Rapid and highly efficient construction of TALE-based transcriptional regulators and nucleases for genome modification
Type
ArticleAuthors
Li, LixinPiatek, Marek J.

Atef, Ahmed
Piatek, Agnieszka Anna

Wibowo, Anjar Tri
Fang, Xiaoyun
Sabir, Jamal Sabir M
Zhu, Jian-Kang

Mahfouz, Magdy M.

KAUST Department
Biological and Environmental Science and Engineering (BESE) DivisionBioscience Program
Center for Desert Agriculture
Laboratory for Genome Engineering
Plant Science
Plant Science Program
Date
2012-01-21Online Publication Date
2012-01-22Print Publication Date
2012-03Embargo End Date
2013-01-21Permanent link to this record
http://hdl.handle.net/10754/562063
Metadata
Show full item recordAbstract
Transcription activator-like effectors (TALEs) can be used as DNA-targeting modules by engineering their repeat domains to dictate user-selected sequence specificity. TALEs have been shown to function as site-specific transcriptional activators in a variety of cell types and organisms. TALE nucleases (TALENs), generated by fusing the FokI cleavage domain to TALE, have been used to create genomic double-strand breaks. The identity of the TALE repeat variable di-residues, their number, and their order dictate the DNA sequence specificity. Because TALE repeats are nearly identical, their assembly by cloning or even by synthesis is challenging and time consuming. Here, we report the development and use of a rapid and straightforward approach for the construction of designer TALE (dTALE) activators and nucleases with user-selected DNA target specificity. Using our plasmid set of 100 repeat modules, researchers can assemble repeat domains for any 14-nucleotide target sequence in one sequential restriction-ligation cloning step and in only 24 h. We generated several custom dTALEs and dTALENs with new target sequence specificities and validated their function by transient expression in tobacco leaves and in vitro DNA cleavage assays, respectively. Moreover, we developed a web tool, called idTALE, to facilitate the design of dTALENs and the identification of their genomic targets and potential off-targets in the genomes of several model species. Our dTALE repeat assembly approach along with the web tool idTALE will expedite genome-engineering applications in a variety of cell types and organisms including plants. © 2012 Springer Science+Business Media B.V.Citation
Li, L., Piatek, M. J., Atef, A., Piatek, A., Wibowo, A., Fang, X., … Mahfouz, M. M. (2012). Rapid and highly efficient construction of TALE-based transcriptional regulators and nucleases for genome modification. Plant Molecular Biology, 78(4-5), 407–416. doi:10.1007/s11103-012-9875-4Publisher
Springer NatureJournal
Plant Molecular BiologyPubMed ID
22271303PubMed Central ID
PMC3580834Additional Links
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3580834http://europepmc.org/articles/pmc3580834?pdf=render
ae974a485f413a2113503eed53cd6c53
10.1007/s11103-012-9875-4
Scopus Count
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