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Transformation of Yeast Using Bioactive Beads With Surface-Immobilized Yeast Artificial Chromosomes

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YAC Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 349))

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Abstract

Yeast artificial chromosomes (YACs) are useful cloning vectors with the capacity to carry large DNA inserts. The largest barrier using such large DNA molecules in transformation experiments has been their physical instability in a solution. We developed a new method for transforming yeast with chromosome-sized DNA. The method uses bioactive beads composed of calcium alginate to immobilize yeast chromosomal DNAs. Chromosomal DNA immobilized on bioactive beads is physically stable when compared with naked chromosomal DNAs. The bead-mediated transformation performed well, not only with respect to the transformation frequency, but also in successful transformation using split chromosomal DNA that exceeded 450 kb in size. In this chapter we introduce a new method for transforming yeast using bioactive beads. In conjunction with genomic YAC libraries and the yeast chromosome-splitting method, this technique will pave the way to stable and effective transfer of YACs into yeast cells.

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Author information

Authors and Affiliations

  1. Venture Business Laboratory Center for Advanced Science, Osaka, Japan

    Shigeki Kawakami

  2. Innovation Osaka University, Osaka, Japan

    Shigeki Kawakami

  3. Laboratory of Molecular Genetics, Department of Biotechnology, Graduate School of Engineering Osaka University, Suita, Osaka, Japan

    Satoshi Harashima

  4. Laboratory of Cell Technology, Department of Biotechnology, Graduate School of Engineering Osaka University, Suita, Osaka, Japan

    Akio Kobayashi

  5. Laboratory of Dynamic Cell Biology, Department of Biotechnology, Graduate School of Engineering Osaka University, Suita, Osaka, Japan

    Kiichi Fukui

Authors
  1. Shigeki Kawakami
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  2. Satoshi Harashima
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  3. Akio Kobayashi
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  4. Kiichi Fukui
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Editor information

Editors and Affiliations

  1. School of Medical Sciences, College of Life Sciences and Medicine, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen, Scotland

    Alasdair MacKenzie

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© 2006 Humana Press Inc., Totowa NJ

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Kawakami, S., Harashima, S., Kobayashi, A., Fukui, K. (2006). Transformation of Yeast Using Bioactive Beads With Surface-Immobilized Yeast Artificial Chromosomes. In: MacKenzie, A. (eds) YAC Protocols. Methods in Molecular Biology™, vol 349. Humana Press. https://doi.org/10.1385/1-59745-158-4:61

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  • DOI: https://doi.org/10.1385/1-59745-158-4:61

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-612-2

  • Online ISBN: 978-1-59745-158-1

  • eBook Packages: Springer Protocols

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