• Journal of Microbiology
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• v.44 no.1
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• pp.64-71
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• 2006

To investigate the effects of the nucleotide sequences in Shine-Dalgarno (SD) and the spacer region (SD-ATG) on bovine growth hormone (bGH) gene expression, the expression vectors under the control of the T7 promoter (pT7-7 vector) were constructed using bGH derivatives (bGH1 & bGH14) which have different 5'-coding regions and were induced in E. coli BL21 (DE3). Oligonucleotides containing random SD sequences and a spacer region were chemically synthesized and the distance between the SD region and the initiation codon were fixed to nine bases in length. The oligonucleotides were annealed and fused to the bGH1 and bGH14 cDNA, respectively. When the bGH gene was induced with IPTG in E. coli BL21(DE3), some clones containing only bGH14 cDNA produced considerable levels of bGH in the range of $6.9\%\;to\;8.5\%$ of total cell proteins by SDS-PAGE and Western blot. Otherwise, the bGH was not detected in any clones with bGH1 cDNA. Accordingly, the nucleotide sequences of SD and the spacer region affect on bGH expression indicates that the sequences sufficiently destabilize the mRNA secondary structure of the bGH14 gene. When the free energy was calculated from the transcription initiation site to the +51 nucleotide of bGH cDNA using a program of nucleic acid folding and hybridization prediction, the constructs with values below -26.3 kcal/mole (toward minus direction) were not expressed. The constructs with the original sequence of bGH cDNA also did not show any expression, regardless of the free energy values. Thus, the disruption of the mRNA secondary structure may be a major factor regulating bGH expression in the translation initiation process. Accordingly, the first stem-loop among two secondary structures present in the 5'-end region of the bGH gene should be disrupted for the effective expression of bGH.

• Korean Journal of Microbiology
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• v.34 no.3
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• pp.169-174
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• 1998

To define the structure and diversity of bacterial communities in the aqutic ecosystem, Lake Soyang, the largest artificial reservoir in Korea, a new method, fluorescent in situ hybridization was applied. This technique relies on the specific hybridization of the nucleic acid probes to the naturally amplified intracellular rRNA. By this method, the bacterial community composition of Lake Soyang and bacterial numbers belong to eubacteria, proteobacteria and Cytophaga-Flavobacterium group were estimated. Total bacterial numbers ranged from $0.3{\times}10^6{\sim}2.0{\times}10^6cells{\cdot}ml^{-1}$, and vertical profile of total bacteria showed the peak at 2 and 5 m depths. The ratio of eubacteria to total bacteria were 22~100% and varied with depth and season. The percentage of Proteobacteria ${\alpha}$-group ranged 2.6~66.7%, ${\beta}$-group 4.5~53.5%, ${\gamma}$-group 4.6~76.7% and Cytophaga-Flavobacterium group 2.1~35.9%. Also, bacteria] community had spatial and temporal characteristics. The dominant groups were ${\beta}$-group in winter, ${\gamma}$-group in spring and early summer and ${\alpha}$-group in summer.

• Journal of Plant Biotechnology
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• v.35 no.2
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• pp.133-140
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• 2008

The modification of DNA and histone plays an important role for gene expression in plant development. The objective of this research is to observe the effects of methylation on the gene expression during dedifferentiation from rice mature seeds to callus and differentiation from callus to shoots. The embryogenic callus with ability to shoot regeneration was not induced on the N6A medium supplemented with 5-azacytidine and abnormal callus with brown color was formed. When the normal rice callus was placed on the regeneration MSRA medium supplemented with 5-azacytidine, the shoot regeneration was inhibited. The results showed that 5-azacytidine, DNA demethylating agent, had negative effects on normal embryogenic callus formation and shoot regeneration. This suggested that DNA methylation of some genes was required for normal cell dedifferentiation and differentiation in tissue culture. The microarray and $GeneFishig^{TM}$ DEG screening were used to observe the gene transcript profile in callus induction and regeneration on N6A (N6 medium + 5-azaC) and MSRA (MS regeneration medium + 5-azaC). Subsets of genes were up-regulated or down-regulated in response to 5-azaC treatments. The genes related with epigenetic regulation, electron transport, nucleic acid metabolism and response to stress were up and down regulated. The different expression of some genes (germin like protein etc.) during callus induction and shoot regeneration was confirmed using RT-PCR and northern blot analysis.

