Teruhisa Shimada

Publications (2)4.44 Total impact

• Article: The Choline Oxidase Gene codA Confers Salt Tolerance to Transgenic Eucalyptus globulus in a Semi-Confined Condition.

  Xiang Yu, Akira Kikuchi, Etsuko Matsunaga, Yoshihiko Morishita, Kazuya Nanto, Nozomu Sakurai, Hideyuki Suzuki, Daisuke Shibata, Teruhisa Shimada, Kazuo N Watanabe
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  ABSTRACT: The performance of tree species is influenced by environmental factors and growth stages. To evaluate the practical performance of transgenic tree species, it is insufficient to grow small, young trees under controlled conditions, such as in a growth chamber. Three transgenic Eucalyptus globulus lines, carrying the choline oxidase gene, were investigated for their salt tolerance and expression of the transgene at the young plantlet stage in a special netted-house. To clarify the characteristics at the young as well during the later stages, salt tolerance and the properties of the transgenic lines at large juvenile and adult stages were evaluated in the special netted-house. All transgenic lines showed high glycinebetaine content, particularly in young leaves. Trees of the transgenic line 107-1 showed low damage because of salinity stress based on the results from the chlorophyll analysis and malondialdehyde content, and they survived the high-salt-shock treatment at the large juvenile and adult stages. Only this line showed salt tolerance at all stages in the special netted-house. In this evaluation in the special netted-house, the tolerant line among young plantlets might perform better at all stages. Since evaluation in these special netted-house mimics field evaluation, line 107-1 is a potential tolerant line.
  Molecular Biotechnology 07/2012; · 2.17 Impact Factor
• Article: Agrobacterium-mediated transformation of Eucalyptus globulus using explants with shoot apex with introduction of bacterial choline oxidase gene to enhance salt tolerance.

  Etsuko Matsunaga, Kazuya Nanto, Masatoshi Oishi, Hiroyasu Ebinuma, Yoshihiko Morishita, Nozomu Sakurai, Hideyuki Suzuki, Daisuke Shibata, Teruhisa Shimada
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  ABSTRACT: Eucalyptus globulus is one of the most economically important plantation hardwoods for paper making. However, its low transformation frequency has prevented genetic engineering of this species with useful genes. We found the hypocotyl section with a shoot apex has the highest regeneration ability among another hypocotyl sections, and have developed an efficient Agrobacterium-mediated transformation method using these materials. We then introduced a salt tolerance gene, namely a bacterial choline oxidase gene (codA) with a GUS reporter gene, into E. globulus. The highest frequency of transgenic shoot regeneration from hypocotyls with shoot apex was 7.4% and the average frequency in four experiments was 4.0%, 12-fold higher than that from hypocotyls without shoot apex. Using about 10,000 explants, over 250 regenerated buds were confirmed as transformants by GUS analysis. Southern blot analysis of 100 elongated shoots confirmed successful generation of stable transformants. Accumulation of glycinebetaine was investigated in 44 selected transgenic lines, which showed 1- to 12-fold higher glycinebetaine levels than non-transgenic controls. Rooting of 16 transgenic lines was successful using a photoautotrophic method under enrichment with 1,000 ppm CO(2). The transgenic whole plantlets were transplanted into potting soil and grown normally in a growth room. They showed salt tolerance to 300 mM NaCl. The points of our system are using explants with shoot apex as materials, inhibiting the elongation of the apex on the selection medium, and regenerating transgenic buds from the side opposite to the apex. This approach may also solve transformation problems in other important plants.
  Plant Cell Reports 01/2012; 31(1):225-35. · 2.27 Impact Factor