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Yeast Protocols pp 45–73Cite as

Targeted Gene Deletion in Saccharomyces cerevisiae and Schizosaccharomyces pombe

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1163))

Abstract

Gene deletion is an important element in the functional characterization of gene and protein function. Efficient tools for gene deletion have been developed in the model yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, all of which rely on the replacement of the endogenous gene of interest with a selectable marker gene by homologous recombination. In order to minimize incidental recombination events between DNA sequences within the marker gene and a chromosomal sequence, gene deletion cassettes consisting entirely of heterologous DNA sequences are preferred. The gene deletion cassettes, which are composed of the marker gene flanked by short DNA segments homologous to the chromosomal sequences lying to the left and right of the gene to be deleted, are generated by PCR and mediate highly efficient one-step gene deletion events. Incorporation of loxP sites flanking the marker gene allows Cre recombinase-mediated rescue, so that the marker can be reused for the next gene deletion. This is particularly useful for the characterization of gene families in S. cerevisiae. The one-step gene deletion method is not limited to the elimination of individual genes, but can also be used for the removal of chromosomal segments exceeding 100 kbp in length. Here we describe a comprehensive set of gene deletion cassettes and outline their use in S. cerevisiae and S. pombe.

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Author information

Authors and Affiliations

  1. Lehrstuhl für Funktionelle Genomforschung der Mikroorganismen, Heinrich-Heine-Universität, Gebäude 25.02.U1.23, Universitätsstr.1, Düsseldorf, 40225, Germany

    Johannes H. Hegemann, Sven Boris Heick, Jennifer Pöhlmann, Marcus M. Langen & Ursula Fleig

Authors
  1. Johannes H. Hegemann
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  2. Sven Boris Heick
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  3. Jennifer Pöhlmann
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  4. Marcus M. Langen
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  5. Ursula Fleig
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Corresponding authors

Correspondence to Johannes H. Hegemann or Ursula Fleig .

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Editors and Affiliations

  1. Capital Normal University College of Life Sciences, Beijing, China

    Wei Xiao

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Hegemann, J.H., Heick, S.B., Pöhlmann, J., Langen, M.M., Fleig, U. (2014). Targeted Gene Deletion in Saccharomyces cerevisiae and Schizosaccharomyces pombe . In: Xiao, W. (eds) Yeast Protocols. Methods in Molecular Biology, vol 1163. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0799-1_5

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  • DOI: https://doi.org/10.1007/978-1-4939-0799-1_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0798-4

  • Online ISBN: 978-1-4939-0799-1

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