• Journal of Ecology and Environment
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• v.48 no.1
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• pp.32-48
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• 2024

Background: Longitudinal connectivity in river systems strongly affects biological components related to ecosystem functioning, thereby playing an important role in shaping local biodiversity and ecosystem health. Environmental DNA (eDNA)-based metabarcoding has an advantage of enabling to sensitively diagnose the presence/absence of species, becoming an efficient/effective approach for studying the community structure of ecosystems. However, little attention has been paid to eDNA-based biomonitoring for river systems, particularly for assessing the river longitudinal connectivity. In this study, by using eDNA we analyzed and compared species diversity and composition among artificial barriers to assess the longitudinal connectivity of the fish community along down-, mid- and upstream in the Hotancheon from the Geum River basin. Moreover, we investigated temporal variation in eDNA fish community structure and species diversity according to season. Results: The results of species detected between eDNA and conventional surveys revealed higher sensitivity for eDNA and 61% of species (23/38) detected in both methods. The results showed that eDNA-based fish community structure differs from down-, mid- and upstream, and species diversity decreased from down to upstream regardless of season. We found that there was generally higher species diversity at the study sites in spring (a total number of species across the sites [n] = 29) than in autumn (n = 27). Nonmetric multidimensional scaling and heatmap analyses further suggest that there was a tendency for community clusters to form in the down-, mid- and upstream, and seasonal variation in the community structure also existed for the sites. Dominant species in the Hotancheon was Rhynchocypris oxycephalus (26.07%) regardless of season, and subdominant species was Nipponocypris koreanus (16.50%) in spring and Odontobutis platycephala (15.73%) in autumn. Artificial barriers appeared to negatively affect the connectivity of some fish species of high mobility. Conclusions: This study attempts to establish a biological monitoring system by highlighting the versatility and power of eDNA metabarcoding in monitoring native fish community and further evaluating the longitudinal connectivity of river ecosystems. The results of this study suggest that eDNA can be applied to identify fish community structure and species diversity in river systems, although some shortcomings remain still need to be resolved.

• Applied Biological Chemistry
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• v.38 no.1
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• pp.63-66
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• 1995

Soybean nuclear extracts were prepared to detect SEF3(soybean embryo factor 3), which is presumed to be a trans-acting factor for the expression of the soybean ${\beta}-conglycinin\;{\alpha}'$ subunit gene. To increase the specific activity of DNA probe during labeling with $[{\alpha}-^{32}P]$dATP, dATP was added to a final concentration of 1.1 mM during the chase reaction. It results in approximately four-fold increase of specific activity of the DNA probe. Effects of several modifications in preparation of soybean nuclear extracts were examined. It was found that glycerol is effective to stabilize SEF3 during the preparation of nuclear extracts and polyethylenimine could be used to increase the specific activity of SEF3 in nuclear extracts.

• BMB Reports
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• v.35 no.3
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• pp.273-282
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• 2002

Sp3 is a bifunctional transcription factor that has been reported to stimulate or repress the transcription of numerous genes. Although the size of Sp3 mRNA is 4.0kb, the size of the known Sp3 cDNA sequence is 3.6kb. Thus, Sp3 functional studies have been performed with an artificially introduced start codon, and thus an amino-terminus that differs from the wild-type. Ideally, full-length cDNA expression vectors with the appropriate start codon should be utilized for these studies. Using 5'rapid amplification of cDNA ends, a full-length Sp3 cDNA clone was generated and the sequence verified in nine cell lines. No AUG initiation codon was present. However, stop codons were present in all three frames 5' to the known coding sequence. In vitro translation of this full-length cDNA clone produced the expected three isoforms-one at 100 kDa and two in the mid 60 kDa range. Electrophoretic mobility shift assays showed that the protein products had the ability to bind to the Sp1/3 consensus sequence. In vitro studies, using our Sp3 clone and site directed mutagenesis, identified the translation initiation site for the larger isoform as AUA. AUA has not been previously described as an endogenous initiation codon in eukaryotes.

