• Title/Summary/Keyword: 실시간 중합효소연쇄반응

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Supraclavicular BCG Lymphadenitis Noted at 21 Months after BCG Vaccination Confirmed by a Molecular Method (분자생물학적 방법으로 확진한 접종 21개월 후에 발생한 BCG 쇄골상부 림프절염 1례)

  • Lee, Min Hyun;Chae, Moon-Hee;Park, Kyoung Un;Cho, Hye-Kyung
    • Pediatric Infection and Vaccine
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    • v.21 no.2
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    • pp.139-143
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    • 2014
  • Bacille Calmette-$Gu\acute{e}rin$ (BCG) lymphadenitis is the most common complication of BCG vaccination. It commonly occurs in infants aged <6 months involving ipsilateral axillary lymph nodes. We described BCG lymphadenitis in a 22-month-old boy presenting swelling of left supraclavicular lymph node that was confirmed by real-time polymerase chain reaction (PCR) and the multiplex PCR targeting the region of difference (RD).

Ethidium monoazide-PCR for the detection of viable Escherichia coli in aquatic environments (수환경에서 살아 있는 대장균의 검출을 위한 ethidium monoazide-중합효소연쇄반응법)

  • Lee, Gyucheol;Kim, Hyunjeong;Lee, Byunggi;Kwon, Soonbok;Kim, Gidon;Lee, Sangtae;Lee, Chanhee
    • Journal of Korean Society of Water and Wastewater
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    • v.23 no.2
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    • pp.199-205
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    • 2009
  • It is very important to differentiate of DNA derived from live or dead bacteria within mixed microbial communities in aquatic environments. Ethidium monoazide (EMA) is a DNA intercalating agent and the treatment of EMA with strong visible light cleaves the genomic DNA of bacteria. In dead bacterial cells, EMA intercalates into the genomic DNA, induces the cleavage of DNA, and inhibits the PCR amplification. In this study, we developed the EMA-PCR and EMA real-time PCR to detect the DNA derived from viable Escherichia coli (E.coli) in mixed cultures of live and dead E.coli. The treatment of EMA, $50{\mu}g/mL$, and 650 W visible halogen light exposure for 2 minutes cleaved the genomic DNA derived from heat killed E.coli but did not those of live E.coli. EMA-PCR could detect the DNA from live E.coli in mixed culture samples of live and dead E.coli at various ratio and there was no DNA amplification in only dead E.coli cultures. Similar results were observed in EMA real-time PCR. Further studies are needed to develop various EMA-PCR methods to detect viable waterborne pathogens such as Helicobacter pylori, Giardia lamblia, and so on.

