• Title/Summary/Keyword: DNA test

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Differentiation of four Mycobacterium Species using DNA-DNA Hybridization Method using Specific Probes

  • Kweon, Tae-Dong;Bai, Sun-Joon;Hong, Seong-Karp
    • Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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    • 2013.05a
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    • pp.1012-1014
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    • 2013
  • DNA-DNA hybridization method with four oligonucleotide-specific probes was used simultaneously for differentiation and identification of four Mycobacterium species (Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii). This DNA-DNA hybridization method with 4 oligonucleotide-specific probes, which targets in the rpoB region of 4 Mycobacteria species, respectively, was tested on 322 clinical isolates. Using DNA-DNA hybridization method, we detected M. tuberculosis (282 strains), M. avim (7 strains), M. intracellulare (9 strains), and M. kansasii (3 strain) from 322 clinical isolates. This result was compared with conventional biochemical test and rpoB DNA sequence analysis of this clinical isolates. We confirmed identification of Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii with high sensitivity (100 %) and specificity (100 %). This DNA-DNA hybridization method could be performed within 4 hours at least. Therefore, we suggest that DNA- DNA hybridization method using 4 rpoB DNA probes of Mycobacteria could be used for accurate, rapid, convenient detection and identification of Mycobacterium tuberculosis, M. avium, M. intracellulare, and M. kansasii in clinical samples.

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Virulence-associated plasmids of Salmonella spp. isolated from animals in Korea (동물에서 분리된 Salmonella균의 병원성 관련 Plasmid에 관한 연구)

  • Choi, Won-pil;Jung, Suk-chan
    • Korean Journal of Veterinary Research
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    • v.32 no.3
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    • pp.369-376
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    • 1992
  • This paper dealt with plasmid DNA profile in 98 Salmonella(S) isolated from pigs and cattle sources in Taegu, Gyeongbook and Gyeongnam during the period from 1984 to 1987. Also we were studied for restriction enzyme analysis of the plasmid DNA, and mouse infection, Sereny test and normal setum resistance test in guinea pig for S typhimurium and S enteritidis harbored or cured 60 megadalton(Md) plasmid and 36 Md plasmid, respectively. Of the 13 Salmonella isolated from cattle, 7 Salmonella harbored one or more plasmids and molecular sizes of the large plasmids were 60 Md for S typhimurium and 36 Md for S enteritidis. Of the 85 Salmonella isolated from pigs, 47 Salmonella were confirmed as being one or more plasmids, and all the S typimurium stains harbored 60 Md plasmid. In enzyme digestion with 8 types of restriction endonuclease for 60 Md plasmid DNA of S typhimurium, cleavage patterns were varied to enzymes, and the DNA was segmented into 4 to 15 fragments. In restriction enzyme analysis of 36 Md plasmid DNA obtained from four strains of S. enteritidis, the DNA showed the same cleavage patterns obtained with Eco RI, Hind III and Bam H I, and was segmented into 3 to 5 fragments. In virulence for mice by measuring the 50% lethal dose ($LD_{50}$), the $LD_{50}$ values obtained for 60 Md virulence-associated plasmid harbored strains of S typhimurium and 36 Md virulence-associated plasmid of S enteritidis were up to $10^4$-fold lower than the values obtained for the plasmid-cured strains of the same serotype. Only the plasmid harbored strains were resistant to the bactericidal activity of 90% guinea pig serum, and only they gave positive responses in sereny test. We suggested that their plasmid DNA might be associated with virulence for mice.

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Effect of Ginseng Extracts on the Binding to DNA of Benzo(a)pyrene Metabolites in uitro in Rats (DNA와 Benzo(a)pyrene 대사물질 결합형성에 미치는 인삼 추출물의 영향)

  • 박진규;고지훈
    • Journal of Ginseng Research
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    • v.13 no.1
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    • pp.37-41
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    • 1989
  • Reactive metabolites generated by benzo(a)pyrene(BP) monooxygenase(AHH) interact with nucleophiles in DNA and cause mutation and carcinogenesis. We studied the effect of Panax ginseng C.A. Meyer, which induce epoxide hydratase(EH) activity without concomitant induction of AHH activity, on the binding of BP metabolites to DNA in uitro in Sprague Dawley rats. DNA-BP metabolite adducts can be resolved into at least five distinct peaks by elution of a Sephadex LH-20 column with a water methanol gradieNt. These peaks are arbitrarily designated A(most polar) through I(least polar). Of the 5 peaks tentatively assigned to 7,8 biol-9,10-oxide(A),7,8·oxide(B),4,5-oxide(C), and further metabolites of 9-OH-BP(D & E), peaks A, C, D, and I were reduced to 70, 85, 80, and 30% of controls, respectively, and there was no significant change in peak B. In connection with this DNA binding study, BP metabolizing enzymes including AHH, EH, demethylase(DM) activity and cyt. P-450 contents were also investigated in order to compare the BP treated control with ginseng and BP treated test groups. The results showed that the EH activity was increased by 139% over the BP control, the Cyt. P-450 content was increased by 180% over the control value, and DM and AHH activities were also increased to some degree for the BP test group, but there was no significant effect of the ginseng treatment.

