| Project/Area Number |
14380271
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
環境保全
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| Research Institution | Mie University |
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Principal Investigator |
KIMURA Tetsuya Mie University, Faculty of Bioresources, Associate professor, 生物資源学部, 助教授 (00281080)
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| Co-Investigator(Kenkyū-buntansha) |
SAKKA Kazuo Mie University, Faculty of Bioresources, Professor, 生物資源学部, 教授 (20154031)
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| Project Period (FY) |
2002 – 2004
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| Project Status |
Completed (Fiscal Year 2004)
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| Budget Amount *help |
¥14,800,000 (Direct Cost: ¥14,800,000)
Fiscal Year 2004: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2003: ¥5,000,000 (Direct Cost: ¥5,000,000)
Fiscal Year 2002: ¥6,200,000 (Direct Cost: ¥6,200,000)
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| Keywords | chlorocatechol / rice / poplar / PHT1 / Arabidopsis / ファイトレメディエーション / 有機塩素系化合物 / Relstonia |
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| Research Abstract |
Utilization of transgenic trees as phytoremediation is considered to be one of the cost-effective ways for successive degradation of chlorinated aromatic compounds such as dioxins which are resistant to degradation for decades. Bacterial genes cbnA encoding chlorocatechol dioxigenase and cbnB encoding chloromuconate cycloisomerase from Ralstonia eutropha NH9, catalyzes one of the important steps for degradation of chlorocatechols and cleaves the aromatic ring of 3-chlorocatechol to produce toxically reduced 2-chloromuconate and muconolactone.
In this study, we constructed a binary vector pCAMBIA-E7131-cbnA-cbnB transcribing cbnA and cbnB gene under the control of CaMV 35S promoter with enhancers, and introduced to rice and hybrid poplar (Populus tremula x tremuloides) by Agrobacterium mediated transformation. Integration of the gene to their chlomosomes was confirmed by PCR and the transgenic lines were isolated for subsequent analyses. Accumulation of cbnA and cbnB translation was visu
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alized by Western bloting and chlorocatechol dioxigenase activity was quantitatively detected by HPLC assay despite its high GC content (65%). When poplar transgenic calli was incubated for 2 hrs in the presence of 1 mM of 3-chlorocatechol, increase of a degradation product 2-chloromuconate peak signal was observed in parallel with reduced peak of 3-chlorocatechol substrate. The results showed that the cbnA gene and cbnB gene were successfully expressed in rice and poplar tree. These kinds of transgenic plants will be used for phytoremediation of environmental pollutants such as chlorinated aromatic compounds.
For phytoremediation of pollutants in contaminated soil, the genes intorduced into plant cells should be expressed in root tissues since most of the contaminated chemicals are in soil. To express the genes in root tissues, strong promoters for root specific expression are necessary. To isolate these kinds of promoters, we selected the PHT1 promoter of Arabidopsis thaliana. GUS reporter analysis of PHT1 promoter in rice and Arabidopsis indicated that this promoter is useful for gene expression in root tissues both in monocot and dicot plants. Less
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