생쥐 초기 배아에서 Aquaporin 8과 9의 발현에 관한 연구

  • 신현상;계명찬;강수만;이성은;이지원;강한승;김문규
    • Proceedings of the Korean Society of Developmental Biology Conference
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    • 2003.10a
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    • pp.68-68
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    • 2003
  • Aquaporin은 막관통 통로 단백질(transmembrane channel protein)로서, 삼투압의 농도구배에 따라 세포막을 가로질러 물분자를 이동시키는 기능을 하고 있다. 포유류 초기배아에서 포배강 형성은 영양외배엽세포에서 $Na^+ / K^+$ATPase에 의한 이온 농도 구배가 형성되면 auqaporin에 의해 물이 포배강으로 유입되면서 이루어진다. 본 연구에서는 생쥐 초기배아에서 반정량적인 역전사 중합효소 연쇄반응 방법(semi-quantitative RT-PCR)과 실시간 역전사 중합효소 연쇄반응 방법(real-time RT-PCR)을 통하여 AQP8과 9의 mRNA발현을 조사하고 다중 면역형광현미경 방법(confocal immunofluorescence microscopy)을 통해 단백질 발현양상을 분석하였다. AQP8 mRNA는 상실기까지 발현되지 않다가 포배기에 이르러 발현되었고 AQP9 mRNA는 수정란에서부터 발현되어 포배기에는 유의할 정도로 증가하였다. 따라서 AQP8 mRNA는 배아유전자가 활성화되어 나타나는 것이고 AQP9 mRNA는 모계유전자 기원임을 알 수 있었다. AQP8 단백질은 상실배 단계까지 발현되지 않다가 포배시기에 영양외배엽세포사이의 접합면에 발현되었고 AQP9 단백질은 상실배 시기에 할구 사이의 인접 부위에서 강하게 발현되었다가 포배시기에는 세포간의 접합면에 약하게 발현하는 경향을 나타내었다. 실시간 역전사 중합효소 연쇄반응 방법으로 조사한 결과 포배에서 물과 글리세롤을 통과시키는 AQP9는 mRNA의 발현양이 AQP8보다 약 4배 정도 많았다. 또한 포배기에 이르러서야 물만을 통과시키는 AQP8의 발현이 나타나는 것을 보아 포배강 형성시 외부에서 영양외배엽을 통해 포배강으로 유입되는 물의 이동(trans- trophectodermal water movements)에 AQP9보다 AQP8이 더 중요하게 관여할 것으로 사료된다., K, Pb, Cd, Cr, Co, Cu, Ni)을 측정하였다. 실험 조건1의 결과로서 각 국의 유아용 일회용 기저귀의 중금속 함량은 거의 유사한 경향을 나타내었으며 Cr, Zn, Pb, Ni, Mn, Mg, Li, K는 detection limit(2 ppm) 이하였고, Cd, Fe, Co, Cu, Ca, Al, Sr는 검출되었지만 기준치 이하였다. 실험 조건2의 결과로서 측정 항목(Cr, Sb, Cd, Pb, Ni, Co, Cu)중 Cr, Cd, Ni, Cu는 detection limit(0.1 ppm) 이하였고, Sb, Pb, Co는 검출되었지만 기준치 이하였다.았다. 4%의 경우에는 8$0^{\circ}C$이하로 온도를 낮추는 것이 좋은 상태를 나타내었다. 이와 같은 결과는 일반적으로 화학적 레팅을 4%, 7%에서한 선행결과와 상당히 다른 결과이다.염 농도가 증가할수록 감소 현상을 보였다.X>, 75BG30은 8.6$\mu\textrm{m}$, 75BG40은 7.02$\mu\textrm{m}$로 나타났다. 따라서 경화제 양에 관계없이 10$\mu\textrm{m}$ 이하로 나타나, 경화제 10$m\ell$만으로 미세한 크기를 얻을 수 있음을 알 수 있다. 젤리 강도 변화에 따른 차이는 300BF는 78.09$\mu\textrm{m}$ 300BG는 56.32$\mu\textrm{m}$로, 75BF나 75BG에 비하여 현저히 증가하여, 젤라틴의 젤리 강도는 캡슐 제조 조건의 주요한 변수임을 알 수 있다.추출물 투여시 혈당강하 및 혈중콜레스테롤 강하가 나타났으며, 상엽복합추출물 투여와 운동을 병행시 이러한 감소 효과가 더 뚜렷하게 나타났다.교육의 적임자로 보는 시각이 비교적 높았고 약 1/2정도는 영양교육에 참여하겠다는 의지를 가지고 있을 뿐만 아니라 실제로 영양지도를

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Development of real-time PCR Detections against 11 Pathogens of Bombus Species (뒤영벌 병원체 11종에 대한 실시간 중합효소 연쇄반응 검출법 개발)

  • Min, Sang-Hyun;Kim, Jung-Min;Lim, Su-Jin;Kim, Byoung-Hee;Lee, Chil-Woo;Yoon, Byoung-Su
    • Journal of Apiculture
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    • v.32 no.2
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    • pp.99-109
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    • 2017
  • The multiple real-time PCRs against pathogens of Bombus species including DWV, IAPV, KBV, SBV, BQCV, kSBV, SBPV and Paenibacillus larvae, Mellisococcus plutonius, Lysinibacillus fusiformis, and Klebsiella oxytoca have been developed. One extracted nucleic acid from Beesample could be applied to 11 different PCRs in same time and condition. Specific PCR-products were amplified qualitative and quantitative manner inner 20 minutes successfully, when each 1000 molecules of pathogen-specific target DNA is existed as template, respectively. The multiple PCR detection that we propose would be expected to apply to quarantine test for international exchange of Bombus species.

