Current Microbiology

, Volume 21, Issue 5, pp 301–306 | Cite as

Increased efficiency of transformation ofBacillus stearothermophilus by a plasmid carrying a thermostable kanamycin resistance marker

  • Hans H. Liao
  • Agnes M. Kanikula
Article

Abstract

Recombinant plasmids carrying either the wildtype kanamycin nucleotidyltransferase gene encoded originally by the mesophilic plasmid pUB110 or the gene encoding the thermostable TK101 mutant were constructed and introduced intoBacillus stearothermophilus by a protoplast transformation procedure. When kanamycin-resistant transformants were selected at 47°C, the transformation efficiency of the plasmid bearing the TK101 gene was nine times higher than that of the plasmid encoding the wildtype enzyme. The difference in transformation efficiencies between the two plasmids was increased when transformants were selected at higher temperatures, reflecting the difference in thermostabilities of the respective kanamycin nucleotidyltransferases. We conclude that, even though the pUB110 enzyme is sufficiently active at 47°C to confer kanamycin resistance toB. stearothermophilus, the additional stability of the TK101 mutant is advantageous in transformation ofB. stearothermophilus. The TK101 gene may also have broad utility as a marker for cloning vectors in other thermophiles.

Keywords

Kanamycin Recombinant Plasmid Transformation Efficiency Cloning Vector Kanamycin Resistance 

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Copyright information

© Springer-Verlag New York Inc 1990

Authors and Affiliations

  • Hans H. Liao
    • 1
  • Agnes M. Kanikula
    • 1
  1. 1.University of Wisconsin Biotechnology CenterMadisonUSA

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