Skip to main content

Advertisement

Log in

Quantitative cDNA-AFLP analysis for genome-wide expression studies

  • Original Paper
  • Published:
Molecular Genetics and Genomics Aims and scope Submit manuscript

Abstract

An improved cDNA-AFLP method for genome-wide expression analysis has been developed. We demonstrate that this method is an efficient tool for quantitative transcript profiling and a valid alternative to microarrays. Unique transcript tags, generated from reverse-transcribed messenger RNA by restriction enzymes, were screened through a series of selective PCR amplifications. Based on in silico analysis, an enzyme combination was chosen that ensures that at least 60% of all the mRNAs were represented by an informative sequence tag. The sensitivity and specificity of the method allows one to detect poorly expressed genes and distinguish between homologous sequences. Accurate gene expression profiles were determined by quantitative analysis of band intensities, and subtle differences in transcriptional activity were revealed. A detailed screen for cell cycle-modulated genes in tobacco demonstrates the usefulness of the technology for genome-wide expression analysis.

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

Access this article

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

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1.
Fig. 2A–D.
Fig. 3A, B.
Fig. 4.

Similar content being viewed by others

References

  • Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25:3389–3402

    PubMed  Google Scholar 

  • Bachem CWB, van der Hoeven RS, de Bruijn SM, Vreugdenhil D, Zabeau M, Visser RGF (1996) Visualization of differential gene expression using a novel method of RNA fingerprinting based on AFLP: analysis of gene expression during potato tuber development. Plant J 9:745–753

    CAS  PubMed  Google Scholar 

  • Breyne P, Zabeau M (2001) Genome-wide expression analysis of plant cell cycle modulated genes. Curr Opin Plant Biol 4:136–142

    CAS  PubMed  Google Scholar 

  • Breyne P, Dreesen R, Vandepoele K, De Veylder L, Van Breusegem F, Callewaert L, Rombauts S, Raes J, Cannoot B, Engler G, Inzé D, Zabeau M (2002) Analysis of the transcriptome during cell division in plants. Proc Natl Acad Sci USA 99:14825–14830

    Article  CAS  PubMed  Google Scholar 

  • Cho RJ, Huang M, Campbell MJ, Dong H, Steinmetz L, Sapinoso L, Hampton G, Elledge SJ, Davis RW, Lockhart DJ (2001) Transcriptional regulation and function during the human cell cycle. Nature Genet 27:48–54

    CAS  PubMed  Google Scholar 

  • Durrant WE, Rowland O, Piedras P, Hammond-Kosack KE, Jones JDG (2000) cDNA-AFLP reveals a striking overlap in race-specific resistance and wound response gene expression profiles. Plant Cell 12:963–977

    Article  CAS  PubMed  Google Scholar 

  • Eisen MB, Spellman PT, Brown PO, Botstein D (1998) Cluster analysis and display of genome-wide expression patterns. Proc Natl Acad Sci USA 95:14863–14868

    CAS  PubMed  Google Scholar 

  • Galbraith DW, Harkins KR, Maddox JM, Ayres NM, Sharma DP, Firoozabady E (1983) Rapid flow cytometric analysis of the cell cycle in intact plant tissues. Science 220:1049–1051

    CAS  Google Scholar 

  • Kornmann B, Preitner N, Rifat D, Fleury-Olela F, Schibler U (2001) Analysis of circadian liver gene expression by ADDER, a highly sensitive method for the display of differentially expressed mRNAs. Nucleic Acids Res 29:e51

    CAS  PubMed  Google Scholar 

  • Laub MT, McAdams HH, Feldblyum T, Fraser CM, Shapiro L (2000) Global analysis of the genetic network controlling a bacterial cell cycle. Science 290:2144–2148

    Article  CAS  PubMed  Google Scholar 

  • Luger K, Mäder AW, Richmond RK, Sargent DF, Richmond TJ (1997) Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389:251–260

    CAS  PubMed  Google Scholar 

  • Meshi T, Taoka K-i, Iwabuchi M (2000) Regulation of histone gene expression during the cell cycle. Plant Mol Biol 43:643–657

    Article  CAS  PubMed  Google Scholar 

  • Nagata T, Nemoto Y, Hasezawa S (1992) Tobacco BY-2 cell line as the "HeLa" cell in the cell biology of higher plants. Int Rev Cytol 132:1–30

    CAS  Google Scholar 

  • Qin L, Overmars H, Helder J, Popeijus H, Rouppe van der Voort J, Groenink W, van Koert P, Schots A, Bakker J, Smant G (2000) An efficient cDNA-AFLP-based strategy for the identification of putative pathogenicity factors from the potato cyst nematode Globodera rostochiensis. Mol Plant-Microbe Interact 13:830–836

    CAS  Google Scholar 

  • Rombauts S, Van de Peer Y, Rouzé P (2003) AFLPinSilico, simulating AFLP fingerprints. Bioinformatics, in press

  • Reichheld JP, Gigot C, Chaubet-Gigot N (1998) Multilevel regulation of histone gene expression during the cell cycle in tobacco cells. Nucleic Acids Res 26:3255–3262

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual (2nd edn). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.

    Google Scholar 

  • Spellman PT, Sherlock G, Zhang MQ, Iyer VR, Anders K, Eisen MB, Brown PO, Botstein D, Futcher B (1998) Comprehensive identification of cell cycle-regulated genes of the yeast Saccharomyces cerevisiae by microarray hybridization. Mol Biol Cell 9:3273–3297

    CAS  PubMed  Google Scholar 

  • Sutcliffe JG, Foye PE, Erlander MG, Hilbush BS, Bodzin LJ, Durham JT, Hasel KW (2000) TOGA: an automated parsing technology for analyzing expression of nearly all genes. Proc Natl Acad Sci USA 97:1976–1981

    CAS  PubMed  Google Scholar 

  • Tavazoie S, Hughes JD, Campbell MJ, Cho RJ, Church GM (1999) Systematic determination of genetic network architecture. Nature Genet 22:281–285

    Article  CAS  PubMed  Google Scholar 

  • Van der Biezen EA, Juwana H, Parker JE, Jones JDG (2000) cDNA-AFLP display for the isolation of Peronospora parasitica genes expressed during infection in Arabidopsis thaliana. Mol Plant-Microbe Interact 13:895-898

    Google Scholar 

  • Vos P, Hogers R, Bleeker M, Reijans M, van de Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M, Zabeau M (1995) AFLP: a new technique for DNA fingerprinting. Nucleic Acids Res 23:4407–4414

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank C. Tréhin for help with the BY2 synchronization experiments, M. Vuylsteke for advice on statistical evaluation of the data and valuable comments, and M. De Cock for help with the manuscript. K.V. is indebted to the Vlaams Instituut voor de Bevordering van het Wetenschappelijk-Technologisch Onderzoek in de Industrie for a predoctoral fellowship

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to D. Inzé.

Additional information

Communicated by R. G. Herrmann

Rights and permissions

Reprints and permissions

About this article

Cite this article

Breyne, P., Dreesen, R., Cannoot, B. et al. Quantitative cDNA-AFLP analysis for genome-wide expression studies. Mol Gen Genomics 269, 173–179 (2003). https://doi.org/10.1007/s00438-003-0830-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00438-003-0830-6

Keywords

Navigation