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DNA Damage and Micronuclei Induced by Di (2-ethylhexyl) phthalate in Human Breast Carcinoma MCF-7 cells (Di(2-ethylhexyl) phthalate에 의해 유도된 DNA손상과 소핵 형성)

  • 김종원;한의식;박미선;엄미옥;김인숙;전혜승;정해관;심웅섭;오혜영
    • Environmental Mutagens and Carcinogens
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    • v.21 no.1
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    • pp.34-43
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    • 2001
  • Di-2-ethylhexyl phthalate (DEHP) is the most commonly used phthalate ester in polyvinyl chloride formulations including food packing and storage of human blood. DEHP is a well known as non-genotoxic carcinogen and endocrine disrupting chemical (EDC). DEHP have shown all negative results in ICH-guildeline recommended standard genotoxicity test battery. In this study, to assess the clastogenic and DNA damaging effect in human-derived tissue specific cells, DEHP was treated in human derived MCE-7 cells, HepG2 cells, LNCap cells, BeWo cells, MCE-10A cells, and female peripheral blood cells using micronucleus assay and in human breast carcinoma MCF-7 cells up to $1.28$\times$10^{-2}$ M using Comet assay. The in vitro micronucleus assay is a mutagenicity test system for the detection of chemicals which induce the formation of small membrane bound DNA fragment i.e. micronuclei in the cytoplasm of interphase cells, originated from clastogenic and/or aneugenic mechanism. The single cell gel electrophoresis assay (Comet assay) is used to detect DNA strand-breaks and alkaline labile site. In our results, DEHP increased significantly and/or dose-depentently and time-dependently micronucleus frequency at the 6 and 24 hr without metabolic activation system only in MCE-7 cells. DEHP treated with 2 hrs in MCF-7 cells using Comet assay induced DNA damage dose-depentantly.

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Validation of Photo-comet Assay as a Model for the Prediction of Photocarcinogenicity

  • Kim, Ji-Young;Koh, Woo-Suk;Lee, Mi-Chael
    • Toxicological Research
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    • v.22 no.4
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    • pp.423-429
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    • 2006
  • Recent reports on the photocarcinogenicity and photogerotoxicity of many compounds led to an increasing awareness for the need of a standard approach to test for photogenotoxicity. The comet assay has been recently validated as a sensitive and specific test system for the quantification of DNA damage. Thus, the objectives of this study are to investigate the utility of photo-comet assay for detecting photo-mutagens, and to evaluate its ability to predict rodent photo-carcinogenicity. Photo-comet assays were performed using L5178Y $Tk^{+/-}$ mouse lymphoma cells on five test substances (8-methoxypsoralen, chlorpromazine, lomefloxacin, anthracene and retinoic acid) that demonstrated positive results in photocarcinogenicity tests. For the best discrimination between the test substance-mediated DNA damage and the undesirable DNA damage caused by direct UV absorption, a UV dose-response of the cells in the absence of the test substances was firstly fnalized. Out of 5 test substances, positive comet results were obtained for chlorpromazine, lomefloxacin, anthracene and retinoic acid while 8-methoxypsoralen found negative. An investigation into the predictive value of this photo-comet assay for determining the photocarcinogenicity showed that photo-comet assay has relatively high sensitivity. Therefore, the photo-comet assay with mammalian cells seems to be a good and sensitive predictor of the photocarcinogenic potential of new substances.

DNA Ploidy as a Predictive Index of Therapeutic Response in Lung Cancer (폐암환자에서 치료에 대한 반응 예측지표로서의 DNA Ploidy)

  • Choi, In-Seon;Lee, Shin-Seok;Yang, Jae-Beom;Park, Kyung-Ok;Jung, Sang-Woo
    • Tuberculosis and Respiratory Diseases
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    • v.39 no.2
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    • pp.150-158
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    • 1992
  • Background: Although many authors have reported that the median survival time of surgically resected non-small cell lung cancer (NSCLC) was shorter in aneuploid than in diploid determined by flow cytometry, there are few reports about DNA ploidy using bronchial brushing material in all types of lung cancer. Method: The DNA ploidy test results of 109 consecutive patients with lung cancer were analyzed to find the relationship of DNA ploidy and anatomic or physiologic stage. And the differences of the response to various therapeutic modalities according to DNA ploidy were evaluated at least 8 weeks after the begining of the therapy. Results: Numbers of patients with DNA aneuploid pattern or high proliferative activity (S+G2M>22%) were not different among the various cell types of lung cancer. The relationship of DNA ploidy and anatomic or physiologic stage was not significant. However, NSCLC patients with high proliferative activity showed more advanced anatomic stage than those without that (p<0.05). The short-term response rate to therapy depended on the anatomic (p<0.005) or physiologic stages (p<0.05) in patients with NSCLC, and not on DNA ploidy or proliferative activity. Conclusion: DNA ploidy test using bronchial brushing material revealed that high proliferative activity means advanced anatomic stage, but it was not useful to predict the therapeutic response.

