• Title/Summary/Keyword: 중합효소연쇄반응법

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Assessment of decontamination of gutta-percha cone and the change of surface texture after rapid chemical disinfection (화학소독제 처리 후 가타파차 콘의 멸균 효과 및 표면 성상의 변화 평가)

  • Pang, Nan-Sim;Jung, Il-Young;Yu, Yoon-Jung;Kum, Kee-Yeon
    • Restorative Dentistry and Endodontics
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    • v.31 no.2
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    • pp.133-139
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    • 2006
  • The purposes of this study were firstly to identify the microbial species on gutta-percha (GP) cones exposed at outpatient clinics using polymerase chain reaction, and secondly to evaluate the rapid sterilization effect of two chemical disinfectants at chair side. It also evaluated the alteration of surface texture of GP cones after 5-min soaking into two chemical disinfectants. A total of 100 GP cones from two endodontic departments were randomly selected for microbial detection using PCR assay with universal primer. After inoculation on the sterilized GP cones with the same microorganism identified by PCR assay, they were soaked in two chemical disinfectants: 5% NaOCl and 2% chlorhexidine for 1, 3, 5, and 10 minutes. The sterilization effect was evaluated by turbidity and subculture. The change of surface textures using a scanning electron microscope was also examined after 5 min-soaking in two chemical disinfectants. Results showed that four bacterial species were detected in 17 GP cones, and all the species belonged to the genus Staphylococcus. Two chemical disinfectants were effective in sterilization with just 1 minute soaking. On the SEM picture of NaOCl-soaked GP cone, a cluster of cuboidal crystals was seen on the cone surface. Present data demonstrate that two chemical disinfectants are useful for rapid sterilization of GP cone just before obturation at chair side, while CHX-soaked GP cone has cleaner surface without crystal precipitation than that of NaOCl-treated cone.

Allele Frequency of the Short Tandem Repeat Locus Human Lipoprotein Lipase(LPL) Gene by Polymerase Chain Reaction in the Korean Population (한국인에서 중합효소연쇄 반응법에 의한 STR 유전좌위 LPL의 유전자빈도 검색)

  • Na, Yun-Ju;Hur, Woong;Yoon, Chang-Lyuk
    • Journal of Oral Medicine and Pain
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    • v.22 no.2
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    • pp.253-260
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    • 1997
  • 한국인 집단에서 개인식별의 기초자료로 활용하고자 한국인 201명을 대상으로 STR 유전좌위 중 하나인 LPL 유전좌위의 유전자 빈도 및 유전자형 분포를 구하였다. 혈액으로부터 추출한 핵 DNA를 중합효소연쇄반응으로 증폭시키고 폴리아크릴아마이드 겔 상에서 전기영동하여 은염색한 후 관찰하여 다음의 결과를 얻었다. 1. 한국인 집단 201명의 LPL 유전자에서 5개의 대립유전자, 7개의 유전자형을 검출하였으며, 이형접합도는 50.7%로 나타났고 대립 유전자다양성 (allelic diversity value)은 0.454, 개 인식 별력 (PD)은 0.674를 보였다. 2. 대립 유전자 및 유전자빈도는 9, 10, 11, 12, 13 대립 유전자에서 각각 0.020, 0.714, 0.100, 0.164, 0.002로 나타났으며, 대립유전자 7, 8, 14는 관찰되지 않았다. 이상의 결과를 볼 때 한국인 집단에서 STR LPL유전좌위의 유전자빈도는 친자감정 등 개인식별에 유용하게 사용할 수 있으나 감정실무에 응용시 다수의 STR유전좌위 및 VNTR유전좌위의 분석을 병행하여야 할 것으로 사료된다.

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Development of a Method to Detect Cattle Material from Processed Meat Products Using a Polymerase Chain Reaction (PCR을 이용한 축산물 가공식품 내 소고기 성분 검출법 개발)

