• Journal of fish pathology
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• v.37 no.1
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• pp.1-8
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• 2024

The advantages of replicon vectors of RNA viruses include a high ability to stimulate innate immunity and exponential amplification of target mRNA leading to high expression of foreign antigens. The present study aimed to construct a DNA-layered nervous necrosis virus (NNV) replicon vector system in which the capsid protein gene was replaced with a foreign antigen gene and to compare the efficiency of foreign antigen expression between the conventional DNA vaccine vector and the present replicon vector. We presented the first report of a nodavirus DNA replicon-based foreign antigen expression system. Instead of a two-vector system, we devised a one-vector system containing both an NNV RNA-dependent RNA polymerase cassette and a foreign antigen-expressing cassette. This single-vector approach circumvents the issue of low foreign protein expression associated with the low co-transfection efficiency of a two-vector system. Cells transfected with a vector harboring hammerhead ribozyme-fused RNA1 and RNA2 (with the capsid gene ORF replaced with VHSV glycoprotein ORF) exhibited significantly higher transcription of the VHSV glycoprotein gene compared to cells transfected with either a vector without hammerhead ribozyme or a conventional DNA vaccine vector expressing the VHSV glycoprotein. Furthermore, the transcription level of the VHSV glycoprotein in cells transfected with a vector harboring hammerhead ribozyme-fused RNA1 and RNA2 showed a significant increase over time. These results suggest that NNV genome-based DNA replicon vectors have the potential to induce stronger and longer expression of target antigens compared to conventional DNA vaccine vectors.

• Natural Product Sciences
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• v.16 no.4
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• pp.223-227
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• 2010

The DNA Topoisomerase I (DNA Topo I) inhibitory effect of ten isolated compounds (1.10) from the leaf and stem of Vitis amurensis were examined. Among them, amurensin G (5) and r-2-viniferin (7) showed high potent inhibitory activity against DNA Topo I. DNA Topo I, an important target for anticancer drugs, can cause DNA breaks and play a key role during cell proliferation, transcription and repair. Thus, the results suggest that the selected compounds (5 and 7) from Vitis amurensis have a possibility as DNA Topo I-targeting anticancer agents.

• Bulletin of the Korean Chemical Society
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• v.33 no.1
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• pp.221-226
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• 2012

DNA-functionalized silver and silver/gold bimetallic nanoparticle superstructure probes with controllable sizes and optical properties are synthesized using monothiol DNA and dithiothreitol. The superstructures exhibit a very narrow size distribution, which can be easily controlled by balancing the ratio of dithiothreitol and DNA. These superstructures assemble reversibly in a highly cooperative manner, and are SERS active. Multiplexed colorimetric detection of DNA targets using these superstructure probes has been demonstrated to identify three different DNA target sequences that are associated with three lethal diseases, respectively.

• Korean Journal of Microbiology
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• v.26 no.3
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• pp.149-154
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• 1988

Japanese Encephalitis Virus(JEV) can be detected conveniently by the use of biotinylated cDNA probe. To prepare biotinylated probe aminoallyl dUTP was first synthesized chemically to reverse transcribe the virial RNA. The allylamine-labeled cDNA was then converted to the biotin-cDNA by the reaction with an activated biotin ester, NHS-ACA-biotin. The JEV genomic RNA was hybridized to the biotinylated cDNA probe on nitrocellulose filter and visualized colorimetrically by streptavidin complexes with alkaline phosphatase polymer. Sensitivity of the detection system was determined by estimating the amount of the JEV genomic RNA through comparison with signals generated from the biotinylated and $^{32/P}$ -labeled probes. It was found that the biotin probe was as sensitive as $^{32/P}$ -cDNA probe which can detect 50pgs of the target RNA.

• Proceedings of the KIEE Conference
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• 2006.07c
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• pp.1319-1321
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• 2006

This research aims to develop the multiple channel electrochemical DNA chip that has the above characteristic and be able to solve the problems. At first, we fabricated a high integration type DNA chip array by lithography technology. It is able to detect a plural genes electrochemically after immobilization of a plural probe DNA and hybridization of non-labeling target DNA on the electrodes simultaneously. It suggested that this DNA chip could recognize the sequence specific genes. It suggested that multichannel electrochemical DNA microarray is useful to develop a portable device for clinical gene diagnostic system.

• The Transactions of the Korean Institute of Electrical Engineers C
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• v.51 no.2
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• pp.92-97
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• 2002

One of the important roles of a DNA chip is the capability of detecting genetic diseases and mutations by analyzing DNA sequence. For a successful electrochemical genotyping, several aspects should be considered including the chemical treatment of electrode surface, DNA immobilization on electrode, hybridization, choice of an intercalator to be selectively bound to double standee DNA, and an equipment for detecting and analyzing the output signal. Au was used as the electrode material, 2-mercaptoethanol was used for linking DNA to Au electrode, and methylene blue was used as an indicator that can be bound to a double stranded DNA selectively. From the analysis of reductive current of this indicator that was bound to a double stranded DNA on an electrode, a normal double stranded DNA was able to be distinguished from a single stranded DNA in just a few seconds. Also, it was found that the peak reduction current of indicator is proportional to the concentration of target DNA to be hybridized with probe DNA. Therefore, it is possible to realize a sim71e and cheats DNA sensor using the electrochemical measurement for genotyping.

