• Journal of Soil and Groundwater Environment
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• v.19 no.5
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• pp.9-17
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• 2014

This paper reported the use of real-time polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), and the culture-based method in the intrinsic bioremediation study at a petroleum contaminated site. The study showed that phenol hydroxylase gene was detected in groundwater contaminated with benzene, toluene, ethylbenzene, xylene isomers (BTEX) and methyl tert-butyl ether (MTBE). This indicated that intrinsic bioremediation occurred at the site. DGGE analyses revealed that the petroleum-hydrocarbon plume caused the variation in microbial communities. MTBE degraders including Pseudomonas sp. NKNU01, Bacillus sp. NKNU01, Klebsiella sp. NKNU01, Enterobacter sp. NKNU01, and Enterobacter sp. NKNU02 were isolated from the contaminated groundwater using the cultured-based method. Among these five strains, Enterobacter sp. NKNU02 is the most effective stain at degrading MTBE without the addition of pentane. The MTBE biodegradation experiment indicated that the isolated bacteria were affected by propane. Biodegradation of MTBE was decreased but not totally inhibited in the mixtures of BTEX. Enterobacter sp. NKNU02 degraded about 60% of MTBE in the bioreactor study. Tert-butyl alcohol (TBA), acetic acid, 2-propanol, and propenoic acid were detected using gas chromatography/mass spectrometry during MTBE degraded by the rest cells of Enterobacter sp. NKNU02. The effectiveness of bioremediation of MTBE was assessed for potential field-scale application.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.41 no.1
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• pp.11-18
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• 2015

Objectives: The goal of this study was to determine the correlation of clinicopathological factors and the up-regulation of vascular endothelial growth factor (VEGF) expression in oral squamous cell carcinoma. Materials and Methods: Immunohistochemical staining of VEGF and quantitative real-time polymerase chain reaction (RT-PCR) of VEGF mRNA were performed in 20 specimens from 20 patients with oral squamous cell carcinoma and another 20 specimens from 20 patients with carcinoma in situ as a controlled group. Results: The results were as follows: 1) In immunohistochemical study of poorly differentiated and invasive oral squamous cell carcinoma, high-level staining of VEGF was observed. Significant correlation was observed between immunohistochemical VEGF expression and histologic differentiation, tumor size of specimens (Pearson correlation analysis, significance r>0.6, P<0.05). 2) In VEGF quantitative RT-PCR analysis, progressive cancer showed more VEGF expression than carcinoma in situ. Paired-samples analysis determined the difference of VEGF mRNA expression level between cancer tissue and carcinoma in situ tissue, between T1 and T2-4 (Student's t-test, P<0.05). Conclusion: These findings suggest that up-regulation of VEGF may play a role in the angiogenesis and progression of oral squamous cell carcinoma.

• Journal of Periodontal and Implant Science
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• v.48 no.4
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• pp.261-271
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• 2018

Purpose: Few studies have examined periodontal pathogens from saliva samples in periodontally healthy young adults. The purposes of this study were to determine the prevalence of periodontopathic bacteria and to quantify periodontal pathogens in saliva samples using real-time polymerase chain reaction (PCR) assays in periodontally healthy Korean young adults under 35 years of age. Methods: Nine major periodontal pathogens were analyzed by real-time PCR in saliva from 94 periodontally healthy young adults. Quantification of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Prevotella intermedia, Fusobacterium nucleatum, Campylobacter rectus, Peptostreptococcus anaerobius, and Eikenella corrodens was performed by DNA copy number measurement. Results: F. nucleatum and E. corrodens were detected in all subjects; the numbers of positive samples were 87 (92.6%), 91 (96.8%), and 90 (95.7%) for P. gingivalis, P. anaerobius, and C. rectus, respectively. Other pathogens were also detected in periodontally healthy subjects. Analysis of DNA copy numbers revealed that the most abundant periodontal pathogen was F. nucleatum, which was significantly more prevalent than all other bacteria (P<0.001), followed by P. anaerobius, P. gingivalis, E. corrodens, C. rectus, and T. denticola. There was no significant difference in the prevalence of each bacterium between men and women. The DNA copy number of total bacteria was significantly higher in men than in women. Conclusions: Major periodontal pathogens were prevalent in the saliva of periodontally healthy Korean young adults. Therefore, we suggest that the development of periodontal disease should not be overlooked in periodontally healthy young people, as it can arise due to periodontal pathogen imbalance and host susceptibility.

