Skip to main content
SpringerLink
Log in

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

  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

Ac/Ds:

Activator/dissociation

EST:

Expressed sequence tag

TIR:

Terminal inverted repeat

TNP:

Transposon insertion line

References

  • Azpiroz-Leehan R, Feldmann KA (1997) T-DNA insertion mutagenesis in Arabidopsis: going back and forth. Trends Genet 13:152–156

    Article  PubMed  CAS  Google Scholar 

  • Baker B, Schell J, Lorz H, Fedoroff N (1986) Transposition of the maize controlling element activator in tobacco. Proc Natl Acad Sci USA 13:4844–4848

    Article  Google Scholar 

  • Balcells L, Coupland G (1994) The presence of enhancers adjacent to the Ac promoter increases the abundance of transposase mRNA and alters the timing of Ds excision in Arabidopsis. Plant Mol Biol 24:789–798

    Article  PubMed  CAS  Google Scholar 

  • Bancroft I, Dean C (1993) Transposition pattern of the maize element Ds in Arabidopsis thaliana. Genetics 134:1221–1229

    PubMed  CAS  Google Scholar 

  • Bancroft I, Jones JD, Dean C (1993) Heterologous tagging of the DRL1 locus in Arabidopsis. Plant Cell 5:631–638

    Article  PubMed  CAS  Google Scholar 

  • Bennetzen JL, Ma J (2003) The genetic colinearity of rice and other cereals on the basis of genomic sequence analysis. Curr Opin Plant Biol 6:128–133

    Article  PubMed  CAS  Google Scholar 

  • Caldwell KS, Langridge P, Powell W (2004) Comparative sequence analysis of the region harboring the hardness locus in barley and its colinear region in rice. Plant Physiol 136:3177–3190

    Article  PubMed  CAS  Google Scholar 

  • Chatterjee S, Starlinger P (1995) The role of subterminal sites of transposable element Ds of Zea mays in excision. Mol Gen Genet 249:281–288

    Article  PubMed  CAS  Google Scholar 

  • Chernyshev AI, Golovkin MV, Milshina NV, Gazumyan AK, Ananyev EV (1988) Molecular-genetic organization of mobile elements of the Ac-Ds family in cereal genomes—identification of barley DNA-sequences homologous to the maize Ac element. Genetika 24:1918–1927

    CAS  Google Scholar 

  • Chin HG, Choe MS, Lee SH, Park SH, Park SH, Koo JC, Kim NY, Lee JJ, Oh BG, Yi GH, Kim SC, Choi HC, Cho MJ, Han C (1999) Molecular analysis of rice plants harboring an Ac/Ds transposable element-mediated gene trapping system. Plant J 19:615–623

    Article  PubMed  CAS  Google Scholar 

  • Colasanti J, Yuan Z, Sundaresan V (1998) The indeterminate gene encodes a zinc finger protein and regulates a leaf-generated signal required for the transition to flowering in maize. Cell 93:593–603

    Article  PubMed  CAS  Google Scholar 

  • Cooley MB, Goldsbrough AP, Still DW, Yoder JI (1996) Site-selected insertional mutagenesis of tomato with maize Ac and Ds elements. Mol Gen Genet 252:184–194

    Article  PubMed  CAS  Google Scholar 

  • Cooper LD, Marquez-Cedillo L, Singh J, Sturbaum AK, Zhang S, Edwards V, Johnson K, Kleinhofs A, Rangel S, Carollo V, Bregitzer P, Lemaux PG, Hayes PM (2004) Mapping Ds insertions in barley using a sequence-based approach. Mol Gen Genomics 272:181–193

    Article  CAS  Google Scholar 

  • Coupland G, Baker B, Schell J, Starlinger P (1988) Characterization of the maize transposable element-Ac by internal deletions. EMBO J 7:3653–3659

    PubMed  CAS  Google Scholar 

  • Cowperthwaite M, Park W, Xu Z, Yan X, Maurais SC, Dooner HK (2002) Use of the transposon Ac as a gene-searching engine in the maize genome. Plant Cell 14:713–726

    Article  PubMed  CAS  Google Scholar 

  • DeLong A, Calderon-Urrea A, Dellaporta SL (1993) Sex determination gene TASSELSEED2 of maize encodes a short-chain alcohol dehydrogenase required for stage-specific floral organ abortion. Cell 74:757–758

    Article  PubMed  CAS  Google Scholar 

  • Fedoroff NV (1989) About maize transposable elements and development. Cell 56:181–191

