Efficient Assembly and Verification of ZFNs and TALENs for Modifying Porcine ApoE gene

Abstract

Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are powerful tools for genome engineering. These synthetic nucleases are assembled with programmable, sequence-specific DNA-binding domain and a non-specific FokI cleavage domain. Apolipoprotein E (ApoE) gene polymorphism is associated with cardiovascular outcomes, including ischaemic stroke and coronary heart disease (CHD). So the objective of this study is to create mutations of AopE gene by ZFNs and TALENs technology. Here, we used the Context-dependent assembly (CoDA) method to design and screen ZFNs specifically targeting with ApoE gene. The targeted cleavage capacity of these ZFNs was validated in yeast system and HEK 293T cells. Meanwhile, an efficient assembled TALENs to target ApoE gene in HEK 293T cells was as a control. The results showed that both ZFNs and TALENs worked on ApoE gene with similar high-efficiency cleavage capability. The result would provide efficient methods for genome editing, so as to get disease model for gene therapy for the further study.