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FRT-seq: amplification-free, strand-specific transcriptome sequencing


We report an alternative approach to transcriptome sequencing for the Illumina Genome Analyzer, in which the reverse transcription reaction takes place on the flowcell. No amplification is performed during the library preparation, so PCR biases and duplicates are avoided, and because the template is poly(A)+ RNA rather than cDNA, the resulting sequences are necessarily strand-specific. The method is compatible with paired- or single-end sequencing.

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Figure 1: Correlation plots for FRT-seq libraries.
Figure 2: Strand-specificity of FRT-seq.


  1. Wang, Z., Gerstein, M. & Snyder, M. Nat. Rev. Genet. 10, 57–63 (2009).

    CAS  Article  Google Scholar 

  2. Wu, J.Q. et al. Genome Biol. 9, R3 (2008).

    Article  Google Scholar 

  3. Ozsolak, F. et al. Nature 461, 814–818 (2009).

    CAS  Article  Google Scholar 

  4. David, L. et al. Proc. Natl. Acad. Sci. USA 103, 5320–5325 (2006).

    CAS  Article  Google Scholar 

  5. Carninci, P. et al. Science 309, 1559–1563 (2005).

    CAS  Article  Google Scholar 

  6. Katayama, S. et al. Science 309, 1564–1566 (2005).

    Article  Google Scholar 

  7. Lister, R. et al. Cell 133, 523–536 (2008).

    CAS  Article  Google Scholar 

  8. Cloonan, N. et al. Nat. Methods 5, 613–619 (2008).

    CAS  Article  Google Scholar 

  9. Croucher, N.J. et al. Nucleic Acids Res. 37, e148 (2009).

    Article  Google Scholar 

  10. He, Y., Vogelstein, B., Velculescu, V.E., Papadopoulos, N. & Kinzler, K.W. Science 322, 1855–1857 (2008).

    CAS  Article  Google Scholar 

  11. Parkhomchuk, D. et al. Nucleic Acids Res. 37, e123 (2009).

    Article  Google Scholar 

  12. Kozarewa, I. et al. Nat. Methods 6, 291–295 (2009).

    CAS  Article  Google Scholar 

  13. Lipson, D. et al. Nat. Biotechnol. 27, 652–658 (2009).

    CAS  Article  Google Scholar 

  14. Chen, D. & Patton, J.T. Biotechniques 30, 574–580, 582 (2001).

    CAS  Article  Google Scholar 

  15. Hubbard, T.J. et al. Nucleic Acids Res. 37, D690–D697 (2009).

    CAS  Article  Google Scholar 

  16. Marioni, J.C., Mason, C.E., Mane, S.M., Stephens, M. & Gilad, Y. Genome Res. 18, 1509–1517 (2008).

    CAS  Article  Google Scholar 

  17. Bentley, D.R. et al. Nature 456, 53–59 (2008).

    CAS  Article  Google Scholar 

  18. Smedley, D. et al. BMC Genomics 10, 22 (2009).

    Article  Google Scholar 

  19. Li, H., Ruan, J. & Durbin, R. Genome Res. 18, 1851–1858 (2008).

    CAS  Article  Google Scholar 

  20. Yang, Y.H. et al. Nucleic Acids Res. 30, e15 (2002).

    Article  Google Scholar 

  21. Smyth, G.K. Stat. Appl. Genet. Mol. Biol. 3, 3 (2004).

    Article  Google Scholar 

  22. Huber, W., von Heydebreck, A., Sultmann, H., Poustka, A. & Vingron, M. Bioinformatics 18 (Suppl. 1), S96–S104 (2002).

    Article  Google Scholar 

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We thank N. Bason and M. Quail for preparing the standard RNA-seq library, J. Burton for coordinating the sequencing of standard RNA-seq libraries, and J. Ule and M. Wollerton for advice on RNA ligation. This work was supported by the Wellcome Trust grant (WT079643).

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Authors and Affiliations



D.J.T. and T.W.B.O. devised the project; L.M. and D.J.T. devised the experimental protocols; L.M. and E.M.S. planned and carried out laboratory work; R.M.A. and K.D.J. performed data analysis; P.D.E. performed microarray work; and C.F.L. oversaw analysis and array work. D.J.T., J.E.C. and L.M. wrote the manuscript.

Corresponding author

Correspondence to Daniel J Turner.

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Competing interests

T.W.B.O. is an Illumina employee, shareholder and share option holder.

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Supplementary Figures 1–8 and Supplementary Tables 1–6 (PDF 2919 kb)

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Mamanova, L., Andrews, R., James, K. et al. FRT-seq: amplification-free, strand-specific transcriptome sequencing. Nat Methods 7, 130–132 (2010).

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