• 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.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.28 no.2
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• pp.95-110
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• 2006

The treatment for oral and maxillofacial carcinoma with chemotherapeutic agents is evaluated by many effective methods to reduce the tumor mass and cancer cell proliferation. However these chemotherapy have many serious side effects, such as bone marrow suppression, renal toxicity, G-I troubles. Therefore a possible approach to develop a clinically applicable chemotherapeutic agent is to screen anticancer activity of Taxol which is known to have very little side effect and have been used to breast cancer and ovarian carcinoma. Taxol is a new anti-microtubular anti-cancer agent extracted from the bark of the Pacific yew, Taxus brevifolia. Paclitaxel(Taxol) acts by promoting tubulin polymerization and over stabilizing microtubules agianst depolymerization. Despite the constant improvements of methods of the cancer treatment especially chemotherapy, the rate of cancer metastasis and recurrent are not decreased. Thus the investigation of new drug which have very little side effect and a possible clinically application continues to be a high priority. Considering that the Taxol have shown very effective chemotherapeutic agent with relatively low toxicity in many solid tumors, it deserves to evaluate its efficacy in oral squamous cell carcinoma. In this study, to investigate the in-vivo and in-vitro anti-cancer efficacy of Taxol in oral squamous cell carcinoma and lastly, the potency of Paclitaxel in the clinical application for oral cancer was evaluated. In vivo study, after HN22 cell line were xenografted in nude mice, the growth of tumor mass was observed, 3 mg/Kg taxol was injected intraperitoneally into nude mice containing tumor mass. The methods of these study were measurement of total volume of tumor mass, histopathologic study, immunohistochemical study, drug resistance assay, growth curve, MTT assay, flow cytometry, cDNA microarray in vivo and in vitro. The results were obtained as following. 1. The visual inspection of the experimental group showed that the volume of the tumor mass was slightly decreased but no significant difference with control group. 2. Ki-67 index was decreased at weeks 4 in experimental group. 3. Microscopic view of the xenografted tumor mass showed well differentiated squamous cell carcinoma and after Taxol injection, some necrotic tissue was seen weeks 4. 4. The growth curve of the tumor cells were decreased after 1day Taxol treatment. 5. According to the MTT assay, HN22 cell line showed relative drug resistancy above $5\;{\mu}g/ml$ concentrations of Taxol. 6. In drug resistance assay, the decrease of cell counts was seen relatively according to concentration. 7. In Flow cytometry, G2M phase cell arrests were seen in low concentration of the Taxol, while S phase cell arrests were seen in high concentration of the Taxol. 8. Using cDNA microarray technique, variable gene expression of ANGPTL4, TXNRD1, FAS, RRAGA, CTGF, CYCLINEA, P19, DUSP5, CEBPG, BTG1 were detacted in the oral squamous cell carcinoma cell after taxol treatment. In this study paclitaxel is effective against oral squamous cell carcinoma cell lines in vitro, but week effect was observed in vivo. So we need continuous study about anticancer effect of taxol in vivo in oral squamous cell carcinoma.

• Tuberculosis and Respiratory Diseases
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• v.50 no.6
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• pp.668-675
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• 2001

Background : Non-smalll lung cancer(NSCLC) develops as a result of the accumulation of multiple genetic abnormalities. Loss of heterozygosity(LOH) is one of the most frequent genetic alterations that is found in NSCLC, and the chromosomal regions that display a high rate of LOH are thought to harbor tumor suppressor genes(TSGs). This study was done to determine the frequency of LOH in 21q with the aim of identifying potential TSG loci. Method : Thirty-nine surgically resected NSCLCs were analysed. Patients peripheral lymphocytes were used as the source of the normal DNA. Five microsatellite Inarkers of 21q were used to study LOH : 21q21.1(D21S1432, and D21S1994); 21q21.2-21.3(D21S1442) ; 21q22.1(21S1445) ; and 21q22.2-22.3(D21S266). The fractional allelic loss(FAL) in a tumor was calculated as the ratio of the number of markers showing LOH to the number of informative markers. Result : LOH for at least one locus was detected in 21 of 39 tumors(53.8%). Among the 21 tumors with LOH, 5(21.8%) showed LOH at almost all informative loci. Although statistically not significant, LOH was found more frequently in squamous cell carcinomas(15 of 23, 65.2%) than in adenocarcinomas(6 of 16, 37.5%). In the squamous cell carcinomas the frequency of LOH was higher in stage II-III (80.0%) than in stage I (53.8%). The FAL value in squamous cell carcinomas($0.431{\pm}0.375$) was significantly higher than that found in adenocarcinomas($0.l92{\pm}0.276$). Conclusion : These results suggest that LOH on 21q may be involved in the development of NSCLC, and that TSG(s) that contribute to the pathogenesis of NSCLC may exist on 21q.

