Skip to main content

PCR-Based Identification of T-DNA Insertion Mutants

  • Protocol
Arabidopsis Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 82))

Abstract

With large-scale sequencing projects (1,2), thousands of unique sequences ESTs; expressed sequence tags) have been generated. In addition, hundreds of sequences have been identified based on then homology with a gene of known interest or from subtractive hybridization procedures. For the vast majority of these genes, the sequence indicates little or nothing about its function in the plant. In thus protocol, we describe a reverse genetics procedure which will be useful for identifying an insertion mutant in Arabidopsis for any sequence of interest.

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

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Newman, T, de Bruijn, F.J., Green, P, Keegstra, K., Kende, H., McIntosh, L., Ohlrogge, J., Raikhel, M., Somerville, S., Thomashow, M., Retzel, E., and Somerville, C (1994) Genes galore a summary of methods for accessing results from large-scale parttal sequencing of anonymous Arabidopsis cDNA clones. Plant Physiol 106, 1241–1255.

    Article  PubMed  CAS  Google Scholar 

  2. Cooke, R, Raynal, M., Laudre, M., Grellet, F, Delseny, M, Morris, P-C., Guerrier, D, Gnaudat, J, Qmgley, F, Clabault, G., Li, Y-F., Mache Knvitzky, M, Gy, I., Krets, M., Lecharny, A, Parmentter, Y, Marbach, J., Fleck, J., Clemnet, B, Philipps, G., Herve, C., Bardet, C., Tremousaygue, D., Lescure, B., Lacomme, C., Roby, D, Jourjon, M-F., Chabrter, P., Charpenteau, J-L., Desprez, T., Amselem, J., Chtapello, H., and Hofte, H. (1996) Further progress towards a catalogue of all Arubidopszs genes, analysis of a set of 5000 non-redundant ESTs. Plant J. 9,101–124

    Article  PubMed  CAS  Google Scholar 

  3. Feldmann, K. A. (1991) T-DNA insertion mutagenesis m Arabtdopsis: mutattonal spectrum. Plant J 1, 71–82.

    Article  CAS  Google Scholar 

  4. Koncz, C., Nemeth, K, Reder, G. P., and Schell, J. (1992) T-DNA insertional mutagenesis in Arabidopsis Plant MoL Biol 20, 963–976 5

    Article  PubMed  CAS  Google Scholar 

  5. Fortsthoefel, N. R, Wu, Y, Schulz, B., Bennett, M. J, and Feldmann, K A (1992) T-DNA insertion mutagenesis m Arabidopsis prospects and perspectives Aust J Plant Physlol 19, 353–

    Article  Google Scholar 

  6. Bechtold, N., Ellis, J, and Pelletter, G. (1993) In planta Agrobacterium mediated gene transfer by mfiltratron of adult Arabidopsis thallana plants CR Acad SCI, Puns 316, 1188–1193

    Google Scholar 

  7. Feldmann, K A, Malmberg, R., and Dean., C. (1994) Mutagenesis in Arabidopsis in Arabidopsis (Meyerowitz, E. and Somerville, C., eds.) Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, pp. 137–172

    Google Scholar 

  8. McKinney, E C, Ali, N., Traut, A., Feldmann, K. A, Belostotsky, D. A., McDowell, J A., and Meagher, R. B Sequence based identification of T-DNA insertion mutatrons m Arabldopsls actin mutants act2‐1 and act4‐1. Plant J. 7,613–622

    Google Scholar 

  9. Azpiroz-Leehan, R and Feldmann, K A (1997) T-DNA insertion mutagenesrs m Arabidposis. going back and forth Trends Genet 13, 152–156

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 1998 Human press Inc, Totowa, NJ

About this protocol

Cite this protocol

Winkler, R.G., Feldmann, K.A. (1998). PCR-Based Identification of T-DNA Insertion Mutants. In: Martinez-Zapater, J.M., Salinas, J. (eds) Arabidopsis Protocols. Methods in Molecular Biology™, vol 82. Humana Press. https://doi.org/10.1385/0-89603-391-0:129

Download citation

  • DOI: https://doi.org/10.1385/0-89603-391-0:129

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-391-7

  • Online ISBN: 978-1-59259-268-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics