• Korean Journal of Microbiology
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• v.44 no.3
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• pp.212-220
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• 2008

A Pn10 DNA probe was introduced as a Prevotella nigrescens ATCC $33563^T$-specific DNA probe. In that study, the specificity of the Pn10 was tested with only type or reference strains of 5 oral bacterial species. The purpose of this study is to evaluate the specificity of the Pn10 using the wild type strains of P. nigrescens and is to develop the P. nigrescens ATCC $33563^T$-specific PCR primers based on the nucleotide sequence of the Pn10. The specificity of the Pn10 DNA probe was determined by Southern blot analysis. The nucleotide sequence of Pn10 DNA probes was determined by chain termination method. The PCR primers were designed based on the nucleotide sequence of cloned DNA fragment. The data showed that Pn10 DNA probe were hybridized with the genomic DNAs from P. nigrescens ATCC $33563^T$ and KB6. The Pn10 homologous region, KB6-Pn10, of P. nigrescens KB6 was cloned by PCR and sequenced. The Pn10 and KB6-Pn10 DNA fragments were consisted of 1,875 bp and 1,873 bp, respectively. The percent identity of the two was 98.8% and the divergence of them was 0.6%. The two primer sets (Pn10-F-AC/ Pn10-R-AC and Pn10-F-A/ Pn10-R-A), designed base on the nucleotide sequences of Pn10 DNA probe, were specific to the P. nigrescens ATCC $33563^T$. The two PCR primer sets could detect as little as 4 pg of genomic DNA of P. nigrescens ATCC $33563^T$. These results indicate that the two PCR primer sets have proven useful for the identification of P. nigrescens ATCC $33563^T$, especially with regard to the maintenance of the strain.

• Korean Journal of Microbiology
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• v.44 no.4
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• pp.346-351
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• 2008

This study develops DNA array which can detect specific sequence of human papilomavirus (HPV) by using lateral flow membrane assay which is usually used for point of care test including pregnant diagnosis. Principle of HPV DNA array is as follow; fixing DNA probe which is peculiar to HPV type 6, 11, 16, 18, 31, 45 on a surface of lateral flow membrane and inducing hybridization response between probe and HPV PCR products which is obtained by using biotin-labeled MY09/l1 primers. And then, we can see the result of DNA hybridization that streptavidin labelled colloidal gold is responded with hybrid biotin. Lateral flow membrane array developed in this study confirms major HPV type economically and conveniently compared with existing HPV DNA chip method.

• Korean Journal of Microbiology
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• v.43 no.3
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• pp.179-185
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• 2007

In this study, we used the method of guanidin isothiocyanate and boiling with Chelex-100 resin to extract genomic DNA of Ralstonia solanacearum from soil. It is more efficient than general protocols to remove inhibitory compounds in soil and R. solanacearum on. Then, we applied polymerase chain reaction and DNA enzyme-linked immunosorbent assay (ELISA) to identify and detect pathogen. The fliC gene of R. solanacearum was selected for specific detection of pathogen and primer sets were designed. Among the primer sets, two specific and sensitive primer sets, RsolfliC(forward: 5-GAACGCCAACGGTGCGAACT-3 and reverse; 5-GGCGGCCTTCAGGGAGGTC-3, designed by J. $Sch\ddot{o}nfeld$ et al.) and RS_247 (forward: 5-GGCGGTCTGTCGGCRG-3 and reverse; 5-CGGTCGCGTTGGCAAC-3 designed by this study), were designed to perform nested PCR. Nested PCR primer was labeled with biotin for hybridization between nested PCR product and probe to analyze with DNA ELISA.

• Journal of KIISE:Software and Applications
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• v.34 no.12
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• pp.1056-1064
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• 2007

Biomolecular computing is a new computing paradigm that uses biomolecules such as DNA for information representation and processing. The huge number of molecules in a small volume and the innate massive parallelism inspired a novel computation method, and various computation models and molecular algorithms were developed for problem solving. In the meantime, the use of biomolecules for information processing supports the possibility of DNA computing as an application for biological problems. It has the potential as an analysis tool for biochemical information such as gene expression patterns. In this context, a DNA computing-based model of a biomolecular perceptron has been proposed and the result of its experimental implementation was presented previously. The weight encoding and weighted sum operation, which are the main components of a biomolecular perceptron, are based on the competitive hybridization reactions between the input molecules and weight-encoding probe molecules. However, thermodynamic symmetry in the competitive hybridizations is assumed, so there can be some error in the weight representation depending on the probe species in use. Here we suggest a generalized model of hybridization reactions considering the asymmetric thermodynamics in competitive hybridizations and present a weight encoding method for the reliable implementation of a biomolecular perceptron based on this model. We compare the accuracy of our weight encoding method with that of the previous one via computer simulations and present the condition of probe composition to satisfy the error limit.

