Summary
The homeologous nitrate reductase (NR) structural genes from tobacco were used to complement nitrate reductase deficient mutants from tobacco and Nicotiana plumbaginifolia. A plasmid conferring kanamycin resistance and lambda genomic clones carrying the tobacco wild-type alleles of the genes were co-electroporated in protoplasts of the tobacco mutant. Among 266 plants regenerated from kanamycin resistant colonies, 3 were able to grow permanently on a medium containing nitrate as sole nitrogen source. One of these three plants was further characterized. The ability to grow on nitrate was transmitted as a new single Mendelian dominant marker linked to kanamycin resistance. Molecular analysis of this clone confirmed the integration of a copy of the wild-type allele, and the synthesis at a low level of an active NR. This NR activity is sufficient to regulate both exogenous wild-type and endogenous mutated alleles of the genes at the transcriptional level. A N. plumbaginifolia mutant carrying a mutation impairing NR mRNA production was transformed by Agrobacterium mediated transfer of the wild-type tobacco nia-2 gene cloned into a binary vector. Similarly, kanamycin resistant calli were tested for their ability to grow on nitrate. Among 70 kanamycin resistant transformants, 7 were restored for nitrate assimilation. Molecular analysis revealed the integration of the tobacco gene, and the synthesis at a low level of the NR mRNA and of a nitrate inducible active NR.
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Vaucheret, H., Chabaud, M., Kronenberger, J. et al. Functional complementation of tobacco and Nicotiana plumbaginifolia nitrate reductase deficient mutants by transformation with the wild-type alleles of the tobacco structural genes. Mol Gen Genet 220, 468–474 (1990). https://doi.org/10.1007/BF00391755
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DOI: https://doi.org/10.1007/BF00391755