• Korean Journal of Microbiology
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• v.42 no.3
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• pp.185-194
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• 2006

The aim of this study was to identify the bacteria isolated from endodontic lesions by cell culture and to determine the antimicrobial susceptibility of them against 8 antibiotics. The necrotic pulpal tissues were collected from 27 infected root canals, which were diagnosed as endodontic infection. Samples were collected aseptically from the infected pulpal tissue of the infected root canals using a barbed broach and a paper point. The cut barbed broaches and paper points were transferred to an eppendorf tube containing $500{\mu}l\;of\;1{\times}PBS$. The sample solution was briefly mixed and plated onto a BHI-agar plate containing 5% sheep blood. The agar plates were incubated in a $37^{\circ}C$ anaerobic chamber for 2 to 5 days. The bacteria grown on the agar plates were identified by comparison of 16S rRNA gene (rDNA) sequencing method at the species level. To test the sensitivity of the bacteria isolated from the infected root canals against 8 antibiotics, minimum inhibitory concentrations (MIC) were determined using broth dilution assay. The data showed that 101 bacterial strains were isolated and were identified. Streptococcus spp. (29.7%) and Actinomyces spp. (21.8%) were predominantly isolated. The 9 strains were excluded in antimicrobial susceptibility test because they were lost during the experiment or were not grown in broth culture. The percentage of bacteria susceptible for each antibiotic in this study was clindamycin, 87.0% (80 of 92); tetracycline, 75.0% (69 of 92); cefuroxime axetil, 75.0% (69 of 92); amoxicillin + clavulanic acid (5:1), 71.7% (66 of 92); penicillin G, 66.3% (61 of 92); erythromycin, 66.3% (61 of 92); amoxicillin, 44.6% (41 of 92); and ciprofloxacin, 31.5% (29 of 92). The susceptibility pattern of 8 antibiotics was dependent on the host of the bacteria strains rather than the kinds of bacterial species. These results indicate that antibiotic susceptibility test should be performed when antibiotics are needed for the treatment of infected root canals.

• Journal of Life Science
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• v.23 no.2
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• pp.167-174
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• 2013

Peroxiredoxin 6 (Prx 6) is a member of the thiol-specific antioxidant protein family, which may play a role in protection against oxidative stress and in regulating phospholipid turnover. The aim of this study was to determine whether a human Prx 6/Luc vector was stably expressed and responded to antioxidants in a lung cell line (NCI-H460). To achieve this, the luciferase signal, hPrx 6 mRNA expression, and superoxide dismutase (SOD) activity were measured in transfectants with a hPrx 6/Luc plasmid after treatment with four antioxidant extracts, including Korea white ginseng (KWG), Korea red ginseng (KRG), Liriope platyphylla (LP), and red Liriope platyphylla (RLP). First, the hPrx 6/Luc plasmid was successfully constructed with DNA fragments of human Prx 6 promoter, amplified by PCR using genomic DNA isolated from NCI-H460 cells, and cloned into the pTransLucent reporter vector. The orientation and sequencing of the hPrx 6/Luc plasmid were identified with restriction enzyme and automatic sequencing. A luciferase assay revealed significant enhancement of luciferase activity in the four treatment groups compared with a vehicle-treated group, although the ratio of the increase was different within each group. The KRG- and LP-treated groups showed higher activity than the KWG- and RLP-treated groups. Furthermore, the luciferase activity against RLP occurred roughly in a dose-dependent manner. However, the level of endogenous hPrx 6 mRNA did not change in any group treated with the four extracts. The SOD activity was in agreement with the luciferase activity. Therefore, these results indicate that the hPrx 6/Luc vector system may successfully express and respond to antioxidant compounds in NCI-H460 cells. The data also suggest that the Prx 6/Luc vector system may be effectively applied in screening the response of hPrx 6 to antioxidant compounds in transgenic mice.

