Zhang, FengCong, LeLodato, SimonaKosuri, SriramChurch, GeorgeArlotta, Paola2018-01-242011Zhang, Feng, Le Cong, Simona Lodato, Sriram Kosuri, George M Church, and Paola Arlotta. 2011. Efficient Construction of Sequence-Specific TAL Effectors for Modulating Mammalian Transcription. Nature Biotechnology 29, no. 2: 149–153. doi:10.1038/nbt1775.1087-0156http://nrs.harvard.edu/urn-3:HUL.InstRepos:34728629The 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.en-USJournal Article10.1038/nbt1775