The Detection and Diagnosis Methods of Infectious Viroids caused Plant Diseases (식물체에 감염성 질병을 유발하는 바이로이드 검출 및 진단 방법)

  • Lee, Se Hee;Kim, Yang-Hoon;Ahn, Ji-Young
    • Journal of Life Science
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    • v.26 no.5
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    • pp.620-631
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    • 2016
  • Viroids are about 250-400 base pair of short single strand RNA fragments have been associated with economically important plant diseases. Due to the lack of protein expression capacity associated with replication, it is very difficult to diagnosis viroid diseases in serological methods. For detecting viroid at plants, molecular-based techniques such as agarose gel electrophoresis, polyacrylamide gel electrophoresis (PAGE), DNA-hybridization, blotting analysis and conventional RT-PCR are reliable. Real-time RT-PCR methods that grafted on RT-PCR methods with improved confirmation methods have been also utilized. However, they are still labor-intensive, time-consuming, and require personnel with expertise. Loop-mediated Isothermal Amplification (LAMP) method is a nucleic acid amplification method under the isothermal condition. The LAMP methodology has been reported to be simple, rapid, sensitive and field applicable in detecting a variety of pathogens. The results of LAMP method can be colorized by adding a visible material such as SYBR green I, Evagreen, Calcein, Berberine and Hydroxy naphthol blue (HNB) with simple equipment or naked eyes. The combination of LAMP method and nucleic pathogens, viroids, can be used to realize simple diagnosis platform for the genetic point-of care testing system. The aim at this review is to summary viroid-caused diseases and the simple visible approach for diagnosing viroids using Loop-mediated Isothermal Amplification (LAMP) method.

Development and Validation Study of Biological Agent Detection Kit (생물학작용제 검출 키트 개발 및 성능시험 연구)

  • Joe, Hae Eun
    • Journal of the Korea Institute of Military Science and Technology
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    • v.22 no.4
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    • pp.575-580
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    • 2019
  • In biological warfare, it is important to identify biological agents for proper treatment. We focused on developing a real-time RT-PCR kit that can detect multiple species of biological agents. AccuPower(R) Biothreat Real-Time RT-PCR Kit(v3.0) could detect Bacillus anthracis, Yersinia pestis, Vibrio cholerae, Francisella tularensis, Salmonella typhi, Rickettsia prowazekii, Variola virus, Hantaan virus, Yellow fever virus, Brucella spp., Shigella dysenteriae in a single reaction. The results showed that the kit was verified to be able to detect at least 0.005 ng of nucleotide and 10,000 CFU/ml of bacteria. Therefore, the kit is expected to be used as a rapid and sensitive detection kit for 11 species of biological agents within 2 hours.

Evaluation of the Effect of Mine Drainage on the Aquatic Environment by Quantitative Real-time PCR (실시간 정량 중합효소연쇄반응을 이용한 광산 배수의 수계 영향 평가)

  • Han, Ji-Sun;Seo, Jang-Won;Ji, Won-Hyun;Park, Hyun-Sung;Kim, Chang-Gyun
    • Journal of Korean Society of Environmental Engineers
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    • v.32 no.2
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    • pp.121-130
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    • 2010
  • Metals and sulfate can be considerably dissolved at low pH condition in the acid mine drainage(AMD) and it would make an environmental problems. There are only few of acid mine drainage treatment systems in Korea which are operating, but these still have an effect on the surrounding stream. In this study, quantification of indicator microorganisms was conducted to judge the environmental impact of AMD on microflora by quantitative real-time PCR in the drainage samples of four mines and the water samples of each surrounding stream. Two species of iron reducing bacteria(Rhodoferax ferrireducens T118 and Acidiphilium cryptum JF-5) were selected for indicator bacteria based on 16S rRNA cloning analysis, and sulfate reducing bacteria(Desulfosporosinus orientus), iron and sulfur oxidizing bacteria(Acidothiobacillus ferrooxidans) and iron oxidizing bacteria(Leptosprillum ferrooxidans) were included into indicator since these were found in the previous studies on the mining area. Thereafter, the comparative analysis of four mines were established by the microbiological variation index and it was determined that the biological environment effect of AMD is highest in Samtan mine which doesn t contain treatment system by the value.

