• Korean Journal of Plant Resources
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• v.20 no.6
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• pp.491-498
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• 2007

It has been hypothesized that efficient exclusion of methylated retrotransposons and repeated DNA region is one of the rapid and cost-effective approaches for comprehensive gene discovery in large genome size of maize. Three kinds of methylation-sensitive restriction enzymes, HapII, MspI and McrBC, were used to identify the restriction frequency of cytosine methylation sites in maize genome. Roughly 60% of total maize genomic DNA was restricted less than 500bp by McrBC, and the most of restricted small size fraction was composed retrotransposon. In order to validate the efficient construction of gene-rich shotgun library, we compare two gene-rich methyl-filtration shotgun libraries using in vivo and in vitro methyl-filtration system. The size selected DNA fraction by Sau3A-McrBC enzyme treated was very stable and has not appeared modification in E. coli, but most insert DNA size of partially digested with Sau3A were decrease less than 500bp by bacterial methylation-modification system. In compare of retroelements portion, A 44.6% of the sequences were retroelement in unmethyl-filtered library, and the most of them was Copia type, such as Prem, Opie and Ji. The portion of retroelement was drastically decreased to 25% and 20% by in vivo and in vitro filtration system, respectively.

• Journal of Periodontal and Implant Science
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• v.25 no.1
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• pp.89-98
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• 1995

P. intermedia are considered an important pathogen in adult periodontitis, rapidly progressing periodontitis, refractory periodontitis, pregnancy gingivitis, acute necrotizing ulcerative gingivitis, pubertal gingivitis. So far 2 DNA homology groups and 3 serotypes of P. intermedia have been reported but there is no data available as yet regarding genetic diversity for the species P. intermedia. The purpose of this study is to investigate, using bacterial DNA restriction endonuclease analysis, genetic diversity between individual strains of P. intermedia which are indistinguishable by serotyping and biotyping, occurrence of an intrafamilial transmission and genetic heterogeneity between P. intermedia strains isolated within a patient and within the same serotypes. The families who have had no systemic disease, no experience of periodontal treatment for the previous 1 year and no experience of antibiotics for the previous 6 months were selected and subgingival plaque was collected at 4 sites in each person and incubated in the anaerobic chamber. P. intermedia were identified by colony shape, gram stain, biochemical test, SK-I03(Sunstar Inc.) test and IIF using monoclonal antibody was perfomed for the determination of serotypes. P. intermedia strains were grown in BHI broth and whole genomic DNA was extracted and digested by restriction endonuclease. The resulting DNA fragments were separated by agarose gel electrophoresis, stained and photographed under UV. As the results of this study, intrafamilial vertial transmissions could be assessed in 2 families and horizintal transmissions in another 2 families. There were different DNA digest patterns within a patient, so P. intermedia showed that individuals could be colonized by multiple clonal types at anyone time. And different serotypes could be found within a patient and in the same serotype within a patient, obvius genetic heterogeneity could not be assessed. But in the same serotype in different famies, there were differences in the DNA digest patterns.

• KSBB Journal
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• v.11 no.2
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• pp.193-200
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• 1996

In molecular biology, type-II restriction endonuclease, which specifically recognize and cleave DNA at a limited number of sites, have been exploited as a means of characterizing DNA fragments, DNA mapping for genetic engineering. Type-II restriction endonucleases have been found to modulate their substrate specificity under modified conditions such as extreme pH, ionic strength, high enzyme concentration, substitution of metallic cofactors or addition of organic solvents. This study was initiated to investigate the modification of recognition specificity of BamHI according to the different pH and organic solvent under the given buffer condition. The specificity of BamHI is highly depends on the presence of hydrophobicity (LogP: partition coefficient) and pH of reaction solution. The specificity of BamHI is changed in range of LogP -1.03∼-1.35(at pH 7.5), -1.03∼-2.5 (at pH 8.0), -0.75∼-0.25(at pH 8.5), 0.32∼-2.5(at pH 8.9), respectively. Alteration of specificity appears in lower concentration of organic solvent when the reaction occurs in more alkali pH. For example, in DMSO solution, alteration of specificity appears in 20% concentration at pH 7.5 but in 4% concentration at pH 8.9.

