Letter | Published:

Efficient construction of sequence-specific TAL effectors for modulating mammalian transcription

Nature Biotechnology volume 29, pages 149153 (2011) | Download Citation


The ability to direct functional proteins to specific DNA sequences is a long-sought goal in the study and engineering of biological processes. Transcription activator–like effectors (TALEs) from Xanthomonas sp. are site-specific DNA-binding proteins that can be readily designed to target new sequences. Because TALEs contain a large number of repeat domains, it can be difficult to synthesize new variants. Here we describe a method that overcomes this problem. We leverage codon degeneracy and type IIs restriction enzymes to generate orthogonal ligation linkers between individual repeat monomers, thus allowing full-length, customized, repeat domains to be constructed by hierarchical ligation. We synthesized 17 TALEs that are customized to recognize specific DNA-binding sites, and demonstrate that they can specifically modulate transcription of endogenous genes (SOX2 and KLF4) in human cells.

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This work was supported by the Harvard Society of Fellows (F.Z.), National Human Genome Research Institute Center for Excellence in Genomics Science (P50 HG003170, G.M.C.), Department of Energy Genomes to Life (DE-FG02-02ER63445, G.M.C.), Defense Advanced Research Projects Agency (W911NF-08-1-0254, G.M.C.), the Wyss Institute for Biologically Inspired Engineering (G.M.C.) and National Institutes of Health Transformative R01 (R01 NS073124-01, F.Z. and P.A.). S.L. was partially supported by a predoctoral fellowship from the European School of Molecular Medicine (S.E.M.M.). We thank the entire Church and Arlotta laboratories for discussion and support.

Author information

Author notes

    • Feng Zhang

    Present address: Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA; McGovern Institute for Brain Research, MIT, Cambridge, Massachusetts, USA; and Department of Brain and Cognitive Sciences, MIT, Cambridge, Massachusetts, USA.

    • Feng Zhang
    •  & Le Cong

    These authors contributed equally to this work.


  1. Society of Fellows, Harvard University, Cambridge, Massachusetts, USA.

    • Feng Zhang
  2. Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA.

    • Feng Zhang
    • , Le Cong
    • , Sriram Kosuri
    •  & George M Church
  3. Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, Massachusetts, USA.

    • Feng Zhang
    • , Le Cong
    • , Sriram Kosuri
    •  & George M Church
  4. Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts, USA.

    • Le Cong
  5. Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts, USA.

    • Feng Zhang
    • , Simona Lodato
    •  & Paola Arlotta
  6. Center for Regenerative Medicine and Department of Neurosurgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.

    • Simona Lodato
    •  & Paola Arlotta


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F.Z. and L.C. conceived the study. F.Z., L.C., S.L. and S.K. designed, performed and analyzed all experiments. P.A. supervised the work of S.L. and G.M.C. supervised the work of F.Z., L.C. and S.K. G.M.C., P.A. and F.Z. provided support for this study. F.Z., L.C. and P.A. wrote the manuscript with support from all authors. G.M.C and P.A. equally contributed to this work.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Feng Zhang or Paola Arlotta.

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    Supplementary Tables 1–3, Supplementary Figs. 1–3, Supplementary Methods and Supplementary Sequences

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