• Korean Journal of Veterinary Service
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• v.39 no.4
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• pp.247-251
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• 2016

Salmonella is one of the most common bacteria that causes heavy losses in swine industry and major causative pathogen of food poisoning in public health. Various methods for the identification of Salmonella such as Gram staining, agglutination test, enzyme-linked immunosorbent assay (ELISA), polymerase chain reaction (PCR) have been used. Several studies have demonstrated that Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI TOF) Mass Spectrometry (MS) identification is an efficient and inexpensive method for the rapid and routine identification of isolated bacteria. In this study, MALDI TOF MS could provide rapid, accurate identification of Salmonella spp. from swine compared with end point PCR and real time PCR.

• The Plant Pathology Journal
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• v.22 no.2
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• pp.155-160
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• 2006

A strain of Pepper mottle virus (PepMov) was isolated from chili pepper plants in Korea. In host range study, this virus, designated PepMoV-SNU1, shared most characteristics with PepMoV isolates reported previously. Thermal inactivation point ($45^{\circ}C\;to\;75^{\circ}C$) and dilution end point ($10^{-1}\;to\;10^{-4}$) of PepMoV-SNU1 showed differences depending on the propagation hosts. Cylindrical and pinwheel-shaped inclusions were always observed in pepper leaf tissues infected with the virus alone. Unexpectedly, a special structure of pinwheel shaped inclusion surrounded with unknown small spots was also observed in the leaf section when co-infected with a strain of pepper mild mottle virus. The partial sequence of coat protein gene and 3' untranslated region of PepMoV-SNU1 showed 98% identity with those of other PepMoV isolates. A primer pair derived from 3' end of the coat protein gene and poly A tail regions were designed. Optimal detection condition of PepMoV-SNU1 by RT-PCR was tested to determine appropriate annealing temperature and additional volumes of oligo-dT (18-mer), dNTP, and Taq polymerase. Under the optimized condition, an expected 500 Up PCR-product was detected in pepper leaves infected with PepMoV-SNU1 but not in healthy plants.

• Research in Plant Disease
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• v.7 no.1
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• pp.1-7
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• 2001

An isolate of Cucumber mosaic cucumovirus(CMV) was isolated from Hydrangea macrophylla for. otaksa(Sieb. et Zucc. ) Wils. showing mosaic symptoms, and designated as Hm-CMV. Hm-CMV was characterized by the tests of host range, physical properties, serological properties, RNA and coat protein compositions, and reverse transcription and polymerase chain reaction (RT-PCR) analysis. Twelve species in 4 families were used in the host range test of Hm-CMV and could be differentiated from Y-CMV used as a control CMV by the ringspot and line pattern on inoculated leaves of several tobacco plants. Thevirus produced local lesions on inoculated leaves of Chenopodium amarticolor, C. quinoa and Vigna unguiculata. The physical properties of the virus were as follows; thermal inactivation point(TIP) was 60$\^{C}$, dilution end point (DEP) was 10$\^$-3/, and longevity in vitro (LIP) was 3∼4 days. Hm-CMV was serologically identical to Y-CMV. SDS-polyaciylamide gel electrophoresis(SDS-PAGE) showed one major protein band of about 28 kDa. In RNA or dsRNA analysis, Hm-CMV consisted of four RNA or dsRNA species, but satellite RNA was not detected. In RT-PCR using CMV-common primer and CMV subgroup I-specific primer, bothe amplified expected size of about 490 bp and 200 bp DNA fragments from Hm-CMV, respectively. Restriction enzyme analysis of the 490 bp RT-PCR products using EcoR I and Msp I showed that Hm-CMV belonged to CMV subgroup I. However, Hm-CMV could be differentiated from other CMV subgroup I isolates by RNA fingerprinting by arbitrarily primed polymerase chain reaction (RAP-PCR).

