• Journal of Life Science
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• v.28 no.11
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• pp.1277-1284
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• 2018

In this study, a repeated yeast integrative plasmid (R-YIp) harboring Cre/loxP system was constructed to integrate various gene expression cassettes into the yeast chromosome. The R-YIp system contains a reusable selective marker (CgTRP1), loxP sequence, and target sequence for integration. Therefore, many gene expression cassettes can be integrated into the same position of the same yeast chromosome. In the present study, several model enzymes involving xylan/xylose metabolism were examined, including endoxylanase (XYLP), ${\beta}$-xylosidase (XYLB), xylose reductase (GRE3) and xylitol dehydrogenase (XYL2). Efficient expression of these genes was obtained using two promoters (GAL10p and ADH1p) and various plasmids (pGMF-GENE and pAMF-GENE plasmids) were constructed. The XYLP, XYLB, GRE3, and XYL2 genes were efficiently expressed under the control of the GAL10 promoter. Subsequently, R-YIps containing the GAL10p-GENE-GAL7t cassette were constructed, resulting in pRS-XylP, pRS-XylB, pRS-Gre3, and pRS-Xyl2 plasmids. These plasmids were sequentially integrated into chromosome VII of a Saccharomyces cerevisiae strain by repeated gene integration and selective marker rescue. These genes were integrated by the R-YIp system and were stably expressed in the yeast transformants to produce active recombinant enzymes. Therefore, we expect that the R-YIp system will be able to overcome current limitations of the host cells and allow selective marker selection for the integration of various genes into the yeast chromosome.

• Journal of Chest Surgery
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• v.41 no.1
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• pp.12-24
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• 2008

Background: It is currently thought that tissue valve degeneration is related to an animal's immune response, which is mainly due to cell surface ${\alpha}$-Gal epitopes. Cell surface ${\alpha}$-Gal epitopes are known to be degraded by the enzyme called green coffee bean ${\alpha}$-Galactosidase. It is also well known that ${\alpha}$-Gal epitopes are immunologically stained by Griffonia Simplicifolia isolectin type B4. We know that many commercially available tissue valves are made of aortic valves and pericardial tissue of pig. So, we investigated whether ${\alpha}$-Gal epitopes of the aortic valve and pericardial tissue of a pig can be removed by green coffee bean ${\alpha}$-Galactosidase, and we did so by comparing immunologic staining of the tissues before and after the enzyme treatment. Material and method: After treating fresh porcine aortic valve and pericardial tissue with green coffee bean ${\alpha}$-Galactosidase at concentrations of 0.5 unit/mL, 1.0 unit/mL, 2.0 unit/mL, respectively, under the condition of pH 6.5, temperature. $4^{\circ}C$ and 24 hours of incubation, each sample was stained with Griffonia Simplicifolia isolectin type B4 immunpfluorescent labeling. We then examined whether the ${\alpha}$-Gal epitopes were reduced or abolished in each consecutive. concentration of green coffee bean ${\alpha}$-Galactosidase by comparing the degree of the Griffonia Simplicifolia isolectin B4 staining in each sample. Result: In the pig aortic valve tissue, a 1.0 unit/mL concentration of green coffee bean ${\alpha}$-Galactosidase at pH 6.5, $4^{\circ}C$ and reaction for 24 hours was enough for complete removal of ${\alpha}$-Gal epitopes from the cell sur face on the immunostaining with Griffonia Simplicifolia isolectin B4. On the other hand, more ${\alpha}$-Gal epitopes were present in the pig pericardial tissue on Griffonia Simplicifolia isolectin B4 staining before the enzyme treatment, and 1.0 unit/mL of galactosidase was not sufficient for complete removal of ${\alpha}$-Gal from the tissue. 2.0 units/mL of green coffee bean ${\alpha}$-Galactosidase was needed to completely remove the ${\alpha}$-Gal epitopes from the pericardial tissue on immunostaining. Conclusion: The ${\alpha}$-Gal epitopes of the pig's aortic valve and pericardial tissue were successfully stained with Griffonia Simplicifolia isolectin B4. We could remove nearly all the ${\alpha}$-Gal epitopes using green coffee bean ${\alpha}$-Galactosidase at the concentration of 1.0 unit/mL in the aortic valve. Of pig, and 2.0 unit/mL was need to nearly completely remove all the ${\alpha}$-Gal epitopes in the pericardial tissue of pig under the condition of pH 6.5, $4^{\circ}C$ and 24 hours of reaction time. In the near future, removal of ${\alpha}$-Gal epitapes in the pig's aortic valve and pericardial tissue will become a powerful tool for the improvement of the tissue valve durability. It needs to be determined if ${\alpha}$-galactosidase treated pig tissue is immune to human anti-Gal antibody or anit-Gal mooclonal antibodies.