• Journal of Microbiology and Biotechnology
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• v.23 no.1
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• pp.22-28
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• 2013

Xo2276 is a putative transcription activator-like effector (TALE) in Xanthomonas oryzae pv. oryzae (Xoo). Xo2276 was expressed with a TAP-tag at the C-terminus in Xoo cells to enable quantitative analysis of protein expression and secretion. Nearly all TAP-tagged Xo2276 existed in an insoluble form; addition of rice leaf extracts from a Xoosusceptible rice cultivar, Milyang23, significantly stimulated secretion of TAP-tagged Xo2276 into the medium. In a T3SS-defective Xoo mutant strain, secretion of TAPtagged Xo2276 was blocked. Xo2276 is a Xoo ortholog of Xanthomonas campestris pv. vesicatoria (Xcv) AvrBs3 and contains a conserved DNA-binding domain (DBD), which includes 19.5 tandem repeats of 34 amino acids. Xo2276- DBD was expressed in E. coli and purified. Direct in vitro recognition of Xo2276-DBD on a putative target DNA sequence was confirmed using an electrophoretic mobility shift assay. This is the first study measuring the homologous expression and secretion of Xo2276 in vitro using rice leaf extract and its direct in vitro binding to the specific target DNA sequence.

• Tuberculosis and Respiratory Diseases
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• v.45 no.6
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• pp.1178-1187
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• 1998

Background : Recently the incidence of tuberculosis is increasing in many countries and control of the disease is further threatened by the emergence of multi-drug resistant tuberculosis. So rapid detection of drug resistance is very important. Pyrazinamide (PZA) is a first-line chemotherapeutic agent for tuberculosis. Now in Korea, we perform PZase activity test instead of actual pyrazinamide susceptibility test for the detection of PZA resistant M. tuberculosis. Recently the pncA gene, encoding the PZase of M. tuberculosis, was completely sequenced. And it was reported that the mutation of pncA gene would be associated with PZA resistance of M. tuberculosis. Therefore we performed this study to evaluate the possibility for the rapid detection of PZA resistant M. tuberculosis using PCR-SSCP of pncA gene. Method : 44 cultured clinical isolates of M. tuberculosis, BCG Tokyo strain. BCG French strain, and one M. bovis isolate were studied. We used H37Rv as the reference strain, The PZase activity test was done at the reference laboratory of Korean Tuberculosis Institute. DNA was extracted by bead-beater method and 561 bp fragment including pncA gene was amplified by PCR. The PCR product were digested by BstB I enzyme. SSCP was done using MDE gel. Of the 44 strains of M. tuberculosis, 22 strains were PZase-positive and other 22 strains were PZase negative. Results : Of the 22 PZase positive strains, 18 strains(82%) showed the same mobility compared with that of H37Rv and 4(18%) showed different mobility. Of the 22 PZase-negative strains, 19(86%) strains showed the same mobility pattern compared with that of H37Rv and 3(14%) showed different mobility. Naturally PZA-resistant BeG-French strain, BCG-Tokyo strain, and one M. bovis isolate showed the same band pattern each other, but their mobility were different from that of H37Rv. The results of PZase activity test and PCR-SSCP of pncA of M. tuberculosis were statistically significantly correlated each other (p<0.01). Conclusion : The PCR-SSCP after BstB I restriction of pncA gene of M. tuberculosis may be a useful method for the rapid detection of PZA-resistant M. tuberculosis.

• Molecular & Cellular Toxicology
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• v.2 no.3
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• pp.202-211
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• 2006

Our objective is to identify molecular factors which contribute to the increased risk of smoke in human. About 677 workers who had control and experimental groups according to their urinary Naphthol levels were enrolled in our study. In the present study, we investigated the effects of smoking on gene expression profiles in human. We determined differential gene expression patterns in smoker versus non-smoker using cDNA microarray. Specific genes were up-or down-regulated according to smoking and age. Inflammatory related genes such as cytokine, interleukin, and tumor necrosis factor were up-regulated, DNA repair related genes such as high-mobility group (nonhistone chromosomal) protein 1, and protein 2 were down-regulated, apoptosis related genes such as myeloperoxidase and Bcl-2-associated athanogene were down-regulated, and cell cycle related genes were down-regulated. In our epidemiological study, notably, inflammatory, DNA repair, apoptosis, signal transduction, metabolism, cell cycle, cell proliferation, transcription related genes were regulated.