• The Korean Journal of Cytopathology
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• v.7 no.1
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• pp.1-11
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• 1996

From the concepts of cellular pathology and of exfoliative cytology, as elucidated by Virchow and Papanicolaou respectively in the late 19th and early 20th century, have evolved the primary methods for the diagnosis of cancer today. From Papanicolaou's concept of exfoliative cytology developed fine needle aspiration biopsy in the early 1960's, this has become a major diagnostic procedure and has contributed to a significant reduction in open biopsies and, therefore, to medical cost-effectiveness immunobiochemical techniques provided us with a supplement to cancer diagnosis in the 1980's. The immunoperoxidase method, using monoclonal antibodies, is applied primarily as an ancillary measure to elucidate the nature of cancers The availability of specific monoclonal antibodies has greatly facilitated the identification of cell products or surface markers. For example, antibodies directed against intermediate filaments have proved to be of value in determining the histogenesis oi poorly differentiated neoplasms. Tumor markers may serve as biochemical indicators of the presence of a neoplasm. They can be detected In plasma and other body fluids. Their concentration can be applied as a diagnostic test, for monitoring the clinical course of known cancer, and as a screening measure to detect certain cancers in a population at risk. Flow cytometry is a useful tool for distinguishing several cell characteristics, such as the immunophenotype of leukemia-lymphoma cells, the DNA content of neoplastic cells, and cell proliferation rate. Molecular biologic techniques provided a giant step for the management of cancer patients encompassing diagnosis, prognostic evaluation, and therapy. Nucleic acid hybridization techniques are utilized as Southern, Northern, and dot blots and in situ hybridization. Molecular biology and its techniques may bring a blight new horizon for understanding cancer biology and in designing therapy on the basis of gene manipulation.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.10
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• pp.1404-1410
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• 2008

This study was performed to clarify whether the variation of stress related heat shock protein 70 (HSP70) (GenBank X68213) gene was associated with the nuclear morphological change of in vitro maturation and in vitro capacitation in oocytes of pig ovaries obtained at the slaughterhouse. The nucleic acid substitution of C to G at the 483rd position was found out in HSP70 K1 (290-512) from X68213. The ovaries were categorized into CC, CG, and GG genotypes using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) (BsiHKA I). After the second in vitro maturation of immature fresh oocytes, the relation of nuclear morphological change in oocytes with the genotype of HSP70 K1 gene was such that the MII ratios of the genotype GG and CG (46.93% and 42.20%, respectively) were significantly higher than that of the CC genotype (10.71%) (p<0.05). With respect to in vitro maturation of frozen-thawed oocytes by an open pulled straw (OPS) method, the percentage of oocytes matured to MII stage of the CG genotype showed a higher trend than CC and GG genotypes. After the in vitro maturation of immature fresh oocytes and frozen-thawed oocytes by the OPS method, the relation of the pronuclei change in oocytes matured in vitro with HSP70 genotype was assessed, and the result showed that the enlarged sperm heads (ESH) of matured fresh oocytes and frozen-thawed oocytes were 80.0% and 60.0% in the CC genotype, respectively. The CC genotype group had a significantly higher rate of ESH than the CG and the GG genotype group (p<0.05). The ratios of polyspermic invasion were not different among HSP70 of the three genotypes. It was considered that the rate of in vitro maturation of fertilized oocytes was expected to differ according to genotype of the stress related gene.

• Applied Biological Chemistry
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• v.38 no.1
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• pp.49-54
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• 1995

To understand the molecular structure and pathogenesis mechanism of Korean garlic viruses, we have isolated cDNA clones for garlic viruses. The partial nucleotide sequences of 24 cDNA clones were determined and those of five clones containing poly(A) tail were compared with sequences of other plant viruses. One of these clones, V9, has a primary structure similar to the carlavirus group, suggesting that the clone V9 derived from a part of garlic latent virus (GLV). Northern blot analysis with the clone V9 as a probe demonstrated that GLV genome is 8.5 knt long and has a poly(A) tail. The clone V9 encodes coat protein (CP) of 33 kDa and nucleic acid binding protein of 10 kDa in different reading frame. The hexanucleotide motif, 5'-ACCUAA, which is conserved in the 3' noncoding region arid was proposed to be a cis-acting element involved in the production of negative strand genomic RNA was noticed. Complementary sequence to the hexanucleotide motif, 5'-TTAGGT, is also found in the positive strand of V9 RNA. The putative CP gene was cloned into the pRSET-A expression vector and expressed in E. coli BL21. The expressed recombinant V9CP protein was purified by $Ni^{2+}$ NTA affinity chromatography. The anti-V9CP antibody recognizes 34 kDa polypeptide which could be CP of GLV in infected garlic leaf extract. Immunoblot and Northern blot analysis of various cultivars shows wide occurrence of GLV in Korean garlic plants.