• Journal of the Korean Society of Surveying, Geodesy, Photogrammetry and Cartography
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• v.13 no.2
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• pp.177-185
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• 1995

Gravity anomalies were recovered on a $5'\times{5'}$grid using the sea surface height data obtained from the combination of Geosat, ERS-1, Topex/Poseidon altimeter data around Korean Peninsula bounded by latitude between $30^\circ{N}\;and\;50^\circ{N}$ and longitude $120^\circ{E}\;to\;140^\circ{E.}$ In order to recover the gravity anomalies from SSH(Sea Surface Height), inverse FFT technique was applied. The estimated gravity anomalies were compared with gravity anomalies measured by shipboard around Korean Peninsula. In comparison with the differences of gravity anomaly between measured data and altimeter data, the mean and the standard deviation were found to be -0.51 mGal and 13.48 mGal, respectively. In case of comparison between the measured data and the OSU91A geopotential model, the mean and the standard deviation were found to be 11.93 mGal and 19.19 mGal, respectively. The comparison of gravity anomalies obtained from the OSU91A geopotential model and the altimeter data was carried out. The results were mean of 5.30 meal and standard deviation of 19.62 mGal. From the results, we could be concluded that the gravity anomalies computed from the altimeter data is used to the geoid computation instead of the measured data.

• Journal of Korean Medicine Rehabilitation
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• v.34 no.2
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• pp.15-28
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• 2024

Objectives We used the D-galactose (D-gal) induced C2C12 myoblast senescence model to investigate whether ethanol extract of Perilla. fructescens leaves (EEPF) could delay cellular senescence and regulate related mechanisms. Methods C2C12 myogenic cells were cultured in an incubator under 37 ℃ and 5% CO₂ conditions. EEPF, dried perilla leaves were pulverized and extracted at 1:10 (v/v) at 50 ℃ for 4 hours. Cell counting kit-8 and western blot analysis was performed. Annexin V-FITC apoptosis detection kit and DAPI staining was applied. Catalase (CAT), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and malondialdehyde analysis kits were used. To measure the level of reactive oxygen species generation, staining and flow cytometry was used. To analyze the mitochondrial activity, membrane potential changes were measured using JC-1. 𝛽-gal activity was analyzed using SA-𝛽-gal staining solution, and DNA damage was analyzed by using 𝛾-H2AX. Quantikine ELISA kit was used to analyze inflammatory cytokine production. Results According to the results of this study, EEPF significantly alleviated the decrease in cell viability in C2C12 cells treated with D-gal and suppressed the decrease in the expression of proliferating cell nuclear antigen. EEPF also markedly blocked D-gal-induced C2C12 cell apoptosis and restored reduced activity of CAT, GSH-Px, T-AOC, SOD. In addition, EEPF suppressed the decrease in 𝛽-galactosidase activity, the induction of DNA damage and the increase in expression of senescence-associated secretory phenotype proteins such as p16, p53 and p21 in D-gal-treated C2C12 cells. Furthermore, EEPF significantly attenuated D-gal-induced production and expression of inflammatory cytokines such as interleukin (IL)-6 and IL-18. Conclusions The results of this study indicate that EEPF can be used as a potential candidate for the prevention and treatment of muscle aging.

• Journal of the Korean Geosynthetics Society
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• v.18 no.4
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• pp.107-114
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• 2019

In this paper, the P-wave multiple detection system for the fast and accurate earthquake early warning nearby the epicenter was developed. The developed systems were installed in five selected public buildings for the validation. During the monitoring, a magnitude 2.3 earthquake occurred in Pohang on 26 September 2019. P-wave initial detection algorithms were operated in three out of four systems installed in Pohang area and recorded as seismic events. At the nearest station, 5.5 km from the epicenter, P-wave signal was detected 1.2 seconds after the earthquake, and S-wave was reached 1.02 seconds after the P-wave reached, providing some alarm time. The maximum accelerations recorded in three different stations were 6.28 gal, 6.1 gal, and 5.3 gal, respectively. The alarm algorithm did not work, due to the high threshold of the maximum ground acceleration (25.1 gal) to operate it. If continuous monitoring and analysis are to be carried out in the future, the developed system could use a highly effective earthquake warning system suitable for the domestic situation.

