4V vs SCE) did not produce the residual current. Thus, it is suggested that a measurement of oxygen concentration in a sample of mitochondria and submitochondrial particles by using dropping mercury electrode should be done with an applied potential of about -0.4V vs SCE. Consumption of oxygen by mitochondria was observed to follow practically zero order kinetics. Its rate constant exhibited the proportional relationship with the respiratory activity of mitochondria. Usefulness of tile instrument was properly demonstrated in the work on the temperature effect on the respiration of mitochondria isolated from several plant 4issues which were selected on the basis of chilling susceptivity.

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Effects of F/M ratio on the EPS production and fouling at MBR (MBR에서 F/M비가 EPS 생성 및 fouling에 미치는 영향)

  • Kim, Yun-Ji;Choi, Yun-Jeong;Hwang, Sun-Jin
    • Journal of Korean Society of Water and Wastewater
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    • v.35 no.3
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    • pp.197-204
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    • 2021
  • In MBR, extracellular polymeric substance (EPS) is known as an important factor of fouling; soluble EPS (sEPS) affects internal contamination of membrane, and bound EPS (bEPS) affects the formation of the cake layer. The production of EPS changes according to the composition of influent, which affects fouling characteristics. Therefore, in this study, the effects of the F/M ratio on the sEPS concentration, bEPS content, and fouling were evaluated. The effects of F/M ratio on the amount and composition of EPS were confirmed by setting conditions that were very low or higher than the general F/M ratio of MBR, and the fouling occurrence characteristics were evaluated by filtration resistance distribution. As a result, it was found that the sEPS increased significantly with the increase of the F/M ratio. When the substrate was depleted, bEPS content decreased because bEPS was hydrolyzed into BAP and seemed to be used as a substrate. In contrast, when the substrate is sufficient, UAP (utilization-associated products) was rapidly generated in proportion with the consumption of the substrate. UAP has a relatively higher Protein/Carbohydrate ratio (P/C ratio) than BAP, and this means, it has a higher adhesive force to the membrane surface. As a result, UAP seems like causing fouling rather than BAP (biomass-associated products). Therefore, Rf (Resistance of internal contamination) increased rapidly with the increase of UAP, and Rc (Resistance of cake layer) increased with the accumulation of bEPS in proportion, and as a result, the fouling interval was shortened. According to this study, a high F/M ratio leads to an increment in UAP generation and accumulation of bEPS, and by these UAP and bEPS, membrane fouling is promoted.

A study on possibility of using CST as a fouling evaluation index at MBR (MBR에서 fouling 평가지표로서 CST의 활용 가능성에 대한 연구)

  • Kim, Yun-Ji;Choi, Yun-Jeong;Sim, Tae-Suk;Hwang, Sun-Jin
    • Journal of Korean Society of Water and Wastewater
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    • v.35 no.6
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    • pp.379-387
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    • 2021
  • In general, trans-membrane pressure (TMP), flux, filtration resistance are used as indicators to evaluate the degree of fouling in MBR. However, they have limitations in determining the level of EPS generation, which is known as an important factor of fouling. Therefore, a new evaluation method is required to monitor the amount of EPS generation. In this study, the applicability of capillary suction time (CST), which is used to measure the dewaterability of sludge, was evaluated as an indirect fouling evaluation index. Statistical analysis was performed to evaluate the effect of EPS on CST, and to determine whether EPS has high similarity with representative fouling evaluation indicators and CST, and quantitatively compared them. As a result, the correlation coefficient between CST and bEPS was 0.7988, which was higher than the correlation coefficient between filtration resistance and bEPS. Since bEPS is a major factor inducing fouling by affecting the formation of the cake layer, it was evaluated that CST, which has a high correlation with bEPS, is suitable to represent EPS. In addition, it was evaluated that the correlation coefficient between filtration resistance and CST was high as 0.7187, which could be used as a fouling evaluation index.

