High-efficiency stable transformation of the model fern species Ceratopteris richardii via microparticle bombardment

Andrew R.G. Plackett, Liandong Huang, Heather L. Sanders, Jane A. Langdale

Research output: Contribution to journalArticlepeer-review

40 Citations (Scopus)

Abstract

Ferns represent the most closely related extant lineage to seed plants. The aquatic fern Ceratopteris richardii has been subject to research for a considerable period of time, but analyses of the genetic programs underpinning developmental processes have been hampered by a large genome size, a lack of available mutants, and an inability to create stable transgenic lines. In this paper, we report a protocol for efficient stable genetic transformation of C. richardii and a closely related species Ceratopteris thalictroides using microparticle bombardment. Indeterminate callus was generated and maintained from the sporophytes of both species using cytokinin treatment. In proof-of-principle experiments, a 35S:β-glucuronidase (GUS) expression cassette was introduced into callus cells via tungsten microparticles, and stable transformants were selected via a linked hygromycin B resistance marker. The presence of the transgene in regenerated plants and in subsequent generations was validated using DNA-blot analysis, reverse transcription-polymerase chain reaction, and GUS staining. GUS staining patterns in most vegetative tissues corresponded with constitutive gene expression. The protocol described in this paper yields transformation efficiencies far greater than those previously published and represents a significant step toward the establishment of a tractable fern genetic model.

Original languageEnglish
Pages (from-to)3-14
Number of pages12
JournalPLANT PHYSIOLOGY
Volume165
Early online date12 Mar 2014
DOIs
Publication statusPublished - May 2014

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

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