• Title/Summary/Keyword: Species-specific PCR

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Detection of Fish Killing Dinoflagellates Cochlodinium polykrikoides and Karlodinium veneficum (Dinophyceae) in the East China Sea by Real-time PCR

  • Park, Tae-Gyu;Kang, Yang-Soon;Park, Young-Tae;Bae, Heon-Meen;Lee, Yoon
    • ALGAE
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    • v.24 no.2
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    • pp.105-110
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    • 2009
  • The rDNAs of figh-killing dinoflagellates Cochlodinium polykrikoides and Karlodinium veneficum were detected from the East China Sea by species-specific real-time PCR probes. Sequence analysesusing the partial ITS sequences from the real-time PCR products showed identical sequences with C. Polykrikoides and K. veneficum, respectively and low expectation values (E-value) of less than 1e-5 suggesting the presence of these organisms in the East Ching Sea shelf water that flows into the Tsushima Strait and the Yellow Sea.

Development of a Multiplex Reverse Transcription-Polymerase Chain Reaction Assay for the Simultaneous Detection of Three Viruses in Leguminous Plants

  • Park, Chung Youl;Min, Hyun-Geun;Lee, Hong-Kyu;Maharjan, Rameswor;Yoon, Youngnam;Lee, Su-Heon
    • Research in Plant Disease
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    • v.24 no.4
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    • pp.348-352
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    • 2018
  • A multiplex reverse transcription-polymerase chain reaction (mRT-PCR) assay was developed for the detection of Clover yellow vein virus (ClYVV), Peanut mottle virus (PeMoV), and Tomato spotted wilt virus (TSWV), which were recently reported to infect soybean and azuki bean in Korea. Species-specific primer sets were designed for the detection of each virus, and their specificity and sensitivity were tested using mixed primer sets. From among the designed primer sets, two combinations were selected and further evaluated to estimate the detection limits of uniplex, duplex, and multiplex RT-PCR. The multiplex RT-PCR assay could be a useful tool for the field survey of plant viruses and the rapid detection of ClYVV, PeMoV, and TSWV in leguminous plants.

Genetic distances of three venerid species identified by PCR analysis

  • Jeon, Jun-Hyub;Yoon, Jong-Man
    • The Korean Journal of Malacology
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    • v.31 no.4
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    • pp.257-262
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    • 2015
  • The seven selected primers BION-13, BION-29, BION-61, BION-64, BION-68, BION-72 and BION-80 generated the total number of loci, average number of loci per lane and specific loci in Meretrix lusoria (ML), Saxidomus purpuratus (SP) and Cyclina sinensis (CS) species. Here, the complexity of the banding patterns varied dramatically between the primers from the three venerid clam species. The higher fragment sizes (> 1,000 bp) are much more observed in the SP species. The primer BION-68 generated 21 unique loci to each species, which were ascertaining each species, approximately 150 bp, 300 bp and 450 bp, in the ML species. Remarkably, the primer BION-80 detected 7 shared loci by the three clam species, major and/or minor fragments of sizes 500 bp, which were matching in all samples. As regards average bandsharing value (BS) results, individuals from CS clam species (0.754) exhibited higher bandsharing values than did individuals from SP clam species (0.607) (P < 0.05). In this study, the dendrogram obtained by the seven oligonucleotides primers indicates three genetic clusters: cluster 1 (LUSORIA01-LUSORIA07), cluster 2 (PURPURATUS08-PURPURATUS14), cluster 3 (SINENSIS15-SINENSIS21). Among the twenty one venerid clams, the shortest genetic distance that displayed significant molecular differences was between individuals 18 and 20 from the CS species (genetic distance = 0.071), while the longest genetic distance among the twenty-one individuals that displayed significant molecular differences was between individuals LUSORIA no. 02 and PURPURATUS no. 09 (genetic distance = 0.778). Relatively, individuals of SP venerid species were appropriately closely related to that of CS species, as shown in the hierarchical dendrogram of genetic distances. Eventually, PCR fragments exposed in the present study may be worthwhile as a DNA marker the three venerid clam species to discriminate.

