Journal
NATURE PROTOCOLS
Volume 7, Issue 1, Pages 171-192Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/nprot.2011.431
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Funding
- Zhang laboratory
- Howard Hughes Medical Institute
- Simons Foundation
- Massachusetts Institute of Technology
- US National Institutes of Health
- McKnight Foundation
- NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE [R01NS073124] Funding Source: NIH RePORTER
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Transcription activator-like effectors (TALEs) are a class of naturally occurring DNA-binding proteins found in the plant pathogen Xanthomonas sp. The DNA-binding domain of each TALE consists of tandem 34-amino acid repeat modules that can be rearranged according to a simple cipher to target new DNA sequences. Customized TALEs can be used for a wide variety of genome engineering applications, including transcriptional modulation and genome editing. Here we describe a toolbox for rapid construction of custom TALE transcription factors (TALE-TFs) and nucleases (TALEs) using a hierarchical ligation procedure. This toolbox facilitates affordable and rapid construction of custom TALE-TFs and TALEs within 1 week and can be easily scaled up to construct TALEs for multiple targets in parallel. We also provide details for testing the activity in mammalian cells of custom TALE-TFs and TALEs using quantitative reverse-transcription PCR and Surveyor nuclease, respectively. The TALE toolbox described here will enable a broad range of biological applications.
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