• Journal of Microbiology and Biotechnology
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• v.18 no.6
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• pp.1164-1169
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• 2008

We developed multiplex RT-PCR assays that can detect and identify 12 hemagglutinin (H1-H12) and 9 neuraminidase (N1-N9) subtypes that are commonly isolated from avian, swine, and human influenza A viruses. RT-PCR products with unique sizes characteristic of each subtype were amplified by multiplex RT-PCRs, and sequence analysis of each amplicon was demonstrated to be specific for each subtype with 24 reference viruses. The specificity was demonstrated further with DNA or cDNA templates from 7 viruses, 5 bacteria, and 50 influenza A virus-negative specimens. Furthermore, the assays could detect and subtype up to $10^5$ dilution of each of the reference viruses that had an original infectivity titer of $10^6\;EID_{50}/ml$. Of 188 virus isolates, the multiplex RT-PCR results agreed completely with individual RT-PCR subtyping results and with results obtained from virus isolations. Furthermore, the multiplex RT-PCR methods efficiently detected mixed infections with at least two different subtypes of influenza viruses in one host. Therefore, these methods could facilitate rapid and accurate subtyping of influenza A viruses directly from field specimens.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2000.04a
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• pp.178-181
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• 2000

In order to develop the novel gene expression system, we introduced new control elements which could influence the foreign gene expression in animal cells. When the foreign genes are introduced into the genome of higher eukaryotic cells, the expressions from these integrated genes are often low and can vary greatly depending on the positions of the integration sites due to the complex nature of the chromatin structures (1). First we screened the various DNA sequence elements which can function as an insulator of gene expression from these position effects and can cooperate with the SV40 enhancer/promoter. Among the several DNA elements from the various sources, we identified the particular DNA element which confers the increased frequency of the positive colonies, assayed by the reporter gene from stable selections indicating significantly reduced position effects. This element also showed the several fold-increased expression level as well as the copy-number dependent expression with host cell specificity. Second we modified the transcription termination element where we introduced the specific terminator in combination with SV40 polyA signal. This modified terminator showed the increased efficiency and the level of the gene expression. By combining these two elements, we made the animal cell expression system and tested successfully for the recombinant protein productions of TGF ${\beta}$-soluble receptor, Antithrombin III, and single chain Pro-Urokinase. [Supported by grants from MOCIE]

• Current Research on Agriculture and Life Sciences
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• v.15
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• pp.25-32
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• 1997

This study was carried out to research the biological characteristics among rhizobia and soybean seed and root lectins, and to obtain a basic imformation of host specificity in biological nitrogen symbiosis system. The results obtained were as follows: Purified seed lectin from soybean varieties of paldal, backwoon and hwangkeum formed immunoprecipitin lines with standard soybean seed lectin and the root lectins from soybean seedlings immunoelectrophoretically. Soybean seed and root lectins interacted with Rhizobium japonicum and Bradyrhizobium japonicum, but didn't interacted with Rhizobium. viceae, whereas pea lectin conjugated with R. viceae, but didn't bind with R. japonicum and B. japonicum. Lipopolysaccharides of B. japonicum and R. viceae were fractionated into LPS I and LPS II on the sephadox G-50. Lipopolysaccharides from B. japonicum showed rhe binding acitivity both with soybean seed lectin and root lectin, but those from R. viceae didn't show it with soybean seed and root lectins.

• The Korean Journal of Mycology
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• v.33 no.2
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• pp.95-97
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• 2005

A rust disease occurred on Veratrum patulum in Deogyusan National Park, Jeollabukdo. The typical symptoms of the disease appeared as small, yellowish spots on leaves at first. The spots then turned brown, their edges rised slightly, and powdery mass of yellow or yellowish brown spores appeared on the lesions. Severely infected leaves blighted and eventually defoliated. Urediniospores were yellowish brown in color, globose, subglobose or ellipsoid in shape and $19{\sim}27\;{\times}\;17{\sim}24\;{\mu}m$ in size. Teliospores were brown in color, ellipsoid, ovate or oblong in shape and $19{\sim}36\;{\times}\;15{\sim}21\;{\mu}m$ in size, the apex mostly rounded, with hyaline or subhyaline conical papilla, the base rounded or attenuated, smooth. Pedicels hyaline or subhyaline, up to $38\;{\mu}m$ long. The causal fungus was identified as Uromyces veratri Schroeter based on mycological characteristics and host specificity.

