• Title/Summary/Keyword: Trans-gene

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Light Regulated Plant Gene Expression (빛에 의한 식물 유전자의 발현)

  • 한태룡
    • Proceedings of the Botanical Society of Korea Conference
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    • 1987.07a
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    • pp.63-79
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    • 1987
  • Light regulates a variety of genes in higher plants. The expression of light-induced plant genes is regulated at the level of transcription via red- light photomorphogenic receptor, phytochrome, as well as unknown blue light photoreceptor(s). Ribulose-5-phosphate carboxylase/oxygenase (Rubisco) small subunit (SSB) and light harvesting chlorophyll a/b (Cab) protein are those of the best understood genes regulated by light. 5'-upstream flanking sequence (- -400) of Rubisco SSB and Cab genes sis known as a light responsive, enhance-like element. It responses to red and blue light in transgenic plant system as a tissue specific manner. Phytochrome gene is also regulated by light. In contrast to most of the light regulated plant genes, it is negatively controlled by red light. Search for the cis- and trans-acting factors responsible for the light signal is in progress to understant photomorphogenesis and development in higher plants.

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Molecular Approaches for Cloning of Important Higher Plant Genes (고등식물의 유용 유전자 크로닝을 위한 분자적 접근)

  • ;Ala
    • KSBB Journal
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    • v.10 no.1
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    • pp.89-96
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    • 1995
  • An Avabidofsis thaliana gene encoding phosphoribosyl anthranilate transferase is shown to be the gene that is defective in blue fluorescent trp 1 mutant plants. This gene, named PAT1, coding region is homologous to those for the phosphoribosyl anthranilate transferase from many microorganisms. This is due to a defect in tryptophan biosynthesis that leads to an accumulation of anthranilate, a fluorescent intermediate in the tryptophan pathway. PAT1 is a single-copy gene that complements all of the visible phenotypes of the different trp1 mutants. Experiments to determine the regulation of the PAT1 gene are in progress. The wild-type PAT1 promoter and several promoter deletions of PAT1 gene have been transformed into Arabidopsis tryptophan mutants. These constructs might identify promoter elements that control this patterns. We have isolated the homozygous lines in T3 seeds and analysed the protein levels using PAT antibody and PAT protein level increased two fold in pHSl07. Finally, the potential of using PAT1 as a selectable marker or visible reporter of gene expression is being explored.

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Molecular cloning, sequences analysis and in vitro expression of the dihydroflavonol 4-reductase gene from Gypsophila paniculata L. (안개초(Gypsophila paniculata L.)로부터 dihydroflavonol 4-reductase 유전자의 분리 및 분석)

  • Min, Byung-Whan;Cheong, Dong-Chun
    • Journal of Plant Biotechnology
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    • v.37 no.1
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    • pp.89-95
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    • 2010
  • Dihydroflavonol 4-reductase (DFR) is a key enzyme of the flavonoid biosynthesis pathway which catalyses the NADPH-dependent reduction of 2R,3R-trans-dihydroflavonols to leucoanthocyanidins. In this study we describe cloning and expression of the genes encoding the flavonoid-biosynthetic enzyme DFR in Gypsophila paniculata L. Inspection of the 1279 bp long sequence revealed an open reading frame 1063 bp, including a 36 bp 5' leader region and 181 bp 3' untranslated region. Comparison of the coding region of this DFR cDNA sequence including the sequences of Arabidopsis thaliana, Citrus sinensis, Dianthus caryophyllus, Ipomoea batatas, Matthiola incana, Nierembergia sp, Petunia hybrida, Solanum tuberosum, Vitis vinifera reveals an identity higher than 69% at the nucleotide level. The function of this nucleotide sequences was verified by comparison with amino acid sequences of the amino-terminus and tryptic peptides from purified plant enzyme, by northern blotting with mRNA from wild type and mutant plants, by in vitro expression yielding and enzymatically active reductase, as indicated by the small leucopelargonidin peak. Genomic southern blot analysis showed the presence of only one gene for DFR in Gypsophila paniculata.

Improved Production of Long-Chain Fatty Acid in Escherichia coli by an Engineering Elongation Cycle During Fatty Acid Synthesis (FAS) Through Genetic Manipulation

