Highly efficient genome editing via 2A-coupled co-expression of two TALEN monomers.

ABSTRACT: Transcription activator-like effector nucleases (TALENs) are a useful tool for targeted gene editing. TALEN monomers are traditionally expressed from two different plasmids. Each encodes a different TALEN arm that binds to a user-defined sequence and mediates gene editing. Expression of TALEN monomers in two separate plasmids requires co-delivery of each plasmid to the cell. Efficacy of gene editing may be increased if each monomer was transcribed from the same reading frame.We developed a TALEN scaffold which expresses both TALEN monomers from a single open reading frame in equal molar amount by linking both monomers with a 2A self-cleaving peptide sequence. This TALEN scaffold, named pTAL10, demonstrates higher levels of genome editing than co-transfected TALENs at similar levels of transfection efficiencies when analyzed for TALEN-induced small insertions and deletions.This protocol for gene editing using 2A-linked TALENs requires transfection of only one plasmid as compared to transfection of two separate plasmids encoding each TALEN monomers.

SUBMITTER: Mariano A 

PROVIDER: S-EPMC4167141 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature