• Title/Summary/Keyword: Poly I:C

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Construction of Complementary DNA Library and cDNA Cloning for Cy Strain of Odontoglossum Ringspot Virus Genomic RNA (오돈토글로썸 윤문 바이러스 Cy계통 게놈 RNA의 cDNA 구축 및 유전자 크로닝)

  • 류기현;박원목
    • Korean Journal Plant Pathology
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    • v.10 no.3
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    • pp.228-234
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    • 1994
  • Genomic RNA was extracted from Cy strain of odontoglossum ringspot tobamovirus (ORSV-Cy) isolated from infected leaves of tobacco cv. Samsun. Size of the genomic RNA was about 6.6 kb in length. The genomic RNA was fractionated using Sephadex G-50 column chromatography into 2 fractions. They were polyadenylated at their 3'-end using E. coli poly(A) polymerase. Polyadenylated viral RNA was recovered by oligo (dT) primer adapter containing NotI restriction site and Moloney murine leukemia virus SuperScript reverse transcriptase (RNase H-). Second-strand cDNA was synthesized by using E. coli DNA ligase, E. coli DNA polymerase I and E. coli RNase H. Recombinant plasmids containing cDNAs for ORSV-Cy RNA ranged from about 800 bp to 3,000 bp. Among the selected 238 recombinants, pORCY-124 clone was the largest one covering 3'-terminal half of the viral RNA. This clone contained two restriction sites for EcoRI and XbaI and one site for AccI, AvaI, BglII, BstXI, HindIII, PstI, and TthIII 1. respectively. The clone contained partial viral replicase, a full-length movement protein and a complete coat protein genes followed by a 3' untranslated region of 414 nucleotides based on restriction mapping and nucleotide sequencing analyses. Clones pORCY-028, -068, -072, -187 and -224 were overlapped with the pORCY-124. Clones pORCY-014 and -095 covered 5' half upstream from the middle region of the viral RNA, which was estimated based on restriction mapping and partial sequence analysis. Constructed cDNA library covered more than 90% of the viral genome.

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(Photosensitive Polymers VII) Mechanism of Photosensitized Curing Reaction of Cinnamoylated Polymers ((感光性 高分子에 關한 硏究 VII) Cinnamoylated Polymers의 光增感 硬化反應機構)

  • Kim, Kwang-Sup;Shim, Jyong-Sup
    • Journal of the Korean Chemical Society
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    • v.10 no.4
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    • pp.166-174
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    • 1966
  • The multistep mechanism of photosensitized curing reaction cinnamoylated photosensitive polymer is proposed from the energy level diagram of cinnamic acid and sensitizer, and from the fact that excess of sensitizer brings the sensitivity to a limiting value etc. Various factors which have effects on the ability of sensitizer are also discussed. The mechanism involves following steps: activation to the first excited singlet states of cinnamoyl group(C) and sensitizer(S) by their absorption of photon, their intersystem crossing to the lowest triplet state, bimolecular internal quenching by formation of excimer of sensitizer, triplet excitation energy transfer and intermolecular addition between cinnamoyl group in ground state and that in triplet state. The rate equation derived from this mechanism is $-\frac{d[C]}{dt} = \frac{K_1[C]}{K_2 + [C]}[\frac{I^c_{abs}}{K_3 + [S]} + \frac{K_4[C]}{(K_5 + [C])(K_6 + [S])}(I^s_{abs} + \frac{K_7I^c_{abs}[S]}{K_8 + [S]})]$ where $I^c_{abs}\;and\;I^s_{abs}$: the rates of absorption of photon by cinnamoyl group and sensitizer $K_n$: Constants. It is proved with the cinnamate of poly(glyceryl phthalate)(PGC) in the absence of sensitizer using the infrared analytical method and successfully applied for the experimental data reported on the effects of the degree of cinnamoyl esterification and the concentration of sensitizer upon the sensitivity.