• Journal of Genetic Medicine
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• v.4 no.1
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• pp.72-79
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• 2007

Purpose : Glycogen storage disease type III (GSD-III) is a rare autosomal recessive disorder of glycogen metabolism. The affected enzyme, amylo-1,6-glucosidase, 4-alpha-glucanotransferase (AGL, glycogen debranching enzyme), is responsible for the debranching of the glycogen molecule during catabolism. The disease shows clinical and biochemical heterogeneity, reflecting genotype-phenotype heterogeneity among different patients. In this study, we aim at analyzing mutations of the AGL gene in three unrelated Korean GSD-III patients, and characterizing their clinical and laboratory findings. Methods : We characterized the clinical features of three unrelated Korean GSD-III patients by biochemical, histological and imaging studies. The 35 exons and part of exon-intron boundaries of AGL were analyzed by direct sequencing using genomic DNA extracted from the peripheral leukocytes of patients. Results : Diverse clinical features were observed in these patients including hepatomegaly (all patients), seizures (patient 2), grow th failure (patients 1 and 2), hyperlipidemia (patients 1 and 3), raised transaminase and creatine kinase concentrations (all patients), and mild cardiomyopathy (patient 2). Liver transplantation w as performed in patient 2 due to progressive hepatic fibrosis. A dministration of uncooked corn starch maintained normoglycemia and improved biochemical and growth profiles. DNA sequence analysis revealed mutations in 5 out of 6 alleles. Patient 1 was a compound heterozygote of c.1282 G>A (p.R428K) and c.1306delA (p.S603PfsX6), patient 2 had c.1510_1511insT (p.Y 504L fsX 10), and patient 3 had c.3416 T >C (p.L 1139P) and c.1735+1 G>T (p.Y 538_R578delfsX 4) mutations. A part from the p.R428K mutation, the 4 other substitutions identified w ere nov el. Conclusion : GSD-III patients display variable phenotypic characteristics resembling those of GSD-Ia. Molecular defects in the AGL gene of Korean GSD-III patients are genetically heterogeneous.

• Journal of Life Science
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• v.18 no.8
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• pp.1090-1095
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• 2008

Marbling (intramuscular fat) is an important factor in determining meat quality in Korean beef market. A grain based finishing system for improving marbling leads to inefficient meat production due to an excessive fat production. Identification of intramuscular fat-specific gene might be achieved more targeted meat production through alternative genetic improvement program such as marker assisted selection (MAS). We carried out ddRT-PCR in 12 and 27 month old Hanwoo steers and detected 300 bp PCR product of the inducible cAMP early repressor (ICER) gene, showing highly gene expression in 27 months old. A 1.5 kb sequence was re-sequenced using primer designed base on the Hanwoo EST sequence. We then predicted the open reading frame (ORF) of ICER gene in ORF finder web program. Tissue distribution of ICER gene expression was analysed in eight Hanwoo tissue using realtime PCR analysis. The highest ICER gene expression showed in Small intestine followed by Longissimus dorsi. Interestingly, the ICER gene expressed 2.5 time higher in longissimus dorsi than in same muscle type, Rump. For gene expression analysis in high- and low marbled individuals, we selected 4 and 3 animal based on the muscle crude fat contents (high is 17-32%, low is 6-7% of crude fat contents). The ICER gene expression was analysed using ANOVA model. Marbling (muscle crude fat contents) was affected by ICER gene (P=0.012). Particularly, the ICER gene expression was 4 times higher in high group (n=4) than low group (n=3). Therefore, ICER gene might be a functional candidate gene related to marbling in Hanwoo.

• Journal of Life Science
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• v.26 no.11
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• pp.1232-1236
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• 2016

Mitochondrial calcium uptake 1 (MICU1) including two canonical EF hands, located in the mitochondrial inner membrane, is known to play a crucial role in the calcium uptake in mitochondria. Mitochondrial calcium uptake in muscular cells is related to post mortem shortening by calcium release from muscles. Therefore, the porcine MICU1 gene has been estimated as a genetic candidate for meat quality traits. In this study, variations on the exonic regions of the porcine MICU1 gene were investigated by sequencing cDNAs and tested for their association with meat quality traits. A total of 667 Berkshire heads (347 sows and 320 castrated boars) were used for this association test. Three SNPs were detected on the 3' untranslated region (UTR) of the porcine MICU1 gene. SNP1 (c.*136G>A) was associated with drip loss (p=0.017) and intramuscular fat content (p=0.039). In addition, SNP2 (c.*222G>A) and SNP3 (c.*485G>A) were associated with drip loss (p=0.018) and intramuscular fat content (p<0.001), respectively. In conclusion, it was verified that three variations on the 3' UTR of the porcine MICU1 gene were significantly associated with meat quality traits. It was also suggested that molecular biological analyses are needed to validate the function of variations on the 3 UTR of the porcine MICU1 gene.

