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Potential risks of gene amplification by PCR as determined by 16S rDNA analysis of a mixed-culture of strict barophilic bacteria

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Abstract

The 16S rDNA genes of an apparently pure culture of a psychrophilic and strict barophilic bacterium (WHB 46) were studied by PCR-mediated amplification and cloning into phage M13 mp18. Sequence analysis of five individual clones revealed the presence of two different 16S rDNA types. The homology value of 90% indicates that culture WHB 46 is actually composed of two closely related species (WHB 46-1 and 46-2). Both strains are members of the γ-subdivision of proteobacteria. Analysis of a sixth clone (WHB 46-1/2) leads to the conclusion that it represents a 16S rDNA hybrid molecule assembled during the PCR reaction. This hypothesis was confirmed by secondary structure analysis of the chimeric rDNA. The appearance of such hybrid molecules point to a potential risk in studies on the diversity of bacterial populations by analysis of rDNA pattern via PCR-mediated amplification because they suggest the existence of organisms that do not actually exist in the sample investigated.

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Authors and Affiliations

  1. Department of Microbiology, University of Queensland, 4072, St. Lucia, Queensland, Australia

    W. Liesack & E. Stackebrandt

  2. Alfred-Wegener-Institut für Polar- und Meeresforschung, W-2850, Bremerhaven, Federal Republic of Germany

    H. Weyland

Authors
  1. W. Liesack
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  2. H. Weyland
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  3. E. Stackebrandt
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Liesack, W., Weyland, H. & Stackebrandt, E. Potential risks of gene amplification by PCR as determined by 16S rDNA analysis of a mixed-culture of strict barophilic bacteria. Microb Ecol 21, 191–198 (1991). https://doi.org/10.1007/BF02539153

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  • DOI: https://doi.org/10.1007/BF02539153

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