• Journal of Microbiology
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• v.44 no.1
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• pp.29-34
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• 2006

The purpose of this study was to develop molecular identification method for medical mushrooms and their preparations based on the nucleotide sequences of nuclear large subunit (LSD) rDNA. Four specimens were collected of each of the three representative medicinal mushrooms used in Korea: Ganoderma Incidum, Coriolus versicolor, and Fomes fomentarius. Fungal material used in these experiments included two different mycelial cultures and two different fruiting bodies from wild or cultivated mushrooms. The genomic DNA of mushrooms were extracted and 3 nuclear LSU rDNA fragments were amplified: set 1 for the 1.1-kb DNA fragment in the upstream region, set 2 for the 1.2-kb fragment in the middle, and set 3 for the 1.3-kb fragment downstream. The amplified gene products of nuclear large subunit rDNA from 3 different mushrooms were cloned into E. coli vector and subjected to nucleotide sequence determination. The sequence thus determined revealed that the gene sequences of the same medicinal mushroom species were more than $99.48\%$ homologous, and the consensus sequences of 3 different medicinal mushrooms were more than $97.80\%$ homologous. Restriction analysis revealed no useful restriction sites for 6-bp recognition enzymes for distinguishing the 3 sequences from one another, but some distinctive restriction patterns were recognized by the 4-bp recognition enzymes AccII and HhaI. This analysis was also confirmed by PCR-RFLP experiments on medicinal mushrooms.

• Korean Journal of Microbiology
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• v.24 no.1
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• pp.7-11
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• 1986

We screened lysates of the laboratory strains of pseudomonads utilizing hydrocarbon by agarose gel electrophoresis and cesium chloride-ethidium bromide equilibrium centrifugation, to find an intrinsic plasmid as a vector and to examine the relationship between the plasmid and hydrocarbon degradation. Only one strain from the examined strains, Pseudomonas putida KU190, contained a plasmid. We named the plasmid pKU41. The molecular size of pKU41 was determined as 41kb, using covalently closed circular forms of RP4 and pSY343 as standard size markers. The restriction sites of pKU41 for BamHI, BglII, EcoRI, HindIII, and SalI were 3, 1, 3, 6 and more than 13, respectively. With double or triple digestion, restriction map of pKU41 was constructed for BamHI, BglII and HindIII. For elucidation on the biological function of the plasmid, test was conducted on the ability of hydrocarbon utilization of the host strain but no apparent relationship was observed.

• Korean Journal of Plant Taxonomy
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• v.41 no.2
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• pp.119-129
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• 2011

Phylogenetic studies were conducted to evaluate the taxonomic relationships among 27 taxa, including 2 outgroups of Korean Campanulaceae, using PCR-RFLP analysis and ITS sequences. In the PCR-RFLP analysis, 15 restriction endonucleases produced 244 restriction sites and size variations from the chloroplast DNA, and 59 restriction sites (24%) showed polymorphism. The length of the ITS regions ranged from 588 bp to 797 bp. The sequence divergence including the outgroups is 0-39.36%. Phylogenetic analyses based on PCR-RFLP and ITS data suggest that Campanulaceae is monophyletic; Codonopsis and Platycodon forms an independent clade; the Peracarpa and Asyneuma clade is a sister to the Adenophora-Hanabusaya clade; Campanula is monophyletic; and Wahlenbergia basally branches within the ITS tree, whereas they are placed between Campanula and the Codonopsis-Platycodon clade in the PCR-RFLP tree; Hanabusaya is placed within the Adenophora clade; and Adenophora is paraphyletic and shows discordance to the infrageneric classifications based on morphological data. The present results show two data sets, largely congruent at the generic level, but their phylogenetic positions, in particular the Wahlenbergia and Hanabusaya and the infrageneric classifications in Adenophora, show some incongruence.

• Korean Journal of Plant Taxonomy
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• v.34 no.1
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• pp.43-61
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• 2004

Molecular taxonomic studies were conducted to evaluate interspecific relationships in Korean Viola 34 taxa including two Japanese populations using RAPD(randornly amplified polymorphic DNA), ISSR(inter simple sequence repeat) and PCR-RFLP(restriction fragment length polymorphism) analysis. Only six and four primers out of 40 arbitrary and 12 ISSR primers were screened for 34 taxa, and were revealed 70 (98.6%) and 28 (96.6%) polymorphic bands, respectively. Fifteen restriction endonucleases produced 80 restriction sites and size variations from the large single copy region of cpDNA, 16 (20%) of which were polymorphic. The separate analyses from the RAPD, ISSR and PCR-RFLP data were incongruent in the relationships among 34 taxa, but combined data was in accordance with previous infrageneric classification system based on morphological characters, especially the subsection and series level. Section Chamaemelanium placed between subsect. Patellares and Vagimtae of section Nomimium was not formed as a distinct group. Viola alb ida complex including three very closely related taxa was recognized independent group within subsect. Patellares in combined data tree. This result strongly suggested that they should be treated to series Pinmtae. RAPD analysis was very useful to clarify the interspecific relationships among the species of Korean Viola than ISSH and PCR-RFLP analyses.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.7
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• pp.942-948
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• 2005

