DOI： 10.5511/plantbiotechnology.19.0807a

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1016/j.scienta.2019.05.003

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記述言語：日本語   掲載種別：研究論文（大学，研究機関等紀要）   出版者・発行元：新潟大学農学部  

黄葉・銅葉・赤葉などカラフルな色彩の葉をもつ植物はカラーリーフプランツと呼ばれ、近年その人気が高まっている。多くの場合、それらの葉の細胞ではアントシアニン類が多量に生合成・蓄積されている。アントシアニン類の生合成にはフラボノイド生合成酵素遺伝子の発現制御が関わっており、その転写因子としてR2R3-MYBが知られている。本研究では、宿根トレニア(Torenia concolor)におけるカラーリーフプランツの育成を目的として、ホトトギス(Tricyrtis sp.)およびシロイヌナズナ(Arabidopsis thaliana)由来R2R3-MYB遺伝子(TrMYB1およびPAP1)を異所発現する形質転換体の作出を試みた。導入したR2R3-MYB遺伝子の種類に関わらず、形質転換体は葉色の変化の程度により3タイプ(ほとんど変化なし、薄い赤紫色に変化、濃い赤紫色に変化)に分けることができた。アントシアニンの定量分析および外来R2R3-MYB遺伝子の発現解析を行ったところ、形質転換体における葉色の変化の程度と葉の総アントシアニン含量および外来遺伝子の発現レベルには相関がみられた。これらの結果から、宿根トレニアへのTrMYB1またはPAP1の導入および異所発現により、葉においてアントシアニン類の生合成・蓄積が誘導され、葉色が改変されることが示唆された。今後は、同様の手法を用いて、他の花き園芸植物におけるカラーリーフプランツの作出を検討する予定である。

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その他リンク： http://hdl.handle.net/10191/50915

記述言語：英語   掲載種別：研究論文（学術雑誌）  

DOI： 10.1007/s13562-019-00487-2

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Elsevier B.V.  

R2R3-MYB transcription factors are known to activate the flavonoid biosynthetic pathway. In the present study, R2R3-MYB gene isolated from the liliaceous ornamental plant Tricyrtis sp. (TrMYB1) under the control of the cauliflower mosaic virus 35S promoter was introduced into Pelargonium crispum via Agrobacterium-mediated transformation in order to alter leaf color phenotype. Ten independent transgenic plants have been obtained, and they could be classified into three types according to the leaf color phenotype: six transgenic plants had deep yellowish-green leaves as non-transgenic plants (Type I)
two had deep red-purple leaves (Type II)
and two had deep red leaves (Type III). Spectrophotometric analysis showed that the amount of total anthocyanins significantly increased in leaves of Type II and Type III transgenic plants compared with non-transgenic and Type I transgenic plants. In addition, several anthocyanins were newly produced in leaves of Type II and Type III transgenic plants as revealed by high performance liquid chromatography analysis. Quantitative real-time reverse transcription-polymerase chain reaction analysis showed that TrMYB1 expression level correlated with the degree of leaf color alteration. Our results indicate the validity of genetic transformation with TrMYB1 for producing colored foliage in heterologous ornamental plants.

DOI： 10.1016/j.scienta.2018.06.029

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Springer Netherlands  

Tricyrtis spp., which belong to the family Liliaceae, produce unique flowers, whose tepals have many reddish-purple spots. Although elucidation of a molecular mechanism of tepal spot formation and molecular breeding for flower colour alteration are desired for Tricyrtis spp., only one flavonoid biosynthesis-related gene, TrCHS encoding chalcone synthase (CHS), has been isolated so far. In the present study, comprehensive isolation and expression analysis of the other flavonoid biosynthesis-related genes were carried out in Tricyrtis sp. Six genes (TrCHI, TrF3H, TrF3′H, TrFLS, TrDFR, and TrANS) encoding biosynthetic enzymes chalcone isomerase (CHI), flavanone-3-hydroxylase (F3H)
flavonoid 3′-hydroxylase (F3′H), flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), and anthocyanin synthase (ANS) as well as three genes (TrMYB1, TrbHLH2 and TrWDR) encoding transcription factors myeloblastosis 1 (MYB1), basic helix-loop-helix (bHLH), and WD40 repeats (WDRs) were newly isolated. Phylogenetic analysis showed that each isolated gene was classified into the monocotyledonous clade. Deduced amino acid sequences of DFRs showed that TrDFR has no substrate specificity. “Early” genes in the flavonoid biosynthetic pathway (TrCHS, TrCHI, and TrF3H) were constantly expressed in tepals during flower development, whereas expression of “late” genes (TrF3′H, TrFLS, TrDFR, and TrANS) varied with the flower developmental stage. Expression patterns of the late genes were mostly correlated with those of transcriptional factor genes, indicating that the late genes may be under the control of a transcription factor complex consisted of TrMYB1, TrbHLH2, and TrWDR. Accumulation of anthocyanins in tepals occurred slightly after transcriptional upregulation of the late genes. Results obtained in the present study may be valuable for further studies on flower colour and flower colour pattern in Tricyrtis spp.

