Nontransgenic genome modification in plant cells

Ira Marton, Amir Zuker, Elena Shklarman, Vardit Zeevi, Andrey Tovkach, Suzy Roffe, Marianna Ovadis, Tzvi Tzfira, Alexander Vainstein

Research output: Contribution to journalArticlepeer-review

126 Scopus citations

Abstract

Zinc finger nucleases (ZFNs) are a powerful tool for genome editing in eukaryotic cells. ZFNs have been used for targeted mutagenesis in model and crop species. In animal and human cells, transient ZFN expression is often achieved by direct gene transfer into the target cells. Stable transformation, however, is the preferred method for gene expression in plant species, and ZFN-expressing transgenic plants have been used for recovery of mutants that are likely to be classified as transgenic due to the use of direct gene-transfer methods into the target cells. Here we present an alternative, nontransgenic approach for ZFN delivery and production of mutant plants using a novel Tobacco rattle virus (TRV)-based expression system for indirect transient delivery of ZFNs into a variety of tissues and cells of intact plants. TRV systemically infected its hosts and virus ZFN-mediated targeted mutagenesis could be clearly observed in newly developed infected tissues as measured by activation of a mutated reporter transgene in tobacco (Nicotiana tabacum) and petunia (Petunia hybrida) plants. The ability of TRV to move to developing buds and regenerating tissues enabled recovery of mutated tobacco and petunia plants. Sequence analysis and transmission of the mutations to the next generation confirmed the stability of the ZFN-induced genetic changes. Because TRV is an RNAvirus that can infect a wide range of plant species, it provides a viable alternative to the production of ZFN-mediated mutants while avoiding the use of direct plant-transformation methods.

Original languageEnglish
Pages (from-to)1079-1087
Number of pages9
JournalPlant Physiology
Volume154
Issue number3
DOIs
StatePublished - 1 Jan 2010
Externally publishedYes

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

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