• Title/Summary/Keyword: KB-C2

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Cloning, Characterization, and Functional Analysis of Maize DEHYDRIN2

  • Paek, Nam-Chon;Jung, Hun-Ki
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.47 no.2
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    • pp.116-122
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    • 2002
  • Dehydrins (LEA Dll proteins) are one of the typical families of plant proteins that accumulate in response to dehydration, cold stress, abscisic acid, or during seed maturation. A 1.3-kb cDNA was cloned from a cDNA expression library of 5-day-old germinating maize scutellums under drought stress. The deduced protein sequence indicated a dehydrin gene encoding SK$_3$ LEA protein typically expressed during cold acclimation, but not by drought stress in barley and wheat. Thus, it was named maize DEHYDRIN2 (ZmDhn2). It accumulates rapidly and highly in drought-stressed scutellum and leaf tissues at any stage, but not under cold stress. ZmDhn2 gene was transformed into Arabidopsis thaliana for functional analysis under drought condition. From electrolyte leakage test, no significant difference showed between wild type and transformants under normal growth condition, but the leakage level of electrolyte in wild type plants was about 3 times as high as that in the transformed plants under drought stress. It suggests that ZmDHN2 playa role in increasing drought tolerance.

Molecular Cloning and Characterization of Catechol 2, 3-Dioxygenase Gene from Aniline-Degrading Psseudomonas acidovorans

  • Lee, Ji-Hyun;Bang, Sung-Ho;Park, Youn-Keun;Lee, Yung-Nok
    • Korean Journal of Microbiology
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    • v.30 no.4
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    • pp.316-321
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    • 1992
  • Catechol 2, 3-dioxygenase (C230) catalyses the oxidative ring cleavage of catechol to 2-hydroxymuconic semialdehyde. This is one of the key reactions in the metabolism of the widespresd pollutant aniline. We have cloned a gene encoding C230 from cells of the aniline degrading bacteria, Pseudomonas acidovorance KCTC2494 strain and expressed in E. coli, A 11.3-kilobase Sau3A partial digested DNA fragment from KCTC2494 was cloned into phagemid vector pBluescript and designated as pLP201. The C230 gene was mapped to a 2.8-kb region, and the derection of transcription was determined. The cloned C230 gene contains its own promoter which can be recognized and employed by E. coli transcriptional apparatus. C230 activities of subclones were identified by enzyme assay and activity staining. The T7 RNA promoter/polymerase system and maxicell analysis showed that a polypeptide with Mw of 35 kDa is the C230 gene product.

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Double-Stranded RNA-Dependent Protein Kinase Gene Expression in Tobacco Plant (연초식물체에서의 dsRNA 의존성 인산화 효소 유전자 발현)

  • 이청호;박희성
    • Korean Journal Plant Pathology
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    • v.11 no.2
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    • pp.173-178
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    • 1995
  • 동물계에서 항바이러스와관련된 dsRNA 의존성 인산화 효소(PKR)의 유전자를 식물체에서 발현시킬 경우 PKR에 의한 단백질합성 및 식물바이러스의 증식조절 가능성에 대한 기초자료를 확보하기 위하여 사람에서 분리된 PKR cDNA를 Agrobacterium 방법에 의하여 연초식물체(Nicotiana tabacum cv. Xanthi-nc)로 형질전환시켰다. HindIII/PstI처리에 의해 얻어지는 약 1.8kb의 phPKR cDNA절편을 일련의 유전자 조작 방법을 통하여 식물발현벡터인 pBI121에 도입하여, p12168을 재조합하였다. 이를 A. tumefaciens LBA 4404에 형질전환시켜 연초식물체형질 전환에 이용하였다. 2mg/l BA와 0.5mg/l NAA가 포함되고 100$\mu\textrm{g}$/ml의 kanamycin이 첨가된 MS배지에서 shooting시킨 후 phytohormone이 첨가되지 않은 MS배지상에서 rooting을 시켜 형질전환 연초식물체를 얻었으며, 형질전환식물체는 정상식물체와 유사한 생육양상을 나타내었다. 형질전환식물체의 유전자도입은 hPKR cDNA의 전사부여는 RT-PCR 방법에 의하여 확인되었다.