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Expression Analysis of the Mx Gene and Its Genome Structure in Chickens

  • Yin, C.G.;Du, L.X.;Li, S.G.;Zhao, G.P.;Zhang, J.;Wei, C.H.;Xu, L.Y.;Liu, T.;Li, H.B.
    • Asian-Australasian Journal of Animal Sciences
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    • v.23 no.7
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    • pp.855-862
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    • 2010
  • Among the known interferon-induced antiviral mechanisms, the Mx pathway is one of the most powerful pathways. The Mx protein has direct antiviral activity and inhibits a wide range of viruses by blocking an early stage of the viral replication cycle. Cloning, characterization, and expression of Mx in vivo and in vitro have been conducted. The chicken Mx gene spans 21 kb and is made up of 14 exons and 13 introns, of which the promoter region was analyzed. The real-time PCR results showed that Mx expression was increased in chicken embryo fibroblasts (CEF) after 12- and 24-h induction with polyI: C. Induction of Mx expression by poly I: C in vivo revealed tissue-specific patterns among the chicken tissues tested. A trace expression of Mx was detected in healthy chicken liver tissues from adult chickens without inducement; the expression levels in the liver, heart, and gizzard were higher than in the muscle and kidney. This is the first report to demonstrate the expression of a glutathione-S-transferase-tagged-Mx fusion protein of 75 KDa, as well as the biological activity tested by SDS-PAGE and western blotting.

Expression of phospholipase C β1 in olive flounder (Paralichthys olivaceus) following external stress stimulation

  • Woo, Soo Ji;Jang, Hee Young;Lee, Hyung Ho;Chung, Joon Ki
    • Fisheries and Aquatic Sciences
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    • v.19 no.4
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    • pp.18.1-18.10
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    • 2016
  • In this study, to clarify the function of $PoPLC-{\beta}1$, in response to stress challenge, we examined the $PoPLC-{\beta}1$ expression pattern in response to external stress (pathogen-associated molecular pathogen challenge and environmental challenge including temperature and salinity). $PoPLC-{\beta}1$ expression analysis of tissue from olive flounder showed that the messenger RNA (mRNA) was predominantly expressed in the brain, heart, eye, liver, spleen, and stomach. We also tested the mRNA expression of the $PoPLC-{\beta}1$ in the spleen and kidney of olive flounder by RT-PCR and real-time PCR following stimulation with lipopolysaccharide (LPS), concanavalin A (ConA), or polyinosinic:polycytidylic acid (PolyI:C) and compared with the inflammatory cytokines IL-1b and IL-6 in the stimulated flounder tissues. Each of the spleen and kidney and mRNA transcripts of $PoPLC-{\beta}1$ were increased 30- and 10-fold than normal tissue at 1-6 h post injection (HPI) with PolyI:C when the expression of $PoPLC-{\beta}1$ transcript was similar to LPS and ConA. We also tested the expression of $PoPLC-{\beta}1$ in response to temperature and salinity stress. The expression of $PoPLC-{\beta}1$ also was affected by temperature and salinity stress. Our results provide clear evidence that the olive flounder $PLC-{\beta}1$ signal pathways may play a critical role in immune function at the cellular level and in inflammation reactions. In addition, $PLC-{\beta}1$ appears to act as an oxidative-stress suppressor to prevent cell damage in fish.

Equilibrium Studies and Thermodynamics for the Mutarotation of Poly(cis-5-methylproline) in Chloroform-Chloroethanol (Chloroform-Chloroethanol 용매중에서 Poly(cis-5-methylproline)의 평형 및 열역학적연구)