DOI： 10.1007/s10535-018-0802-7

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記述言語：英語   掲載種別：研究論文（大学，研究機関等紀要）  

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

A considerable portion of agricultural land in central-east Japan has been contaminated by radioactive material, particularly radioactive Cs, due to the industrial accident at the Fukushima Daiichi nuclear power plant. Understanding the mechanism of absorption, translocation and accumulation of Cs+ in plants will greatly assist in developing approaches to help reduce the radioactive contamination of agricultural products. At present, however, little is known regarding the Cs+ transporters in rice. A transporter-enriched yeast expression library was constructed and the library was screened for Cs+ transporter genes. The 1452 full length cDNAs encoding transporter genes were obtained from the Rice Genome Resource Center and 1358 clones of these transporter genes were successively subcloned into yeast expression vectors; which were then transferred into yeast. Using this library, both positive and negative selection screens can be performed, which have not been previously possible. The constructed library is an excellent tool for the isolation of novel transporter genes. This library was screened for clones that were sensitive to Cs+ using a SD-Gal medium containing either 30 or 70 mM CsCl; resulting in the isolation of 13 Cs+ sensitive clones. 137 Cs absorption experiments were conducted and confirmed that all of the identified clones were able to absorb 137 Cs. A total of 3 potassium transporters, 2 ABC transporters and 1 NRAMP transporter were among the 13 identified clones.

DOI： 10.1111/ppl.12569

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：Springer Netherlands  

There was a typing error in the title of the original publication. The title is correct in this erratum.

DOI： 10.1007/s11240-017-1269-5

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

B class MADS-box genes play important roles in petal and stamen development. Some monocotyledonous species, including liliaceous ones, produce flowers with petaloid tepals in whorls 1 and 2. A modified ABCE model has been proposed to explain the molecular mechanism of development of two-layered petaloid tepals. However, direct evidence for this modified ABCE model has not been reported to date. To clarify the molecular mechanism determining the organ identity of two-layered petaloid tepals, we used chimeric repressor gene-silencing technology (CRES-T) to examine the suppression of B function in the liliaceous ornamental Tricyrtis sp. Transgenic plants with suppressed B class genes produced sepaloid tepals in whorls 1 and 2 instead of the petaloid tepals as expected. In addition, the stamens of transgenic plants converted into pistil-like organs with ovule- and stigma-like structures. This report is the first to describe the successful suppression of B function in monocotyledonous species with two-layered petaloid tepals, and the results strongly support the modified ABCE model.

DOI： 10.1038/srep24549

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記述言語：英語   掲載種別：研究論文（大学，研究機関等紀要）   出版者・発行元：新潟大学農学部  

