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  • 1
    ISSN: 1573-5028
    Keywords: Nicotiana tabacum ; plant transformation ; gene expression ; bacterial lysine decarboxylase ; protein transport ; chloroplasts ; cadaverine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A possible approach for altering alkaloid biosynthesis in plants is the expression of genes encoding key enzymes of a pathway such as lysine decarboxylase (ldc) in transgenic plants. Two strategies were followed here: one focused on expression of the gene in the cytoplasm, the other on subsequent targeting of the protein to the chloroplasts. Theldcgene fromHafnia alvei was therefore (a) placed under the control of the 1′ promoter of the bidirectional Tr promoter fromAgrobacterium tumefaciens Ti- plasmid, and (b) cloned behind therbcS promoter from potato fused to the coding region of therbcS transit peptide. Bothldc constructs, introduced intoNicotiana tabacum with the aid ofA. tumefaciens, were integrated into the plant genome and transcribed as shown by Southern and northern hybridization. However, LDC activity was only detectable in plants expressing mRNA under the control of therbcS promoter directing the LDC fusion protein into chloroplasts with the aid of the transit peptide domain. In plants expressing the processed bacterial enzyme cadaverine levels increased from nearly zero to 0.3–1% of dry mass.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-9368
    Keywords: Nicotiana tabacum ; transformed root cultures ; metabolic engineering ; lysine decarboxylase ; protein targeting ; Rubisco transit peptide ; anabasine ; cadaverine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The gene of a bacterial lysine decarboxylase (ldc) fused to arbcS transit peptide coding sequence (tp), and under the control of the CaMV 35S promoter, was expressed in hairy root cultures ofNicotiana tabacum. The fusion of theldc to the targeting signal sequence improved the performance of the bacterial gene in the plant cells in many respects. Nearly all transgenic hairy root cultures harbouring the35S-tp-ldc gene contained distinctly higher lysine decarboxylase activity (from 1.5 to 30 pkat LDC per mg protein) than those which had been transformed with constructs in which the gene had been directly cloned behind the CaMV 35S promoter. The higher enzyme activity led to the accumulation of up to 0.7% cadaverine on a dry mass basis. In addition, part of the cadaverine pool was used for increased biosynthesis of anabasine, an alkaloid which was hardly detectable in control cultures. The best line contained anabasine levels of 0.5% dry mass, which could further be enhanced by feeding of lysine.
    Type of Medium: Electronic Resource
    Library Location Call Number Volume/Issue/Year Availability
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