Plastid transformation of sporelings and suspension-cultured cells from the liverwort Marchantia polymorpha L

Methods Mol Biol. 2014:1132:439-47. doi: 10.1007/978-1-62703-995-6_30.

Abstract

We describe simple and efficient plastid transformation methods for suspension-cultured cells and sporelings of the liverwort, Marchantia polymorpha L. Use of rapidly proliferating cells such as suspension-cultured cells and sporelings, which are immature thalli developing from spores, as targets made plastid transformation by particle bombardment efficient. Selection on a sucrose-free medium and linearization of the transformation vector significantly improved the recovery rate of plastid transformants. With the methods described here, a few plastid transformants are obtained from a single bombardment of sporelings, while more efficient plastid transformation is expected in suspension-cultured cells, ~60 transformants from a single bombardment. Homoplasmic transformants of thalli are obtained immediately after primary selection, whereas homoplasmic transformants from suspension-cultured cells are obtained after 12-16 weeks of repeated subculture.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology
  • Biolistics / methods*
  • Cell Culture Techniques
  • Cells, Cultured
  • Chloroplasts / genetics*
  • DNA, Plant / genetics
  • Drug Resistance / genetics
  • Genetic Vectors / genetics
  • Homologous Recombination
  • Marchantia / genetics*
  • Nucleotidyltransferases / genetics
  • Plants, Genetically Modified / genetics
  • Plasmids / genetics
  • Spectinomycin / pharmacology
  • Spores / growth & development
  • Transformation, Genetic*

Substances

  • Anti-Bacterial Agents
  • DNA, Plant
  • Spectinomycin
  • Nucleotidyltransferases
  • gentamicin 2''-nucleotidyltransferase