Plant Molecular Biology

, Volume 62, Issue 6, pp 937–950

High-frequency Ds remobilization over multiple generations in barley facilitates gene tagging in large genome cereals

  • Jaswinder Singh
  • Shibo Zhang
  • Calvin Chen
  • Laurel Cooper
  • Phil Bregitzer
  • Anne Sturbaum
  • Patrick M. Hayes
  • Peggy G. Lemaux
Article

DOI: 10.1007/s11103-006-9067-1

Cite this article as:
Singh, J., Zhang, S., Chen, C. et al. Plant Mol Biol (2006) 62: 937. doi:10.1007/s11103-006-9067-1

Abstract

Transposable elements have certain advantages over other approaches for identifying and determining gene function in large genome cereals. Different strategies have been used to exploit the maize Activator/dissociation (Ac/Ds) transposon system for functional genomics in heterologous species. Either large numbers of independent Ds insertion lines or transposants (TNPs) are generated and screened phenotypically, or smaller numbers of TNPs are produced, Ds locations mapped and remobilized for localized gene targeting. It is imperative to characterize key features of the system in order to utilize the latter strategy, which is more feasible in large genome cereals like barley and wheat. In barley, we generated greater than 100 single-copy Ds TNPs and determined remobilization frequencies of primary, secondary, and tertiary TNPs with intact terminal inverted repeats (TIRs); frequencies ranged from 11.8 to 17.1%. In 16% of TNPs that had damaged TIRs no transposition was detected among progeny of crosses using those TNPs as parental lines. In half of the greater than 100 TNP lines, the nature of flanking sequences and status of the 11 bp TIRs and 8-bp direct repeats were determined. BLAST searches using a gene prediction program revealed that 86% of TNP flanking sequences matched either known or putative genes, indicating preferential Ds insertion into genic regions, critical in large genome species. Observed remobilization frequencies of primary, secondary, tertiary, and quaternary TNPs, coupled with the tendency for localized Ds transposition, validates a saturation mutagenesis approach using Ds to tag and characterize genes linked to Ds in large genome cereals like barley and wheat.

Keywords

Ac/DsBarleyRemobilizationTransposon tagging

Abbreviations

Ac/Ds

Activator/dissociation

EST

Expressed sequence tag

TIR

Terminal inverted repeat

TNP

Transposon insertion line

Copyright information

© Springer Science+Business Media B.V. 2006

Authors and Affiliations

  • Jaswinder Singh
    • 1
  • Shibo Zhang
    • 1
  • Calvin Chen
    • 1
  • Laurel Cooper
    • 2
  • Phil Bregitzer
    • 3
  • Anne Sturbaum
    • 3
  • Patrick M. Hayes
    • 2
  • Peggy G. Lemaux
    • 1
  1. 1.Department of Plant and Microbial BiologyUniversity of CaliforniaBerkeleyUSA
  2. 2.Department of Crop and Soil SciencesOregon State UniversityCorvallisUSA
  3. 3.USDA-ARSAberdeenUSA