File:The schematic diagram of ZFNs.png

English: ZFNs are the fusion of artificially modified zinc-finger protein (ZFP), having specific binding activity, and Fok Ⅰ endonuclease, having non-specific cutting activity. Several amino acid residues on the α helix of ZFP play a direct role in the recognition of the target site and can pair with bases adjacent to it. For example, the −1, +3, +6 locus on the α helix of Zif268 (a kind of ZFP) can directly recognize and bind to three adjacent bases on the target sequence. In addition, the binding domain and cutting domain of the Fok Ⅰ endonuclease can be separated, and the non-specific cutting function of the Fok Ⅰ endonuclease can be obtained when the binding domain is removed. Finally, under the action of Fok Ⅰ dimer, the target site will be cut, and DSBs will be generated. ZFNs, as the first generation of gene editing technology, have been successfully applied in animals at first and have attracted wide attention. Subsequently, ZFNs were applied in corn, rice, and other crops. However, there are inherent flaws in this system. For example, the inter-ZFP context effect, that is, it is difficult to achieve high efficiency by simply linking specific ZFPs together. In addition, there are shortcomings such as unsuitability for multiple editing, high off-target efficiency, and high cytotoxicity, which are gradually being overcome by the later gene editing tools. Therefore, its application in crops is not well developed.