The family Colchicaceae contains some important ornamental plants, such as Gloriosa spp. and Sandersonia aurantiaca. We have produced various intergeneric hybrid plants in this family in order to obtain wide variability in horticultural traits and to develop novel cultivars. In the present study, we examined chromosome doubling of a diploid intergeneric hybrid between L. modesta and G. superba 'Lutea' (Lit×Gsu-1; 2n=2x=22) for fertility restoration and for further widening the variability. Shoot apical segments harvested from in vitro shoot cultures were treated with various concentrations of amiprophos-methyl (APM; 10, 20 or 40 mg L^<-1>), colchicine (COL; 100, 500 or 1000 mg L^<-1>) or oryzalin (ORY; 5, 10 or 20 mg L^<-1>) for 24 h. Two months after spindle toxin treatment, the highest percentage apical segments with shoot elongation (80.0%) was obtained in 10 mg L^<-1> APM treatment. Flow cytometry analysis of elongated shoots showed that a solid tetraploid (4x) was obtained from 40 mg L^<-1> APM treatment and ploidy chimeras (2x+4x) were obtained from treatments with 10 or 20 mg L^<-1> APM and 5 or 10 mg L^<-1> ORY. Tetraploid and polyploidy chimera shoots showed compact forms with reduced internodes compared with diploid shoots.コルチカム科には、グロリオーサやサンダーソニアなど、重要な花き園芸植物が含まれている。我々は、園芸形質の拡大および新奇品種の育成のために、これまでコルチカム科内で様々な属間雑種を作出してきた。本研究では、稔性の回復および更なる園芸形質の拡大を目的として、コルチカム科の二倍体属間雑種であるリットニア × グロリオーサ'ルテア'（Littonia modesta × Gloriosa superba 'Lutea'；Lit × Gsu-1；2n=2x=22）の染色体倍加を試みた。シュート増殖培養物のシュートから茎頂を含む切片を調製し、様々な濃度のアミプロホスメチル（APM；10、20 または 40 mg L^<-1>）、オリザリン（ORY；5、10 または 20mg L^<-1>）またはコルヒチン（COL；100、500 または 1000 mg L^<-1>）を添加した液体培地で 24 時間振とう処理した。紡錘糸形成阻害剤処理２ヵ月後、10 mg L^<-1> APM 処理区において最も効率的に茎頂切片からシュートが伸長した。伸長したシュートの葉組織を用いてフローサイトメトリー分析を行ったところ、40 mg L^<-1> APM 処理区で四倍体（4x）のシュートが、また、10 または 20 mg L^<-1> APM および 5 または 10 mg L^<-1> ORY 試験区で倍数性キメラ（2x+4x）のシュートが確認された。これらの四倍体および倍数性キメラのシュートは、二倍体のシュートと比較して、節間が短縮したコンパクトな草姿を示した。

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その他リンク： http://hdl.handle.net/10191/39812

記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER  

Artemisia annua L. is a commercial source of artemisinin. Nevertheless, artemisinin content within the plant is relatively low and varies depending on genotype and environment. To broaden the genetic variability, the mutation effect of C-12-ion beam irradiation on A. annua was examined. Irradiation at 2.5 Gy had a slight lethal effect to nodal segments while a noticeable lethal effect was observed at 5 and 10 Gy. Furthermore, at higher doses (20 and 50 Gy), a severe lethal effect was observed. Mutations at the DNA level of axillary bud-derived shoots were performed by RAPD. The mutation frequency at 10 Gy was about 1.7 and 2.1 times higher than that at 2.5 and 5 Gy, respectively. After growth and artemisinin production observation of 72 irradiated mutants, around 14 and 7 % of them showed higher artemisinin content and artemisinin yield compared to the controls, respectively. The highest artemisinin content in a mutant was 1.43 % DW, which was 3.2-fold higher than the original wild type. Additionally, the highest artemisinin yield in mutants was 3.68 mg/plant, which was around 1.4-fold higher than in the wild type. Moreover, irradiated mutants exhibited antibacterial activity against S. aureus, but the wild types did not. This study presents an effective application of heavy ion beam irradiation to create variations and improve artemisinin production in A. annua.

DOI： 10.1007/s11240-014-0519-z

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記述言語：英語   掲載種別：研究論文（国際会議プロシーディングス）   出版者・発行元：International Society for Horticultural Science  

Gibberellins (GAs) are plant hormones controlling many aspects of plant growth and development including stem elongation, germination and transition from vegetative growth to flowering. GA 20-oxidase (GA 20ox) and GA 3-oxidase (GA 3ox), classes of 2-oxoglutarate-dependent dioxygenases, catalyze the conversion of precursor GAs to their bioactive forms, and therefore play a direct role in determining the levels of bioactive GAs in plants. Transgenic plants of the liliaceous ornamental Tricyrtis sp. 'Shinonome' overexpressing the GA 20ox or GA 3ox gene from Torenia fournieri (TfGA20ox2 and TfGA3oxl) were produced. After 3 years of cultivation, 4 and 2 independent transgenic plants containing TfGA20ox2 and TfGA3oxl, respectively, were subjected to morphological characterization at the flowering stage. Because GA 20ox and GA 3ox catalyze the last step in the formation of bioactive GAs, overexpression of TfGA20ox2 or TfGA3oxl was initially expected to induce a GA-overproduction phenotype in transgenic plants, such as internode elongation. However, on the contrary, all the transgenic plants exhibited reduced plant height, reduced internode length and reduced stem diameter compared with the control, non-transgenic plants, irrespective of the kind of transgene. In addition, all the transgenic plants had slender leaves and narrow flower tepals. Exogenous treatment of transgenic plants with gibberellic acid and a GA biosynthesis inhibitor, uniconazol, resulted in increased and decreased plant height, respectively. Possible factors leading to morphological alterations, observed in the present study, of transgenic Tricyrtis sp. were proposed.

