Abstract:

TALEN (Transcription activator-like effector nuclease) is a newly developed family of artificially engineered sequence-specific endonucleases, which consists of a highly specific and repetitive DNA-binding domain derived from TALE (Transcription activator-like effector) and fused with the non-specific endonuclease domain of FokⅠ. TALENs have been reported to be able to induce site specific genome modification in quite a few species and in vi-tro cultured cells. Here, we introduced the principles for target site selection and confirmation and described a brief experimental protocol for the easy construction of customized TALENs using unit assembly (UA) method and also for gen-eration of and screening for TALEN-mediated targeted zebrafish mutants through microinjection of TALEN mRNAs into zebrafish embryos. Theoretically, the principles and methods we described here are also applicable to gene targeting in other species.

Key words: transcription activator-like effector nuclease (TALEN), zebrafish, site specific genome modification, gene targeting, reverse genetics approach