• Title/Summary/Keyword: specific probe

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Application of Species-specific DNA Probe to Field Samples of Alexandrium tamarense (Lebour) Balech (자연 시료로부터 Alexandrium tamarense을 위한 종 특이적 DNA탐침의 응용)

  • Cho, Eun-Seob;Kim, Gi-Young;Park, Hyung-Sik;Kim, Hak-Gyoon;Moon, Sung-Ki;Lee, Jae-Dong
    • Journal of Life Science
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    • v.12 no.3
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    • pp.250-255
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    • 2002
  • Fluorescent species-specific DNA probe (AT1) of toxic dinoflagellate Arexandrium tamarense was tested on several other species, on comparison of binding activity at different preservatives for fixation of the cells, at different culture age and estimation of cell density by light microscope or epifluorescent microscope using whole cell hybridization. Th AT1 probe specifically bound to Alexandrium tamarense, whereas it did not bind to other phytoplankton, in particular Alexandrium catenella, morphologically similar to Alexandrium tamarense, could not react to AT1 probe. When cells were fixed with all three preservatives, labeling cells of Alexandrium tamarense emitted strong fluorescent signal intensity. In addition, regardless culture days, binding activity with AT1 probe was strong. The tell densities estimated by epifluorescent microscope were than those estimated by light microscope. The enumeration and identifying of Arexandriurn tamarense using DNA probe method will be contributed to a new biotoxin monitoring and prediction system in field.

Identification of Brucella melitensis isolates originating from Mongolia and diagnostic real-time PCR evaluation using a specific SNP (몽골 유래 Brucella melitensis 동정 및 특이 SNP를 이용한 real-time PCR법에 의한 진단 평가)

  • Kang, Sung-Il;Kim, Ji-Yeon;Kim, Suk Mi;Lee, Jin Ju;Sung, So-Ra;Kim, Yeon-Hee;Jung, Suk Chan;Her, Moon
    • Korean Journal of Veterinary Research
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    • v.55 no.2
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    • pp.105-110
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    • 2015
  • A real-time PCR assay using hybridization probe (HybProbe) has been developed to detect Brucella (B.) melitensis strains. The primer and HybProbe sets were designed based on the gap gene of chromosome I with a specific single nucleotide polymorphism of B. melitensis. Specificity of the assay was confirmed by comparison to reference Brucella species and other related strains. In the melting curve analysis, B. melitensis generated a peak at $67^{\circ}C$ unlike those for other Brucella species observed at $61^{\circ}C$. Sensitivity of the assay for B. melitensis ranged from 20 ng to 200 fg of genomic DNA. The ability to identify 94 Mongolian B. melitensis isolates using the real-time PCR assay was identical to that of classical biotyping methods and differential multiplex PCR. These data showed that this new molecular technique is a simple and quick method for detecting B. melitensis, which will be important for the control and prevention of brucellosis.

Quality Control Probes for Spot-Uniformity and Quantitative Analysis of Oligonucleotide Array

  • Jang, Hyun-Jung;Cho, Mong;Kim, Hyung-Hoi;Kim, Cheol-Min;Park, Hee-Kyung
    • Journal of Microbiology and Biotechnology
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    • v.19 no.7
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    • pp.658-665
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    • 2009
  • Quality control QC for spot-uniformity is a critical point in fabricating an oligonucleotide array, and quantification of targets is very important in array analysis. We developed two new types of QC probes as a means of confirming the quality of the uniformity of attached probes and the quantification of targets. We compared the signal intensities and fluorescent images of the QC and target-specific probes of arrays containing only target-specific probes and those containing both QC and target-specific probes. In a comparison of quality control methods, it was found that the arrays containing QC probes could check spot-uniformity or spot defects during all processes of array fabrication, including after spotting, after washing, and after hybridization. In a comparison of quantification results, the array fabricated by the method using QC probes showed linear and regular results because it was possible to normalize variations in spot size and morphology and amount of attached probe. This method could avoid errors originating in probe concentration and spot morphology because it could be normalized by QC probes. There were significant differences in the signal intensities of all mixtures (P<0.05). This result indicates that the method using QC probes is more useful than the ordinary method for quantification of mixed target. In the quantification of mixed targets, this method could determine a range for mixed targets of various amounts. Our results suggest that methods using QC probes for array fabrication are very useful to the quality control of spots in the fabrication processes of quantitative oligonucleotide arrays.

