Transcription activator-like effectors (TALEs) contain programmable DNA-binding domains that can be fused to various effectors to manipulate genetic sequences or transcriptional state. However, the construction of plasmids encoding the modular DNA-binding domain remains challenging due to their repetitive nature. Here, we describe methods for a simple TALE assembly reaction (STAR) that uses a 68-part plasmid library to create TALEs binding to 17 bp target sequences. Manual production of many tens of TALEs can be achieved using a simple 8 h protocol, with full length sequence-verified plasmids available within a few days. This simple tale assembly reaction (STAR) provides a convenient method for generating tens to hundreds of TALENs or TALE-TFs without the need for large plasmid libraries or expensive liquid handling.
Keywords: Gibson assembly; Mammalian; Plasmid; TAL effector; TALEN.