Segmented Filamentous Bacteria Induce Divergent Populations of Antigen-Specific CD4 T Cells in the Small Intestine

  • Yi, Jaeu;Jung, Jisun;Han, Daehee;Surh, Charles D.;Lee, You Jeong
    • Molecules and Cells
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    • v.42 no.3
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    • pp.228-236
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    • 2019
  • CD4 T cells differentiate into $ROR{\gamma}t/IL$-17A-expressing cells in the small intestine following colonization by segmented filamentous bacteria (SFB). However, it remains unclear whether SFB-specific CD4 T cells can differentiate directly from naïve precursors, and whether their effector differentiation is solely directed towards the Th17 lineage. In this study, we used adoptive T cell transfer experiments and showed that naïve CD4 T cells can migrate to the small intestinal lamina propria (sLP) and differentiate into effector T cells that synthesize IL-17A in response to SFB colonization. Using single cell RT-PCR analysis, we showed that the progenies of SFB responding T cells are not uniform but composed of transcriptionally divergent populations including Th1, Th17 and follicular helper T cells. We further confirmed this finding using in vitro culture of SFB specific intestinal CD4 T cells in the presence of cognate antigens, which also generated heterogeneous population with similar features. Collectively, these findings indicate that a single species of intestinal bacteria can generate a divergent population of antigen-specific effector CD4 T cells, rather than it provides a cytokine milieu for the development of a particular effector T cell subset.

Development and Application of PCR-based Markers for the Discrimination of Bang-Poong and Related Species (방풍류의 감별을 위한 분자마커의 탐색과 활용)

  • Hong, Seong-Mi;Lee, Mi-Young;Koh, Jae-Chul;Ko, Byoung-Soeb
    • Journal of Plant Biotechnology
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    • v.31 no.1
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    • pp.1-6
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    • 2004
  • Bang-Poong and related species are an important herbal medicine. However, it is difficult to determine the commercial dry material through anatomical and chemotaxonomical characteristics. Here, we used a PCR-based technique for an accurate discrimination of Bang-Poong and related species. With the RAPD primers, 215 RAPDSs(random amplified polymorphic DNAs) were obtained, and 98% of them showed polymorphic patterns. RAPDs from the four primers were appropriate for the discrimination of S. divaricata $(T_{URCZ{\cdot}})\;S_{CHISKIN}$, those from the six primers for P. japonicum $T_{HUNBERG}$, those from the four primers for P. terebinthaceum $F_{ISHER}$, and those from the six primers for G. littoralis Fr. $S_{CHMIDT}$. The specific bands from the primer 425 were obtained and used to develop SCAR (sequence characterized amplified region) markers, based on the sequence information of the RAPD markers. The SCAR primers generated a 215 bp fragment specific to Peucedanum terebinthaceum $F_{ISHER}$, and a 177 bp and a 300 bp fragment specific to G. littoralis Fr. $S_{CHMIDT}$. As a result, the three SCAR markers were able to discriminate from two Bang-Poong related species.

A PCR Method to Distinguish Matsumuraeses phaseoli from M. falcana Based on the Difference of Nucleotide Sequence in the Mitochondrial Cytochrome c Oxidase Subunit I (미토콘드리아 COI 영역의 뉴클레오티드 서열 차이를 이용한 팥나방과 어리팥나방의 PCR 판별법)

  • Seo, Bo Yoon;Jung, Jin Kyo;Cho, Jum Rae;Kim, Yonggyun;Park, Chang Gyu
    • Korean journal of applied entomology
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    • v.51 no.4
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    • pp.365-370
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    • 2012
  • The two closely related major leguminous crop pests in Korea, Matsumuraeses phaseoli and M. falcana (Lepidoptera: Tortricidae) have very similar morphological characters, which occasionally give rise to a failure in distinguishing between the two. In this study, we report an easy PCR-SSP method to distinguish between them, with a sequence specific primer set (P-SF2, F-SF3, and C-SR3) based on single nucleotide mismatch in 3' terminal base of a primer, which is found in the mitochondrial cytochrome c oxidase subunit I DNA (mtCOI). Through application of this method, each species may be clearly identified in terms of its PCR band size and pattern, only one band (245 bp) for M. falcana and one (409 bp) or two bands (409 bp & 245 bp) for M. phaseoli.

Development of Differential Media and Multiplex PCR Assays for the Rapid Detection of Listeria monocytogenes (Listeria monocytogenes의 신속검출을 위한 선택배지 및 multiplex PCR 기법 개발)

