• 제목/요약/키워드: melting curve

검색결과 93건 처리시간 0.019초

Real-time PCR을 이용한 원유시료 유래 황색포도상구균의 신속 검출 (SYBR Green I-based Real-time PCR Assay and Melting Curve Analysis for Rapid Detection of Staphylococcus aureus from Raw Milks Samples)

  • 정재혁;정순영;이상진;최성숙
    • 한국식품위생안전성학회지
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    • 제23권2호
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    • pp.121-128
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    • 2008
  • 본 연구는 Lightcycler (Roche)를 이용한 Real-Time PCR(LC-PCR)기법을 통하여 원유시료에서 신속, 정확하게 황색포도상구균을 검출하는 기법을 개발하고자 하였다. coagulase 전구체를 coding하는 113 bp의 coa 유전자의 증폭, melting curve 분석 및 DNA염기서열을 분석하여 황색포도상구균 특유의 유전자 검출하는 기법을 개발하였다. 또한 분리된 균주중 메치실린에 내성을 나타내는 균주를 검출하고자 penicillin-binding protein, PBP2a (mecA)를 coding 하는 209 bp의 mecA 유전자의 증폭, melting curve 분석 및 DNA염기서열을 분석하여 메치실린내성 황색 포도상구균을 real-time PCR 기법으로 검출하는 기술을 개발하였다. 본 실험에 따르면 647개의 원유시료중 6개의 시료에서 황색포도상구균이 검출되었으며 이중 2개의 시료에서 분리된 황생포도상구균이 메치실린내성 황색포도상구균임을 확인하였다. 또한 DNA 검출한계는 10 fg으로 기존 PCR에 비해 매우 감도가 우수한 것을 확인하였다. 또한 3개의 원유시료에서 돼지나 소의 삼출성 피부염의 원인균인 Staphylococcus chromogenes가 분리되었다.

Detection of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella spp. and Staphylococcus aureus using duplex real-time PCR assay with melting curve analysis on fresh lettuce

  • Lee, Na-Ri;Kwon, Kyung-Yoon;Choi, Sung-Wook;Koo, Min-Seon;Chun, Hyang-Sook
    • 한국식품위생안전성학회지
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    • 제26권2호
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    • pp.114-119
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    • 2011
  • 본 연구에서는 신선편의 채소와 과일에서 주로 검출되는 대장균(E. coli Ol577:H7), 리스테리아(L. monocytogenes), 살모넬라(Salmonella, spp.), 형색포도상구균(S. aureus)을 검출하고 동정할 수 있는 real time PCR 방법을 melting curve 분석을 활용하여 single tube 반응으로 두 종의 식중독균을 동시 검출하는 방법을 개발하고자 하였다. 대장균의 ${\beta}$-glucuronidase (uidA), 황색포도상구균의 thermonuclease (nuc), 리스테리아의 hemolycin (hly), 살모넬라의 tetrahionate reductase (ttr) 를 특이적으로 검출할 수 있는 4종의 프라이머 세트에 대한 real-time PCR의 melting curve 분석을 통하여 황색포도상구균과 대장균 동시분석 시 $T_m$ 값이 $80.6{\pm}0.9^{\circ}C$$86.9{\pm}0.5^{\circ}C$, 리스테리아와 살모넬라 동시분석 시 Tm 값이 $80.4{\pm}0.6^{\circ}C$$88.1{\pm}0.11^{\circ}C$로 확인하였고, 그 결과 정제되어진 각 식중독균의 genomic DNA를 주형으로 한 duplex real-time PCR 방법이 uniplex real-time PCR 방법과 마찬가지로 10 genome equivalents 까지 검출할 수 있는 민감도를 나타내었다. 또한, 양배추에 네 종의 식중독균을 접종하고, 증균배양 없이 DNA를 추출하여 duplex real-time PCR 을 수행한 결과 모든 식종목균에서 $10^3$ CFU/g의 검출 한계를 나타내었다. 결과적으로 개발된 melting curve 분석을 이용한 duplex real-time PCR 방법은 식품안전 증진을 위한 시간, 노동력, 비용 절감에 있어서 유효한 방법이 될 것으로 판단된다.