• Reproductive and Developmental Biology
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• v.35 no.3
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• pp.385-390
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• 2011

The purpose of this study was undertaken to evaluate of cryopreservation efficiency in ${\alpha}$ 1,3-galactosyltransferase knock-out(GalT KO) cloned miniature pig sperm. To compare ability of frozen-thawed sperm characteristics, three different pig strains (GalT KO) cloned miniature pig, PWG miniature pig and Duroc were used. The ejaculated semen from the three pig species was diluted with same volume extender and added to LEY solution for freezing. The diluted semen was placed in 0.5 ml straws, and freezing was initiated by exposing the straws to liquid nitrogen ($LN_2$) vapours for 10 min before placing them into $LN_2$ for cryopreservation. After thawing, the sperm ability were assessed for viability (SYBR-14/PI staining), abnormality (Rose Bengal staining), and acrosome status (intactness, intensity and capacitation) (chlorotetracycline, CTC staining). The viability of frozen-thawed GalT KO pig sperm had no significant difference as compared with Duroc and PWG miniature pig sperm. However, The CTC pattern of frozen-thawed GalT KO cloned miniature pig spermatozoa showed significantly lower rates in F pattern and AR pattern (p<0.05) and significantly higher rates in B pattern than Duroc and PWG miniature pig (p<0.05). The abnormality of GalT KO cloned miniature pig sperm was significantly lower as compared to Duroc and PWG miniature pig sperm (p<0.05). In conclusion, GalT KO cloned miniature pig semen can be cryopreserved successfully and used for artificial insemination reasonably.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.23
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• pp.10217-10223
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• 2015

Hyperactivated ${\alpha}2$-6-sialylation on N-glycans due to overexpression of the Golgi enzyme ${\beta}$-galactoside: ${\alpha}2$-6-sialyltransferase (ST6Gal-I) often correlates with cancer progression, metastasis, and poor prognosis. This study was aimed to determine the association between ST6Gal-I expression and the risk of recurrence and survival of patients with localized clear-cell renal cell carcinoma (ccRCC) following surgery. We retrospectively enrolled 391 patients (265 in training cohort and 126 in validation cohort) with localized ccRCC underwent nephrectomy at a single center. Tissue microarrays were constructed for immunostaining of ST6Gal-I. Prognostic value and clinical outcomes were evaluated. High ST6Gal-I expression was associated with Fuhrman grade (p<0.001 and p=0.016, respectively) and the University of California Los-Angeles Integrated Staging System (UISS) score (p=0.004 and p=0.017, respectively) in both cohorts. Patients with high ST6Gal-I expression had significantly worse overall survival (OS) (p<0.001 and p<0.001, respectively) and recurrence free survival (RFS) (p<0.001 and p=0.002, respectively) than those with low expression in both cohorts. On multivariate analysis, ST6Gal-I expression remained associated with OS and RFS even after adjusting for the UISS score. Stratified analysis suggested that the association is more pronounced among patients with low and intermediate-risk disease defined by the UISS score. High ST6Gal-I expression is a potential independent adverse predictor of survival and recurrence in ccRCC patients, and the prognostic value is most prominent in those with low and intermediate-risk disease defined by the UISS score.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.3
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• pp.215-218
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• 2000

Sialytrisaccharides based on $\beta$-galactosyldisaccharides were synthesized using $\beta$-galactosidase and trans-sialidase in one pot. Using $\beta$-galactosidase from Bacillus Ciculans and trans-sialidase from Trypanosoma cruzi simulaneously, 6mM sialyltrisaccharides composed of about 95% NeuAc$\alpha$(2,3)Gal$\beta$(1,4)GlcNAc and 5% NeuAc$\alpha$(2,3)Gal$\beta$(1,6)GlcNAc were produced from a reaction mixture containing 25mM o-nitropheny1-$\beta$-D-galsctolneuraminic acid. One beauty of this reaction was that a secondary hydrolysis of the disaccharide intermediate occurring between the activated galactopyranoside and N-acetylgucosamine was prevented. Using $\beta$-galactosidase from Escherichia cloi and the same trans-sialidase, 15mM sialyltrisaccharides composed of about 90% NeuAc$\alpha$(2,3)Gal$\beta$(1,6)GlcNac and 10% NeuAc$\alpha$(2,3)Gal$\beta$(1,4)GlcNAc were produced from a reaction misture containing 400nM galactose, 800nM N-acetylglucosylation rection between galactose and N-actylgucosamine was diminant since the disaccharide intermediate mainly resulted sreulted in the silylated product.