  • M. J. Han
    • Journal of the Korean Chemical Society
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    • v.23 no.4
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    • pp.187-197
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    • 1979
  • The molar optical rotation of poly(cis-5-methylproline) was measured in solvent mixtures of chloroform and chloroethanol. After proper allowance for time-dependent mutarota-tions, equilibrium states between form A and form B were observed to occur with a solvent composition of 0.5~10 % chloroethanol in chloroform by volume. From the equilibrium constants, which were calculated by optical rotations at equilibrium measured at three different temperatures (5, 25, and 45 $^{circ}$C), the thermodynamic parameters-free enthalpy, enthalpy and entropy changes for the mutarotation-were evaluated. It was found that starting with equimolar concentrations of form A and form B, the forward mutarotation occurred in the solvent compositions of chloroethanol greater than 3 % by volume, whereas the reverse mutarotation resulted in solvent compositions of chloroethanol less than 3 % by volume. The changes in enthalpy and entropy for the forward mutarotation were found to be positive, while those were for the reverse mutarotation were negative. The driving forces for the forward mutarotation were found to be the increase in entropy, whereas that for the reverse mutarotation was the negative enthalpy change. The thermodynamic data were explained by the interaction between polymer and solvent, i.e., preferential hydrogen bonding of chloroethanol with the carbonyl group in form B over form A, and by difference in conformational energies between form A and form B.

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Preset State of Thermoreversible Poly(vinylidene fluoride)/propylene Carbonate Gel System: 1. Core-Shell Model (열가역적인 Poly(vinylidene fluoride)/Propylene Carbonate(PC) 겔 시스템에서의 Pregea 상태 : 1. Core-Shell 모델)

  • 박일현
    • Polymer(Korea)
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    • v.26 no.2
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    • pp.227-236
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    • 2002
  • The structure of pregel state in thermoreversible poly(vinylidene fluoride)(PVDF) /propylene carbonate(PC) system was investigated by laser light scattering. It was found that the PVDF chain did not exist as a separate chain even in a very dilute concentration(i.e. 100 times more dilute than the gel formation concentration) but as a large spherical aggregate with the radius of gyration $R_G$, of 232 nm and the effective hydrodynamic radius $R_H$= of 407 nm at $40^{\circ}C$. Based upon experimental results such as $R_H/R_G$=ratio of 1.75 and the pattern of scattering intensity with a minimum, a core-shell type sphere model was suggested as a structure of the aggregate. According to this model, the radius of core part was estimated as 215 nm, the shell thickness as 192 nm, and the ratio of monomer density of the shell part to that of the core part as about 0.075.

Preparation and Release Behaviors of Poly(ε-caprolactone) Microcapsules Containing SiO2 and Nifedipine (실리카와 니페디핀을 함유한 Poly(ε-caprolactone) 마이크로캡슐의 제조와 방출 거동)

  • Park, Soo-Jin;Lee, Yun-Mok;Han, Mijeong
    • Applied Chemistry for Engineering
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    • v.16 no.4
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    • pp.588-593
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    • 2005
  • In this study, biodegradable poly(${\varepsilon}$-caprolactone) (PCL) microcapsules containing chemically treated $SiO_2$ and nifedipine were prepared by oil-in-water (O/W) emulsion solvent evaporation method. The microcapsules containing drugs were confirmed using FT-IR spectra. The morphologies of the microcapsules were observed with scanning electron microscope (SEM). The nifedipine's release behaviors from the microcapsules were also examined with UV/vis spectroscopy. As a result, the inclusion of nifedipine into the microcapsules was determined by the presence of nifedipine's specific peak, i.e., C=O stretch vibration at $1682cm^{-1}$. The average particle size of the microcapsules decreased with increasing stirring rate. The nifedipine adsorption capacity and release rate of treated $SiO_2$ that was treated with basic solution decreased because with the increased basicity it lowered the specific surface area of $SiO_2$ and promoted stronger acid-base interactions between $SiO_2$ and nifedipine.