• Parasites, Hosts and Diseases
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• v.32 no.4
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• pp.231-242
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• 1994

Pnragonimus westermnni, the lung fluke, is known to migrate to the pulmonary tissue of mammalian hosts and causes pathological changes in the lungs. An acidic thiol-dependent proteinase with a molecular weight of approximately 20,000 daltons was purified to homogeneity using ion-exchange chromatography and gel filtration chromatography. On SDS-PAGE, the molecular weight of the enzyme was 17,500 daltons. Isoelectric point was 6.45. The enzyme was similar to the acidic cysteine proteinase of vertebrates in the properties of pH optimum, substrate specificity and inhibitor sensitivity. Enzymatic activity was stable at pH 5.5 for at least two days when stored at 4℃. The cysteine proteinase was capable of degrading collagen and hemoglobin. Sera of patients with paragonimiasis and mice infected with R westermani reacted in immunoblots with the partially purified proteinase. This result suggested that the cysteine proteinase of P. westermnni may play a role in migration in tissues, and in acquisition of nutrients by parasites from the host. It is also potentially an antigen for the serodiagnosis of paragonimiasis.

• Parasites, Hosts and Diseases
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• v.51 no.5
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• pp.503-510
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• 2013

Toxoplasma gondii is an apicomplexan parasite with a broad host range of most warm-blooded mammals including humans, of which one-thirds of the human population has been infected worldwide which can cause congenital defects, abortion, and neonatal complications. Here, we developed a rapid diagnostic test (RDT) for T. gondii infection. Antigenic N-terminal half of the major surface antigen (SAG1) was linked with intrinsically unstructured domain (IUD) of dense granule protein 2 (GRA2). The recombinant GST-GRA2-SAG1A protein was successfully expressed and purified as 51 kDa of molecular weight. Furthermore, antigenicity and solubility of the rGST-GRA2-SAG1A protein were significantly increased. The overall specificity and sensitivity of GST-GRA2-SAG1A loaded RDT (TgRDT) were estimated as 100% and 97.1% by comparing with ELISA result which uses T. gondii whole cell lysates as the antigen. The TgRDT tested with Uganda people sera for field trial and showed 31.9% of seroprevalence against T. gondii antibody. The TgRDT is proved to be a kit for rapid and easy to use with high accuracy, which would be a suitable serodiagnostic tool for toxoplasmosis.

• Journal of Microbiology
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• v.43 no.3
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• pp.219-227
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• 2005

Little is known about the bacterial communities associated with the plants inhabiting sand dune ecosystems. In this study, the bacterial populations associated with two major sand dune plant species, Calystegia soldanella (beach morning glory) and Elymus mollis (wild rye), growing along the costal areas in Tae-An, Chungnam Province, were analyzed using a culture-dependent approach. A total of 212 bacteria were isolated from the root and rhizosphere samples of the two plants, and subjected to further analysis. Based on the analysis of the 16S rDNA sequences, all the bacterial isolates were classified into six major phyla of the domain Bacteria. Significant differences were observed between the two plant species, and also between the rhizospheric and root endophytic communities. The isolates from the rhizosphere of the two plant species were assigned to 27 different established genera, and the root endophytic bacteria were assigned to 21. Members of the phylum Gammaproteobacteria, notably the Pseudomonas species, comprised the majority of both the rhizospheric and endophytic bacteria, followed by members of Bacteroidetes and Firmicutes in the rhizosphere and Alphaproteobacteria and Bacteroidetes in the root. A number of isolates were recognized as potentially novel bacterial taxa. Fifteen out of 27 bacterial genera were commonly found in the rhizosphere of both plants, which was comparable to 3 out of 21 common genera in the root, implying the host specificity for endophytic populations. This study of the diversity of culturable rhizospheric and endophytic bacteria has provided the basis for further investigation aimed at the selection of microbes for the facilitation of plant growth.