• Journal of Ginseng Research
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• v.43 no.3
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• pp.482-487
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• 2019

Background: The mixed-cultivation of different Panax ginseng cultivars can cause adverse effects on stability of yield and quality. K-1 is a superior cultivar with good root shape and stronger disease resistance. DNA markers mined from functional genes are clearly desirable for K-1, as they may associate with major traits and can be used for marker-assisted selection to maintain the high quality of Korean ginseng. Methods: Five genes encoding pathogenesis-related (PR) proteins of P. ginseng were amplified and compared for polymorphism mining. Primary, secondary, and tertiary structures of PR5 protein were analyzed by ExPASy-ProtParam, PSSpred, and I-TASSER methods, respectively. A coding single nucleotide polymorphism (SNP)-based specific primer was designed for K-1 by introducing a destabilizing mismatch within the 3' end. Allele-specific polymerase chain reaction (PCR) and real-time allele-specific PCR assays were conducted for molecular discrimination of K-1 from other cultivars and landraces. Results: A coding SNP leading to the modification of amino acid residue from aspartic acid to asparagine was exploited in PR5 gene of K-1 cultivar. Bioinformatics analysis showed that the modification of amino acid residue changed the secondary and tertiary structures of the PR5 protein. Primer KSR was designed for specific discrimination of K-1 from other ginseng cultivars and landraces. The developed real-time allele-specific PCR assay enabled easier automation and accurate genotyping of K-1 from a large number of ginseng samples. Conclusion: The SNP marker and the developed real-time allele-specific PCR assay will be useful not only for marker-assisted selection of K-1 cultivar but also for quality control in breeding and seed programs of P. ginseng.

• Korean Journal of Veterinary Service
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• v.39 no.4
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• pp.247-251
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• 2016

Salmonella is one of the most common bacteria that causes heavy losses in swine industry and major causative pathogen of food poisoning in public health. Various methods for the identification of Salmonella such as Gram staining, agglutination test, enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR) have been used. Several studies have demonstrated that Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI TOF) Mass Spectrometry (MS) identification is an efficient and inexpensive method for the rapid and routine identification of isolated bacteria. In this study, MALDI TOF MS could provide rapid, accurate identification of Salmonella spp. from swine compared with end point PCR and real time PCR.

• Journal of Korean society of Dental Hygiene
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• v.24 no.2
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• pp.131-139
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• 2024

Objectives: Oral bacterial samples included subgingival, supragingival, and saliva plaques. As the diversity and number of microorganisms deffer depending on the area of the oral cavity and the method used, an appropriate and reliable collection method is important. The present study investigated oral bacterial sampling methods. Methods: Supragingival dental plaque was collected from the buccal and lingual tooth surfaces of study participants using sterilized cotton swabs. Plaques were collected from the subgingival area using a sterilized curette. Bacterial genomic DNA was extracted using MagNA Pure 96 DNA and Viral NA low-volume kits. Real-time polymerase chain reaction (PCR) was performed using the PowerCheck^TM Periodontitis Pathogens Multiplex Real-time PCR kit. Results: Aggregatibacter actinomycetemcomitans, Prevotella intermedia, and Fusobacterium nucleatum of the orange complex were not observed in the subgingival biofilms of all study participants. For Porphyromonas. gingivalis, a significant correlation was observed between supragingival, subgingival, and total tooth surface biofilms. Compared to the supragingival and subgingival biofilmss, total tooth surface biofilm exhibited the highest bacterial count when the inswabbing method was used. Conclusions: Based on these findings, the supragingival swab method is recommended for oral bacterial research.

