Mar 1, 1994

Alpha-domain of human metallothionein IA can bind to metals in transgenic tobacco plants

Molecular & General Genetics : MGG
A PanB Ru

Abstract

With a view to exploring its use as a metal-binding factor in transgenic plants we prepared the alpha-domain of metallothionein by reconstitution of rabbit apometallothionein and proteolysis of MT-1 and MT-2 with subtilisin. The isolated alpha-domains were characterised by UV and CD spectroscopy Double-Stranded. DNA encoding the alpha-domain (106 bp) of the human MT-IA was constructed from chemically synthesized oligomers by repair synthesis and enzymatic ligation, cloned into pUC19 and sequenced. A expression construct containing the cloned alpha-domain was introduced into tobacco cells on a disarmed Agrobacterium tumefaciens Ti-plasmid. Transformed tobacco cells were selected and regenerated on medium containing cadmium and kanamycin. The growth of roots and shoots of transformants was unaffected by up to 100 microM cadmium, whereas control plants showed severe inhibition of root and shoot growth, and chlorosis of leaves on medium containing only 10 microM cadmium. Southern hybridization confirmed the presence of the transgene in the transformed plant tissues. The concentration of human alpha-domain peptides in transgenic tobacco leaves was determined by the Cd/hemoglobin saturation assay and polarography using the rabbit alp...Continue Reading

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Citations

Mentioned in this Paper

Shuttle Vectors
Agrobacterium tumefaciens
Plants, Toxic
Metals
MT1A
Plants, Genetically Modified
Peptide Fragments
Nicotiana tabacum
Metallothionein IIA
Protein Engineering

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