Study on the Safety of Firefly Luciferase in Human as a Transient Reporter Gene of Oncolytic Virotherapy (항암 바이러스 치료제의 보고유전자로써 반딧불이 루시퍼레이즈의 인체 내 안전성에 대한 연구)

  • Hong, Young Mi;Yoon, Woong Hee;Lee, You Ra;Kim, Soo Ji;Ngabire, Daniel;Narayanasamy, Badrinath;Ornella, Mefotse Saha Cyrelle;Kim, Myunghee;Cho, Euna;Lee, Bora;Hwang, Tae-Ho
    • Journal of Life Science
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    • v.31 no.11
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    • pp.1028-1036
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    • 2021
  • Firefly luciferase (FLuc) can function as an efficient marker in the gene and viral therapies. Nonetheless, its clinical translation has been unaccomplished with the concerns on its exogenous nature and the similarity with human fatty acyl-CoA synthetase. In this study, we aimed to show safety of FLuc by conducting a set of preclinical experiments and a human use. Initially, FLuc permeability across the plasma membrane was investigated by delivering the FLuc-carrying viral vector, OTS-412, or the FLuc recombinant protein. After in vitro infection of OTS-412 into different cancer cell lines, FLuc activity was detected only in the cell lysates, but not in culture media. In addition, recombinant FLuc protein further showed the impermeability against the plasma membrane. Similar result was also observed in the in vivo experiment. After being injected into the VX2 tumor-bearing rabbit, the FLuc exclusively resided within the tumor tissue without being detected in the blood plasma or other organs. Human cancer cell lines originated from various organs were lysed and treated to the FLuc, and none of the human substrates was reactive against the FLuc. As a final step, FLuc recombinant protein was intravenously injected into a human. The luciferase was degraded with the half-life of 20 to 30 minutes in blood, and was untraceable from 1.5 hr after the injection. In addition, the blood plasma was nonresponsive against the fatty acids. Hematological analysis was also comparable between the pre- and post-injection. Altogether, our study collectively demonstrates the safety of the firefly luciferase.

Preparations of Universal, Functionalized Long-Chain Alkylthiol Linkers for Self-assembled Monolayers (자기조립단분자막을 위한 보편적이고 기능화된 긴 사슬 알킬티올 연결자의 제조)

  • Yoo, Dong-Jin;Lee, Kyong-Sub;Kim, Ae-Rhan;Nahm, Kee-Suk
    • Korean Chemical Engineering Research
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    • v.49 no.3
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    • pp.330-337
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    • 2011
  • In this research, the preparation processes for making a series of $\omega$-mercapto alkylamine 1 and $\omega$-mercapto alkanoic acid 2 useful for studying of the self-assembled monolayer(SAM) are described. The preparation methods of the first goal materials, $\omega$-mercapto alkylamines 1 were carried out as follows: First, $\omega$-phthalimide alkanol 3 was synthesized from commercially available potassium phthalimide derivatives and $\omega$-bromoalkanol in DMF at $80{^{\circ}C}$ via substitution reaction. After refluxing $\omega$-phthalimide alkanol 3 with hydrazine hydrate in ethanol followed by treating with c-HCl, $\omega$-aminoalkanol 4 was obtained in 76-98% yield, accompanied with side-product 5. Bromination of hydroxyl moiety of $\omega$-aminoalkanol 4 using aqueous hydrobromic acid furnished $\omega$-bromoamine 6 in 34-97% yields. Substitution reaction 6 with thiourea in 95% ethanol gave $\omega$-aminoalkanthiuronium 7, which was treated with aqueous strong base and aqueous strong sulfuric acid gave desired products, $\omega$-mercapto alkylamines 1 through overall 5 steps. The second target material, $\omega$-mercapto alkanoic acid 2 was prepared via 2 steps. $\omega$-bromo alkanoic acid was reacted with thiourea to give $\omega$-thiourea alkanoic acid 7 in 69-85%, which was treated with aqueous strong base and strong acid to furnish $\omega$-mercapto alkanoic acid 2 in 50-98%. The fabricated long-chain alkylthiol(LCAT) can be used as linkers to immobilize protein, enzyme and various kinds of biomolecules on the surface of metallic materials(Au, Pt, Ti) by SAM, and can be useful chemical tools for the application study on the surface modification of metallic materials.