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Molecular Cloning and Expression of dapA, the Gene for Dihydrodipicolinate Synthetase of Corynebacterium glutamicum (Dihydrodipicolinate Synthetase를 코딩하는 Corynebacterium glutamicum의 dapA 유전자의 클로닝 및 발현)

  • 오종원;한종권;이현환;현형환;이재흥;스테판정
    • Korean Journal of Microbiology
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    • v.29 no.4
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    • pp.203-208
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    • 1991
  • The dapA-complementing gene (L-2, 3-dihydrodipicolinate synthetase: DHDP synthetase, dapA) has been cloned by using a cosmid genomic bank of Corynebacterium glutamicum JS231 that is a lysine overproducer, AEC (s-(2-aminoethyl)-L-cysteine) resistant mutant. By enzymatic deletion analysis, the DNA region complementing the escherichia coli dapA host could be confined to 4.5kb SalI-generated DNA fragment. This DNA fragment was inserted into the C. glutamicum/E. coli shuttle vector pECCG117 to construct pDHDP5812. The specific activity of DHDP synthetase detected in C. glutamicum JS231/pDHDP5812 was increased about 10 fold above that of C. glutamicum JS231. The addition of leucine during growth did not repress the expressin of dapA, and the enzyme activity was not inhibited by lysine.

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CLONING AND LIGHT-DEPENDENT EXPRESSION OF A cDNA FOR PEA CYTOSOLIC FRUCTOSE-1,6-BISPHOSPHATASE

  • Son, Tae-Jong;Hahn, Tae-Ryong
    • Journal of Photoscience
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    • v.4 no.3
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    • pp.141-145
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    • 1997
  • Polymerase chain reaction(PCR) was conducted with a pea cDNA library using two primers synthesized from homology analysis of amino acid sequences for animal and plant cytosolic FBPases. A PCR product with 650 bp long was cloned into pGEM-T vector and sequenced. The deduced amino acid sequence of the cDNA fragment was 98, 91, and 85% homologous with those of cytosolic FBPases from spinach, sugarbeet, and sugarcane, respectively. It was 51% homologous with amino acid sequence of FBPase from pea chloroplasts. Northern blot analysis was proceeded with the cDNA clone resulting that 1.2 kb transcript was highly expressed in light-grown pea leaves but almost not expressed in dark-grown etiolated pea seedlings. When peas grown in the light for 10 days were transferred to darkness, the transcript was gradually decreased with dark treatment, indicating that the expression of the enzyme was induced by continuous white light but suppressed by dark treatment. Pea cytosolic FBPase was highly expressed in leaves with trace amounts in stems. but almost not expressed in roots.

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Gene Cloning and Nucleotide Sequence of Human Dihydrolipoamide Dehydrogenase-Binding Protein

  • Lee, Jeongmin;Ryou, Chongsuk;Jeon, Bong Kyun;Lee, Poongyeon;Woo, Hee-Jong;Kwon, Moosik
    • Asian-Australasian Journal of Animal Sciences
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    • v.15 no.3
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    • pp.421-426
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    • 2002
  • The pyruvate dehydrogenase complex (PDC), a member of $\alpha$-keto acid dehydrogenase complex, catalyzes the oxidative decarboxylation of pyruvate with the formation of $CO_2$, acetyl-CoA, NADH, and $H^+$. This complex contains multiple copies of three catalytic components including pyruvate dehydrogenase (E1), dihydrolipoamide acetyltransferase (E2), and dihydrolipoamide dehydrogenase (E3). Two regulatory components (E1-kinase and phospho-E1 phosphatase) and functionally less-understood protein (protein X, E3BP) are also involved in the formation of the complex. In this study, we have partially cloned the gene for E3BP in human. Nine putative clones were isolated by human genomic library screening with 1.35 kb fragment of E3BP cDNA as a probe. For investigation of cloned genes, Southern blot analysis and the construction of the restriction map were performed. One of the isolated clones, E3BP741, has a 3 kb-SacI fragment, which contains 200 bp region matched with E3BP cDNA sequences. The matched DNA sequence encodes the carboxyl-terminal portion of lipoyl-bearing domain and hinge region of human E3BP. Differences between yeast E3BP and mammalian E3BP coupled with the remarkable similarity between mammalian E2 and mammalian E3BP were confirmed from the comparison of the nucleotide sequence and the deduced amino acid sequence in the cloned E3BP. Cloning of human E3BP gene and analysis of the gene structure will facilitate the understanding of the role(s) of E3BP in mammalian PDC.