• Bulletin of the Korean Chemical Society
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• v.27 no.4
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• pp.535-538
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• 2006

In our previous studies, we have observed that curcumin and momordin I isolated from Ampelopsis radix inhibit the formation of Fos-Jun-activation protein-1 (AP-1) DNA complex. We have screened more effective compounds which have a 5-membered ring framework like momordin I and have modified disaccharide or carboxylic acid portions in momordin I. We synthesized momordin I derivatives according to the published method with slight modification. Synthetic momordin I derivatives showed remarkable inhibitory activities on Fos-Jun-AP-1 DNA complex formation results in in vitro assays. The $IC_{50}$ values of momordin I derivatives were about 4.0 $\mu$M in an electrophoretic mobility shift assay (EMSA). This value is about 125 times higher than that of curcumin and about 12 times higher than that for curcumin derivative C1, and moreover about 30 times higher than that for momordin I. We found momordin I derivatives (a) and (b) are the strongest inhibitory compound for Fos-Jun-AP-1 DNA complex formation.

• The Journal of Korean Medicine
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• v.21 no.4
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• pp.64-72
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• 2000

Objectives : This study was designed to investigate the neuroprotective effect of Hwansodan(Huanshao-dan) on the apoptosis induced by withdrawal of neurotrophic support. Methods : PCl2 pheochromocytoma cells have been used extensively as a model for studying the cellular and molecular effects of neuronal cells. The viability of cells was measured by MTT assay. We used DNA fragmentation and caspase 3-like protease activation assay. Results : The water extract of Hwansodan(Huanshao-dan) significantly showed protective effects on serum and glucose deprivation-induced apoptotic death. Hwansodan(Huanshao-dan) also prevents DNA fragmentation and caspase 3-like protease activation, representing typical neuronal apoptotic phenomena in PCl2 pheochromocytoma cells and induces tyrosine phosphorylation of proteins around 44 kDa, which was identified as ERK1 with electrophoretic gel mobility shift by Western blot. In addition, MAPK kinase(MEK) inhibitor PD98059 and Ras inactivator, ${\alpha}-hydroxyfarnesylphosphonic$ acid attenuated the neuroprotective effects of Hwansodan(Huanshao-dan) in serum-deprived PCl2 cells. Conclusions : These results indicate that Ras/MEK/ERK signaling pathway plays a key role in neuroprotective effects of Hwansodan(Huanshao-dan) in serum-deprived PCl2 cells. Taken together, we suggest the possibility that Hwansodan(Huanshao-dan) might provide a neurotrophic-like activity in PCl2 cells.

• Journal of Microbiology and Biotechnology
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• v.11 no.2
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• pp.287-293
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• 2001

A fast and easy PCR-SSCP method was developed and assessed for the early detection of rifampin-resistant Mycobacterium leprae in skin biopsy samples from Korean leprosy patients. The 190 bp of the rpoB gene, in which mutation is known to cause resistance to rifampin, was amplified by PCR and then analyzed by SSCP and DNA sequencing, All PCR products showing mobility shift on PCR-SSCP contained mutations, demonstrating that this method can be used for an early diagnositic method to detect a putative rifampin-resistant M. leprae strain. DNA sequence analysis revealed that 19 of 34 patient samples contained M. leprae strains with missense mutations in the rpoB gene: five were the same mutations previously reported to cause rifampin resistance and eight were the new type of mutatios that likely cause rifampin resistance. These newly identified dmutations, whose all five cytosine bases of four amino acids were substitued with thymine, were found at different sites from those reported in Mycobacterium tuberculosis or M. leprae. Therefore, they may provide additional clues to understand the molecular biological basis on the rifampin resistance of M. leprae.

• Journal of Microbiology and Biotechnology
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• v.30 no.1
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• pp.127-135
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• 2020

Small RNAs (sRNAs) are widespread and play major roles in regulation circuits in bacteria. Previously, we have demonstrated that transcription of esrE is under the control of its own promoter. However, the regulatory elements involved in EsrE sRNA expression are still unknown. In this study, we found that different cis-regulatory elements exist in the promoter region of esrE. We then screened and analyzed seven potential corresponding trans-regulatory elements by using pull-down assays based on DNA affinity chromatography. Among these candidate regulators, we investigated the relationship between the ferric uptake regulator (Fur) and the EsrE sRNA. Electrophoresis mobility shift assays (EMSAs) and β-galactosidase activity assays demonstrated that Fur can bind to the promoter region of esrE, and positively regulate EsrE sRNA expression in the presence of Fe²⁺.