• Journal of Crop Science and Biotechnology
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• v.11 no.2
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• pp.91-100
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• 2008

Melting curve analysis of fluorescently labeled DNA fragments is used extensively for genotyping single nucleotide polymorphism(SNP). Here, we evaluated a SNP genotyping method by melting curve analysis with the two probe chemistries in a 384-well plate format on a Roche LightCycler 480. The HybProbe chemistry is based on the fluorescence resonance energy transfer(FRET) and the SimpleProbe chemistry uses a terminal self-quenching fluorophore. We evaluated FRET HybProbes and SimpleProbes for two SNP sites closely linked to two quantitative trait loci(QTL) for southern root-knot nematode resistance. These probes were used to genotype the two parents and 94 $F_2$ plants from the cross of PI 96354$\times$Bossier. The SNP genotypes of all samples determined by the LightCycler software agreed with previously determined SSR genotypes and the SNP genotypes determined on a Luminex 100 flow cytometry instrument. Multiplexed HybProbes for the two SNPs showed a 98.4% success rate and 100% concordance between repeats two of the same 96 DNA samples. Also, we developed a HybProbe assay for the Rcs3 gene conditioning broad resistance to the frogeye leaf spot(FLS) disease. The LightCycler 480 provides rapid PCR on 384-well plate and allows simultaneous amplification and analysis in approximately 2 hours without any additional steps after amplification. This allowed for a reduction of the potential contamination of PCR products, simplicity, and enablement of a streamlined workflow. The melting curve analysis on the LightCycler 480 provided high-throughput and rapid SNP genotyping and appears highly effective for marker-assisted selection in soybean.

• The Korean Journal of Malacology
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• v.27 no.4
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• pp.387-393
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• 2011

The objective of this study was to develop a real-time PCR method for the rapid detection and quantification of the protozoan pathogen Perkinsus olseni using a TaqMan probe. For the standard, genomic DNA was extracted from $10^5$ in vitro-cultured P. olseni trophozoites, and then 10-fold serial dilutions to the level of a single cell were prepared. To test the reliability of the technique, triplicates of genomic DNA were extracted from $5{\times}10^4$ cells and 10-fold serial dilutions to the level of 5 cells were prepared. The standards and samples were analyzed in duplicate using an $Exicycler^{TM}$ 96 real-time quantitative thermal block. For quantification, the threshold cycle ($C_T$) values of samples were compared with those obtained from standard dilutions. There was a strong linear relationship between the $C_T$ value and the log concentration of cells in the standard ($r^2$ = 0.996). Detection of DNA at a concentration as low as the equivalent of a single cell showed that the assay was sensitive enough to detect a single cell of P. olseni. The estimated number of P. olseni cells was similar to the original cell concentrations, indicating the reliability of P. olseni quantification by real-time PCR. Accordingly, the designed primers and probe may be used for the rapid detection and quantification of P. olseni from clam tissue, environmental water, and sediment samples.

• Proceedings of the Korean Society for Noise and Vibration Engineering Conference
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• 2014.10a
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• pp.358-360
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• 2014

For the several decades, various nanomaterials are broadly used in industry and research. With the growth of nanotechnology, the study of nanotoxicity is being accelerated. Particularly, mercury ion is widely used in real life. Because the mercury is representative high toxic material, it is highly recommended to detect the mercury ion. In previous reported work, thymine-thymine mismatches (T-T) capture mercury ion and create very stable base pair ($T-Hg^{2+}-T$). Here, we performed the high sensitive sensing method for direct label free detection of mercury ions and DNA binding using Kelvin Probe Force Microscope (KPFM). In this method, 30 base pairs of thymine (T-30) is used for mercury specific DNA binding ($T-Hg^{2+}-T$). KPFM is able to detect the mercury ion because there is difference between bare T-30 DNA and mercury mediated DNA ($T-Hg^{2+}-T$).