• Tuberculosis and Respiratory Diseases
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• v.49 no.4
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• pp.453-465
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• 2000

Background : Lung carcinogenesis is a multistage process involving alterations in multiple genes and diverse pathway. Mutational activation of oncogenes and inactivation of tumor suppressor genes, and subsequent increased genetic instability are the major genetic events. The p53 gene and FHIT gene as tumor suppressor genes contribute to the pathogenesis of lung cancer, evidenced by mutation, microsatellite instability(MI) and loss of heterozygosity(LOH). Methods : We analysed genetic mutations of p53 and FHIT gene in 29 surgical specimens of nonsmall cell lung cancer using PCR-single strand conformation polymorphism, DNA sequencing and RT-PCR. MI and LOH were analyzed in loci of D3S1285, D9S171, and TP53. Results : In 2 cases, point mutation of p53 gene was observed on exon 5. MI of 3 times and LOH of 14 times were observed in at least one locus. In terms of the location of microsatellite, D3S1285 as a marker of FH1T was observed in 5 cases out of 26 specimens; D9S171 as a marker of p16 in 5 out of 17; and TP53 as a marker of p53 in 7 out of 27. In view of histologic type, squamous cell carcinoma presented higher frequency of microsatellite alteration, compared to others. Mutation of FHIT gene was observed in 11 cases and 6 cases of those were point mutation as a silent substitution on exon 8. FHIT mRNA expression exhibited deletion on exon 6 to 9 in 4 cases among 15 specimens, presenting beta-actin normally. Conclusion : Our results show comparable frequency of genetic alteration in nonsmall cell lung cancer to previous studies of Western countries. Microsatellite analysis might have a role as a tumor marker especially in squamous cell carcinoma. Understanding molecular abnormalities involved in the pathogenesis could potentially lead to prevention, earlier diagnosis and the development of novel investigational approaches to the treatment of lung cancer.

• Nuclear Medicine and Molecular Imaging
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• v.43 no.5
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• pp.468-477
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• 2009

Purpose: Hydrodynamic-based procedure is a simple and effective gene delivery method to lead a high gene expression in liver tissue. Non-invasive imaging reporter gene system has been used widely with herpes simplex virus type 1 thymidine kinase (HSV1-tk) and its various substrates. In the present study, we investigated to image the expression of HSV1-tk gene with 5-(2-iodovinyD-2'-deoxyuridine (IVDU) in mouse liver by the hydrodynamicbased procedure. Materials and Methods: HSV1-tk or enhanced green fluorescence protein (EGFP) encoded plasmid DNA was transferred into the mouse liver by hydrodynaminc injection. At 24 h post-injection, RT-PCR, biodistribution, fluorescence imaging, nuclear imaging and digital wholebody autoradiography (DWBA) were performed to confirm transferred gene expression. Results: In RT-PCR assay using mRNA from the mouse liver, specific bands of HSV1-tk and EGFP gene were observed in HSV1-tk and EGFP expressing plasmid injected mouse, respectively. Higher uptake of radiolabeled IVDU was exhibited in liver of HSV1-tk gene transferred mouse by biodistribution study. In fluorescence imaging, the liver showed specific fluorescence signal in EGFP gene transferred mouse. Gamma-camera image and DWBA results showed that radiolabeled IVDU was accumulated in the liver of HSV1-tk gene transferred mouse. Conclusion: In this study, hydrodynamic-based procedure was effective in liver-specific gene delivery and it could be quantified with molecular imaging methods. Therefore, co-expression of HSV1-tk reporter gene and target gene by hydrodynamic-based procedure is expected to be a useful method for the evaluation of the target gene expression level with radiolabeled IVDU.