Expression of Human Lactoferricin in HC11 Cells (HC11 세포에서 인체 락토페리신의 발현)

  • Nam, Myoung-Soo
    • Korean Journal of Agricultural Science
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    • v.28 no.2
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    • pp.92-98
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    • 2001
  • Lactofenicin is an antibacterial peptide fragment (about 5 kD) derived from lactoferrin (80 kD) that displays the various biological functions. The production of a human lactoferricin (Lactoferricin H) in mouse HC11 mammary epithelial cells was achieved by placing its cDNA under the control of the bovine ${\beta}$-casein gene. To express lactoferricin H in this cell culture system, constructed a hybride-splice signal consisting of bovine ${\beta}$-casein intron I and rabbit ${\beta}$-globin intron II, and a DNA fragment spanning intron 8 of the bovine ${\beta}$-casein gene. Expression of lactofenicin H from this expression vector was identified by RT-PCR, northern and dot blot analysis. RT-PCR using total RNA of HC11 cells transfected with pBL1-cin expression vector yielded a product identified as having a size of the 150bp. Northern blot analysis was identified about 2.3 kb. In dot blot analysis, recombinant lactofenicin H was recognized with anti-human lactofrrnin polyclonal antibody.

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Biochemical and molecular characterization of a tetrachloroethylene (PCE) dechlorinating Clostridium bifermentans DPH-1

  • Chang, Young-Cheol;Toyama, Tadashi;Kikuchi, Shintaro
    • Journal of environmental and Sanitary engineering
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    • v.23 no.2
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    • pp.1-18
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    • 2008
  • The tetrachloroethylene (PCE) dehalogenase of Clostridium bifermentans DPH-1 (a halorespiring organism) was purified, cloned, and sequenced. This enzyme is a homodimer with a molecular mass of ca. 70 kDa and exhibits dehalogenation of dichloroethylene isomers along with PCE and trichloroethylene (TCE). Broad range of substrate specificity for chlorinated aliphatic compounds (PCE, TCE, cis-1,2-dichloroethylene, trans-1,2-dichloroethylene, 1,1-dichloroethylene, 1,2-dichloropropene, and 1,1,2-trichloroethane) for this enzyme was also observed. A mixture of propyl iodide and titanium citrate caused a light-reversible inhibition of enzymatic activity suggesting the involvement of a corrinoid cofactor. A partial sequence (81 bp) of the encoding gene for PCE dehalogenase was amplified and sequenced with degenerateprimers designed from the N-terminal sequence (27 amino acid residues). Southern analysis of C. bifermentans genomic DNA using the polymerase chain reaction product as a probe revealed restriction fragment bands. A 5.0 kb ClaI fragment, harboring the relevant gene (designated pceC) was cloned (pDEHAL5) and the complete nucleotide sequence of pceC was determined. The gene showed homology mainly with microbial membrane proteins and no homology with any known dehalogenase, suggesting a distinct PCE dehalogenase. So, C. bifermentans could play some important role in the initial breakdown of PCE and other chlorinated aliphatic compounds in sites contaminated with mixtures of halogenated substances.