• Proceedings of the Korean Vacuum Society Conference
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• 2013.08a
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• pp.92.1-92.1
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• 2013

Molecular electronics has been the subject of intese research for many years because of the fundamental interest in molecular charge transport and potential applications, such as (bio)nanosensors and molecular memory devices. Molecular electronics requires a method for making reliable eletrical contacts to singlemolecules. To date, several approaches have been reported: scanning-probe microscopy, mechanical break junctions, nano patterning, and direct deposition of electrode on a self-assembled monolayers. However, most methods are laborious and difficult for large-scale application and more importantly, cannot control the number of moleucles in the junction. Recently, DNA has been used as a template for metallic nanostructures (e.g., Ag, Pd, and Au nanowires) through DNA metallization process. Furthermore, oligodeoxynucleotides have been tethered to organic molecules by using conventional organic reactions. Collectively, these techniques should provide an efficient route toward reliable and reproducible molecular electronic devices with large-scale fabrication. Therefore, I will present a paradigm for the fabrication of moleuclar electronic devices by using micrometer-sized DNA-singe organic molecule and DNA triblock structures.

• The Transactions of the Korean Institute of Electrical Engineers C
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• v.51 no.6
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• pp.265-270
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• 2002

The determination of DNA hybridization reaction can apply the molecular biology research, clinic diagnostics, bioengineering, environment monitoring, food science and application area. So, the improvement of DNA hybridization detection method is very important for the determination of this hybridization reaction. Several molecular biological techniques require accurate predictions of matched versus mismatched hybridization thermodynamics, such as PCR, sequencing by hybridization, gene diagnostics and antisense oligonucleotide probes. In addition, recent developments of oligonucleotide chip arrays as means for biochemical assays and DNA sequencing requires accurate knowledge of hybridization thermodynamics and population ratios at matched and mismatched target sites. In this study, we report the characteristics of the probe and matched, mismatched target oligonucleotide hybridization reaction using thermodynamic method. Thermodynamic of 5 oligonucleotides with central and terminal mismatch sequences were obtained by measured UV-absorbance as a function of temperature. The data show that the nearest-neighbor base-pair model is adequate for predicting thermodynamics of oligonucleotides with average deviations for $\Delta$H$^{0}$ , $\Delta$S$^{0}$ , $\Delta$G$_{37}$ $^{0}$ and T$_{m}$, respectively.>$^{0}$ and T$_{m}$, respectively.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.130-130
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• 2003

Gummy stem blight pathogen is very difficult not only to monitor the inoculum levels prior to host infection, and also it is destructive and hard to control in field condition. We have applied RAPD technique to elucidate the genetic diversity of the genomic DNA of Didymella bryoniae and also to generate specific diagnostic DNA probe useful for identification and detection. The 40 primers produced clear bands consistently from the genomic DNA of twenty isolates of Didymella bryoniae, and two hundred seventy-three amplified fragments were produced with 40 primers. The combined data from 273 bands was analyzed by a cluster analysis using UPGMA method with an arithmetic average program of NTSYS-PC (Version 1.80) to generate a dendrogram. At the distance level of 0.7, two major RAPD groups were differentiated among 20 strains. RAPD group (RG) I included 8 isolates from watermelon except one isolate from melon. RAPD group (RG) IV included 12 isolates from squash, cucumber, watermelon and melon.. In amplification experiment with SCAR specific primer RG1F-RG1R resulted in a single band of 650bp fragment only for 8 isolates out of 20 isolates that should be designated as RAPD Group 1. However, same set of experiment done with RGIIF-RGIIR did not result in any amplified product.. Our attempts to detect intraspecific diversity of ITS region of rDNA by amplifying ITS region and 17s rDNA region for 20 isolates and restriction digestion of amplified fragment with 12 enzymes did not reveal polymorphic band. In order to develop RAPD markers for RGIV specific primer, a candidate PCR fragment( ≒1.4kb) was purified and Southern hybridized to the amplified fragment RGIV isolates. This promising candidate probe recognized only RGIV isolates