• Journal of fish pathology
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• v.30 no.2
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• pp.79-88
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• 2017

At the present, fish farms are suffering a lot of economic losses due to infectious diseases caused by various pathogens including aeromonad. Aeromonad is ubiquitous bacteria that causes infectious diseases. At least 26 species in the genus Aeromonas have been reported to cause fatal infections not only in salmonid fishes, but also in other freshwater and seawater fishes. Molecular techniques based on nucleic acid sequences of 16S rDNA and housekeeping genes can be used to identify the Aeromonas species. In this study, The genus Aeromonas was isolated from salmonid fishes of sixteen fish farms in Gangwon-Do, Korea and phylogenetically identified based on the sequences of 16S rDNA and housekeeping genes for Aeromonad, i.e. RNA polymerase sigma factor ${\sigma}^{70}$ (rpoD) or DNA gyrase subunit B (gyrB). Consequently, 96 strains were collected from Atlantic salmon (Salmo salar), coho salmon (Oncorhynchus kisutch), masou salmon (Oncorhynchus masou) and rainbow trout (Oncorhynchus mykiss), and 36 isolates were identified as the genus Aeromonas by 16S rDNA analysis. Thirty six Aeromonad isolates were further analysed based on rpoD or gyrB gene sequences and found Aeromonas salmonicida (24 isolates), A. sobria (10 isolates), A. media (1 isolates) and A. popoffii (1 isolates), indicating that A. salmonicida is a main infectious bacteria in Salmonid fishes in Gangwon-Do. It was also proved that the phylogenetic identification of Aeromonas species based on the sequences of housekeeping gene is more precise than the 16S rDNA sequence.

• BMB Reports
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• v.30 no.1
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• pp.26-32
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• 1997

Of all HLA class I molecules, HLA-C gene products are most poorly understood because they express at a low level on the cell surface compared to HLA-A and -B. In order to identify serologically detectable and undetectable HLA-C antigens, we have established a DNA-based tissue typing method for the HLA-C locus by PCR-SSP (polymerase chain reaction-sequence specific primers). Genomic DNA prepared from Iymphoblastoid 21 B-cell lines and 120 Korean individuals by proteinase K digestion and pheno/chloroform extractions have been typed by PCR-SSP (23 primer mixes were used). The PCR-SSP results of control cell lines were discrepant from serology in 1 case among 21 cases: Cw6 which was negative by serology but positive by PCR-SSP (cell line: MANIKA). Twenty four HLA-Cw "blank" antigens among fifty Korean individuals were completely determined by PCR-SSP DNA typing. HLA-Cw*0101 (15.3%), Cw*1401 (12.3%) and Cw*0701 (11.7%) alleles were frequently found in 120 Korean individual samples. In conclusion. the high level of discrimination for HLA-C alleles may prove useful and informative in the study of transplant survival, and identify the importance of allelic differences, not readily detectable by serology, on host and donor compatibility.

• International Journal of Oral Biology
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• v.38 no.1
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• pp.29-36
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• 2013

Mitis group streptococci (MGS) were classified based on the nucleotide sequences 16S rRNA gene (16S rDNA) and comprised 13 Streptococcus species. However, 16S rDNA homogeneity among MGS was too high to discriminate between clinical strains at the species level, notably between Streptococcus mitis, Streptococcus oralis, Streptococcus pneumoniae, and Streptococcus pseudopneumoniae. The purpose of this study was to discriminate between 37 strains of MGS isolated from Korean oral cavities using phylogenetic analysis of the DNA-dependant RNA polymerase beta-subunit gene (rpoB). 16S rDNA and rpoB from clinical strains of MGS were sequenced using the dideoxy chain termination method and analyzed using MEGA version 5 software. The resulting phylogenetic data showed that the rpoB sequences could delineate clinical strains of MGS at the species level. Phylogenetic analysis of rpoB is therefore a useful approach for identifying MGS at the species level.

• BMB Reports
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• v.33 no.1
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• pp.59-62
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• 2000

Hepatitis C virus (HCV) nonstructural 5B (NS5B) protein is an RNA-dependent RNA polymerase (RdRp). To determine whether it can contribute to viral replication by interaction with cellular proteins, the yeast two-hybrid screening system was employed to screen a human liver cDNA library. Using the HCV NS5B as a bait, we have isolated positive clones encoding a cellular protein. The NS5B interacting protein, 5BIP, is a novel cellular protein of 170 amino acids. Interaction of the HCV NS5B protein with 5BIP was confirmed by a protein-protein blotting assay. Recently, we have demonstrated that NS5B possesses an RdRp activity and thus it is possible that 5BIP, in association with NS5B, plays a role in HCV replication.

• Biomedical Science Letters
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• v.9 no.3
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• pp.167-170
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• 2003

Cytochrome $b_{5}$($b_{5}$) can be found in a variety of tissues and plays a role in the electron transfer pathways. Several retropseudogenes (numbered as I, II, II, IV, V) have been identified and well investigated for their structures. However, retropseudogene I is not clear in terms of its location on the chromosome. In addition the structure and the exression of retropseudogene V have not been confirmed. To examine the structure of bs retropseudogenes V and to see whether it is expressed in human blood we applied recombinant DNA technologies including polymerase chain reaction (PCR) and DNA sequencing. Retropseudogene V turned out to contain open reading frame (ORF) within its structure, however, no evidence of its expression was detected. Retropseudogene I was also found on the chromosome V. This study should contribute to the understanding of the structure of bs gene family.