Development of Repetitive DNA Probes for Genetic Analysis of Phytophthora capsici (Phytophthora capsici의 유전적 특성 분석을 위한 Repetitive DNA Probe의 개발)

  • Song, Jeong-Young;Kim, Hong-Gi
    • The Korean Journal of Mycology
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    • v.30 no.1
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    • pp.66-72
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    • 2002
  • To develop DNA markers for analysis of genetic characteristics of Phytophthora capsici population, randomly selected clones from HindIII-digested genomic DNA library of P. capsici 95CY3119 were surveyed by hybridizing to Southern blots of HindIII-digested total genomic DNA of P. capsici. Probe DNAs inserted into selected individual clones strongly hybridized with HindIII digests of P. capsici. Among probes examined, PC9 revealed the repetitive and highly polymorphic bands to HindIII digests of inter-and intra-field P. capsici isolates. Genetic diversity of individual isolates was also clearly revealed in cluster analysis based on its band patterns. The other probe, PC22, was hybridized only to DNA from P. capsici and this was highly repetitive. However, there was no response to other Phytophthora species and Pythium sp. These DNA probes could be used as very useful markers in analysing genetic diversity and identification for P. capsici population throughout the world.

Evaluation of a novel TaqMan probe-based real-time polymerase chain reaction (PCR) assay for detection and quantitation of red sea bream iridovirus

  • Kim, Guk Hyun;Kim, Min Jae;Choi, Hee Ju;Koo, Min Ji;Kim, Min Jeong;Min, Joon Gyu;Kim, Kwang Il
    • Fisheries and Aquatic Sciences
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    • v.24 no.11
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    • pp.351-359
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    • 2021
  • The red sea bream iridovirus (RSIV) belonging to genus Megalocytivirus is responsible for red sea bream iridoviral disease (RSIVD) in marine and freshwater fishes. Although several diagnostic assays for RSIV have been developed, diagnostic sensitivity (DSe) and specificity (DSp) of real-time polymerase chain reaction (PCR) assays are not yet evaluated. In this study, we developed a TaqMan probe-based real-time PCR method and evaluated its DSe and DSp. To detect RSIV, the probe and primers were designed based on consensus sequences of the major capsid protein (MCP) genes from megalocytiviruses including RSIV, infectious spleen and kidney necrosis virus (ISKNV), and turbot reddish body iridovirus (TRBIV). The probe and primers were shown to be specific for RSIV, ISKNV, and TRBIV-types megalocytiviruses. A 95% limit of detection (LOD95%) was determined to be 5.3 viral genome copies/µL of plasmid DNA containing the MCP gene from RSIV. The DSe and DSp of the developed real-time PCR assay for field samples (n = 112) were compared with those of conventional PCR assays and found to be 100% and 95.2%, respectively. The quantitative results for SYBR Green and TaqMan probe-based real-time PCR were not significantly different. The TaqMan probe-based real-time PCR assay for RSIV may be used as an appropriate diagnostic tool for qualitative and quantitative analysis.

Flow characteristics at the Impeller Exit of a Centrifugal Pump (원심펌프의 회전차 출구 유동 특성)

  • Hong, Soon-Sam;Kang, Shin-Hyoulg
    • 유체기계공업학회:학술대회논문집
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    • 2000.12a
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    • pp.36-44
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    • 2000
  • The flow at the impeller exit is important to validate engineering design and numerical analysis of pumps. We installed axisymmetric collector instead of the volute casing, so there is no interaction between the impeller and casing. A hot-film probe and a high response pressure transducer are used to investigate the flow at impeller exit and vaneless diffuser region for design and off design flow rate. For a single suction centrifugal pump of low specific speed, the flow field such as velocity, flow angle, and total pressure are measured by traversing the probe across the vaneless diffuser. These data can be used for performance prediction, design, and numerical analysis of pumps.

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Measurement Data Comparison of Fast SAR Measurement System by Probe Arrays with Robot Scanning SAR Measurement System

  • Kim, Jun Hee;Gimm, Yoon-Myoung
    • Journal of electromagnetic engineering and science
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    • v.14 no.4
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    • pp.336-341
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    • 2014
  • Dosimetry of radiating electromagnetic wave from mobile devices to human body has been evaluated by measuring Specific Absorption Rate (SAR). Usual SAR measurement system scans the volume by robot arm to evaluate RF power absorption to human body from wireless devices. It is possible to fast estimate the volume SAR by software deleting robot moving time with the 2D surface SAR data acquired by arrayed probes. This paper shows the principle of fast SAR measurement and the measured data comparison between the fast SAR system and the robot scanning system. Data of the fast SAR is well corresponding with data of robot scanning SAR within ${\pm}3$ dB, and its dynamic range covers from 10 mW/kg to 30 W/kg with 4.8 mm probe diameter.