  • Jung, Byeong-yeal;Lim, Hyun-sook;Jung, Suk-chan
    • Korean Journal of Veterinary Research
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    • v.43 no.2
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    • pp.231-237
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    • 2003
  • Listeria (L.) monocytogenes in samples could not be detected occasioally by faster growth of other Listeria spp. especially L. innocua. The aim of this study was to develop the differential media and multiplex polymerase chain reaction (PCR) assays for the rapid detection of L. monocytogenes. L. monocytogenes colonies were characterized by their ${\beta}$-hemolysis with fluorescence under 366 nm UV light on the Listeria hemolysis agar (LHA). L. innocua, a species commonly present in foods, did not produce ${\beta}$-hemolysis on LHA. Therefore, one or more colonies of L. monocytogenes were easily distinguished from large populations of L. innocua. The multiplex PCR assays were developed to distinguish from L. monocytogenes and other Listeria spp. with two pairs of primers. The primers were designed in 16S rRNA and listeriolysin O gene for specific amplification of all members of the genus Listeria and L. monocytogenes, respectively. The multiplex PCR assays produced 560 and 938 bp products in L. monocytogenes; only 938 bp products in the genus Listeria. The multiplex PCR assays could detect as little as 50 pg of L monocytogenes DNA. These results indicated that the differential media and multiplex PCR assays might be useful diagnostic tools for the rapid detection of L. monocytogenes.

Development of glufosinate-tolerant GMO detection markers for food safety management (식품안전관리를 위한 제초제 glufosinate 특이적 GM 작물 검출마커 개발)

  • Song, Minji;Qin, Yang;Cho, Younsung;Park, TaeSung;Lim, Myung-Ho
    • Korean Journal of Food Science and Technology
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    • v.52 no.1
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    • pp.40-45
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    • 2020
  • Over 500 genetically modified organisms (GMOs) have been developed since 1996, of which nearly 44% have glufosinate herbicide-tolerant traits. Identification of specific markers that can be used to identify herbicide-tolerant traits is challenging as the DNA sequences of the gene(s) of a trait are highly variable depending on the origin of the gene(s), plant species, and developers. To develop specific PCR marker(s) for the detection of the glufosinate-tolerance trait, DNA sequences of several pat or bar genes were compared and a diverse combination of PCR primer sets were examined using certified reference materials or transgenic plants. Based on both the qualitative and quantitative PCR tests, a primer set specific for pat and non-specific for bar was developed. Additionally, a set of markers that can detect both pat and bar was developed, and the quantitative PCR data indicated that the primer pairs were sensitive enough to detect 0.1% of the mixed seed content rate.

Multiple Symbiotic Associations Found in the Roots of Botrychium ternatum

  • Lee, Jun-Ki;Eom, Ahn-Heum;Lee, Sang-Sun
    • Mycobiology
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    • v.30 no.3
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    • pp.146-153
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    • 2002
  • Two types of mycorrhizae, orchid(OM) and arbuscular mycorrhizae(AM), were observed in the cortical cells of Botrychium ternatum roots. The vesicles or arbuscules of AM fungi were examined and the fresh or digestive pelotons by other species of basidiomycetes were also observed in the roots under light microscope. These symbioses were, as the genomic DNAs extracted from roots of B. ternatum reacted with the specific primers, confirmed with PCR technique, being added to more strong evidences. These discoveries were rarely happened in the roots, especially a fern in nature. OM was observed in the roots of B. ternatum collected from the nationwide areas, whereas AM was only in the roots of B. ternatum collected from Chung-Buk areas. It is speculated that OM are associated with the nitrogen cycle in Islands and the growth of B. ternatum in the inland of Central Korea is related to both the phosphate and nitrogen cycle in the nature. The results suggest that B. ternatum is a typical species with two types of mycorrhizae under various growing conditions.

The occurrence trend of the RSV and its coining of coat protein of korean strain.

  • Park, Jo-Im;Lee, Bong-Choon;Hong, Yeon-Kyu;Kwak, Do-Yeon;Oh, Byeong-Geon;Park, Sung tae
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.114.2-115
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    • 2003
  • Rice stripe virus causes severe damage to rice in Korea, Japan and China. RSV is a type member of the tenuivirus group and transmitted by the small brown planthopper, Laodeiphax striatellus, in a persistant manner. Until now, occurrence of RSV is limited in of southern part of Korea. But recently occurrence of RSV is increasing and spreading in central part of Korea including Chungcheong and Kyonggi province. So we analyzed recent occurrence trend of RSV which is increased and cloned and sequenced coat protein gene for isolating of RSV strain. Infected rice of each species(Ilpumbyeo, Sindongjinbyeo, Keumobyeo-2, Dongjinbyeo, Jongnambyeo, Samcheonbyeo, etc.)is collected, we extracted total RNA from infected leaves and detected RSV viral RNA by reverse transcription(RT)-PCR using specific primer of coat protein gene. The result of RT-PCR, we observed specific band. We already cloned cDNA from the band, is analyzing sequence variety and homology of each species.

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