• Korean Journal of Veterinary Research
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• v.49 no.2
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• pp.105-111
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• 2009

In this study, occurrence of canine brucellosis was surveyed in kennels, indoor dogs and stray dogs in Korea, and infrequent restriction site-polymerase chain reaction (IRS-PCR) was applied to analyze DNA polymorphism of Brucella canis (B. canis) isolates. Among a total of 501 dogs tested, B. canis antibodies by both rapid screening agglutination with 2-mercaptoethanol (2-ME RSAT) and immunochromatographic assay were detected in only 14.1% of kennel dogs. There were no seropositive cases in indoor dogs and stray dogs. DNA polymorphism was observed in 16 B. canis isolates by the IRS-PCR. Sixteen isolates were tested with primers, PsalA, PsalC, PsalG and PsalT, and different primers produced different DNA patterns. In regard to the IRS-PCR pattern of 16 isolates, 9 (56.3%) belonged to the IRS-PCR type I. The remaining 7 were differentiated as type II, III and IV. An application of the primer PsalC provided discrimination between B. canis isolated in 2005 and others.

• Clinical and Experimental Pediatrics
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• v.53 no.3
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• pp.397-407
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• 2010

Purpose : The F9 gene is known to be the causative gene for hemophilia B, but unfortunately the detection rate for restriction fragment length polymorphism-based linkage analysis is only 55.6%. Direct DNA sequencing can detect 98% of mutations, but this alternative procedure is very costly. Here, we conducted multiplex polymerase chain reactions (PCRs) and conformation sensitive gel electrophoresis (CSGE) to perform a screened DNA sequencing for the F9 gene, and we compared the results with direct sequencing in terms of accuracy, cost, simplicity, and time consumption. Methods : A total of 27 unrelated hemophilia B patients were enrolled. Direct DNA sequencing was performed for 27 patients by a separate institute, and multiplex PCR-CSGE screened sequencing was done in our laboratory. Results of the direct DNA sequencing were used as a reference, to which the results of the multiplex PCR-CSGE screened sequencing were compared. For the patients whose mutation was not detected by the 2 methods, multiplex ligation-dependent probe amplification (MLPA) was conducted. Results : With direct sequencing, the mutations could be identified from 26 patients (96.3%), whereas for multiplex PCRCSGE screened sequencing, the mutations could be detected in 23 (85.2%). One patient's mutation was identified by MLPA. A total of 21 different mutations were found among the 27 patients. Conclusion : Multiplex PCR-CSGE screened DNA sequencing detected 88.9% of mutations and reduced costs by 55.7% compared with direct DNA sequencing. However, it was more labor-intensive and time-consuming.

• Herbal Formula Science
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• v.19 no.2
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• pp.83-92
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• 2011

Objectives : This study was carried out to moniter microbial flora on freeze-dried herbal powder and identify isolated bacteria. Methods : We measured the total number of bacteria and fungi in 29 herbal powder which had made according to the guideline of KFDA. For the identification, we observed microscopic properties and carried out polymerase chain reaction(PCR). The purified DNA was analyzed by DNA sequencer. Results : Among the 29 herbal powders, the fungi were detected only one sample as unacceptable range of total aerobic bacteria. Isolated bacteria were identified as Bacillus cereus, B. subtilis, B. megaterium, B. licheniformis, Erwinia tasmaniensis, E. amylovora, and Pantoea agglomerans by 16S rDNA analysis. E. tasmaniensis was observed 20 herbal samples. Conclusions : According to above results, further studies for the effective sterilization of low herbal materials should be needed.

• BMB Reports
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• v.38 no.6
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• pp.690-694
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• 2005

Transcription termination of the human mitochondrial genome requires specific binding to termination factor mTERF. In this study, mTERF was produced in E. coli and purified by two-step chromatography. mTERF-binding DNA sequences were isolated from a pool of randomized sequences by the repeated selection of bound sequences by gel-mobility shift assay and polymerase chain reaction. Sequencing and comparison of the 23 isolated clones revealed a 16-bp consensus sequence of 5'-GTG$\b{TGGC}$AGANCCNGG-3' in the light-strand (underlined residues were absolutely conserved), which nicely matched the genomic 13-bp terminator sequence 5'-$\b{TGGC}$AGAGCCCGG-3'. Moreover, mTERF binding assays of heteroduplex and single-stranded DNAs showed mTERF recognized the light strand in preference to the heavy strand. The preferential binding of mTERF with the light-strand may explain its distinct orientation-dependent termination activity.

• The Plant Pathology Journal
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• v.19 no.2
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• pp.106-110
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• 2003

Stolbur Phytoplasma was detected from Lilium Oriental hybrids showing flattened stem and flower clustering. The presence of phytoplasma was demonstrated using polymerase chain reaction（PCR) assays with phyto-plasma-universal（P1/P6）and stolbur phytoplasma-specific 16F1/R1-S primer pairs amplifying phytoplasma 16S rDNA regions. Nucleotide suquences of the phytoplasma 16S rDNA were determined. Nucleic acid extracted from lily amplified 1.5 kb DNA with a phytoplasma universal primer pair. In nested PCR, 1.1 kb PCR product was obtained using specific primer pair, indicating an isolate of stolbur phytoplasma. Nucleotide sequence of phytoplasma 16S rDNA reported in this study showed 99.5% and 99.1％ identities with two known stolbur phytoplamas (16Sr XII-A). Also, it exhibited a sequence homology of 98.0％ with phormium yellow leaf (16Sr XII-B), and 97.9％ with Australian grapevine yellows (16Sr XII-B). Meanwhile, it showed 98.1% identity with strawberry green petal phytoplama, (16Sr1-C), and 94.7 % with American aster yellows (16Sr1-B). Homology percentage of the 16S rDNA nucleotide sequence suggests that this phytoplama could be classified into the stolbur phytoplasma, subgroup A (16Sr XII-A), as a type strain stolbur.