High-throughput SNP Genotyping by Melting Curve Analysis for Resistance to Southern Root-knot Nematode and Frogeye Leaf Spot in Soybean

  • Ha, Bo-Keun;Boerma, H. Roger
    • Journal of Crop Science and Biotechnology
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    • 제11권2호
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    • pp.91-100
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    • 2008
  • Melting curve analysis of fluorescently labeled DNA fragments is used extensively for genotyping single nucleotide polymorphism(SNP). Here, we evaluated a SNP genotyping method by melting curve analysis with the two probe chemistries in a 384-well plate format on a Roche LightCycler 480. The HybProbe chemistry is based on the fluorescence resonance energy transfer(FRET) and the SimpleProbe chemistry uses a terminal self-quenching fluorophore. We evaluated FRET HybProbes and SimpleProbes for two SNP sites closely linked to two quantitative trait loci(QTL) for southern root-knot nematode resistance. These probes were used to genotype the two parents and 94 $F_2$ plants from the cross of PI 96354$\times$Bossier. The SNP genotypes of all samples determined by the LightCycler software agreed with previously determined SSR genotypes and the SNP genotypes determined on a Luminex 100 flow cytometry instrument. Multiplexed HybProbes for the two SNPs showed a 98.4% success rate and 100% concordance between repeats two of the same 96 DNA samples. Also, we developed a HybProbe assay for the Rcs3 gene conditioning broad resistance to the frogeye leaf spot(FLS) disease. The LightCycler 480 provides rapid PCR on 384-well plate and allows simultaneous amplification and analysis in approximately 2 hours without any additional steps after amplification. This allowed for a reduction of the potential contamination of PCR products, simplicity, and enablement of a streamlined workflow. The melting curve analysis on the LightCycler 480 provided high-throughput and rapid SNP genotyping and appears highly effective for marker-assisted selection in soybean.

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대용량 이차전지 보호 시스템용 전류 감지 동작형 보호소자의 저융점 금속 가용체 설계 (Design of Low-Melting Metal Fuse Elements of Current Sensing Type Protection Device for Large Capacity Secondary Battery Protection System)

  • 김은민;강창룡
    • 한국전기전자재료학회논문지
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    • 제31권6호
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    • pp.427-432
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    • 2018
  • High-capacity secondary batteries can cause explosion hazards owing to microcurrent variations or current surges that occur in short circuits. Consequently, complete safety cannot be achieved with general protection that is limited to a mere current fuse. Hence, in the case of secondary batteries, it is necessary for the protector to limit the inrush current in a short circuit, and to detect the current during microcurrent variations. To serve this purpose, a fuse can be employed for the secondary battery protection circuit with current detection. This study aims at designing a protection device that can stably operate in the hazardous circumstances associated with high-capacity secondary batteries. To achieve the said objective, a detecting fuse was designed from an alloy of low melting point elements for securing stability in abnormal current states. Experimental results show that the operating I-T and V-T characteristic constraints can be satisfied by employing the proposed current detecting self-contained low melting point fuse, and through the resistance of the heating resistor. These results thus verify that the proposed protection device can prevent the hazards of short circuit current surges and microcurrent variations of secondary batteries.

저압배선용 전선퓨즈의 I-t곡선 시뮬레이션 (I-t Curve Simulation of the Low Voltage Distriubtion Line Fuse)

  • 박두기;이세현;박영범;구경완;김종식;한상옥
    • 한국전기전자재료학회:학술대회논문집
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    • 한국전기전자재료학회 1996년도 추계학술대회 논문집
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    • pp.214-217
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    • 1996
  • In this paper. we deal wish the I-t characteristic of law voltage distribution fuse. It is used to be thermal characteristic in being produced at fuse element part. The elements are divided low temperature melting element(LTME) by high temperature melting element(HTME). Those parts make of coordination. The characteristic of fuse is decided by material and design etc. used at element. We analysis I-t characteristic curve by using the numerical method. And we compared the curve of simulation with that of experiment

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휴대용 이차전지 보호 시스템용 전류 감지 동작형 보호소자의 퓨즈 가용체 설계 (Design of Fuse Elements of Current Sensing Type Protection Device for Portable Secondary Battery Protection System)

  • 강창룡;김은민
    • 전기학회논문지
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    • 제67권12호
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    • pp.1619-1625
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    • 2018
  • Portable electronic devices secondary batteries can cause fire and explosion due to micro-current change in addition to the situation of short-circuit inrush current, safety can not be secured with a general operation limited current fuse. Therefore, in secondary battery, it is necessary for the protector to satisfy both the limit current type operation in the open-short-circuit inrush current and the current detection operation characteristic in the micro current change situation and for this operation, a fuse for the current detection type secondary battery protection circuit can be applied. The purpose of this study is to design a protection device that operates stably in the hazardous situation of small capacity secondary battery for portable electronic devices through the design of low melting fuse elements alloy of sensing type fuse and secures stability in abnormal current state. As a result of the experiment, I-T and V-T operation characteristics are satisfied in a the design of the alloy of the current sensing type self-contained low melting point fuse and the resistance of the heating resistor. It is confirmed that it can prevent accidents of short circuit over-current and micro current change of secondary battery.