    Article  PubMed  CAS  Google Scholar 

  • Goodrich J, Puangsomlee P, Martin M, Long D, Meyerowitz EM, Coupland G (1997) A polycomb-group gene regulates homeotic gene expression in Arabidopsis. Nature 386:44–51

    Article  PubMed  CAS  Google Scholar 

  • Greco R, Ouwerkerk PBF, Sallaud C, Kohli A, Colombo L, Puigdomenech P, Guiderdoni E, Christou P, Hoge JHC, Pereira A (2001) Transposon insertional mutagenesis in rice. Plant Physiol 125:1175–1177

    Article  PubMed  CAS  Google Scholar 

  • Greco R, Ouwerkerk PB, De Kam RJ, Sallaud C, Favalli C, Colombo L, Guiderdoni E, Meijer AH, Hoge Dagger JH, Pereira A (2003) Transpositional behaviour of an Ac/Ds system for reverse genetics in rice. Theor Appl Genet 108:10–24

    Article  PubMed  CAS  Google Scholar 

  • Grossniklaus U, Vielle-Calzada JP, Hoeppner MA, Gagliano WB (1998) Maternal control of embryogenesis by MEDEA, a polycomb group gene in Arabidopsis. Science 280:446–450

    Article  PubMed  CAS  Google Scholar 

  • Gu Q, Ferrandiz C, Yanofsky MF, Martienssen R (1998) The FRUITFULL MADS-box gene mediates cell differentiation during Arabidopsis fruit development. Development 125:1509–1517

    PubMed  CAS  Google Scholar 

  • Hake S, Vollbrecht E, Freeling M (1989) Cloning knotted, the dominant morphological mutant in maize using Ds2 as a transposon tag. EMBO J 8:15–22

    PubMed  CAS  Google Scholar 

  • Hannon GJ (2002) RNA interference. Nature 418:244–251

    Article  PubMed  CAS  Google Scholar 

  • Hehl R, Baker B (1989) Induced transposition of Ds by a stable Ac in crosses of transgenic tobacco plants. Mol Gen Genet 217:53–59

    Article  PubMed  CAS  Google Scholar 

  • Ito T, Motohashi R, Kuromori T, Mizukado S, Sakurai T, Kanahara H, Seki M, Shinozaki K (2002) A new resource of locally transposed Dissociation elements for screening gene-knockout lines in silico on the Arabidopsis genome. Plant Physiol 129:1695–1699

    Article  PubMed  CAS  Google Scholar 

  • Izawa T, Ohnishi T, Nakano T, Ishida N, Enoki H, Hashimoto H, Itoh K, Terada R, Wu C, Miyazaki C, Endo T, Iida S, Shimamoto K (1997) Transposon tagging in rice. Plant Mol Biol 35:219–229

    Article  PubMed  CAS  Google Scholar 

  • Jeon JS, Lee S, Jung KH, Jun SH, Jeong DH, Lee J, Kim C, Jang S, Yang K, Nam J, An K, Han MJ, Sung RJ, Choi HS, Yu JH, Choi JH, Cho SY, Cha SS, Kim SI, An G (2000) T-DNA insertional mutagenesis for functional genomics in rice. Plant J 22:561–570

    Article  PubMed  CAS  Google Scholar 

  • Jones DA, Thomas CM, Hammond-Kosack KE, Balint-Kurti PJ, Jones JD (1994) Isolation of the tomato Cf-9 gene for resistance to Cladosporium fulvum by transposon tagging. Science 266:789–793

    Article  PubMed  CAS  Google Scholar 

  • Keller B, Feuillet C (2000) Colinearity and gene density in grass genomes. Trends Plant Sci 5:246–251

    Article  PubMed  CAS  Google Scholar 

  • Kohli A, Prynne MQ, Miro B, Pereira A, Twyman RM, Capell T, Christou P (2004) Dedifferentiation-mediated changes in transposition behavior make the Activator transposon an ideal tool for functional genomics in rice. Mol Breed 13:177–191

    Article  CAS  Google Scholar 

  • Kolesnik T, Szeverenyi I, Bachmann D, Kumar CS, Jiang S, Ramamoorthy R, Cai M, Ma ZG, Sundaresan V, Ramachandran S (2004) Establishing an efficient Ac/Ds tagging system in rice: large-scale analysis of Ds flanking sequences. Plant J 37:301–314

    PubMed  CAS  Google Scholar 

  • Koprek T, McElroy D, Louwerse J, Williams-Carrier R, Lemaux PG (2000) An efficient method for dispersing Ds elements in the barley genome as a tool for determining gene function. Plant J 24:253–263

