Knocking Out the Wall: Protocols for Gene Targeting in Physcomitrella patens

  • Alison W. Roberts
  • Christos S. Dimos
  • Michael J. BudziszekJr.
  • Chessa A. Goss
  • Virginia Lai
Part of the Methods in Molecular Biology book series (MIMB, volume 715)


The moss Physcomitrella patens has become established as a model for investigating plant gene function due to the feasibility of gene targeting. The chemical composition of the P. patens cell wall is similar to that of vascular plants and phylogenetic analyses of glycosyltransferase sequences from the P. patens genome have identified genes that putatively encode cell wall biosynthetic enzymes, providing a basis for investigating the evolution of cell wall polysaccharides and the enzymes that synthesize them. The protocols described in this chapter provide methods for targeted gene knockout in P. patens, from constructing vectors and maintaining cultures to transforming protoplasts and analyzing the genotypes and phenotypes of the resulting transformed lines.

Key words

Gene targeting Immunocytochemistry Physcomitrella patens Gateway® cloning 



We thank the members of the moss community for their collegiality and helpful suggestions. Magdalena Bezanilla and the members of her group provided invaluable advice and assistance. We thank Didier Schaefer for the gift of pBHSNR. This project was supported by the National Research Initiative Competitive Grant no. 2007-35318-18389 from the USDA National Institute of Food and Agriculture.


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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Alison W. Roberts
    • 1
  • Christos S. Dimos
    • 1
  • Michael J. BudziszekJr.
    • 1
  • Chessa A. Goss
    • 1
  • Virginia Lai
    • 1
  1. 1.Department of Biological SciencesCenter for Biotechnology and Life Sciences, University of Rhode IslandKingstonUSA

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