New polymorphism of major histocompatibility complex B-G genes was investigated by amplification and digestion of a 401bp fragment including intron 1 and exon 2 using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique with two restriction enzymes of Msp I and Tas I in eight breeds of Chinese indigenous chickens and one exotic breed. In the fragment region of the gene, three novel single nucleotide polymorphisms (SNPs) were detected at the two restriction sites. We found the transition of two nucleotides of A294G and T295C occurred at Tas I restriction site, and consequently led to a non-synonymous substitution of asparagine into serine at position 54 within the deduced amino acid sequence of immunoglobulin variable-region-like domain encoded by the exon 2 of B-G gene. It was observed at rare frequency that a single mutation of A294G occurring at the site, also caused an identical substitution of amino acid, asparagine 54-to-serine, to that we described previously. And the transversion of G319C at Msp I site led to a non-synonymous substitution, glutamine 62-to-histidine. The new alleles and allele frequencies identified by the PCR-RFLP method with the two enzymes were characterized, of which the allele A and B frequencies at Msp I and Tas I loci were given disequilibrium distribution either in the eight Chinese local breeds or in the exotic breed. By comparison, allele A at Msp I locus tended to be dominant, while, the allele B at Tas I locus tended to be dominant in all of the breeds analyzed. In Tibetan chickens, the preliminary association analysis revealed that no significant difference was observed between the different genotypes identified at the Msp I and Tas I loci and the laying performance traits, respectively.

• Korean Journal of Environmental Biology
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• v.36 no.3
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• pp.352-358
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• 2018

This study aimed to develop a species identification method for the egg and fry of the three Korean bitterling fishes (Pisces: Acheilognathinae), including Acheilognathus signifer, Acheilognathus yamatsutae and Rhodeus uyekii based on the PCR-based Restriction Fragment Length Polymorphism (RFLP) markers. We conducted a field survey on the Deokchicheon River from the North Han River basin, where the three Acheilognathinae species co-occur, and also analyzed the existing sequence dataset available from the GenBank. We found coexistence of the three species at the study site. The egg and fry were obtained from the host mussels (Unio douglasiae sinuolatus) by hand from May to June 2015 and in May 2017. To develop PCR-based RFLP markers for species identification of the three Acheilognathinae fish species, restriction enzymes pinpointing species-specific single nucleotide variation (SNV) sites in mitochondrial DNA COI (cytochrome oxidase I) and cyt b (cytochrome b) genes were determined. Genomic DNA was extracted from the egg and fry and RFLP experiments were carried out using restriction enzymes Apal I, Stu I and EcoR V for A. signifer, A. yamatsutae and R. uyekii, respectively. Consequently, unambiguous discrimination of the three species was possible, as could be seen in DNA band patterns from gel electrophoresis. Our developed PCR-based RFLP markers will be useful for the determination of the three species for the young and would assist in studying the spawning patterns and reproductive ecology of Acheilognathinae fishes. Furthermore, we believe the obtained information will be of importance for future maintenance, management and conservation of these natural and endangered species.

• Management Science and Financial Engineering
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• v.14 no.2
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• pp.45-66
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• 2008

Refuse collection, one of important elements in reverse logistics, is an activity rendering recyclables or wastes and moving them to some points where further treatment is required. Among various decisions in the collection activity, we focus on network design, which is the problem of locating collection points as well as allocating refuses at demand points to collection points while satisfying the capacity restriction at each collection point. Here, the collection point is the place where recyclables or wastes near the point are gathered, and locating the collection points is done by selecting them from a given set of potential sites. The objective is to minimize the sum of fixed costs to open collection points and transportation costs to move refuses from demand points to collection points. An integer programming model is developed to represent the problem mathematically and due to the complexity of the problem, two types of heuristics, one with simultaneous and the others with separate location and allocation, are suggested. Computational experiments were done on test problems up to 500 potential sites, and the results are reported. In particular, some heuristics gave near optimal solutions for small-size test problems, i.e., 2% gaps in average from the optimal solution values.