高效精确的基因工程技术对于发展基因治疗，建立细胞和动物疾病模型具有极为重要的价值。近年来特异性定点核酸酶的研究取得了长足的进步。类转录激活因子效应物核酸酶（TALEN）和常间回文重复序列丛集关联蛋白系统(CRISPR/Cas)具有精确高效，易于设计合成的优点，已经被广泛应用于多种模式生物及人类细胞系中。但其对不同基因位点基因修饰的效率具有很大差异，而我们对此差异产生的机制了解非常有限。这一局限导致我们在选择和设计目标位点时只能采用试错法，不仅影响效率，而且对一些难以修饰的位点没有好的解决办法。在本项目中，我们拟针对CRISPR和TALEN系统对不同位点基因修饰效率的差异，从遗传序列组成和表观遗传修饰两个方面进行系统的研究，从而阐明差异产生的根本机制。对这一差异产生机制的了解将使我们能够针对具体目标位点更有效的选择和设计基因修饰系统，从而提高基因工程的效率，促进新型基因治疗和动物模型的建立。

Efficient and accurate genome engineering technologies are essential for the establishment of novel gene therapy approaches and animal models. Recent development of two types of site-specific nucleases, transcription activator-like effector nucleases (TALENs) and Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins (CRISPR/Cas), greatly expanded our capability of targeted gene modification. Because of the ease of design and application, both systems have been successfully applied to major model organisms and human cell lines.However, using either system, there are vast variations of the efficiency when different genomic loci are targeted and the underlying mechanism is poorly understood. Evolved in bacteria and archaea, CRISPR/Cas and TALEN systems have never been challenged by the complex genomic composition, the formation of chromatin, and the epigenetic modifications in mammalian genomes. Therefore, when introduced into mammalian cells, CRISPR/Cas system's activity and specificity could be significantly affected by these variables. Indeed, studies on a very limited number of genomic loci suggested that TALEN binding to DNA is sensitive to the presence of 5-methylated cytosine. Here, we propose to systematically characterize the efficiency of CRISPR/Cas and TALEN mediated genome editing across major epigenetic marks as well as the complex genomic sequence space, to provide an important reference for future applications and improvements of TALEN and CRISPR/Cas technologies.