Pervaporation Separation of Ethanol Aqueous Solution through Carbonate-type Polyurethane Membrane II. The Effect of Pendent Anionic Group (카보네이트형 폴리우레탄막을 이용한 에탄올 수용액의 투과증발분리 II. 음이온성기에 의한 영향)

  • Han, In Ki;Oh, Boo Keum;Lee, Young Moo;Noh, Si Tae
    • Applied Chemistry for Engineering
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    • v.3 no.4
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    • pp.595-604
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    • 1992
  • Carbonate-type polyurethane resins containing anionic moieties were systhesized from NCO-terminated prepolymer method. Membranes were manufactured from the polymer solution and the separation of aqueous ethanol solution was investigated. To enhance the property of urethane resin, carbonate-type polyol(PTMCG) was used. ${\alpha}^{\prime},{\alpha}^{{\prime}{\prime}}$-dimethylolpropionic acid was used as a chain extender to increase the hydrophilicily of the urethane membrane. The ionization of the pendent carboxylic groups in urethane resin was carried out using trimthylamine. To confirm the formation of anionic groups in urethane resin, IR spectra of model compounds were compared with those of urethane resins. It was confirmed that the concentration of hard segment and hydrogen bond contributed to the property of the concentration of hard segment and hydrogen bond contributed to the property of urethane resin in which the mole ratio of chain extender and polyol was from 3:1 to urethane resin in which the mole ratio of chain extender and polyol was from 3:1 to 5:1. The carbonate-type polyurethane containing pendent carboxylic grop(PU) had Tg of around-$25^{\circ}C$ and Tm, $45^{\circ}C$ measured by DSC. Transition temperatures of one containing pendent anionic group(APU) prepared from the ionization of PU shifted to $8{\sim}10^{\circ}C$ lower temperature region than those of PU. Pervaporation membrane was prepared through the casting method. N, N-dimethylformamide (DMF) were used as a solvent and hexamethylene diisocyanate(HMDl) as a crosslinking agent. Swelling degree increased with ethanol concentration in mixure and the control of the swelling degree of the membrane could be achieved by crossliking. The results of pervaporation were as follows : separation factor, 2.3~9.8 ; flux, $27{\sim}79.5g/m^2hr$. Pervaporation separation capacity could be enhanced by reducing the molecular weight of polyol from 2,000 to 1,000.

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Comparison of Behavioral Response between Intranasal and Submucosal Midazolam Adminstration (소아 진정 치료 시 구강 점막 하와 비점막 Midazolam 투여의 행동 반응 비교)

  • Kim, Yun-Hee;Jung, Sang-Hyuk;Baek, Kwang-Woo
    • Journal of the korean academy of Pediatric Dentistry
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    • v.35 no.3
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    • pp.427-436
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    • 2008
  • Purpose. The objective of this study was to evaluate the behavioral response and assess the effectiveness of additional intranasal (IN) and submucosal (SM) administration of midazolam during pediatric sedation for dental procedure. Material and methods. Thirty-three cases of healthy (ASAⅠ), uncooperative children aged from 24 to 72 month old at pediatric dental clinic of Ewha Womans University Hospital were selected for this study. Children received oral chloral hydrate 50 mg/kg with hydroxyzine 1.0 mg/kg. After waiting for 45 minutes, midazolam 0.2 mg/kg was administrated via IN route and via SM route randomly maintaining 50% of $N_2O$. A pulse oximeter and a capnograph were used for measuring vital signs ($SpO_2$, PR, RR, $EtCO_2$) throughout the sedation. Behavioral response was evaluated as Quiet (Q), Crying (C), Movement (M) or Struggling (S) in every 2 minutes for 40 minutes. Results. There were also no statistically significant differences in vital signs of the two groups. The behavioral response for the first ten minutes during sedation was a statistically significant difference (P < 0.05) between the two groups. After the first ten minutes, it was revealed that there was no significant difference. Conclusion. This study demonstrated that the addition of IN midazolam to the combination of oral chloral hydrate with hydroxyzine and nitrous oxide/oxygen inhalation is as safe and effective as that of SM midazolam in pediatric sedation for dental procedure.

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