Fig. 1

The principle of the transient expression system used to assess the relative cleavage activity of customized endonucleases. a The incorporation of a target site for sequence-specific endonucleases generates a frame shift in the yfp sequence. b Upon co-transformation of the target vector with TALENs or RGENs, double- strand DNA breaks at the target site are induced. c The imperfect repair of these breaks via non-homologous end-joining can restore the wild type reading frame, thereby leading to expression of yfp and the emission of a YFP signal. The elements shown are not drawn to scale. 2x35SP: doubled enhanced CaMV 35S promoter; LeB4: Vicia faba legumin B4 signal peptide; YFP: synthetic yellow fluorescent protein gene; NOST: A. tumefaciens NOPALINE SYNTHASE termination sequence; FokI: DNA cleavage domain of Flavobacterium okeanokoites type IIS restriction endonuclease; gRNA: guide-RNA; Cas9: Streptococcus pyogenes Cas9; TALEN: transcription activator-like effector nucleases; NHEJ: non-homologous end joining