Molecular Cloning and Expression in Escherichia coli of a Rabbit Globin Gene (유전공학적 방법에 의한 토끼 글로빈 유전자의 재조합과 대장균에서의 발현)

  • Jang, Sung-Key;Park, Hyune-Mo
    • The Korean Journal of Zoology
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    • v.27 no.2
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    • pp.103-116
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    • 1984
  • The structural gene of rabbit hemoglobin was cloned into Pst I site of pBR322 in E. coli. The complementary DNA (cDNA) was synthesized from rabbit globin mRNA with avian myeloblastosis viral reverse transcriptase, and then RNA was destroyed at pH 11. The double stranded cDNA was synthesized with both Klenow fragment of E. coli DNA polymerase I and reverse transcriptase and then the hairpin loop was opened with Sl nuclease. Double stranded cDNA was subsequently tailed with dCTP and annealed to dGMP-tailed vector DNA. After transformation and initial screening of appropriate clones by plasmid size, the cloned colonies were identified by in situ colony hybridization using by plasmid size, the cloned colonies were identified by in situ colony hybridization using $[^32P]$-labeled cDNA probes and characterized the inserts with restriction endonucleases. The expression of cloned globin gene was investigated by standard radioimmunoassay using rat anti-rabbit Hb serum as primary antibody and goat antirat IgG serum as secondary antibody. The result suggested that the chimeric proteins (the part of $\\beta$-lactamase from the vector pBR322 and globin from rabbit) were supposedly produced in E. coli and the product had the antigenic determinant of rabbit hemoglobin.

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Growth and Properties of GaN by Vapor Transport Epitaxy (Vapor Transport Epitaxy에 의한 GaN의 성장과 특성)

  • Lee, Jae-Bum;Kim, Seon-Tai
    • Korean Journal of Materials Research
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    • v.16 no.8
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    • pp.479-484
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    • 2006
  • Highly c-axis oriented poly-crystalline GaN with a dimension of $1{\sim}3\;{\mu}m$ was deposited on $c-Al_2O_3$ substrate by vapor transport epitaxy (VTE) method at the temperature range of $900{\sim}1150^{\circ}C$. XRD intensities from (00'2) plane of grown GaNs were increased with reaction conditions which indicate the improvement of the crystal quality. In the PL spectra measured at 10 K, the spectrum composed with the neutral-donor bound exciton-related emission at 3.47 eV, crystal defect-related emission band at 3.42 eV and with its phonon replicas. The fact that intensity of $I_2$ were increased and FWHM were decreased with growth conditions means that the quality of GaN crystals were improved. With this simple VTE technology, we confirm that the GaNs were simply deposited on sapphire substrate and crystal quality related to optical properties of GaN grown by VTE were relatively good. PL emission without deep level emission in spite of polycrystalline structure can be applicable to the fabrication of large area and low cost optical devices using poly-GaN grown by VTE.

Characterization of a Low Molecular Weight Heat-Shock Protein cDNA Clone from Nicotiana tabacum

  • Park, Soo-Min;Joe, Myung-Kuk;Hong, Choo-Bong
    • Proceedings of the Botanical Society of Korea Conference
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    • 1999.04a
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    • pp.18-18
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    • 1999
  • We characterized a cDNA clone for a low molecular weight heat-shock protein (LMW HSP) from tobacco named TLHS-l. Nucleotide sequence determination of TLHS-1 identified an open reading frame for 159 amino acids. To the upstream of the open reading frame, a sequence of 124 nucleotides was determined. To the 3' downstream of the open reading frame, 212 nucleotides were identified which carried poly(A)-tail. Comparison of the open reading frame and hydropathy plot of TLHS-1 with the previously reported class I LMW HSPs showed high identity which classified TLHS-1 as a class I LMW HSP cDNA clone. We proposed that there are six consensus regions in class I LMW HSPs. RNA blot hybridization for TLHS-1 showed a typical expression pattern of heat-shock-inducible gene from three common tobacco cultivars. The open reading frame of TLHS-1 was overexpressed in Escherichia coli. TLHS-1 protein confers thermal protection of other proteins in vitro and in vivo. Thermal induced aggregation of citrate synthase was reduced by purified TLHS-1 protein, and thermal death rate at $50^{\circ}C$ was reduced in E. coli expressing TLHS-l. From these data, we can expect that TLHS-1 acts as a molecular chaperone.perone.

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