• Transactions of the Korean Society of Mechanical Engineers A
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• v.28 no.8 s.227
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• pp.1135-1139
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• 2004

This paper presents a microextractor for the separation of DNA molecules by their sizes. The DNA microextractor immobilizes the DNA molecules of specific size in the micropillar array by adjusting the period of the crossed electric field, thus providing a starting-point independent target DNA extraction method without separation process monitoring. The DNA microextractor has been fabricated by a three-mask micromachining process. The velocity of three different DNA molecules has been measured at the electric field of E=5V/0.8cm in the fabricated DNA microextractor, resulting in the reorientation times of $4.80{\pm}0.44sec,\;7.12{\pm}0.75sec$, and $9.88{\pm}0.30sec$ for ${\lambda}$ DNA, micrococcus DNA, and T4 DNA, respectively. T4 DNA is trapped in the micropillar array when the crossed electric field of 5V/0.8cm is applied alternately at a 10 second time interval. The present DNA microextractor filters the DNA in a specific size range by adjusting the magnitude and/or the period of the crossed electric field applied in the micropillar array.

• Korean Journal of Veterinary Service
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• v.35 no.3
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• pp.215-222
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• 2012

Species-specific $TaqMan^{(R)}$ probe-based real-time PCR assays were developed for detection of beef, pork, chicken, duck, goose and turkey. The primer and probe sets used in this study were designed to be complementary to fibroblast growth factor (FGF) for cattle and pig, mitochondrial NADH dehydrogenase (ND) subunit 3 and ND2 for chicken and duck, 12S rRNA for goose and turkey, respectively. As internal positive control we used conserved region in the ribosomal 18S RNA gene to ensure the accuracy of the detection of target DNA by real-time PCR. We confirmed that real-time PCR assays with the primer and probe sets were positive for cattle, pig and chicken intended target animal species with no cross-reactivity with other non-target animal species. Only >50 ng DNA of beef show cross-reactivity in the determination of duck. Using species-specific primer and probe sets, it was possible to detect amounts of 0.1 ng DNA of cattle and pig, 1.0 pg DNA of chicken, duck and turkey, and 0.1 pg DNA of goose for raw samples, respectively. The detection limits were 0.1 ng DNA of cattle, 1.0 ng DNA of pig and 1.0 pg DNA of chicken for DNA mixtures (beef, pork and chicken) extracted from heat-treated ($121^{\circ}C$/5 min) meat samples. In conclusion, it can be suggested that the $TaqMan^{(R)}$ probe-based assay developed in this study might be a rapid and specific method for the identification of meat species in raw or cooked meat products.

• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.3
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• pp.225-229
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• 2005

To investigate impediments to plasmid transformation in Brevibacterium flavum BF4 and B. lactofermentum BL1, cell surface barriers were determined by measuring growth inhibition whilst enzymatic barriers were determined by comparing DNA methylation properties. B. lactofermentum was more sensitive to growth inhibition by glycine than B. flavum. Release of cellular proteins during sonication was more rapid for B. lactofermentum than for B. flavum. Plasmid DNA (pCSL 17) isolated from B. flavum transformed recipient $McrBC^+$ strains of Escherichia coli with lower efficiency than $McrBC^-$. McrBC digestion of this DNA confirmed that B. flavum contain methylated cytidines in the target sequence of McrBc sequences but B. lactofermentum contained a different methylation pattern. DNA derived from the B. lactofermentum transformed recipient $EcoKR^+$ strains of E. coli with lower efficiency than $EcoKR^-$, indicating the presence of methylated adenosines in the target sequence of EcoK sequences. The present data describe the differences in the physical and enzymatic barriers between two species of corynebacteria and also provide some insight into the successful foreign gene expression in corynebacteria.

• Biomedical Science Letters
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• v.10 no.2
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• pp.65-73
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• 2004

I report that single-stranded antisense as a part of large circular (LC-) genomic DNA of recombinant M13 phage exhibits enhanced stability, sequence specific antisense activity, and no need for target site search. A cDNA fragment (708 bp) of rat TNF-$\alpha$ was inserted into a phagemid vector, and TNF-$\alpha$ antisense molecules (TNF$\alpha$-LCAS) were produced as single-stranded circular DNA. When introduced into a rat monocyte/macrophage cell line, WRT7/P2, TNF$\alpha$-LCAS was able to ablate LPS-induced TNF-$\alpha$ mRNA to completion. The antisense effect of TNF$\alpha$-LCAS was shown to be sequence-specific because expressions of three control genes ($\beta$-actin, GAPDH and IL-1$\beta$) were not significantly altered by the antisense treatment. Further, TNF$\alpha$-LCAS was found to be highly efficacious as only 0.1 $\mu$g (0.24 nM) of TNF$\alpha$-LCAS was sufficient to block TNF-$\alpha$ expression in 1$\times10^5$ WRT7/P2 cells. I have also observed specific antisense activity in reduction of NF-$\kappa$B gene expression. The results suggest that an antisense sequence as a part of single-stranded circular genomic DNA has a specific antisense activity.