• Reproductive and Developmental Biology
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• v.39 no.3
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• pp.59-67
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• 2015

Galactose-${\alpha}1,3$-galactose (${\alpha}1,3$-Gal) epitope is synthesized at a high concentration on the surface of pig cells by ${\alpha}1,3$-galactosyltransferase gene (GGTA1). The ${\alpha}1,3$-Gal is responsible for hyperacute rejection in pig-to-human xenotransplantation. The generation of transgenic pigs as organ donors for humans is necessary to eliminate the GGTA1 gene that synthesize $Gal{\alpha}$(1,3)Gal. To prevent hyperacute graft rejection in pig-to-human xenotransplantation, previously, we developed ${\alpha}1,3$-galactosyltransferase gene-knock-out somatic cell by homologous recombination. In this study, we established cell lines of ${\alpha}1,3$-GT knock-out expressing hDAF and hHT gene from minipig fibroblasts to apply somatic cell nuclear transfer. The hDAF and hHT mRNA were expressed in the knock-in somatic cells and ${\alpha}1,3$-GT mRNA was suppressed. However, the knock-in somatic cells were increased resistance to human serum-mediated cytolysis.

• Journal of Acupuncture Research
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• v.17 no.2
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• pp.95-117
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• 2000

The purpose of this morphological studies was to investigate the relation between the meridian, acupoints and viscera using neuroanatomical tracers. The common locations of the spinal ganglia, sympathetic chain ganglia, spinal cord and brain projecting to the large intestine meridian were observed following injection of transganglionic tracer, WGA-HRP and transsynaptic neurotropic virus, pseudorabies virus(PRV), Bartha strain(Ba) and PRV-Ba-Gal (Galactosidase)) into the the large intestine(cecum, colon and rectum), ST37 and LI4. After survival times of 96 hours following injection into the thirty rats with WGA-HRP, PRV-Ba and PRV-Ba-Gal. They were perfused, and their spinal ganglia, sympathetic chain ganglia, spinal cord and brain were frozen sectioned($30{\mu}m$). These sections were stained by HRP and X-gal histochemical and PRV immunohistochemical staining method, and observed with a light microscope. The results were as follows : 1. WGA-HRP labeled neurons innervating the large intestine were observed bilaterally within the T13-L4 sympathetic chain ganglia, and T9-11 spinal ganglia. WGA-HRP labeled neurons innervating ST37 were observed within the L3-5 sympathetic chain ganglia, and L2-4 spinal ganglia. WGA-HRP labeled neurons innervating LI4 were observed in the middle cervical ganglion and stellate ganglion, and C5-8 spinal ganglia. 2. In spinal cord, PRV-Ba labeled neurons projecting to the large intestine, ST37 and LI4 were found in thoracic, lumbar and sacral spinal segments. Densely labeled areas of each spinal cord segment were founded in lamina N, V, VII(intermediolateral nucleus), Ⅸ, X and dorsal nucleus. 3. In medulla oblongata, PRV-Ba and PRV-Ba-Gal labeled neurons projecting to the large intestine, ST37 and LI4 were commonly found in the A1 noradrenalin cells/C1 adrenalin cells/caudoventrolateral reticular nucleus, dorsal motor nucleus of vagus nerve, nucleus tractus solitarius, raphe obscurus nucleus, raphe pallidus nucleus, raphe magnus nucleus and gigantocellular nucleus. 4. In pons, PRV-Ba and PRV-Ba-Gal labeled neurons were commonly found in locus coeruleus, Kolliker-Fuse nucieus and A5 cell group. 5. In midbrain, PRV-Ba and PRV-Ba-Gal labeled neurons were commonly found in central gray matter. 6. In diencephalon, PRV-Ba and PRV-Ba-Gal labeled neurons were commonly found in paraventricular hypothalamic nucleus. These results suggest that PRV-Ba and PRV-Ba-Gal labeled common areas projecting to the large intestine may be correlated to that of the large intestine meridian, ST37 and LI4. Especially, These morphological results provide that interrelationship of meridian-acupoints -viscera may be related to the central autonomic pathways.