  • Kwon, Young Chul;Hah, Do-Yun;Heo, Yunwi;Kim, Tae-Kyu;Choi, Yoo-Jeong;Jo, Dae-Hoon;Nam, Sang-Yun;Son, Byeong-Guk;Hwang, Bo-Won;Yang, Byoung-Seon;Kim, Euikyung
    • Korean Journal of Clinical Laboratory Science
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    • v.49 no.2
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    • pp.135-140
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    • 2017
  • Polymerase chain reaction (PCR) was used to detect cattle material from processed meat products. Seventy-eight different commercial processed meat products were purchased from several big food marts. Among them, 17 products contained cattle material (10 samples contained only cattle, 5 samples mixed with cattle and porcine, 2 samples mixed with cattle, porcine and chicken). The genomic DNA was extracted directly from the processed meat products, and strain-specific primer targeting the 16S ribosomal RNA mitochondrial gene was used. All PCR products were cloned into the pGEM-T easy vector and sequenced. Consequently, the PCR products were amplified from 10 processed meat products, which contained only cattle material in our conditions. Furthermore, PCR reactions showed the same results at mixed samples. The DNA sequence obtained from pGEM-T easy/PCR products showed more than 95% identity with Bos taurus 16S rRNA gene using homology analysis. In conclusion, we suggest that the method using PCR, as performed in this study, could be useful in detecting cattle material in processed meat products. Moreover, our system could be applicable in inspection procedures to improve the verification of correct labeling for import and export processed meat products.

Diagnosis of Bovine Theileriosis by Direct PCR and Electrophoresis from Whole Blood Without DNA Extraction (DNA 추출없이 전혈을 이용한 PCR-전기영동법에 의한소의 타일레리아병 진단)

  • Seong Ho Kang;Sangmin Jang;Joon-Seok Chae;Yongseong Kim
    • Journal of the Korean Chemical Society
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    • v.47 no.2
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    • pp.127-132
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    • 2003
  • We have developed a direct polymerase chain reaction (PCR) and electrophoresis method for the diagnosis of bovine theileriosis from whole blood DNA analysis without DNA extraction. The technique empolyed a FoLT (formamide, low temp.) technique and was utilized in the diagnosis of bovine theileriosis. Formamide solubilize the blood cells, and the lowered incubation temperatures reduced protein coagulation. 100-200 nL of whole blood and PCR reagents were introduced directly into a PCR tube. After the amplification, the PCR product (816-bp DNA) was introduced into the electrophoresis system. The results of this analysis were consist with those obtained using purified DNA.

Development and Evaluation of a SYBR Green Real-time PCR Assay for Canine Cytokine Gene Expression (SYBR Green 실시간 역전사 중합효소연쇄반응을 이용한 개 싸이토카인 유전자 발현의 정량)

  • Yu, Do-Hyeon;Ihn, Dong-Chul;Park, Chul;Park, Jin-Ho
    • Journal of Veterinary Clinics
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    • v.27 no.5
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    • pp.508-513
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    • 2010
  • Cytokines are important mediators of the immune response, and quantitating cytokine mRNA is a highly sensitive and attractive method for measuring cytokine production. The objective of the current study was to develop and validate a SYBR green quantitative real-time reverse transcriptase PCR (qRT-PCR) assay for measuring canine cytokine mRNA. The optimal annealing temperatures ($T_a$) of the designed primers were $62^{\circ}C$ for interleukin (IL)-$1{\beta}$, IL-6 and IL-10; $60^{\circ}C$ for glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and tumor necrosis factor (TNF)-${\alpha}$; and $58^{\circ}C$ for high mobility group box 1 (HMGB1). Primer efficiencies of all primers calculated for standard curve samples were between 97.1% and 102.6%. No evidence of secondary structure or primer-dimer formation was seen via melt-curve analysis or gel electrophoresis. The developed qRT-PCR assays are highly specific and sensitive and can be used to quantify gene expression levels of canine cytokines.

The Evaluation of Recornbinant Circumsporozoite Protein in Malaria Diagnosis (말라리아 진단시 재조합 Circumsporozoite 단백질의 유용성 평가)

  • 이형우;이종수;이원자;조신형;이호자
    • Korean Journal of Microbiology
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    • v.36 no.2
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    • pp.142-149
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    • 2000
  • It had been evaluated the recombinant Circumsporozoite(CS) protein of Plasmodium viva in serologic diagnosis of vivax malaria. Western blot was done to analyse the sera of malaria patients according to the days after onset. The sera which have the terms within 15 days were shown 43.8%(14/32) of positive rates and the sera over the 16 days were shown 94.4%(17/18) of positive rates. So the total positive rate was 62%(31/50). It was 22.6%(7/31) which was shown negative response in Western blot, even though they were shown positive response in Immuuofluorescent antibody test(1FAT) using whole blood stage antigens. The positive rate of non-epidemic area(Yechon-gun, Kyongsangbuk-do) was 10.7%(3/28), and epidemic area(Kangwha-gun, Inchon-shi) was 27.6%(13/47) in Western blot analysis using recombinant CS protein. In order to applicate the recombinant CS protein in seroepidemiological survey, blood samples of 422 inhabitants were collected who lived in malaria epidemic areas, Chosm-ri, Majeong-ri, Hyangyang-ri and Noejo-n in Paju-shi, Kyonggi-do. All of them were negative in microscopic examination and two(0.5%) of them were positive in Polymerase Chain Reaction. 42(10.0%) of them were seropositive in FAT using whole blood antigens and 71(16.8%) of them were seropositive in Enzyme-linked immunosorbent assay using recombinant CS protein. It was figured out the positive rates were much higher according to the distances of villages which were closed to the demilitalized zone(DMZ) in all kind of diagnostic methods, respectively.