Development of prevotella intermedia ATCC 49046 Strain-Specific PCR Primer Based on a Pig6 DNA Probe (Pig6 DNA probe를 기반으로 하는 Prevotella intermedia ATCC 49046 균주-특이 PCR primer 개발)

  • Jeong Seung-U;Yoo So-Young;Kang Sook-Jin;Kim Mi-Kwang;Jang Hyun-Seon;Lee Kwang-Yong;Kim Byung-Ok;Kook Joong-Ki
    • Korean Journal of Microbiology
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    • v.42 no.2
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    • pp.89-94
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    • 2006
  • The purpose of this study is to develop the strain-specific PCR primers for the identification of prevotella inter-media ATCC 49046 which is frequently used in the pathogenesis studies of periodontitis. The Hind III-digested genomic DNA of P. intermedia ATCC 49046 were cloned by random cloning method. The specificity of cloned DNA fragments were determined by Southern blot analysis. The nucleotide sequence of cloned DNA probes was determined by chain termination method. The PCR primers were designed based on the nucleotide sequence of cloned DNA fragment. The data showed that Pig6 DNA probe were hybridized with the genomic DNA from P. intermedia strains (ATCC $25611^T$ and 49046) isolated from the Westerns, not the strains isolated from Koreans. The Pig6 DNA probe were consisted of 813 bp. Pig6-F3 and Pig6-R3 primers, designed base on the nucleotide Sequences Of Pig6 DNA Probe, were 3150 specific to the only both P. intermedia ATCC $25611^T$ and P. intermedia ATCC 49046. In the other hand, Pig6-60F and Pig6-770R primers were specific to the only P. intermedia ATCC 49046. The two PCR primer sets could detect as little as 4 pg of chromosomal DNA of P. intermedia. These results indicate that Pig6-60F and Pig6-770R primers have proven useful for the identification of P. intermedia ATCC 49046, especially with regard to the maintenance of the strain.

Molecular Identification of the Toxic Alexandrium tamiyavanichii (Dinophyceae) by the Whole-cell FISH Method

  • Kim Choong-Jae;Yoshimatsu Sada-Akfi;Sako Yoshihiko;Kim Chang-Hoon
    • Fisheries and Aquatic Sciences
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    • v.7 no.4
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    • pp.175-183
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    • 2004
  • The dinoflagellate Alexandrium tamiyavanichii Balech, a producer of toxins causing paralytic shellfish poisoning (PSP), has recently been considered as one of main organisms responsible for toxication of shellfish in Japan. In this study, A. tamiyavanichii was subjected to a molecular phylogenetic analysis inferred from 28S rDNA D1-D2 sequences and a species-specific LSU rRNA-targeted oligonucleotide DNA probe was designed to identify A. tamiyavanichii using the whole cell-FISH (fluorescence in situ hybridization). The sequences of the 28S rDNA D1-D2 region of A. tamiyavanichii showed no difference from A. cohorticular AF1746l4 (present name A. tamiyavanichii) and formed a distinct clade from the 'tamarensis species complex'. The probe, TAMID2, reacted specifically with A. tamiyavanichii cultured cells, without any cross-reaction with other species belonging to the same genus, including A. tamarense, A. catenella, A. affine, A. fraterculus, A. insuetum and A. pseudogonyaulax. In a test of cross-reactivity with a field sample, TAMID2 reacted consistently with only A. tamiyavanichii, indicating that the present protocol involving the TAMID2 probe might be useful for detecting toxic A. tamiyavanichii in a simple and rapid manner.

Probe Diffusion in Polymer Solutions by Forced Rayleigh Scattering

  • Jaeyung Lee;Taiho Park;Jungmoon Sung;Sangwook Park;Taihyun Chang
    • Bulletin of the Korean Chemical Society
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    • v.12 no.5
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    • pp.569-574
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    • 1991
  • Methyl red diffusion in polymer solutions was studied by a transient holographic method, forced Rayleigh scattering. In semi-dilute solutions of a polystyrene, where no specific interaction with the probe exists, we found within experimental uncertainty that the retardation of diffusion rate of methyl red is independent of the solvents used. This indicates that the hydrodynamic interaction in polymer coils is not affected by the nature of solvents enough to exhibit a detectable change in the diffusion rate of the probe. On the other hand, a substantial reduction of diffusion rate was observed in poly(methyl methacrylate) solutions in toluene. Together with the similar observation reported with poly(vinyl acetate), it is confirmed that hydrogen bond between the probe and the polymer is responsible for the retarded diffusion. The decay-growth-decay profile found in this system reveals a finite difference in diffusion coefficients of cis and trans isomer of methyl red. We estimate the difference and suggest that the cis isomer interacts with the polymer more strongly than the trans isomer.