• Tuberculosis and Respiratory Diseases
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• v.55 no.1
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• pp.88-97
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• 2003

Background : Although smoking is a major cause of chronic obstructive pulmonary disease (COPD), only 10-20% of cigarette smokers develop symptomatic COPD, which suggests the presence of genetic susceptibility. This genetic susceptibility to COPD might depend on variations in the activities of the enzyme that detoxify hazardous chemical products, such as microsomal epoxide hydrolase (mEPHX) and glutathione-S transferase M1 subunit (GSTM1) genes. Methods : The genotypes of 58 patients with COPD, and 79 age matched control subjects, were determined by a polymerase chain reaction, followed by restriction fragment length polymorphism (PCR-RFLP) for the mEPHX, and multiplex PCR for the GSTM1. Results : GSTM1 was deleted in 53.3% of the subjects. There was no difference in GSTM1 deletion rates between the COPD patients (32/58, 55.2%) and the control subjects (41/79, 51.9%). The combination patterns of two polymorphisms of mEPHX showed slow enzyme activity in 29(21.2%), normal in 73(53.3%) and fast in 32(23.4%). The COPD group (7/57, 12.3%) showed a significantly lower incidence of slow enzyme activity compared to the control subjects (22/77, 28.6%, p<0.05). However, when the COPD and control groups were compared with smokers only, there were no significant differences in the genotypes of GSTM1 and mEPHX. Conclusion : The genotypes of GSTM1 and mEPHX were not significant risk factors of COPD in this cohort of study.

• Journal of Food Hygiene and Safety
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• v.28 no.1
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• pp.50-55
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• 2013

Since 1 June 2012, it is prohibited to sell oilfish as a food material but there are still many illegal cases of selling oilfish as if it is tuna or grilled Patagonian toothfish. So it is absolutely crucial to construct the system to distinguish the real food material from oilfish. There are two sorts of oil fish called Ruvettus pretiosus and Lepidocybirium flavobrunneum involved in Percifomes order and Gempylidae class. 16S DNA gene region in mitochondria was selected to design the specific primers. For design species-specific primer, the theoretical experiment were performed for the sequences of R. pretiosus, L. flavobrunneum, Thunnus thynnus, Thunnus albacores, Makaira mitsukurii and Xiphias gladius, registered at the Gene bank from the National Centre for Biotechnology Information, using BioEdit 7.0.9.0. program. Through the analysis of the result from experiments, it was possible to design the 4 kinds of primers to distinguish R. pretiosus and L. flavobrunneum. As a comparison group, 3 kinds of tuna and 4 kinds of billfishes were selected and experimental verification was performed. As a result, for R. pretiosus and L. flavobrunneum, R.P-16S-006-F/R.P-16S-008-R and L.F-16S-004-F/L.F-16S-006-R primers were selected eventually and PCR condition was established. In addition, 178bp and 238bp of PCR products were confirmed from the established condition and non-specific band was not amplified among similar species. Therefore, the species-specific primers developed in this study would be very useful and used in various ways such as internet shopping mall and illegal distributions with fast and scientific results.

