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A nitrate reductase gene of the cyanobacterium Synechococcus PCC6301 inferred by heterologous hybridization, cloning and targeted mutagenesis

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Abstract

DNA probes from the narG gene of Escherichia coli, which encodes the large polypeptide of respiratory nitrate reductase, show cross-hybridization at low stringency to a single region of the genome of the cyanobacterium Synechococcus PCC6301. This segment of cyanobacterial DNA was cloned as the insert of plasmid pDN1 and characterized. RNA complementary to pDN1 was shown to be substantially more abundant in nitrate grown cells of Synechococcus PCC6301 than in ammonium grown cells, thus parallelling the nitrate induction and ammonium repression of nitrate reductase activity in cultures of this cyanobacterium. A mutant of Synechococcus PCC6301 deficient in nitrate reductase activity was obtained after a potentially mutagenic transformation treatment using pDN1 as a donor. This mutant was restored to the wild type phenotype following stable integrative transformation with pDN1 DNA. Taken together these data suggest that pDN1 might encode a polypeptide of nitrate reductase. pDN1 is distinct from three clones of genes involved in nitrate assimilation that were isolated previously from the related cyanobacterium Synechococcus PCC7942 (Kuhlemeier et al., 1984a, J.Bact. 159, 36–41, and 1984b, Gene 31, 109–116).

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Authors and Affiliations

  1. Department of Genetics, University of Leeds, LS2 9JT, Leeds, UK

    D. A. Lightfoot, A. J. Baron, J. M. Cock & J. C. Wootton

  2. Department of Plant and Soil Science, University of Southern Illinois, 62901, Carbondale, IL, USA

    D. A. Lightfoot

  3. Laboratoire de Biologie Moleculaire des Relations Plantes-Microorganismes, INRA CNRS, 31326, Castanet-Tolosan Cedex, France

    A. J. Baron & J. M. Cock

  4. National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, 20894, Bethesda, MD, USA

    J. C. Wootton

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  1. D. A. Lightfoot
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  2. A. J. Baron
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  3. J. M. Cock
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  4. J. C. Wootton
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Lightfoot, D.A., Baron, A.J., Cock, J.M. et al. A nitrate reductase gene of the cyanobacterium Synechococcus PCC6301 inferred by heterologous hybridization, cloning and targeted mutagenesis. Genetica 85, 107–117 (1992). https://doi.org/10.1007/BF00120317

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  • DOI: https://doi.org/10.1007/BF00120317

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