    Article  PubMed  CAS  Google Scholar 

  • Koprek T, Rangel S, McElroy D, Louwerse JD, Williams-Carrier RE, Lemaux PG (2001) Transposon-mediated single-copy gene delivery leads to increased transgene expression stability in barley. Plant Physiol 125:1354–1362

    Article  PubMed  CAS  Google Scholar 

  • Krysan PJ, Young JC, Sussman MR (1999) T-DNA as an insertional mutagen in Arabidopsis. Plant Cell 11:2283–2290

    Article  PubMed  CAS  Google Scholar 

  • Kunze R, Starlinger P (1989) The putative transposase of transposable element Ac from Zea mays L. interacts with subterminal sequences of Ac. EMBO J 8:3177–3185

    PubMed  CAS  Google Scholar 

  • Kuromori T, Hirayama T, Kiyosue Y, Takabe H, Mizukado S, Sakurai T, Akiyama K, Kamiya A, Ito T, Shinozaki K (2004) A collection of 11 800 single-copy Ds transposon insertion lines in Arabidopsis. Plant J 37:897–905

    Article  PubMed  CAS  Google Scholar 

  • May BP, Martienssen RA (2003) Mutagenesis in the study of plant development. Crit Rev Plant Sci 22:1–35

    CAS  Google Scholar 

  • McClintock B (1949) Mutable loci in maize. Carnegie Inst Wash Year Book 48:142–154

    Google Scholar 

  • McElroy D, Louwerse JD, McElroy SM, Lemaux PG (1997) Development of a simple transient assay for Ac/Ds activity in cells of intact barley tissue. Plant J 11:157–165

    Article  PubMed  CAS  Google Scholar 

  • Meng L, Ziv M, Lemaux PG (2006) Nature of stress and transgene locus influences transgene expression stability in barley. Plant Mol Biol (in press)

  • Nakagawa Y, Machida C, Machida Y, Toriyama K (2000) Frequency and pattern of transposition of the maize transposable element Ds in transgenic rice plants. Plant Cell Physiol 41:733–742

    PubMed  CAS  Google Scholar 

  • Parinov S, Sevugan M, Ye D, Yang WC, Kumaran M, Sundaresan V (1999) Analysis of flanking sequences from dissociation insertion lines: a database for reverse genetics in Arabidopsis. Plant Cell 11:2263–2270

    Article  PubMed  CAS  Google Scholar 

  • Raina S, Mahalingam R, Chen F, Federoff N (2002) A collection of sequenced and mapped Ds transposon insertion sites in Arabidopsis thaliana. Plant Mol Biol 50:93–110

    Article  PubMed  CAS  Google Scholar 

  • Schmidt E, Guzzo F, Toonen MAJ, de Vries SC (1997) A leucine-rich repeat containing receptor-like kinase marks somatic plant cells competent to form embryos. Development 124:2049–2062

    PubMed  CAS  Google Scholar 

  • Scholz S, Lorz H, Lutticke S (2001) Transposition of the maize transposable element Ac in barley (Hordeum vulgare L.). Mol Gen Genet 264:653–661

    Article  PubMed  CAS  Google Scholar 

  • Scott L, LaFoe D, Weil CF (1996) Adjacent sequences influence DNA repair accompanying transposon excision in maize. Genetics 142:237–246

    PubMed  CAS  Google Scholar 

  • Shimamoto K, Miyazaki C, Hashimoto H, Izawa T, Itoh K, Terada R, Inagaki Y, Iida S (1993) Trans-activation and stable integration of the maize transposable element Ds cotransfected with the Ac transposase gene in transgenic rice plants. Mol Gen Genet 239:354–360

    Article  PubMed  CAS  Google Scholar 

  • Shirasu K, Schulman AH, Lahaye T, Schulze-Lefert P (2000) A contiguous 66-kb barley DNA sequence provides evidence for reversible genome expansion. Genome Res 10:908–915

    Article  PubMed  CAS  Google Scholar 

  • Smith D, Yanai Y, Liu YG, Ishiguro S, Okada K, Shibata D, Whittier RF, Fedoroff NV (1996) Characterization and mapping of Ds-GUS-T-DNA lines for targeted insertional mutagenesis. Plant J 10:721–732

    Article  PubMed  CAS  Google Scholar 

  • Szeverenyi I, Ramamoorthy R, Wei Teo Z, Luan HF, Gang Ma Z, Ramachandran S (2006) Large-scale systematic study on stability of the Ds element and timing of transposition in rice. Plant Cell Physiol 47:84–95

