Yuki Asakura

University of Toyama, Тояма, Toyama, Japan

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Publications (5)5.55 Total impact

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    ABSTRACT: Agrobacterium-mediated transformation of leaf tissues of Atropa belladonna with an adenosine 5'-diphosphate (ADP)-ribosylation factor gene of carrot, arf-001, was performed employing pBCR82 as an expression vector. This vector co-expresses rol gene cluster together with arf-001, and thus, the transformed host cells were obtained as hairy roots. Two cell lines of the transformed belladonna were established as the liquid cultures of hairy root tissues, and expression of arf-001 and accumulation of its product in the cells were confirmed by RT-PCR and Western blot analyses, respectively. A marked increase in extracellular protein concentrations was observed in the transformed belladonna root cultures as compared with the controls transformed with an empty vector. However, the secretion of the proteins of the transformants was markedly reduced in the presence of a physiological concentration of monensin. These results suggest that over-expression of arf-001 in belladonna results in the enhancement of secretory activity in the transformed cells.
    Biological & Pharmaceutical Bulletin 08/2008; 31(7):1465-8. · 1.85 Impact Factor
  • Yuki Asakura, Fumiya Kurosaki
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    ABSTRACT: A homology-based cloning strategy yielded a cDNA clone designated Dcga, presumably encoding alpha subunit of GTP-binding protein, from carrot (Daucus carota) seedlings. Molecular phylogenetic tree analysis of G protein alpha subunits from various biological sources suggested that, unlike in animal cells, classification of Galpha proteins into specific subfamilies could not be applicable to the proteins from higher plants. The restriction digests prepared from genomic DNA of carrot showed one or two hybridized signals in Southern blot analyses, and the expression level of Dcga was appreciably decreased upon the exposure of carrot to high temperature or the prolonged treatment with salt. These results suggest that Dcga occurs as single or double copy genes in carrot genome, and its transcript might play specific roles in heat- and salt-induced responses of the plant.
    Biological & Pharmaceutical Bulletin 10/2007; 30(9):1800-4. · 1.85 Impact Factor
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    ABSTRACT: A homology-based cloning strategy yielded a cDNA clone, designated Sd-cam, encoding calmodulin protein from Scoparia dulcis. The restriction digests of genomic DNA of S. dulcis showed a single hybridized signal when probed with the fragment of this gene in Southern blot analyses, suggesting that Sd-cam occurs as a sole gene encoding calmodulin in the plant. The reverse-transcription polymerase chain reaction analysis revealed that Sd-cam was appreciably expressed in leaf, root and stem tissues. It appeared that transcription of this gene increased transiently when the leaf cultures of S. dulcis were treated with methyl jasmonate and calcium ionophore A23187. These results suggest that transcriptional activation of Sd-cam is one of the early cellular events of the methyl jasmonate-induced responses of S. dulcis.
    Biological & Pharmaceutical Bulletin 07/2007; 30(6):1161-3. · 1.85 Impact Factor
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    Yuki Asakura, Fumiya Kurosaki
    Biological & Pharmaceutical Bulletin - BIOL PHARM BULL. 01/2007; 30(9):1800-1804.
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    ABSTRACT: A homology based strategy yielded two cDNA clones designated arf-001 (1097 bp) and arf-002 (897 bp) from carrot seedlings. These genes contained open reading frames which encode the proteins of 192 and 181 amino acid residues, respectively, with the significant homology to ADP-ribosylation factors of plants, animals and microbial sources. Genomic Southern blot hybridization analyses revealed that both genes showed one main signal, respectively; however, several related genes might be present in carrot genome. The products of these genes obtained by over-expression in Escherichia coli showed the specific binding activity toward GTP. The expression of arf-001 was mainly observed in leaf and stem tissues while arf-002 was appreciably expressed in roots as analyzed by RT-PCR. The transcriptional level of arf-001 showed the transient increase by the exposure of carrot seedlings to ethylene. In contrast, arf-002 appeared to be a house-keeping gene, and its expression level was maintained at constant level upon the treatment with various stimuli and under stress conditions. These results suggest that the arf-001 and 002 play the distinct physiological roles in carrot cells.
    Plant Science. 01/2007;