Skip to main content

Abstract

TILLING (Targeted Induced Local Lesions in Genomes; McCallum et al. 2000) is a reverse genetic tool for the identification of point mutations in genes of interest within EMS mutagenised populations (Till et al. 2003, Wienholds et al. 2003, Smits et al. 2004), facilitating the efficient screening of a considerable range of mutant alleles for their relation to gene function. TILLING employs the mismatch specific endonuclease CELI which enzymatically identifies any single nucleotide polymorphism between PCR products by recognising sites of mismatch and cleaving the DNA. Several TILLING projects have been instigated in a variety of organisms such as Arabidopsis thaliana, Medicago truncatula, pea, soybean, maize, nematode, rat and zebrafish. In the following chapter, an outline of the procedure and methodology is described, and the initial results from the Lotus japonicus TILLING project are presented.

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

Access this chapter

Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  • The C. elegans Sequencing Consortium (1998) Genome Sequence of the Nematode C. elegans: A Platform for Investigating Biology Science 282, 2012–2018.

    Google Scholar 

  • Colbert T, Till BJ, Tompa R, Reynolds S, Steine MN, Yeung AT, McCallum CM, Comai L, Henikoff S. (2001) High-throughput screening for induced point mutations. Plant Physiology 126, 480–484.

    Article  CAS  PubMed  Google Scholar 

  • The EU Arabidopsis Genome project (1998) Analysis of 1.9 Mb of contiguous sequence from chromosome 4 of Arabidopsis thaliana Nature 391, 485–488.

    Google Scholar 

  • Hayashi M, Miyahara A, Sato S, Kato T, Yoshikawa M, Taketa M, Hayashi M,. Pedrosa A, Onda R, Imaizumi-Anraku H, Bachmair A, Sandal N, Stougaard J, Murooka Y, Tabata S, Kawasaki S, Kawaguchi M, and Harada K. (2001) Construction of a genetic linkage map of the model legume Lotus japonicus using an interspecific F2 population. DNA Research 8, 301–310.

    CAS  PubMed  Google Scholar 

  • International Human Genome Sequencing Consortium (2001) Initial Sequencing and analysis of the human genome. Nature 309, 860–921.

    Google Scholar 

  • Mc Callum CM, Comai L, Greene EA and Henikoff S. (2000) Targeted screening for induced mutations Nature Biotechnology 18, 455–457.

    Google Scholar 

  • Perry JA, Wang TL, Welham TJ, Gardner S, Pike JM, Yoshida S and Parniske M. (2003) A TILLING reverse genetics tool and a web accessible collection of mutants of the legume Lotus japonicus. Plant Physiology 131, 866–871.

    Article  CAS  PubMed  Google Scholar 

  • Oleykowski CA, Bronson Mullins CR, Godwin AK and Yeung AT (1998) Mutation detection using a novel plant endonuclease Nucleic Acids Research 26, 204597–4602.

    Article  Google Scholar 

  • Sambrook J, Fritsch E, and Maniatis T. (2001) Molecular Cloning: a Laboratory Manual. 3rd edition Cold Spring Harbor Press, Cold Spring Harbor, New York A8.19.

    Google Scholar 

  • Smits BM, Mudde J, Cuppen E, Plasterk RH. (2004 )Target-selected mutagenesis of the rat. Genomics 83, 332–334.

    Article  CAS  PubMed  Google Scholar 

  • Stracke S, Kistner C, Yoshida S, Mulder L, Sato S, Kaneko T, Tabata S, Sandal N, Stougaard J, Szczyglowski K, Parniske M. (2002) A receptor-like kinase required for both bacterial and fungal symbiosis. Nature 417, 959–962.

    Article  CAS  PubMed  Google Scholar 

  • Till BJ, Reynolds SH, Greene EA, Codomo CA, Enns LC, Johnson JE, Burtner C, Odden AR, Young K, Taylor NE, Henikoff JG, Comai L, Henikoff S. (2003) Large-scale discovery of induced point mutations with high-throughput TILLING. Genome Research 13, 524–530.

    Article  CAS  PubMed  Google Scholar 

  • Till BJ, Colbert T, Tompa R, Enns L, Codomo C, Johnson J, Reynolds SH, Henikoff JG, Greene EA, Steine M, Comai N, and Henikoff S. (2003) High-throughput TILLING for functional genomics. In: Plant Functional Genomics: Methods and Protocols (Grotewald E, Ed.), Humana Press, 236; 205–220.

    Google Scholar 

  • Wienholds E, Van Eeden F, Kosters M, Mudde J, Plasterk RH, and Cuppen E (2003) Efficient target-selected mutagenesis in zebrafish. Genome Research 13, 2700–2007.

    Article  CAS  PubMed  Google Scholar 

  • Yang B, Wen X, Kodali NS, Oleykowski CA, Miller CG, Kulinski J, Besak D, Yeung JA, Kowalski D, and Yeung AT (2000) Purification, Cloning and Characterization of the CEL I Nuclease. Biochemistry 39, 3533–3541.

    CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Trevor Wang .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2005 Springer

About this chapter

Cite this chapter

Perry, J., Welham, T., Cheminant, S., Parniske, M., Wang, T. (2005). Tilling. In: Márquez, A.J. (eds) Lotus japonicus Handbook. Springer, Dordrecht. https://doi.org/10.1007/1-4020-3735-X_20

Download citation

Publish with us

Policies and ethics