• Proceedings of the Korean Vacuum Society Conference
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• 2013.08a
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• pp.273-273
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• 2013

Recently, Point-of-care (POC) testing microdevices enable to do the patient monitoring, drug screening, pathogen detection in the outside of hospital. Immunochromatographic strip (ICS) is one of the diagnostic technologies which are widely applied to POC detection. Relatively low cost, simplicity to use, easy interpretations of the diagnostic results and high stability under any circumstances are representative advantages of POC diagnosis. It would provide colorimetric results more conveniently, if the genetic analysis microsystem incorporates the ICS as a detector part. In this work, we develop a reverse transcriptase-polymerase chain reaction (RT-PCR) microfluidic device integrated with a ROSGENE strip for colorimetric influenza H1N1 virus detection. The integrated RT-PCR- ROSGENE device is consist of four functional units which are a pneumatic micropump for sample loading, 2 ${\mu}L$ volume RT-PCR chamber for target gene amplification, a resistance temperature detector (RTD) electrode for temperature control, and a ROSGENE strip for target gene detection. The device was fabricated by combining four layers: First wafer is for RTD microfabrication, the second wafer is for PCR chamber at the bottom and micropump channel on the top, the third is the monolithic PDMS, and the fourth is the manifold for micropump operation. The RT-PCR was performed with subtype specific forward and reverse primers which were labeled with Texas-red, serving as a fluorescent hapten. A biotin-dUTP was used to insert biotin moieties in the PCR amplicons, during the RT-PCR. The RT-PCR amplicons were loaded in the sample application area, and they were conjugated with Au NP-labeled hapten-antibody. The test band embedded with streptavidins captures the biotin labeled amplicons and we can see violet colorimetric signals if the target gene was amplified with the control line. The off-chip RT-PCR amplicons of the influenza H1N1 virus were analyzed with a ROSGENE strip in comparison with an agarose gel electrophoresis. The intensities of test line was proportional to the template quantity and the detection sensitivity of the strip was better than that of the agarose gel. The test band of the ROSGENE strip could be observed with only 10 copies of a RNA template by the naked eyes. For the on-chip RT-PCR-ROSGENE experiments, a RT-PCR cocktail was injected into the chamber from the inlet reservoir to the waste outlet by the micro-pump actuation. After filling without bubbles inside the chamber, a RT-PCR thermal cycling was executed for 2 hours with all the microvalves closed to isolate the PCR chamber. After thermal cycling, the RT-PCR product was delivered to the attached ROSGENE strip through the outlet reservoir. After dropping 40 ${\mu}L$ of an eluant buffer at the end of the strip, the violet test line was detected as a H1N1 virus indicator, while the negative experiment only revealed a control line and while the positive experiment a control and a test line was appeared.

• Research in Plant Disease
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• v.12 no.2
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• pp.139-143
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• 2006

Virion captured reverse transcription polymerase chain reaction (VC/RT-PCR) could detect plant virus quickly and accurately. In the VC/RT-PCR, no antibody is needed unlike immuno-captured RT-PCR (IC/RT-PCR) which had been improved method of RT-PCR for plant viruses, and virus nucleic acids can be obtained easily within 30minutes by property of polypropylene PCR tube which is hold and immobilized viral particles on its surface. For the virion capture of Tomato spotted wilt virus (TSWV), the extraction buffer was tested. The optimum macerating buffer for TSWV was 0.01M potassium phosphate buffer, pH 7.0, containing 0.5% sodium sulfite. The viral crude sap was incubated for 30 min at $4^{\circ}C$. The virions in the PCR tubes were washed two times with 0.01M PBS containing 0.05% Tween-20. The washed virions were treated at $95^{\circ}C$ immediately for 1 min containing RNase free water and chilled quickly in the ice. Disclosed virions' RNAs by heat treatment were used for RT-PCR. Dilution end point of $10^{-5}$ from plant's crude sap infected with TSWV showed relatively higher detection sensitivity for VC/RT-PCR. During multiple detection using two or more primers, interference was arisen by interactions between primer-primer and plant species. The result of multiplex RT-PCR was influenced by combinations of primers and the kind of plant, and the optimum extraction buffer for the multiplex detection by VC/RT-PCR should be developed.

• The Plant Pathology Journal
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• v.26 no.4
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• pp.328-339
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• 2010

A tobamovirus, Ribgrass mosaic virus (RMV), was identified newly from chinese cabbage (Brassica campestris L. pekinensis) in Korea. Virus disease incidence of RMV on chinese cabbage was 37.9% in alpine area on August in 1993. RMV induced the symptoms of necrotic ring spots, necrotic streak on midrib and malformation. RMV, Ca1 and Ca3 isolate, could infect 35 species out of 45 plants including Chenopodium amaranticolor. Physical properties of RMV Ca1 isolate were very stable as 10.8 over for dilution end point, $95^{\circ}C$ for temperature inactivation point and 18 weeks for longevity in vitro. RMV had the soil transmission rate of 75.0% for the chinese cabbages, 'Chunhawang' and 'Seoul' cultivars. The purified virions of RMV had the typical ultraviolet absorption spectrum of maximum at 260 nm and minimum at 247 nm. RMV of Ca1 isolate was related serologically with antisera of Tobacco mosaic virus (TMV)-Cym, TMV-O and Pepper mottle virus, but not related with antiserum of Odontoglossum ring spot virus. coat protein gene of RMV-Ca1, sized 473 nucleotides, encoded 158 amino acid residues. Nucleotide identity of RMV-Ca1 CP gene was 96.4% with RMV-Shanghai (GenBank accession No. of AF185272) from China and 96.0% with RMV-Impatiens (GenBank accession No. of AM040974) from Germany. Identity of amino acids between RMV-Ca1 and the two RMV isolates was 96.8%. Specific three primers were selected for rapid and easy genetic detection of RMV using Virion Captured (VC)/RT-PCR method.

• Journal of Microbiology and Biotechnology
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• v.21 no.6
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• pp.659-664
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• 2011

Staphylococcus aureus (SA) bacteremia is associated with high mortality, and often results in metastatic infections. The methicillin-resistant SA (MRSA) is an urgent health care issue, as nosocomial infections with these bacteria represent limited treatment alternatives. Samples of whole blood containing challenge inoculums of SA and MRSA strains were passed through columns packed with surfaceheparinized polyethylene beads. The bound bacteria were eluted and quantitatively determined by culturing and by real-time PCR. Significant amounts of both SA and MRSA adhered to the heparinized beads (more than 65% of inoculated bacteria). After rinsing with buffer at high ionic strength, viable bacteria or bacterial DNA were eluted from the columns, indicating that the binding was specific. The conclusions that can be made from these experiments are that, as earlier reported in the literature, the high affinity of SA to heparin is retained in whole blood, and MRSA in whole blood binds to heparin with similar or higher affinity than SA. It should be possible to lower the amount of SA and/or MRSA from the blood of infected patients to levels that could be taken care of by the immune system. In previous studies, we have shown that passing blood from septic patients over beads coated with end-point-attached, biologically active heparin is a useful technique for regulating the levels of heparinbinding cytokine. These findings in combination with the present findings indicate the possibility of creating an apheresis technology for treatment of sepsis caused by SA and/or MRSA.

• Korean Journal of Biological Psychiatry
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• v.8 no.1
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• pp.106-110
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• 2001

Background : Dopamine receptors have been regarded as a strong candidate involved in etiology of schizophrenia and a target for various antipsychotic drugs. The purpose of our study was to investigate whether dopamine $D_1$ receptor(DRD1) gene polymorphisms would predict the treatment response to antipsychotics in schizophrenia. Method : One hundred thirty-four schizophrenic patients, who met DSM-IV criteria for schizophrenia were entered into a 48 -week study. The psychopathology of the patients was assessed at baseline, 12th, 24th 48th weeks of treatment by PANSS. Responders were defined by a 20% of the reduction in total PANSS score at end point. The genomic DNA fragment corresponding to nucleotides of dopamine $D_1$ receptor gene was amplified by polymerase chain reaction(PCR). Result: Neither allelic frequencies nor genotypes for dopamine $D_1$ receptor differed significantly between responders and non-responders. Also, there was no difference of changes of PANSS scores among three genotype groups of the dopamine $D_1$ receptor. Conclusion : Allelic variation in the dopamine $D_1$ gene is not associated with individual differences in antipsychotic response.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.12
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• pp.6155-6157
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• 2012