• Journal of Life Science
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• v.23 no.6
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• pp.757-769
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• 2013

A microorganism producing carboxymethylcellulase (CMCase) was isolated from seawater, identified as Bacillus velezensis by analyses of 16S rDNA and partial sequences of the gyrA, and designated as B. velezensis A-68. The optimal conditions for production of CMCase by B. velezensis A-68 were established using response surface methodology (RSM). The optimal concentrations of rice hulls and yeast extract, and initial pH of the medium for cell growth were 60.2 g/l, 7.38 g/l, and 7.18, respectively, whereas those for production of CMCase were 50.0 g/l, 5.00 g/l, and 7.30. The analysis of variance (ANOVA) implied that the most significant factor for cell growth as well as production of CMCase was yeast extract. The optimal concentrations of $K_2HPO_4$, NaCl, $MgSO_4{\cdot}7H_2O$, and $(NH_4)_2SO_4$ in the medium for cell growth were 7.50, 1.00, 0.10, and 0.80 g/l, respectively, which were the same as those for production of CMCase. The optimal temperatures for cell growth and production of CMCase were 30 and $35^{\circ}C$, respectively. The maximal production of CMCase under optimized conditions was 83.8 U/ml, which was 3.3 times higher than that before optimization. In this study, rice hulls, agro-byproduct, were developed as a substrate for production of CMCase and time for production of CMCase was reduced to 3 days using a newly isolated marine bacterium.

• Journal of Plant Biotechnology
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• v.43 no.4
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• pp.450-456
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• 2016

Rhizoctonia leaf blight (large patch) has become a serious problem in Korean lawn grass, which is extremely hard to treat and develops mostly from the roots of lawn grass to wither it away. Rhizoctonia leaf blight (large patch) is caused by Rhizoctonia solani AG2-2 (IV). To develop zoysia japonica with strong disease tolerance against this pathogenic bacterium, ${\beta}-1,3-glucanase$ was cloned from zoysia japonica, which is one of the PR-Proteins known to play a critical role in plant defense reaction. ${\beta}-1,3-glucanase$ is known to be generated within the cells when plant tissues have a hypersensitive reaction due to virus or bacterium infection and secreted outside the cells to play mainly the function of resistance against pathogenic bacteria in the space between the cells. This study utilized the commonly preserved part in the sequence of corn, wheat, barley, and rice which had been researched for their disease tolerance among the ${\beta}-1,3-glucanase$ monocotyledonous plants. Based on the part, degenerate PCR was performed to find out the sequence and full-length cDNA was cloned. E.coli over-expression was conducted in this study to mass purify target protein and implement in vitro activation measurement and antibacterial test. In addition, to interpret the functions of ZjGlu1 gene, each gene-incorporating plant transformation vectors were produced to make lawn grass transformant. Based on ZjGlu1 protein, antibacterial activity test was conducted on 9 strains. As a result, R. cerealis, F. culmorum, R.solani AG-1 (1B), and T. atroviride were found to have antibacterial activity. The gene-specific expression amount in each organ showed no huge difference in the organs based upon the transformant and against 18s gene expression amount.

• Microbiology and Biotechnology Letters
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• v.41 no.2
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• pp.190-197
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• 2013

Three slime-forming lactic acid bacteria were isolated from traditional Korean fermented soy sauce and soybean paste and shown to produce exopolysaccharides (EPS) in sucrose media. By isolating the strains, examining their morphological characteristics and determining their 16S rDNA sequences, N58-5 and K6-7 were identified as Leuconostoc mesenteroides and N45- 10 as Leuconostoc citreum. The acid and bile tolerances of these three strains were investigated. Amongst the three lactic acid bacteria, Leuc. citreum N45-10 exhibited the highest viability ($10^5-10^6$ CFU/ml) in 0.05 M sodium phosphate buffer (pH 0.3) for 2 h, in artificial gastric juice for 2 h and in 0.3%, 0.5% oxgall for 24h. Leuc. mesenteroides K6-7, N58-5 and Leuc. citreum N45- 10 were grown in sucrose liquid medium and 8.16 g/L, 3.65 g/L, 16.17 g/L of EPS was collected, respectively. The hydrolyzed EPS was analyzed by HPLC in order to determine the sugar composition of EPS. Leuc. mesenteroides K6-7 and N58-5 showed two peaks indicating glucose and fructose, thus they were determined to be hetero-type polysaccharides. Leuc. citreum N45-10 showed only the glucose polymer, indicating it to be a homo-type polysaccharide. In addition, all three lactic acid bacterial hemolysis did not demonstrate a clear zone in blood agar in the area surrounding a lactic acid bacteria colony.