  • Jeon, Eunyoung;Lee, Sunhee;Lee, Seunghan;Han, Sung Ok;Yoon, Yeo Joon;Lee, Jinwon
    • Journal of Microbiology and Biotechnology
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    • v.22 no.7
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    • pp.990-999
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    • 2012
  • The microbial biosynthesis of fatty acid of lipid metabolism, which can be used as precursors for the production of fuels of chemicals from renewable carbon sources, has attracted significant attention in recent years. The regulation of fatty acid biosynthesis pathways has been mainly studied in a model prokaryote, Escherichia coli. During the recent period, global regulation of fatty acid metabolic pathways has been demonstrated in another model prokaryote, Bacillus subtilis, as well as in Streptococcus pneumonia. The goal of this study was to increase the production of long-chain fatty acids by developing recombinant E. coli strains that were improved by an elongation cycle of fatty acid synthesis (FAS). The fabB, fabG, fabZ, and fabI genes, all homologous of E. coli, were induced to improve the enzymatic activities for the purpose of overexpressing components of the elongation cycle in the FAS pathway through metabolic engineering. The ${\beta}$-oxoacyl-ACP synthase enzyme catalyzed the addition of acyl-ACP to malonyl-ACP to generate ${\beta}$-oxoacyl-ACP. The enzyme encoded by the fabG gene converted ${\beta}$-oxoacyl-ACP to ${\beta}$-hydroxyacyl-ACP, the fabZ catalyzed the dehydration of ${\beta}$-3-hydroxyacyl-ACP to trans-2-acyl-ACP, and the fabI gene converted trans-2-acyl-ACP to acyl-ACP for long-chain fatty acids. In vivo productivity of total lipids and fatty acids was analyzed to confirm the changes and effects of the inserted genes in E. coli. As a result, lipid was increased 2.16-fold higher and hexadecanoic acid was produced 2.77-fold higher in E. coli JES1030, one of the developed recombinants through this study, than those from the wild-type E. coli.

Effect of Transgenic Genotype on Transgene Expression in Mud Loach (Misgurnus mizoIepis): I. Copy Number-Dependent Expression in Gynogenetically Derived Homozygous Transgenics

  • Nam Yoon Kwon;Noh Jae Koo;Kim Dong Soo
    • Fisheries and Aquatic Sciences
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    • v.4 no.1
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    • pp.39-46
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    • 2001
  • To examine the effect of copy number-dependent transgenic genotype on the expression of foreign gene, stable hemizygous and homozygous transgenic breeding line was established using artificial parthenogenesis. For this purpose, induced diploid gynogenetic transgenesis was optimized in mud loach (Misgurnus mizolepis) using UV-irradiated cyprinid loach (M. anguillicaudatus) sperm and thermal shocks. Optimum UV range for inactivation of cyprinid loach sperm was between 3,150 to $4,050\;ergs/mm^2$ The UV-irradiated sperm were inseminated into eggs from recessive color strain (yellow) or heterozygous transgenic mud loach containing CAT gene. Cold shock at $2^{\circ}C$ for 60 min, 5 min post fertilization successfully restored the diploidy of eggs inseminated with UV-irradiated sperm. Restoration to diploidy was confirmed by flow cytometry and gynogenetic status was verified by examining maternal exclusive inheritance of multi-locus DNA fingerprints, body color and transgenic marker. Putative isogenic transgenic fish clearly showed homozygous status at trans gene locus based on Southern blot hybridization and progeny testing. Further, such homozygous gynogenetic diploids revealed the increased levels of transgene expression, when compared to those of heterozygous (hemizygous) transgenic fish.

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Identification of a Gene Encoding Adenylate Kinase Involved in Antifungal Activity Expression of the Biocontrol Strain Burkholderia pyrrocinia CH-67

  • Lee, Kwang Youll;Kong, Hyun-Gi;Lee, Seon-Woo
    • The Plant Pathology Journal
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    • v.28 no.4
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    • pp.373-380
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    • 2012
  • Burkholderia pyrrocinia CH-67 is a biocontrol bacterium with strong antifungal activity against several plant pathogenic fungi. Transposon mutagenesis was performed to identify the genes responsible for the antifungal activity of B. pyrrocinia CH-67. Of the 2,500 mutants tested using the Fulvia fulva spore screening method, a mutant deficient in antifungal activity, M208, was selected. DNA sequence analysis of the transposon-inserted region revealed that a gene encoding an adenylate kinase-related kinase was disrupted in M208. Antifungal activity was restored in M208 when a full-length adenylate kinase gene with its promoter was introduced in trans. The deduced amino acid sequence of adenylate kinase from CH-67 was 80% identical to that of B. cenocepacia MCO-3. Adenosine diphosphate supplementation or high levels of adenosine triphosphate and adenosine monophosphate together restored antifungal activity in M208, suggesting that adenylate kinase of B. pyrrocinia CH-67 is involved in antifungal activity expression.

Genetic Transformation of Chlamydomonas reinhardtii with the RNAi Suppressor p19 Gene of Tombus Virus (Tombus 바이러스의 RNAi Suppressor p19 유전자에 의한 Chlamydomonas reinhardtii의 형질전환)

  • Jeong, Won-Joong;Liu, Jang-Ryol;Cerutti, Heriberto
    • Journal of Plant Biotechnology
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    • v.34 no.4
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    • pp.307-312
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    • 2007
  • Chlamydomonas reinhardtii was transformed with the coding sequence of the Tombus virus gene p19 to determine whether the gene functions as an RNAi suppressor in C. reinhardtii. Transformants were confirmed to have 1 to several copies of p19 gene in their chromosomes. When an RNAi strain of C. reinhardtii generated by transforming the inverted repeat (IR) sequence homologous to the 3'UTR region of the MAA7 gene was re-transformed with the gene p19, MAA7 transcript levels of transformants fluctuated and proliferation of trans-formants on the medium containing 5-FI was suppressed. Overall results suggest that p19-mediated silencing suppression works at a low level in C. reinhardtii because of difference in codon usage resulting in weak P19 expression unless p19-mediated silencing suppression in C. reinhardtii works in a different manner from higher plants.