Background: Mutations in the MAPK (Mitogen Activated Protein Kinase) signaling pathway - EGFR/Ras/RAF/MEK have been associated with the development of several carcinomas. ERK2, a downstream target of the MAPK pathway and a founding member of the MAPK family is activated by cellular signals emanating at the cell membrane. Activated ERK2 translocates into the nucleus to transactivate genes that promote cell proliferation. MKP - a dual specific phosphatase - interacts with activated ERK2 via the common docking (CD) domain of the later to inactivate (dephosphorylate) and effectively terminate further cell proliferation. A constitutively active form of ERK2 carrying a single point mutation - E322K in its CD domain, was earlier reported by our laboratory. In the present study, we investigated the prevalence of this CD domain E322K mutation in 88 well differentiated OSCC tissue samples. Materials and Method: Genomic DNA specimens isolated from 88 oral squamous cell carcinoma tissue samples were amplified with primers flanking the CD domain of the ERK2 gene. Subsequently, PCR amplicons were gel purified and subjected to direct sequencing to screen for mutations. Results: Direct sequencing of eighty eight OSCC samples identified an E322K CD domain mutation in only one (1.1%) OSCC sample. Conclusions: Our result indicates that mutation in the CD domain of ERK2 is rare in OSCC patients, which suggests the role of genetic alterations in other mitogenic genes in the development of carcinoma in the rest of the patients. Nevertheless, the finding is clinically significant, as the relatively rare prevalence of the E322K mutation in OSCC suggests that ERK2, being a common end point signal in the multi-hierarchical mitogen activated signaling pathway may be explored as a viable drug target in the treatment of OSCC.

• Journal of Soil and Groundwater Environment
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• v.10 no.2
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• pp.52-58
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• 2005

In Korea, little attention has been paid to microbial perchloroethylene (PCE) and/or trichloroethylene (TCE) dechlorination activity and identification of microorganisms involved in PCE reductive dechlorination at a PCE-contaminated aquifer. We performed microcosm tests using the groundwater samples from 4 different contaminated sites (i.e. Changwon A, Changwon B, Bucheon and Yangsan) to assess PCE reductive dechlorination activity. We also adapted molecular techniques to screen what types of known reductive dechlorinators are present at the PCE-contaminated aquifers. In the Changwon A and Changwon B active microcosms where potential electron donors such as sodium propionate, sodium lactate, sodium butyrate, and sodium fumarate, were added, ethylene, an end-product of complete reductive dechlorination of PCE, was detected after a period of 90 days of incubation. In the Bucheon and Yangsan active microcosms, cis-1,2-dichloroethylene (c-DCE) was accumulated without the production of vinyl chloride (VC) and ethylene. Molecular techniques were used to evaluate the microbial community structures in the Changwon B and Yangsan aquifer. We found two sequence types that were closely related to a known PCE to ethylene dechlorinator, named uncultured bacterium clone DCE47, in the Changwon B site clone library. However, in the Yangsan site clone library, no sequence type was closely related to known PCE dechlorinators reported. It is plausible that microorganisms being capable of completely dechlorinating PCE to ethylene may be present in the Changwon B site aquifer. In this study we find that complete PCE reductive dechlorinators are present at some PCE-contaminated sites in Korea. In an engineering point of view this information makes it feasible to apply a biological reductive dechlorination process for remediating PCE- and/or TCE-contaminated aquifers in Korea.