Taurine exerts neuroprotective effects via anti-apoptosis in hypoxic-ischemic brain injury in neonatal rats (신생 흰쥐의 저산소성 허혈성 뇌손상에서 항세포사멸사를 통한 taurine의 신경보호 효과)

  • Jeong, Ji Eun;Kim, Tae Yeol;Park, Hye Jin;Lee, Kye Hyang;Lee, Kyung Hoon;Choi, Eun Jin;Kim, Jin Kyung;Chung, Hai Lee;Seo, Eok Su;Kim, Woo Taek
    • Clinical and Experimental Pediatrics
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    • v.52 no.12
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    • pp.1337-1347
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    • 2009
  • Purpose:Taurine (2-aminoethanesulfonic acid) is a simple sulfur-containing amino acid. It is abundantly present in tissues such as brain, retina, heart, and skeletal muscles. Current studies have demonstrated the neuroprotective effects of taurine, but limited data are available for such effects during neonatal period. The aim of this study was to determine whether taurine could reduce hypoxic-ischemic (HI) cerebral injury via anti-apoptosis mechanism. Methods:Embryonic cortical neurons isolated from Sprague-Dawley (SD) rats at 18 days gestation were cultured in vitro. The cells were divided into hypoxia group, taurine-treated group before hypoxic insult, and taurine-treated group after HI insult. In the in vivo model, left carotid artery ligation was performed in 7-day-old SD rat pups. The pups were exposed to hypoxia, administered an injection of 30 mg/kg of taurine, and killed at 1 day, 3 days, 1 week, 2 weeks, and 4 weeks after the hypoxic insult. We compared the expressions of Bcl-2, Bax, and caspase-3 among the 3 groups by using real- time polymerase chain reaction (PCR) and western blotting. Results:The cells in the taurine-treated group before hypoxic insult, although similar in appearance to those in the normoxia group, were lesser in number. In the taurine-treated group, Bcl-2 expression increased, whereas Bax and caspase-3 expressions reduced. Conclusion:Taurine exerts neuroprotective effects onperinatal HI brain injury due to its anti-apoptotic effect. The neuroprotective effect was maximal at 1-2 weeks after the hypoxic injury.

Heat Shock Protein 90 Gene Expression in Juvenile Sea Cucumber Apostichopus japonicus (Echinodermata; Holothuroidea) according to Releasing Methods (어린 돌기해삼 Apostichopus japonicus (Echinodermata; Holothuroidea) 방류 방법에 따른 열충격단백질90 유전자의 발현 분석)

  • Lee, Dong-Han;Lee, Seungheon;Jeong, Dong-Bin;Sohn, Young Chang
    • Journal of Marine Life Science
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    • v.7 no.1
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    • pp.29-36
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    • 2022
  • Sea cucumber, Aposticopus japonicus, is a major invertebrate species in the coastal regions of Korea. To evaluate the short-term stress levels according to the releasing methods, this study investigated the gene expression profiles of heat shock protein 90 (HSP90) by real-time quantitative polymerase chain reaction. When the juvenile sea cucumbers were packed in the vinyl bag with oxygen followed by transportation for 30 min or air-exposed for 1 h, the HSP90 gene expression levels in the experimental groups were significantly increased compared to those of the control groups (transported group, p=0.001; air-exposed group, p=0.032). The experimental group at 6 h post-release by seed-spreading method and at 2~6 h post-release by underwater hose-releasing method on board a fishing boat showed that the levels of HSP90 gene expression were not statistically significant but decreased slightly compared to the control group (seed-spreading group, p=0.069; hose-releasing group, p=0.093). On the other hand, the HSP90 gene expression showed an increasing pattern as the time passed (~6 h) after underwater release of juvenile sea cucumbers by divers (p=0.061). These results suggest that HSP90 gene expression can be used to investigate short-term stress response and effective releasing methods of juvenile sea cucumbers.

Design of an Inexpensive Heater using Chip Resistors for a Portable Real-time Microchip PCR System (저항소자를 이용한 휴대형 Real-time PCR 기기용 히터 제작)

  • Choi, Hyoung-jun;Kim, Jeong-tae;Koo, Chi-wan
    • Journal of IKEEE
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    • v.23 no.1
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    • pp.295-301
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    • 2019
  • A heater in a portable real-time polymerase chain reaction(PCR) system is one of the important factors for controlling the PCR thermocycle precisely. Since heaters are integrated on a small-sized PCR chip for rapid heating and fabricated by semiconductor processes, the cost of producing PCR chips is high. Here, we propose to use chip resistors as an inexpensive and accurate temperature control method. The temperature distribution was simulated using one or two chip resistors on a real-time PCR chip and the PCR chip with uniform temperature distribution was fabricated. The temperature rise and fall rates were $18^{\circ}C/s$ and $3^{\circ}C/s$, respectively.