IN2001 Regulates CYP3A4 Gene Expression in Hep G2 Cells

  • Ahn, Mee-Ryung;Kim, Dae-Kee;Sheen, Yhun-Yhong
    • Environmental Mutagens and Carcinogens
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    • v.24 no.4
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    • pp.171-179
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    • 2004
  • Cytochrome P4503A4(CYP3A4) is the most abundnat CYPs in human liver, comparising approximately 30% of the total liver CYPs contents ans is involbed in the metabolism of more than 60% of currently used therapeutic drugs. The expression of CYP3A4 is induced by a variety of structurally unrelated xonobiotics including the antibiotic rifampicin and endogenous hormones, and might be mediated through steroid and xenobiotic receptor(SXR) system. The molecular mechanisms underlying regulation of CYP3A4 gene expression hae not been understood. In order to gain the insight of the molecular mechanism of CYP3A4 gene expression, study has been undertaken to investigate if the histone deacelylation is involved in the regulation of CYP3A4 gene expression by proximal promoter or not. Also SXR was investigated to see if they were involved in the regulation of CYP3A4 proximal promoter activity. HepG2 or Hena-I cells were transfected with a plasmid containing~1kb of the CYP3A4 proximal promoter region (-863 to +64bp) cloned in front of a reporter gene, luciferase, in the presence or absence of SXR or hER. Transfected cells were treated with CYP3A4 inducers such as rifampicin, PCN and RU 486, or with estradiol, in order to exmine to regulation of CYP3A4 gene expression in the presence or absence of trichostatin A (TSA). In HepG2 cells, CYP3A4 inducers and estradiol increased significantly the luciferase activity by CYP3A4 proximal promoter, only when TSA was co-treated after SXR cotransfection. In the case of Hepa-I cells CYP3A4 inducers and estradiol incressed modestly the luciferase activity when TSA was co-treated, but this increment was not enhanced by SXR cotransfection in contrast to HepG2 cells. Taken together, these results indicated that the inhibition of histone deacetylation was required to SXR-mediated increase in CYP3A4 proximal promoter region when rifampicin, or PCN was treated. Futher a trans-activation by SXR may demand other species-specific transcription factors.

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Positive Regulation of Pyoluteorin Biosynthesis in Pseudomonas sp. M18 by Quorum-Sensing Regulator VqsR

  • Huang, Xianqing;Zhang, Xuehong;Xu, Yuquan
    • Journal of Microbiology and Biotechnology
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    • v.18 no.5
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    • pp.828-836
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    • 2008
  • The biocontrol rhizobacterium Pseudomonas sp. M18 can produce two kinds of antibiotics, namely pyoluteorin (Plt) and phenazine-1-carboxylic acid (PCA), and is antagonistic against a number of soilborne phytopathogens. In this study, a luxR-type quorum-sensing regulatory gene, vqsR, was identified and characterized immediately downstream of the Plt gene cluster in strain MI8. A vqsR-inactivated mutant led to a significant decrease in the production of Plt and its biosynthetic gene expression. However, this was restored when introducing the vqsR gene by cloning into the plasmid pME6032 in trans. The vqsR mutation did not exert any obvious influence on the production of PCA and its biosynthetic gene expression and the production of N-acylhomoserine lactones (C4 and C8-HSLs) and their biosynthetic gene rhlI expression. Accordingly, these results introduce VqsR as a regulator of Plt production in Pseudomonas spp., and suggest that the regulatory mechanism of vqsR in strain M18 is distinct from that in P. aeruginosa. In addition, it was demonstrated that vqsR mutation did not have any obvious impact on the expression of Plt-specific ABC transporters and other secondary metabolic global regulators, including GacA, RpoS, and RsmA.

Genome-wide analysis of Solanum lycopersicum L. cyclophilins

  • Khatun, Khadiza;Robin, Arif Hasan Khan;Islam, Md. Rafiqul;Jyoti, Subroto Das;Lee, Do-Jin;Kim, Chang Kil;Chung, Mi-Young
    • Journal of Plant Biotechnology
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    • v.49 no.1
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    • pp.15-29
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    • 2022
  • Cyclophilins (CYPs) are highly conserved ubiquitous proteins belong to the peptidyl prolyl cis/trans isomerase (PPIase) superfamily. These proteins are present in a wide range of organisms; they contain a highly conserved peptidyl-prolyl cis/trans isomerase domain. A comprehensive database survey identified a total of 35 genes localized in all cellular compartments of Solanum lycopersicum L., but largely in the cytosol. Sequence alignment and conserved motif analyses of the SlCYP proteins revealed a highly conserved CLD motif. Evolutionary analysis predicted the clustering of a large number of gene pairs with high sequence similarity. Expression analysis using the RNA-Seq data showed that the majority of the SlCYP genes were highly expressed in mature leaves and blooming flowers, compared with their expression in other organs. This study provides a basis for the functional characterization of individual CYP genes in the future to elucidate their role(s) in protein refolding and long-distance signaling in tomatoes and in plant biology, in general.