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Direct detection of cariogenic streptococci in metal brackets in vivo using polymerase chain reaction (교정용 메탈 브라켓에서 자가중합효소연쇄반응을 통한 치아우식증 원인균의 탈출)

  • Ahn, Sug-Joon;Lee, Shin-Jae;Baek, Seung-Hak;Kim, Tae-Woo;Chang, Young-Il;Nam, Dong-Seok;Lim, Bum-Soon
    • The korean journal of orthodontics
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    • v.35 no.4 s.111
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    • pp.312-319
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    • 2005
  • Streptococcus mutans and Streptococcus sobrinus are major etiological agents in enamel demineralization around orthodontic appliances. This study was designed to examine the prevalence of these streptococci on orthodontic brackets in vivo using polymerase chain reaction. Four incisor brackets in the upper and lower arches were removed and collected from 80 patients at the time of debonding. The genomic DMA of adhered bacteria was extracted and each dextranase gene of S. mutans and S. sobrinus was amplified using the specific oligonucleotide primers. The results showed that the maxillary incisor brackets were colonized by both cariogenic streptococci to a somewhat higher degree than that taken from the mandible. The prevalence of S. mutans was $50.0\%$ on the maxillary incisor brackets and $33.8\%$ on the mandibular incisor brackets, and that of S. sobrinus was $17.5\%$ and $15.0\%$, respectively. Both species were detected on the maxillary incisor brackets of 7 patients $(8.8\%)$ and the mandibular incisor brackets of 5 patients $(6.3\%)$. These results suggest that cariogenic streptococci can adhere to the incisor brackets and may be resident species on the incisor brackets.

Detection of Poliovirus in Water by Cell Culture and PCR Methods (세포배양법과 PCR 방법에 의한 물에서의 폴리오 바이러스 검출)

  • 조연희;이찬희
    • Korean Journal of Microbiology
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    • v.38 no.3
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    • pp.198-204
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    • 2002
  • Poliovirus is a member of enterovirus which causes paralytic poliomyelitis, encephalitis and aseptic meningitis. Since poliovirus is spread by the fecal-oral route and poliovirus-contaminated water could be a potential threat for public health, detection of poliovirus in drinking water resource is important. Infectious poliovirus and poliovirus inactivated by heat or UV were used to test three detection methods such as cell culture method, reverse transcription-polymerase chain reaction (RT-PCR) and integrated cell culture (ICC)-PCR. Infectious poliovirus was detected by all three methods and ICC-PCR was the most sensitive and fast in detecting poliovirus. Inactivated polioviruses could not be detected by cell culture or ICC-PCR methods. On the other hand, heat- inactivated viruses could be detected by RT-PCR. Thus it is suggested that ICC-PCR method is the most sensitive and effective in detecting infectious polioviruses in water sample.

Genetic comparison between Spirometra erinacei and S. mansonoides using PCR-RFLP analysis (만손열두조충과 북미열두조충의 중합효소연쇄반응-마디길이여러꼴 분석법을 이용한 유전 형질 비교)

  • LEE, Soo-Ung;HUH, Sun;PHARES, C. Kirk
    • Parasites, Hosts and Diseases
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    • v.35 no.4
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    • pp.277-282
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    • 1997
  • The only observed morphological difference between Spirometra erinqsei and S. mcnsonoides is the uterine shape of the mature proglottid. Two species of worms are thought to be evolutionarily closely related. Biomolecular colnparison of the ho worms by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was conducted to observe the genetic distance. The 285 rDNA, mitochondrial cytochrome c oxidase subunit I (mCOI), and ribosomal internal transcribed spacer 1 (ITSI) fragments were obtained from the worms by PCR. The PCR products were cleaved by 5 four-base pair restriction enzyme combinations (Msp I, Hae III, Alu I, Cfo I, Rsa I) , electrophoresed and analyzed with PAUP 3.1.1. The fragment Patterns or 285 rDNA and Lni demonstrated that two worms were in identical systematic tree with bootstrap number 94 and 100, respectively As for mCOI, bootstrap number was 74 in a different tree. Above results are indicative of recent common ancestry between S. etinocei and S. mansonoides.

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