• Journal of Embryo Transfer
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• v.21 no.1
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• pp.13-20
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• 2006

The purpose of this study was to develope the transgenic cattle expressing hFSH into the urine using the nuclear transfer. To produce the interest gene in urine, the specific vector was ligated with hFSH gene undo. maUII promoter. The fetal fibroblast cells (KbFF) were isolated from a 45-day male fetus. The hFSH gene was co-transfected with pcDNA3 (neo) vector to KbFF cells by electroporation. The gene-transfected cells were cultured with G-418 selection medium for 2 weeks. Selected colonies were confirmed by PCR. For nuclear transfer, enucleated bovine oocytes were transferred with hFSH transfected or nontransfected fetal fibroblasts. The cleavage and blastocyst formation rates were significantly lower (p<0.05) in cloned embryos transfected with hFSH gene (68.7% and 15.7%) than in those non-transfected (67.6% and 24.5 %), respectively. Apoptosis analysis showed no difference between hFSH transfected and non-transfected blastocysts (p>0.05). The blastocysts were transfected to 77 (control 24, hFSH 53) recipient cows. Two calves were born (1.9%) following transfer with NT embryos transfected with hFSH gene, but they were confirmed not to be transgenic calves. This result shows that the hFSH colonies were mixed with transfected and non transfected cells. Further research will be needed for selection and establishment of gene transfected cells.

• Journal of Animal Science and Technology
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• v.45 no.5
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• pp.703-710
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• 2003

The purpose of this study was to detect association between genetic variation and economic trait in the porcine heart type fatty acid-binding protein gene as a candidate gene for the traits related with growth and meat quality in pigs. The H-FABP is a 15-kDa protein expressed in several tissues with high demand for fat metabolism such as cardiac and skeletal muscle and lactating mammary gland. H-FABP is small intracellular protein involved in fatty acid transport from the plasma membrane to the site of $\beta$-oxidation and/or triacylglycerol or phospholipid synthesis. In this study, H-FABP PCR-RFLP was performed in F$_2$ population composed of 214 individuals from an intercross between Korean Native Boars and Landrace sows. PCR products from two primer sets within H-FABP gene were amplified in 850bp and 700bp. Digestion of PCR products with the restriction digestion enzymes HaeⅢ and HinfⅠ, revealed fragment length polymorphisms(RFLPs). The genotype frequencies from H-FABP/HaeⅢ was .29 for genotype DD, .53 for genotype Dd, and .15 for genotype dd, respectively. The genotype frequencies of HH, Hh, and hh from H-FABP/HinfⅠ was .38, .41 and .20, respectively, in the population. Relationships between their genotypes and economic traits were estimated. In H-FABP/HaeⅢ locus, there were specific genotypes(Dd and dd) associated with economic traits such as body weights at 3, 5, 12, and 30 week of age (p〈.05 to .001). The ‘d’ allele was associated with gaining of body weight. In H-FABP/HinfⅠ locus, Genotypes of HH and Hh associated with growth traits such as body weights at 5, 12, and 30 week of age (p〈.05 or p〈.001) and back fat thickness, body fat including abdominal and trimmed fat (p〈.001) and intramuscular fat(p〈.05) The ‘H’ allele was positively associated with gaining of body weight and fatness deposition. In conclusion, a significant association of the H-FABP gene from its genetic variation was found on body weight, intramuscular fat and backfat thickness.

• 한국균학회소식:학술대회논문집
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• 2014.10a
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• pp.9-9
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• 2014

Null mutants generated by targeted gene replacement are frequently used to reveal function of the genes in fungi. However, targeted gene deletions may be difficult to obtain or it may not be applicable, such as in the case of redundant or lethal genes. Constitutive expression system could be an alternative to avoid these difficulties and to provide new platform in fungal functional genomics research. Here we developed a novel platform for functional analysis genes in Magnaporthe oryzae by constitutive expression under a strong promoter. Employing a binary vector (pGOF1), carrying $EF1{\beta}$ promoter, we generated a total of 4,432 transformants by Agrobacterium tumefaciens-mediated transformation. We have analyzed a subset of 54 transformants that have the vector inserted in the promoter region of individual genes, at distances ranging from 44 to 1,479 bp. These transformants showed increased transcript levels of the genes that are found immediately adjacent to the vector, compared to those of wild type. Ten transformants showed higher levels of expression relative to the wild type not only in mycelial stage but also during infection-related development. Two transformants that T-DNA was inserted in the promotor regions of putative lethal genes, MoRPT4 and MoDBP5, showed decreased conidiation and pathogenicity, respectively. We also characterized two transformants that T-DNA was inserted in functionally redundant genes encoding alpha-glucosidase and alpha-mannosidase. These transformants also showed decreased mycelial growth and pathogenicity, implying successful application of this platform in functional analysis of the genes. Our data also demonstrated that comparative phenotypic analysis under over-expression and suppression of gene expression could prove a highly efficient system for functional analysis of the genes. Our over-expressed transformants library would be a valuable resource for functional characterization of the redundant or lethal genes in M. oryzae and this system may be applicable in other fungi.

