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. 2012 Jan 24;1(1):e3.
doi: 10.1038/mtna.2011.5.

Targeting DNA With Fingers and TALENs

Daniel F Carlson  1 , Scott C Fahrenkrug, Perry B Hackett
Affiliations

Targeting DNA With Fingers and TALENs

Daniel F Carlson et al. Mol Ther Nucleic Acids. .
No abstract available

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Figures

Figure 1
Figure 1
Comparison of zinc-finger nuclease (ZFN) and Transcription Activator-Like Effector Nuclease (TALEN) architecture. (a) ZFNs. Each ZFN polypeptide consists of two functional domains, a DNA-binding domain comprising a chain of finger modules (ZFs) that each typically recognize a unique 3-base pair sequence of DNA and a DNA-cleaving domain composed of the nuclease domain of the FokI nuclease. FokI functions as a dimer, hence when two FokI nucleases bind to DNA proximal to one another they can dimerize and introduce a double-strand break. Targeted double-strand DNA cutting can be obtained by designing zinc fingers for specific sequences that flank the desired cleavage site; in the example 12 base pairs per ZFN are targeted with polypeptides containing four zinc-finger modules each (ZF-1 through ZF-4 and ZF-5 through ZF-8). (b) Model of a TALEN. A TAL Effector (TALE) polypeptide contains a series of typically 34-amino acid repeats, of which residues 12 and 13 [repeat variable diresidues (RVDs) shown in orange] are responsible for recognition of a specific base as shown in the box (note that there is some discussion about the precision of the RVD NK recognition of G and other RVDs can specify base contacts). FokI nuclease is fused to the C-terminal end of the protein using wild-type TALE sequence as a spacer. Several spacer lengths between the TALE binding core and FokI have demonstrated activity. The number of tandem 34-amino acid repeats in the binding core defines the length of the recognition sequence, and the end of the functional DNA-binding motif. Each target sequence must be preceded by a T nucleotide. Two TALENs are shown to assemble on a genomic sequence in the opposite polarity to ZFNs to form a heterodimeric cleavage complex.
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