• Korean Journal of Microbiology
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• v.24 no.4
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• pp.352-356
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• 1986

A study was carried out to determine the effect of ginseng saponin an its related materials on aflatoxin production by Aspergillus parasiticus NRRL2999 in glucose-salts(GS) medium. Maximal growth of the mold and AF froduction in the medium occurred after 5 and 9 days at $28^{\circ}C$, respectively. When various concentrations of saponin added to the medium aflatoxin synthesis were significantly reduced (p<0.05) compared to the control after 9 days at $28^{\circ}C$. 0.05% of saponin inhibited aflatoxin production most effectively in the low concerntrations of saponin (0.01-0.2%) and the toxin synthesis reduced with an increasing concentrations of saponin in the high concentrations (0.03-5.0%). Various concentrations (0.01-1.0%) of saponin diol and triol in the media also caused to reduce aflatoxin synthesis by the mold (p<0.05). All saponin fractions were found to decrease aflatoxin production significantly. Saponin fraction numbers of 1,2,4 and 6 were shown to reduce aflatoxin production effectively, and the number 1 was the most effective. Addition of 0.05% of nucleic acid related materials to the medium reduced aflatoxin production (p<0.05). Aflatoxins could not be found in broth at all, but in mycelia when 0.05% of caffeine was added to the medium. Aflatoxin synthesis was well correlated with total lipid synthesis, growth and glucose uptake. When aflatoxin synthesis inhibited (5.0% of saponin) both total lipid synthesis and growth were stimulated and the efficiency of glucose utilization was reduced.

• 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.

• The Korean Journal of Food And Nutrition
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• v.17 no.3
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• pp.254-259
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• 2004

Rapid detection of foodbome pathogens is becoming increasingly important. The requirement for faster, more reliable tests has lead to the development of a wide range of rapid methods. Among these methods, the use of systems based on nucleic acid based detection has been increasing since they offer advantages of reduction in test time and more reliable detection or identification. Random Amplification Polymorphic DNA(RAPD) method has been used to fingerprint foodbome microorganisms; Listeria monocytogenes. In this study, 10-mer primer OPG-13(5'-CTCTCCGCCA-3') was used to generate RAPD-PCR for detection of pathogenic L. monocytogenes of Listeria spp. Among 20 primers tested, OPG-13 showed on acceptable result for the differentiation of a pathogenic Listeria from non-pathogenic microorganisms. Pathogenic Listeria, L. monocytogenes(ATCC 15313, 19111, 19112, 19113) showed two bands for 700 bp and 1,500 bp while non-pathogenic bacteria, L. ivanovii, L. grayi, L. murrayi, L. innocua, L. welshimeri, and L. seeligeri had only one band sizing from 2,000 to 2,300 bp. This RAPD method proved to be a valuable to gain important information on sources of pathogenic bacteria in food industry.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.129-129
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• 2017

The first key point to the successful pollination and fertilization in plants is the pollen pistil interaction, referring to the cellular and molecular levels, which mainly play active roles in limiting gene flow among maize populations and between maize and teosinte. This study was carried out to identify proteins and investigate the mechanism of gametophytic factors using protein analysis. W22 (ga1); which didn't carry a gametophytic factor and W22 (Ga1), a near iso-genic line were used for the proteome investigation. SDS-PAGE was executed to investigate proteins in the pollen and pistil of W22 (ga1) and W22 (Ga1). A total of 44 differentially expressed proteins were identified in the pollen and pistil on SDS-PAGE using LTQ-FTICR MS. Among the 44 proteins, a total of 24 proteins were identified in the pollen of W22 (ga1) and W22 (Ga1) whereas 20 differentially expressed proteins were identified from the pistil of W22 (Ga) and W22 (Ga1). However, in pollen, 2 proteins were identified only in the W22 (ga1) and 12 proteins only in the W22 (Ga1) whereas 10 proteins were confirmed from the both of W22 (ga1) and W22 (Ga1). In contrary, 10 proteins were appeared only in the pistil of W22 (ga1) and 7 proteins from W22 (Ga1) while 3 proteins confirmed in the both of W22 (ga1) and W22 (Ga1). Moreover, the identified proteins were generally involved in hydrolase activity, nucleic acid binding and nucleotide binding. These results help to reveal the mechanism of gametophytic factors and provide a valuable clue for the pollen and pistil research in maize. In addition, it might provide a comprehensive insight on the proteins that were involved in the regulation of pollen-pistil interaction.