• 한국생물공학회:학술대회논문집
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• 2002.04a
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• pp.457-460
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• 2002

In an effort to isolate novel strains expressing a thermostable esterase that hydrolyzed the rac-ketoprofen ethyl ester to ketoprofen in the stereospecific manner, we screened various soils and composts from broad ecological niches in which the activity was expected to be found. Three hundreds of microbial strains were tested to determine their ester-hydrolyzing activity by using an agar plate containing insoluble tributyrin as an indicative substrate, and then further screened by activity on the (R,S)-ketoprofen ethyl ester. Twenty-six strains were screened primarily at high growth and incubation temperature and further compared the ability to ethyl ester-hydrolyzing activity in terms of conversion yield and chiral specificity. Consequently, a strain JYl44 was isolated as a novel strain that produced a (R)-stereospecific esterase with high stability and systematically identified as a Bacillus stearothermophilus JY144. The enzyme indeed stables at a broad range of temperature, upto 65 $^{\circ}C$, and pH ranging from 6.0 to 10.0. The optimal temperature and pH for enzymatic conversion were 50 $^{\circ}C$ and 9.0, respectively. Based on the observations that resulted a poor cell growth, and enzyme expression in wild type strain, we further attempted the gene cloning into a general host Escherichia coli and determined its primary structure, concomitantly resulting a high level expression of the enzyme. The cloned gene had an open reading frame (250 amino acids) with a calculated molecular mass of 27.4 kDa, and its primary structure showed a relative high homology (45-52 %) to the esterases from Streptomyces and Bacillus strains. The recombinant whole cell enzyme could efficiently convert the rac-ketoprofen ethyl ester to (R)-ketoprofen, with optical purity of 99 % and yield of 49 %.

• Journal of Microbiology and Biotechnology
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• v.14 no.4
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• pp.885-890
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• 2004

A survey for vesicular-arbuscular mycorrhizae (VAM) occurrence was undertaken in three endangered vegetation sites in the area of Kudankulam atomic power station. Fifteen VAM fungal species were isolated from the root-zone soils of fourteen different plant species. There was a significant correlation observed between the number of spores and of percentage root colonization as exemplified by Phyllanthus niruri and Paspalum vaginatum (450, 95%; 60, 25%). Although VAM species are not known to be strictly site specific, the fact that Acaulospora elegans was observed only in site 1, Glomus pulvinatum in site 2 only, and Gl. intraradices in site 3 only, showed site-specificity in this study. To confirm the infection efficiency, two host plant species in the sites, P. niruri and Eclipta alba, were selected and inoculated in field with three selected VAM fungal spores. Gl. fasciculatum was found to be the most efficient VAM species in percentage root colonization, number of VAM spores, and dry matter content. When the nutrients in roots of P. niruri and E. alba were analyzed, there was higher uptake of K (4.2 and 3.4 times, respectively) and Ca (5.3 and 4.9 times, respectively), the analogues for $^{137}Cs$ and $^{90}Sr$, respectively. From the results, it might be concluded that VAM association helps the plants survive in a disturbed ecosystem and enhances uptake and cycling of radionuclides from the ecosystem.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2001.06a
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• pp.35-39
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• 2001

The Integration Host factor (IHF) of Escherichia coli is a small, basic protein that is required for a variety of functions including site-specific recombination, transposition, gene regulation, plasmid replication, and DNA packaging. It ,is composed of two subunits that are encoded by the ihfA ($\alpha$-subunit) and ihjB ($\beta$-subunit) genes. IHF binding sites are composed of three elements called the WATCAR, TTG, and poly (dAT) elements. We have characterized IHF binding to the H site of bacteriophage λ. We have isolated suppressors that bind to altered H' sites using a challenge phage selection. Two different suppressors were isolated that changed the adjacent $\alpha$P64 and $\alpha$K65 residues. The suppressors recognized both the wild-type site and a site with a change in the WATCAR element. Three suppressors were isolated at $\beta$-E44. These suppressors bound the wild-type and a mutant site with a T：A to A：T change (H44A) in the middle of the TIR element. Site-directed mutagenesis was used to make several additional changes at $\beta$E44. The wild-type and $\beta$E44D mutant could not bind the wild-type site but were able to bind the H44A mutant site. Other mutants with neutral, polar, or a positive charge at $\beta$E44 were able to repress both the wild-type and H44A sites. Examination of the IHF crystal structure suggests that the ability of the wild-type and $\beta$E44D proteins to discriminate between the T：A and A：T basepairs is due to indirect interactions. The $\beta$-E44 residue does not contact the DNA directly. It imposes binding specificity indirectly by interactions with residues that contact the DNA. Details of the proposed interactions are discussed.