Geochemistry and Metamorphism of the Amphibolite in the Odesan Gneiss Complex (오대산편마암복합체내에 산출되는 앰피볼라이트의 지화학적 특성과 변성작용)

  • 권용완
    • The Journal of the Petrological Society of Korea
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    • v.7 no.2
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    • pp.111-131
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    • 1998
  • The migmatitic gneiss in the Odesan Gneiss Complex has small amount of quartzite, amphibolite and marble and the Kuryong Group which contact with migmatitic gneiss unconformitly, also contains some amphibolite. Preview studies of this area had regarded that the amphibolites contact with marble had been produced by metasomatism from the pelitic and calcareous sediments mixtures, but the amphibolite is reinterpreted as igneous origin. $SiO_2$ content of the amphibolite is 45.9~52.7 wt%, which corresponds to basaltic composition. MgO content has narrow range (4.6~6.87 wt%) and major and trace element are plotted against MgO,$TiO_2, P_2O_5$, Hf, Zr are reduced and Cr and Ni are increased their content with increasing MgO. This phenomenon indicates that the basaltic magma as the protolith of the amphibolite had frationated with the crystallization of the pyroxene and/or olivine. REE pattern has smoothly decrease from LREE to HREE. Eu/Eu(0.83~1.19) show the flat Eu anomaly, which indicate small fractional crystallization of plagioclase. HREE is enriched in the garnet-bearing amphibolites. Several discrimination diagram for the basaltic magma show that the amphibolite of the study area is originated tholeiitic basaltic magma indicating continental rift environment. Due to determine the metamorphic condition garnet-hornblende geothermometry and hornblende-plagioclase geobarometry are used. Peak metamorphic temperature range of the amphibolite $788~870^{\circ}C$ and is deduced toward the northeastern part. The calculated temperature from the amphibolite has slightly higher than the temperature of the metapelites but the trend of metamorphic grade which decrease from western to eastern part progradly is similar to each other. The metamorphic pressure calculated by garnet- hornblede-plagioclase geobarometry is 4~5kb. But ilmenite-plagioclase pair enclosed in garnet show 8 kb at $700^{\circ}C$ by garnet-ilmenite-rutile-plagioclase geobarometery. The zonal profile of garnet in sample 84 shows the bell-shape profile, which grossular content decreases whereas pyrope content increases progressively. This means that the amphibolite has undergone the clockwise P-T-t path which is shown in the migmatitic gneiss of the Odesan Gneiss Complex.

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Ore Geology of Skarn Ore Bodies in the Kasihan Area, East Java, Indonesia (인도네시아 까시한지역 스카른광체의 광상학적 특성)

  • Han, Jin-Kyun;Choi, Sang-Hoon
    • Economic and Environmental Geology
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    • v.45 no.1
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    • pp.1-8
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    • 2012
  • Copper-zinc-bearing skarns of the Kasihan area developed at limestone layers in the sedimentary facies of the Late Oligocene Arjosari Formation. The skarns consist mainly of fine-grained, massive clinopyroxene-garnet, garnet, garnet-epidote, and epidote skarns. Most copper and zinc(-lead) ore mineralization occur in the clinopyroxene-garnet and garnetepidote skarn, respectively. Clinopyroxene occurs as a continuous solid solution of diopside and hedenbergite (from nearly pure diopside up to ${\approx}34$ mole percent hedenbergite), with a maximum 28.2 mole percent johannsenite component. The early and late pyroxenes of Kasihan skarns are diopsidic and salitic, respectively. They fall in the fields typical Cu- and Zn-dominated skarns, respectively. Garnet displays a relatively wide range of solid solution between grossular and andradite with up to ${\approx}2.0$ weight percent MnO. Garnet in early pyroxene-garnet skarn ranges from 49.1 to 91.5 mole percent grossular (mainly ${\geq}78$ mole % grossular). Garnets in late garnet and garnet-epidote skarns range from 2.8 to 91.4 mole percent grossular (mainly ${\geq}70$ mole % for garnet skarn). Epidote compositions indicate solid solutions of clinozoisite and pistacite varying from 65.8 to 76.2 mole percent clinozoisite. Phase equilibria indicate that skarn evolution was the result of interaction of water-rich fluids ($X_{CO_2}{\leq}0.1$) with original lithologies at ${\approx}0.5$ kb with declining temperature (early clinopyroxene-garnet and garnet skarn, ${\approx}450$ to $370^{\circ}C$; late garnet-epidote and epidote skarn, ${\approx}370$ to $300^{\circ}C$).