Sequence Distribution and Thermal Property of PEN/PBN Copolymers

  • Park, Sang-Soon;Hwang, Jeong-Jun;Jun, Ho-Wook;Im, Seung-Soon
    • Bulletin of the Korean Chemical Society
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    • 제18권1호
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    • pp.38-43
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    • 1997
  • Poly(ethylene 2, 6-naphthalate-co-tetramethylene 2, 6-naphthalate) (PEN/PBN) copolymers were synthesized and studied by 13C NMR spectroscopy, DSC analysis and X-ray diffraction. A minimum in the melting point vs. composition curve was found at approximately 60 mol% tetramethylene 2, 6-naphthalate. The PEN/PBN copolymers were shown to be statistically random throughout the range of 1, 4-butanediol compositions. The melting point depression behavior of annealed PEN/PBN copolymers depended upon the sequence propagation probability, PS, which is suggested by indivisual crystal formation of two pure comonomers; that is, ethylene-naphthalate-ethylene, EE, and tetramethylene-naphthalate-tetramethylene, BB. However, it can be seen from the X-ray curve that the peaks of PEN/PBN copolymers appear from a crystal lattice which is governed only by the rich component between two different aliphatic units in the copolymer composition.

$^{3}$He의 고체-액체 융해압력 측정 ($^{3}$He Solid-Liquid Melting Pressure Thermomety)

  • 김상보;최창호;이일수;김동락
    • 한국초전도저온공학회:학술대회논문집
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    • 한국초전도저온공학회 1999년도 제1회 학술대회논문집(KIASC 1st conference 99)
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    • pp.161-164
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    • 1999
  • $^{3}$He melting curve thermometer is made, tested, and compared with Greywall scale. The experimental results show that we can identify the temperature within a few percentage. To improve the accuracy of our thermometer we need temperature fixed points like $^{3}$He superfluid phase transition temperature.

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가스압력조절식 나트륨 열관전기로를 이용한 은 고정점 실현과 고온백금저항온도 계측에의 응용 (Realization of silver fixed-point and application for the high temperature platinum resistance thermometry using the gas-pressure controlled sodium heat-pipe furnace)

  • 감기술;최인묵;양인석;김용규
    • 센서학회지
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    • 제18권2호
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    • pp.116-121
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    • 2009
  • Pressure-controlled sodium heat pipe furnace was designed and fabricated, and its characteristics was investigated. Pressure control system was controlled within ${\pm}0.5\;Pa$ at 150 kPa and the stability of pressure was decreased to ${\pm}2.5\;Pa$, when the pressure-controlled system connected with the heat pipe. The melting curve of Ag fixed-point realized well by the adiabatic method using the pressure-controlled sodium heat pipe furnace and its accuracy showed ${\pm}2.27\;mK$ from the calculation of 20% to 80% at the plateau. The freezing curve of Ag fixed-point also realized and its plateau value was 2.23 mK lower than that of the melting curve.

Genotypic Identification of Cystoisospora in Immunocompromised Patients Using Tm-Variation Analysis

  • Basyoni, Maha M.A.;Elghobary, Hany Ahmed Fouad
    • Parasites, Hosts and Diseases
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    • 제55권6호
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    • pp.601-606
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    • 2017
  • Cystoisospora is responsible for morbidity in immunocompromised patients. PCR is sensitive for diagnosing Cystoisospora; however, it needs reevaluation for differential molecular diagnosis of cystoisosporiasis. We aimed at evaluating melting curve analysis (MCA) after real-time PCR (qPCR) in diagnosis and genotyping of Cystoisospora as an alternative to conventional PCR. We included 293 diarrheic stool samples of patients attending the Department of Clinical Oncology and Nuclear Medicine of Cairo University Hospitals, Egypt. Samples were subjected to microscopy, nested PCR (nPCR), and qPCR targeting the internal transcribed spacer 2 region (ITS2) of the ribosomal RNA (r RNA) gene followed by melting temperatures ($T_ms$) analysis and comparing the results to PCR-RFLP banding patterns. Using microscopy and ITS2-nPCR, 3.1% and 5.8% of cases were Cystoisospora positive, respectively, while 10.9% were positive using qPCR. Genotyping of Cystoisospora by qPCR-MCA revealed 2 genotypes. These genotypes matched with 2 distinct melting peaks with specified $T_ms$ at $85.8^{\circ}C$ and $88.6^{\circ}C$, which indicated genetic variation among Cystoisospora isolates in Egypt. Genotype II proved to be more prevalent (65.6%). HIV-related Kaposi sarcoma and leukemic patients harbored both genotypes with a tendency to genotype II. Genotype I was more prevalent in lymphomas and mammary gland tumors while colorectal and hepatocellular tumors harbored genotype II suggesting that this genotype might be responsible for the development of cystoisosporiasis in immunocompromised patients. Direct reliable identification and differentiation of Cystoisospora species could be established using $qPCR-T_ms$ analysis which is useful for rapid detection and screening of Cystoisospora genotypes principally in high risk groups.