Portal de Periódicos da CAPES

Repeating pattern of non-RVD variations in DNA-binding modules enhances TALEN activity

2013; Nature Portfolio; Volume: 3; Issue: 1 Linguagem: Inglês

10.1038/srep03379

2045-2322

Tetsushi Sakuma, Hiroshi Ochiai, Takehito Kaneko, Tomoji Mashimo, Daisuke Tokumasu, Yuto Sakane, Kenichi Suzuki, Tatsuo Miyamoto, Naoaki Sakamoto, Shinya Matsuura, Takashi Yamamoto,

Plant Virus Research Studies

Transcription activator-like effector (TALE) nuclease (TALEN) is a site-specific nuclease, which can be freely designed and easily constructed. Numerous methods of constructing TALENs harboring different TALE scaffolds and repeat variants have recently been reported. However, the functionalities of structurally different TALENs have not yet been compared. Here, we report on the functional differences among several types of TALENs targeting the same loci. Using HEK293T cell-based single-strand annealing and Cel-I nuclease assays, we found that TALENs with periodically-patterned repeat variants harboring non-repeat-variable di-residue (non-RVD) variations (Platinum TALENs) showed higher activities than TALENs without non-RVD variations. Furthermore, the efficiencies of gene disruption mediated by Platinum TALENs in frogs and rats were significantly higher than in previous reports. This study therefore demonstrated an efficient system for the construction of these highly active Platinum TALENs (Platinum Gate system), which could establish a new standard in TALEN engineering.