Targeted deletion and inversion of tandemly arrayed genes in Arabidopsis thaliana using zinc finger nucleases

G3 (Bethesda). 2013 Oct 3;3(10):1707-15. doi: 10.1534/g3.113.006270.


Tandemly arrayed genes (TAGs) or gene clusters are prevalent in higher eukaryotic genomes. For example, approximately 17% of genes are organized in tandem in the model plant Arabidopsis thaliana. The genetic redundancy created by TAGs presents a challenge for reverse genetics. As molecular scissors, engineered zinc finger nucleases (ZFNs) make DNA double-strand breaks in a sequence-specific manner. ZFNs thus provide a means to delete TAGs by creating two double-strand breaks in the gene cluster. Using engineered ZFNs, we successfully targeted seven genes from three TAGs on two Arabidopsis chromosomes, including the well-known RPP4 gene cluster, which contains eight resistance (R) genes. The resulting gene cluster deletions ranged from a few kb to 55 kb with frequencies approximating 1% in somatic cells. We also obtained large chromosomal deletions of ~9 Mb at approximately one tenth the frequency, and gene cluster inversions and duplications also were achieved. This study demonstrates the ability to use sequence-specific nucleases in plants to make targeted chromosome rearrangements and create novel chimeric genes for reverse genetics and biotechnology.

Keywords: Arabidopsis; deletion; inversion; tandemly arrayed genes (TAGs); zinc finger nuclease (ZFN).

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Arabidopsis / genetics*
  • Base Sequence
  • Chromosome Deletion*
  • Endodeoxyribonucleases / chemistry
  • Endodeoxyribonucleases / genetics*
  • Gene Targeting / methods*
  • Genes, Plant*
  • Molecular Sequence Data
  • Multigene Family
  • Mutagenesis, Site-Directed / methods
  • Plants, Genetically Modified / genetics
  • Tandem Repeat Sequences*
  • Zinc Fingers


  • Endodeoxyribonucleases