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Pathological Factors Affecting DNA Quality in BRAF, EGFR, and KRAS Gene Molecular Tests (BRAF, EGFR, KRAS 유전자 분자병리검사에서 DNA 품질에 영향을 미치는 병리학적인 인자에 관한 연구)

  • Yun, Hyon-Goo;Kim, Bo-Ra;Lee, Joo-Mi;Song, Eun-Ha;Kim, Dong-Hoon
    • Korean Journal of Clinical Laboratory Science
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    • v.52 no.4
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    • pp.381-388
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    • 2020
  • The quality control of pathological specimens is important for accurate molecular pathology testing. This study evaluated that specimen factors affecting the DNA quality during tissue processing and sample types for BRAF, EGFR, and KRAS mutations tests. One thousand seven hundred and seventy-two molecular pathology tests were investigated for the factors influencing the DNA quality, such as sample type, formalin fixation time, and reexamination status. Cytology samples stored in a saline solution had better DNA quality than commercial cytology preservation. Tissue samples fixed in formalin within 24 hours had better DNA quality than the samples fixed over 24 hours. Between the types of samples, fresh tissue samples and tissue samples with a high tumor cell density had relatively better DNA quality than the formalin-fixed paraffin-embedded (FFPE) tissues and cytology specimens. Of real-time PCR, the non-PNA Ct value increased proportionally with samples held for longer than 24 hours in formalin, and that the formalin-fixed time affects the sample DNA quality. In conclusion, the appropriate tumor cellularity and 10% neutral formalin fixation time are the most important factors for maintaining the DNA quality. These factors should be managed properly for an accurate pathological molecular test to ensure optimal DNA quality.

Isolation of Plasmid DNA in Pediococci from Kimchi (김치의 Pediococci에 존재하는 Plasmid DNA 분리)

  • Park, Yun-Hee;Ryu, Uk-Sang;Jo, Do-Hyun
    • Applied Biological Chemistry
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    • v.31 no.1
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    • pp.33-37
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    • 1988
  • Three species of Pediococci, Pediococcus pentosaceus, Pediococcus acidilactici and Pediococcus halophilus were isolated from Kimchi. P. pentosaceus and P. acidilactici showed inhibitory activity against Streptococcus faecalis, Pseudomonas sp., P20 and Vibrio parahaemolyticus. However, the growth of all test organisms was not inhibited by P. halophilus. Ten strains contained one to seven plasmids, ranging in size from 1 to 60 megadaltons.

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한우 6번 염색체의 Bootstrap기법을 이용한 우수 DNA 탐색

  • Lee, Je-Yeong;Yeo, Jeong-Su;Kim, Jae-Woo;Lee, Yong-Won;Kim, Mun-Jeong
    • 한국데이터정보과학회:학술대회논문집
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    • 2003.05a
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    • pp.41-47
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    • 2003
  • 한우 6번 염색체 유전자 지도에서 한우의 질을 높이기 위한 QTL(quantitative trait loci)분석을 실시하여 선별된 Loci 값을 Permutation Test를 이용하여 계산하였다. 한편, 경제적으로 주요한 한우의 특성부위(질적부위와 육량등)에 따른, 우수 경제형질 DNA marker를 K-평균 군집법을 실시 파악하였다. 이들 QTL과 K-평균법에 의해 한우의 염색체 6번, ILST035의 주요 경제 형질별 DNA marker들을 선별하여, Bootstrap BCa방법을 이용하여 각 DNA marker들의 신뢰구간을 구했다.

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Advantages of the single nucleotide polymorphism-based noninvasive prenatal test

  • Kim, Kunwoo
    • Journal of Genetic Medicine
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    • v.12 no.2
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    • pp.66-71
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    • 2015
  • Down syndrome screening with cell-free DNA (cfDNA) in the maternal plasma has recently received much attention in the prenatal diagnostic field. Indeed, a large amount of evidence has already accumulated to show that screening tests with cfDNA are more sensitive and specific than conventional maternal serum and/or ultrasound screening. Globally, more than 1,000,000 of these noninvasive prenatal tests (NIPTs) have been performed to date. There are several different methods for NIPTs that are currently commercially available, including shotgun massively parallel sequencing, targeted massively parallel sequencing, and single nucleotide polymorphism (SNP)-based methods. All of these methods have their own advantages and disadvantages. In this review, I will focus specifically on the SNP-based NIPT.