• Journal of Veterinary Clinics
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• v.32 no.2
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• pp.154-157
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• 2015

A major cause of diarrhea in a dog is an infection with bacteria which include Salmonella spp., Campylobacter (C.) spp., and Clostridium (Cl.) spp.. It is fastidious to identify these bacteria by the culture. The purpose of this experiment is to devise the method for detecting Cl. perfringens, C. jejuni, C. coli, and Salmonella spp. with rapid and high sensitivity. The fecal samples collected from 71 normal and 66 diarrheic dog feces were used to compare the prevalence of the enteric pathogens and to develop a multiplex real-time polymerase chain reaction (PCR) assay for clinical use. Detection of Cl. perfringens, C. coli, and C. jejuni in diarrhea feces was higher than normal feces. A developed multiplex real-time PCR is useful for determining the presence and quantity of pathogen-specific or other unique sequences with in a fecal sample.

• Journal of Chest Surgery
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• v.44 no.2
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• pp.123-130
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• 2011

Background: The aim of the present study was to identify chromosomal loci that contribute to the pathogenesis of aortic dissection (AD) in a Korean population using array comparative genomic hybridization (CGH) and to confirm the results using real-time polymerase chain reaction (PCR). Materials and Methods: Eighteen patients with ADs were enrolled in this study. Genomic DNA was extracted from individual blood samples, and array CGH analyses were performed. Four corresponding genes with obvious genomic changes were analyzed using real-time PCR in order to assess the level of genomic imbalance identified by array CGH. Results: Genomic gains were most frequently detected at 8q24.3 (56%), followed by regions 7q35, 11q12.2, and 15q25.2 (50%). Genomic losses were most frequently observed at 4q35.2 (56%). Real-time PCR confirmed the results of the array CGH studies of the COL6A2, DGCR14, PCSK6, and SDHA genes. Conclusion: This is the first study to identify candidate regions by array CGH in patients with ADs. The identification of genes that may predispose an individual to AD may lead to a better understanding of the mechanism of AD formation. Further multicenter studies comparing cohorts of patients of different ethnicities are warranted.

• Journal of Microbiology and Biotechnology
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• v.23 no.7
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• pp.993-1003
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• 2013

Methanotrophs are the most important sink of $CH_4$, which is a more highly potent greenhouse gas than $CO_2$. Methanotrophic abundance and community diversity in cover soils from two typical semi-aerobic landfills (SALs) in China were detected using real-time polymerase chain reaction (real-time-PCR) and denaturing gradient gel electrophoresis (DGGE) based on 16S rRNA genes, respectively. Real time-PCR showed that Type I methanotrophs ranged from $1.07{\times}10^6$ to $2.34{\times}10^7$ copies/g soil and that of Type II methanotrophs from $1.51{\times}10^7$ to $1.83{\times}10^8$ copies/g soil. The ratio of Type II to Type I methanotrophic copy numbers ranged from 5.61 to 21.89, indicating that Type II methanotrophs dominated in SAL. DGGE revealed that Type I methanotrophs responded more sensitively to the environment, changing as the community structure varied with different soil types and locations. Methylobacter, Methylosarcina, and Methylomicrobium for Type I, and Methylocystis for Type II were most prevalent in the SAL cover layer. Abundant interflow $O_2$ with high $CH_4$ concentration in SALs is the reason for the higher population density of methanotrophs and the higher enrichment of Type II methanotrophs compared with anaerobic landfills and other ecosystems, which proved a conclusion that increasing the oxygen supply in a landfill cover layer would greatly improve $CH_4$ mitigation.

• Korean Journal of Veterinary Service
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• v.38 no.2
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• pp.107-116
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• 2015

In this study, we developed a rapid, sensitive and specific reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) assay for detection of swine influenza viruse (SIV) including major subtypes of swine influenza viruses H1N1, H1N2 and H3N2, and a novel subtype of influenza A virus that accidentally infected in pig population. The RT-LAMP was completed in 40 min at $58^{\circ}C$ and the sensitivity of the RT-LAMP ($1copy/{\mu}L$) was 10-fold higher than conventional reverse transcription-polymerase chain reaction (RT-PCR) ($10copy/{\mu}L$) and the same to real time RT-PCR ($1copy/{\mu}L$). Also, the result of the RT-LAMP can be confirmed without any detection system. Therefore, the RT-LAMP could be a alternative diagnostic method for SIV detection in national SIV monitoring system and clinical diagnostic laboratory in the future.