DOI： 10.17660/ActaHortic.2014.1025.2

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記述言語：英語   掲載種別：研究論文（国際会議プロシーディングス）   出版者・発行元：International Society for Horticultural Science  

The class B genes, which belong to the MADS-box gene family, play important roles in regulating petal and stamen development in flowering plants. These genes exist in two different types termed DEFLCLENS (DEF)- and GLOBOSA (GLO)-like genes, and the B-function is provided by heterodimers of DEF- and GLO-like gene products. In order to understand the molecular mechanism of floral development in the agapanthaceous ornamental Agapanthus praecox ssp. orientalis, we produced and characterized transgenic A. praecox ssp. orientalis plants ectopically expressing the DEF- or GLCMike gene of the same plant, ApDEF or ApGLO. No visible morphological alterations were observed both in vegetative and floral organs of all the 7 independent transgenic plants containing ApDEF. On the other hand, in 4 out of 6 independent transgenic plants containing ApGLO, organs developed in whorl 4 showed noticeable morphological alteration: they were thick compared with carpels of non-transgenic plants, and had a branch tip. No apparent morphological alterations were observed in floral organs of the other 3 whorls. Scanning electron microscopic observations showed that the tip surface of whorl 4 organs of non-transgenic plants was covered with papilla cells, while there were few papilla cells on the tip surface of the morphologically altered whorl 4 organs of transgenic plants. In addition, epidermal cells of the morphologically altered whorl 4 organs of transgenic plants showed an intermediate morphology between those of ovaries and filaments of non-transgenic plants. Since it has been reported that ApDEF is expressed in all the 4 whorls of non-transgenic A. praecox ssp. orientalis, endogenous ApDEF products and transgenic ApGLO products may form heterodimers causing homeotic conversion in whorl 4 of transgenic plants.

DOI： 10.17660/ActaHortic.2014.1025.14

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：ELSEVIER SCIENCE BV  

Agapanthus praecox ssp. orientalis (Leighton) Leighton, a monocotyledonous ornamental plant belonging to Agapanthaceae, has recently become popular as,a potted plant for landscaping and as a cut flower. As a first step toward molecular breeding for flower color alteration in this plant, we isolated and characterized a gene encoding dihydroflavonol 4-reductase (DFR), a pivotal enzyme of the flavonoid biosynthetic pathway. A full-length cDNA clone for DFR was isolated from flower tepals of a cultivar with deep-blue flowers, and its genomic clone, designated ApDFR1 (accession number AB099529 in the GenBank/EMBL/DDBJ databases) was isolated from leaves by a polymerase chain reaction (PCR)-based strategy. Nucleotide sequence analysis revealed that ApDFR1 contains four introns and an open reading frame encoding a polypeptide of 378 amino acid residues. Deduced amino acid sequence shows 59-75% identities with those of previously reported DFR genes. Southern blot analysis showed that there are one or two copies of the DFR gene in the genome of A. praecox ssp. orientalis. ApDFR1 transcripts were detected in young flower tepals, stamens, pistils and bracts, but not in pedicles, scapes and leaves as revealed by reverse transcription-PCR analysis. When ApDFR1 was expressed under the control of the cauliflower mosaic virus 35S promoter in transgenic Petunia hybrida 'W85', a dfr-recessive line, some transgenic plants showed drastic flower color alteration froth white to red purple. High-performance liquid chromatography analysis showed that colored flower limbs of transgenic plants accumulate anthocyanidins, mainly cyanidin and petunidin. These results indicate that ApDFR1 encodes DFR and is active in a heterologous plant species. (C) 2013 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.scienta.2013.12.009

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：JAPANESE SOC PLANT CELL & MOLECULAR BIOLOGY  

The gibberellin 2-oxidase catalyzes the bioactive gibberellins or their immediate precursors to inactive forms. We have previously produced transgenic plants of the liliaceous plant Tricyrtis sp. containing the GA2ox gene from the linderniaceous plant Torenia fournieri. These transgenic plants showed dwarf phenotypes as expected but unfortunately had no flowers or only small, unopened flowers. Recently, one newly produced transgenic line (G2-55) formed fully opened flowers. G2-55 showed a moderately dwarf phenotype and the shoot length decreased to 63.4% of that of the control, non-transgenic plants. No significant differences in the number of flowers per shoot and in the flower size were observed between G2-55 and the control. Flow cytometry analysis and chromosome observation showed that G2-55 was tetraploid (2n=4x=52), whereas the other transgenic lines producing no or only small flowers were diploid (2n=2x=26) as the mother plant. Pollen fertility of G2-55 was 81.2% as determined by acetocarmine staining. The tetraploidy in G2-55 might be resulted from somaclonal variation of embryogenic calluses used as a target material for Agrobacterium-mediated transformation. The tetraploid transgenic plant G2-55 may be useable not only directly as a potted plant, but also as a material for further breeding of Tricyrtis spp.

DOI： 10.5511/plantbiotechnology.14.0916a

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記述言語：英語   掲載種別：研究論文（学術雑誌）  

Gibberellins (GAs) are the plant hormones that control many aspects of plant growth and development, including stem elongation. Genes encoding enzymes related to the GA biosynthetic and metabolic pathway have been isolated and characterized in many plant species. Gibberellin 2-oxidase (GA2ox) catalyzes bioactive GAs or their immediate precursors to inactive forms
therefore, playing a direct role in determining the levels of bioactive GAs. In the present study, we produced transgenic plants of the liliaceous monocotyledon Tricyrtis sp. overexpressing the GA2ox gene from the linderniaceous dicotyledon Torenia fournieri (TfGA2ox2). All six transgenic plants exhibited dwarf phenotypes, and they could be classified into two classes according to the degree of dwarfism: three plants were moderately dwarf and three were severely dwarf. All of the transgenic plants had small or no flowers, and smaller, rounder and darker green leaves. Quantitative real-time reverse transcription-polymerase chain reaction (PCR) analysis showed that the TfGA2ox2 expression level generally correlated with the degree of dwarfism. The endogenous levels of bioactive GAs, GA1 and GA4, largely decreased in transgenic plants as shown by liquid chromatography-mass spectrometry (LC-MS) analysis, and the level also correlated with the degree of dwarfism. Exogenous treatment of transgenic plants with gibberellic acid (GA3) resulted in an increased shoot length, indicating that the GA signaling pathway might normally function in transgenic plants. Thus, morphological changes in transgenic plants may result from a decrease in the endogenous levels of bioactive GAs. Finally, a possibility of molecular breeding for plant form alteration in liliaceous ornamental plants by genetically engineering the GA metabolic pathway is discussed. © 2013 Elsevier GmbH.

DOI： 10.1016/j.jplph.2013.05.002

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SPRINGER  

Chalcone synthase (CHS) is the key enzyme in an early stage of the flavonoid biosynthetic pathway. In the present study, a full-length cDNA clone for CHS was isolated from flower tepals of the liliaceous ornamental Tricyrtis sp., in which tepals have many reddish-purple spots resulting from accumulation of cyanidin derivatives. The deduced amino acid sequence of the isolated cDNA clone, designated TrCHS1 (accession number AB478624 in the GenBank/EMBL/DDBJ databases), shows 79.4-91.4% identity with those of previously reported CHS genes. An RNA interference (RNAi) construct targeting TrCHS1 was introduced by Agrobacterium-mediated transformation in order to alter the flower color of Tricyrtis sp. Seven transgenic plants that produced flowers could be classified into three types according to flower color phenotype: one transgenic plant had tepals with as many reddish-purple spots as non-transgenic plants (Type I); one had tepals with reduced numbers of reddish-purple spots (Type II); and five had completely white tepals without any spots (Type III). High-performance liquid chromatography analysis showed that tepals of Type III transgenic plants did not accumulate detectable amounts of anthocyanidins. In addition, TrCHS1 mRNA levels in tepals of Type II and Type III transgenic plants decreased substantially compared with non-transgenic plants, as determined by quantitative real-time reverse transcription-polymerase chain reaction analysis. Our results indicate the validity of RNAi suppression of the flavonoid biosynthetic pathway genes for flower color alteration in Tricyrtis sp. To the best of our knowledge, this is the first report on flower color alteration by genetic transformation in monocotyledonous ornamentals.

DOI： 10.1007/s11032-011-9653-z

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS LTD  

An understanding of the cadmium (Cd) transport system in rice can serve as a basis for coping with Cd-related problems in rice and human health. To identify a new gene involved in Cd transport in rice, we screened our yeast library, expressing 140 kinds of rice ATP binding cassette (ABC)-type transporters. From the screening, we found that OsABCG43/PDR5 conferred high Cd tolerance on yeast. The Cd concentration of yeast carrying OsABCG43 was similar to that of the vector control. The OsABCG43 transcript was detected both in shoots and roots. Accumulation of it was elevated by Cd treatment in the roots but not in the shoots. This study indicates that OsABCG43 is a Cd inducible-transporter gene capable of conferring Cd tolerance on yeast.

DOI： 10.1271/bbb.110193

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