• Title/Summary/Keyword: hybridization

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Molecular Cloning of Serratia rnarcescens Metalloprotease Gene into Escherichia coli (Serratia marcescens Metalloprotease 유전자의 대장균에로의 클로닝)

  • 김기석;이창원;이상열;이병룡;신용철
    • Microbiology and Biotechnology Letters
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    • v.20 no.3
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    • pp.280-288
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    • 1992
  • Molecular cloning of metalloprotease gene from Serratia marcescens ATCC 21074 into Escherichia coli JM109 was carried out. Chromosomal DNA of S. marcescens was completely digested with Hind111 and southern hybridization with a synthetic oligonucleotide probe revealed that a 50 KD metalloprotease gene was contained in 4.0 Kb chromosomal DNA fragment, 4.0 Kb chromosomal DNA fragments eluted from agarose gel were ligated with pUC19 and transformed into E. coli JM109. Nine positive clones were obtained from about $1\times 10^3$ transformants by colony hybridization. Their recombinant plasmids, pSPl and pSP2 have same chromosomal DNA fragments in pUC19 in opposite-orientations. When cloned metalloprotease gene was expressed in E. coli, about 52 KD precursor protein of metalloprotease was detected by western blot analysis from E. coli harboring a recombinant plasmid pSP2. Plasmid pSP2 showed no protease activities in E. coli but overproduced the active metalloprotease in S. rnarcescens ATCC 27117.

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Detection of Plant RNA Viruses by Hybridization Using In Vitro Transcribed RNA Probes (In Viro 전사 RNA Probe를 이용한 식물 바이러스병의 진단)

  • 최장경;이종희;함영일
    • Korean Journal Plant Pathology
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    • v.11 no.4
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    • pp.367-373
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    • 1995
  • The cDNAs derived from the coat protein (CP) genes of six plant RNA viruses, tobacco mosaic virus-pepper strains (TMV-P) and -ordinary strain (TMV-OM), potato virus Y (PVY), turnip mosaic virus (TuMV), cucumber mosaic virus (CMV) and potato leafroll virus (PLRV), were subcloned into the transcription vector, pSPT18, containing SP6 and T7 promoters. The digoxigenin (DIG)-labeled RNA polymerase after linearlization of the cloned pSPTs with XbaI or SacI, and were tested for their sensitivities for the detection of the six viruses. In slot-blot hybridization, dilution end points for the detection of TMV-P and TMV-OM were 10-4, while those of PVY, TuMV and CMV were 10-3. PLRV was detected at the dilution of 10-2. When each RNA probe was applied for the detection of the viruses in the preparations from the leaf disks (8 mm in diameter, and 12 to 15 mg in weight) of infected natural host plants, TMV-P, TMV-OM and TuMV could be detected from one disk, while PVY from 1 or 2 disks. CMV was detected in the preparation from two disks, and PLRV from three disks. With DIG-labeled RNA probe, PVY was detected at 5 days after inoculation, but with ELISA the virus was detected at 8 days after inoculation to tobacco (Nicotiana tabacum cv. Xanthi nc) plants on which symptoms appeared at 9 days after inoculation. No difference was observed in cross reaction between the RNA probes for the detection of TMV-P and TMV-OM.

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Campylobacter jejuni 의 열충격 반응과 그유전자에 관한 연구

  • 김치경;임채일;이길재
    • Korean Journal of Microbiology
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    • v.30 no.3
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    • pp.232-238
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    • 1992
  • Canz~~j~lohuc;tc.~jurn i werc studied for their heat shock responses at several elevated temperatures and their heat shock genes were detected by the technique of Southern hybridization. (.. ,jc\ulcorneruni sy~>thesized the major heat shock proteins of hsp90. hsphh. and hsphO at 48$^{\circ}$C . ant1 their w~u.ival rates were maintained as the same level at optimal temperature. '1-hc heat shock genes in chromosome of C ,jc:jutii werc determined to be homologous to the heat shock genes or E. t,oli. by showing strong signals in Southern hybridization analysis using clnaK and groESL- as DNA probe But the restriction sites for thc fragmcnts including heat shock genes were different betueen E. c,oli and C ,jtjuni.

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Control of Tensile Behavior of Ultra-High Performance Concrete Through Artificial Flaws and Fiber Hybridization

  • Kang, Su-Tae;Lee, Kang-Seok;Choi, Jeong-Il;Lee, Yun;Felekoglu, Burak;Lee, Bang Yeon
    • International Journal of Concrete Structures and Materials
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    • v.10 no.sup3
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    • pp.33-41
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    • 2016
  • Ultra-high performance concrete (UHPC) is one of the most promising construction materials because it exhibits high performance, such as through high strength, high durability, and proper rheological properties. However, it has low tensile ductility compared with other normal strength grade high ductile fiber-reinforced cementitious composites. This paper presents an experimental study on the tensile behavior, including tensile ductility and crack patterns, of UHPC reinforced by hybrid steel and polyethylene fibers and incorporating plastic beads which have a very weak bond with a cementitious matrix. These beads behave as an artificial flaw under tensile loading. A series of experiments including density, compressive strength, and uniaxial tension tests were performed. Test results showed that the tensile behavior including tensile strain capacity and cracking pattern of UHPC investigated in this study can be controlled by fiber hybridization and artificial flaws.

Application of Epstein-Barr Virus Cell Lines (CCL85 EB-3) in Performing the EBER mRNA In Situ Hybridization as a Positive Control (Epstein-Barr 바이러스 인사이투 보결합 시행시 양성대조표지로서의 버키트 림프종 세포주 (CCL85 EB-3)의 응용)

  • Kim, Sung-Sook;Han, Woon-Sup;Suh, Joo-Young;Huh, Joo-Ryung
    • The Korean Journal of Cytopathology
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    • v.7 no.1
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    • pp.38-43
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    • 1996
  • Epstein-Barr virus(EBV) is associated with a wide spectrum of benign and malignant disorders including leukoplakia, Hodgkln's lymphoma, central nervous system lymphoma, peripheral T cell lymphoma and nasopharyngeal undifferentiated carcinoma. There are several distinctive aspects of biology of the virus that are important in investigation of virus in clinical specimens. The abundant expression of the EBER mRNA transcripts makes possible the sensitive detection of latent expression in EBV-associated tumors. Although there has been a dramatic increased interest in the direct characterization of EBV in clinical specimens, there have been few studios about the effective and reliable positive controls in performing in situ hybridization technique for EBV, especially on paraffin-em bedded tissue. We applied Burkitts lymphoma ceil line as positive control in EBV in situ hydridization using Oncor Kit. The cell block of Burkitt lymphoma cell line(CCL85 EB-3) showed strong and specific positivity for EBER in situ in nuclei of EBV infected cells.

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Changes of the Level of G Protein ${\alpha}-subunit$ mRNA by Withdrawal from Morphine and Butorphanol

  • Oh, Sei-Kwan
    • The Korean Journal of Physiology and Pharmacology
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    • v.4 no.4
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    • pp.291-299
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    • 2000
  • Morphine or butorphanol was continuously infused into cerebroventricle (i.c.v.) with the rate of $26\;nmol/{\mu}l/h$ for 3 days, and the withdrawal from opioid was rendered 7 hrs after the stopping of infusion. The expression of physical dependence produced by these opioids was evaluated by measuring the naloxone-precipitated withdrawal signs. The withdrawal signs produced in animals dependent on butorphanol (kappa opioid receptor agonist) were similar to those of morphine (mu opioid receptor agonist). Besides the behavioral modifications, opioid withdrawal affected G protein expression in the central nervous system. The G-protein ${\alpha}-subunit$ has been implicated in opioid tolerance and withdrawal. The effects of continuous infusion of morphine or butorphanol on the modulation of G protein ${\alpha}-subunit$ mRNA were investigated by using in situ hybridization study. In situ hybridization showed that the levels of $G\;{\alpha}s$ and $G\;{\alpha}i$ were changed during opioid withdrawal. Specifically, the level of $G\;{\alpha}s$ mRNA was decreased in the cortex and cerebellar granule layer during the morphine and butorphanol withdrawal. The level of $G\;{\alpha}i$ mRNA was decreased in the dentate gyrus and cerebellar granule layer during the morphine withdrawal. However, the level of $G\;{\alpha}i$ mRNA was significantly elevated during the butorphanol withdrawal. These results suggest that region-specific changes of G protein ${\alpha}-subunit$ mRNA were involved in the withdrawal from morphine and butorphanol.

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The Preparation of Mockeoseuk(China Fossil) Composite by Hybridization Technique and Evaluation of Its Efficacy (복합화기술을 응용한 목어석 복합체의 제조 및 이의 효능에 관한 연구)

  • Kwon, Sun-Sang;Yi, Seung-Hwan;Kim, Duck-Hee;Kim, Jun-Oh;Chang, Ih-Seop
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.33 no.3
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    • pp.153-157
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    • 2007
  • Mockeoseuk(China fossil) contains the various kinds of minerals and radiates far infrared light. In order to apply mockeoseuk to the cosmetic formulation, hybridization technique was adapted and modified by selecting a spherical silicone powder as substrate. The resultant composite improved the physical properties such as skin feeling and apparent color and still sustained the efficacy of mockeoseuk. In a clinical test, the cosmetic formulation with 10 wt% mockeoseuk composite raised the temperature of facial skin through enhancement of skin blood flow.

Two-Stage Logistic Regression for Cancer Classi cation and Prediction from Copy-Numbe Changes in cDNA Microarray-Based Comparative Genomic Hybridization

  • Kim, Mi-Jung
    • The Korean Journal of Applied Statistics
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    • v.24 no.5
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    • pp.847-859
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    • 2011
  • cDNA microarray-based comparative genomic hybridization(CGH) data includes low-intensity spots and thus a statistical strategy is needed to detect subtle differences between different cancer classes. In this study, genes displaying a high frequency of alteration in one of the different classes were selected among the pre-selected genes that show relatively large variations between genes compared to total variations. Utilizing copy-number changes of the selected genes, this study suggests a statistical approach to predict patients' classes with increased performance by pre-classifying patients with similar genetic alteration scores. Two-stage logistic regression model(TLRM) was suggested to pre-classify homogeneous patients and predict patients' classes for cancer prediction; a decision tree(DT) was combined with logistic regression on the set of informative genes. TLRM was constructed in cDNA microarray-based CGH data from the Cancer Metastasis Research Center(CMRC) at Yonsei University; it predicted the patients' clinical diagnoses with perfect matches (except for one patient among the high-risk and low-risk classified patients where the performance of predictions is critical due to the high sensitivity and specificity requirements for clinical treatments. Accuracy validated by leave-one-out cross-validation(LOOCV) was 83.3% while other classification methods of CART and DT performed as comparisons showed worse performances than TLRM.

Electrophoretic Karyotyping by PFGE in the Genus Fusarium (Fusarium속에서 PFGE를 이용한 Electrophoretic Karyotyping)

  • Min, Byung-Re;Jung, Jin-Sook;Choi, Yong-Keel
    • The Korean Journal of Mycology
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    • v.26 no.2 s.85
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    • pp.135-143
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    • 1998
  • Contour-clamped homogeneous electric field gel electrophoresis was used to establish electrophoretic karyotype for 10 species of Fusarium sections Sporotrichiella, Liseola, Gibbosum, Discolor and Martiella. Intact chromosomal DNA was isolated from fungal protoplast and separated under various conditions according to their size in order to improve DNA separation. The numbers of chromosome-sized DNA molecules for individual species ranged from 5-13, with individual chromosomes ranging from 0.78 Mb to 7.20 Mb in size. The total genome DNA size of each species was estimated at about 18.32 Mb to 48.20 Mb. Comparison of karyotype profiles following Southern hybridization analysis with a randomly selected genomic probe of F. oxysporum formae speciales litii was carried out.

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Cloning of hexavalent chromium reductase gene from E.coli ATCC 33456

  • Lee, Han-Ki;Ahn, Min-Jung;Bae, Woo-Chul;Jeong, Byeong-Chul
    • 한국생물공학회:학술대회논문집
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    • 2000.11a
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    • pp.672-675
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    • 2000
  • E.coli ATCC 33456 has relatively higher activity of Cr(VI) reduction than other microorganism. The purpose of this research is cloning of Cr(V) reductase from E.coli ATCC 33456. Using colony and southern hybridization, we selected two condidates. Among candidates, pNCR9 is higher Cr(VI) reduction activity than E.coli ATCC 33456. Purified Cr(VI) reductase antibody was reacted at estimated 42Kda protein band of candidate's crude extract on 12% SDS-PAGE. This results showed cloned gene's product is very similar to purified Cr(VI) reductase from E.coli ATCC 33456.

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