• Microbiology and Biotechnology Letters
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• v.26 no.2
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• pp.122-129
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• 1998

It were reported that antifungal mechanism of Enterobacter cloacae is a volatile ammonia that produced by the strain in soil, and the production of ammonia is related to the bacterial urease activity. A powerful bacterium SH14 against soil-borne pathogen Fusarium solani, which cause root rot of many important crops, was selected from a ginseng pathogen suppressive soil. The strain SH14 was identified as Bacillus subtilis by cultural, biochemical, morphological method, and $API^{circledR}$ test. From several in vitro tests, the antifungal substance that is produced from B. subtilis SH14 was revealed as heat-stable and low-molecular weight antibiotic substance. In order to construct the multifunctional biocontrol agent, the urease gene of Bacillus pasteurii which can produce pathogenes-suppressive ammonia transferred into antifungal bacterium. First, a partial BamH I digestion fragment of plasmid pBU11 containing the alkalophilic B. pasteurii l1859 urease gene was inserted into the BamH I site of pEB203 and expressed in Escherichia coli JM109. The recombinant plasmid was designated as pGU366. The plasmid pGU366 containing urease gene was introduced into the B. subtilis SH14 with PEG-induced protoplast transformation (PIP) method. The urease gene was very stably expressed in the transformant of B. subtilis SH14. Also, the optimal conditions for transformation were established and the highest transformation frequency was obtained by treatment of lysozyme for 90 min, and then addition of 1.5 ${mu}g$/ml DNA and 40% PEG4000. From the in vitro antifungal test against F. solani, antifungal activity of B. subtilis SH14(pGu366) containing urease gene was much higher than that of the host strain. Genetical development of B. subtilis SH14 by transfer of urease gene can be responsible for enhanced biocontrol efficacy with its antibiotic action.

• Korean journal of applied entomology
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• v.55 no.4
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• pp.445-452
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• 2016

Exorista japonica is one of the major natural enemies of noctuid larvae, Mythimna separata and Spodoptera litura. The examined parasitoid was obtained from host species M. separata, collected at Gimje city and identified by DNA sequences (partial cytochrome oxidase I, 16S, 18S, and 28S). For purposed of this study, laboratory reared S. litura served as the host species for the development of the E. japonica. The developmental period of E. japonica immature stages were investigated at seven constant temperatures (16, 19, 22, 25, 28, 31, $34{\pm}1^{\circ}C$, RH 20~30%). Temperature-dependent developmental rates and development completion models were developed. E. japonica was successfully developed from egg to adult in $16{\sim}31^{\circ}C$ temperature regimes. Developmental duration was the shortest at $34^{\circ}C$ (8.3 days) and the longest at $16^{\circ}C$ (23.4 days) from egg to pupa development. Pupal development duration was the shortest at $28^{\circ}C$ (7.3 days). Total immature-stage development duration decreased with increasing temperature, and was the shortest at $31^{\circ}C$ (16.3 days) and the longest at $16^{\circ}C$ (45.4 days). The lower developmental threshold was $7.8^{\circ}C$ and thermal constant required to complete total immature-stage development was 370.4 degree days. Among four non-linear temperature-dependent developmental rate models, Briere 1 model had the highest adjusted R-squared (0.96). The distribution model of development completion for total immature stage development of E. japonica was well described by all model ($r^2_{adj}=0.90$) based on the standardized development duration. These results of study would be necessary not only to develop population dynamics model but also to understand fundamental biology of E. japonica.