    Article  PubMed  CAS  Google Scholar 

  • Upadhyaya NM, Zhou XR, Ramm K, Zhu QH, Wu L, Eamens A, Sivakumar R, Kato T, Yun DW, Kumar S, Narayanan KK, Peacock WJ, Dennis ES (2002) An iAc/Ds gene and enhancer trapping system for insertional mutagenesis in rice. Funct Plant Biol 29:547–559

    Article  CAS  Google Scholar 

  • Van Sluys MA, Tempe J, Fedoroff N (1987) Studies on the introduction and motility of the maize activator element in Arabidopsis thaliana and Daucus carota. EMBO J 6:3881–3889

    PubMed  Google Scholar 

  • Varagona M, Wessler SR (1990) Implications for the cis-requirements for Ds transposition based on the sequence of the wxB4 Ds element. Mol Gen Genet 220:414–418

    Article  PubMed  CAS  Google Scholar 

  • Walbot V (2000) Saturation mutagenesis using maize transposons. Curr Opin Plant Biol 3:103–107

    Article  PubMed  CAS  Google Scholar 

  • Waterhouse PM, Helliwell CA (2003) Exploring plant genomes by RNA-induced gene silencing. Nat Rev Genet 4:29–38

    Article  PubMed  CAS  Google Scholar 

  • Wicker T, Zimmermann W, Perovic D, Paterson AH, Ganal M, Graner A, Stein N (2005) A detailed look at 7 million years of genome evolution in a 439 kb contiguous sequence at the barley Hv-eIF4E locus: recombination, rearrangements and repeats. Plant J 41:187–194

    Article  Google Scholar 

  • Xiao YL, Peterson T (2002) Ac transposition is impaired by a small terminal deletion. Mol Genet Genomics 266:720–731

    Article  PubMed  CAS  Google Scholar 

  • Yang WC, Ye D, Xu J, Sundaresan V (1999) The sporocyteless gene of Arabidopsis is required for the initiation of sporogenesis and encodes a novel nuclear protein. Genes Dev 13:2108–2117

    PubMed  CAS  Google Scholar 

  • Yoder JI, Palys J, Alpert K, Lassner M (1988) Ac transposition in transgenic tomato plants. Mol Gen Genet 213:291–296

    Article  CAS  Google Scholar 

  • Zhu QH, Ramm K, Shivakkumar R, Dennis ES, Upadhyaya NM (2004) The ANTHER INDEHISCENCE1 gene encoding a single MYB domain protein is involved in anther development in rice. Plant Physiol 135:1514–1525

    Article  PubMed  CAS  Google Scholar 

  • Zhu QH, Hoque MS, Dennis ES, Upadhyaya NM (2003) Ds tagging of BRANCHED FLORETLESS 1 (BFL1) that mediates the transition from spikelet to floret meristem in rice (Oryza sativa L). BMC Plant Biol 3:6

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

The authors thank David Bae, Greg East, Michael Freudiger, Chris Gates, Chris Lowe, and Mark Ou for assistance in DNA isolation and care of the plants in the greenhouse and Barbara Alonso for excellent graphics assistance on the figures. The authors also thank Dr. Damon Lisch for critical reading of the manuscript. This work was supported by NSF Award #0110512 to PGL and an NSF REU award for CC; PGL is also supported by USDA Cooperative Extension through the University of California.

Author information

Authors and Affiliations

  1. Department of Plant and Microbial Biology, University of California, Berkeley, CA, 94720, USA

    Jaswinder Singh, Shibo Zhang, Calvin Chen & Peggy G. Lemaux

  2. Department of Crop and Soil Sciences, Oregon State University, Corvallis, OR, 97331, USA

    Laurel Cooper & Patrick M. Hayes

  3. USDA-ARS, Aberdeen, ID, 83210, USA

    Phil Bregitzer & Anne Sturbaum

Authors
  1. Jaswinder Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Shibo Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Calvin Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Laurel Cooper
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Phil Bregitzer
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Anne Sturbaum
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Patrick M. Hayes
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Peggy G. Lemaux
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jaswinder Singh.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Singh, J., Zhang, S., Chen, C. et al. High-frequency Ds remobilization over multiple generations in barley facilitates gene tagging in large genome cereals. Plant Mol Biol 62, 937–950 (2006). https://doi.org/10.1007/s11103-006-9067-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11103-006-9067-1

Keywords

Search

Navigation