体细胞核移植可以将终末分化的体细胞重编程为具有全能性的胚胎，但是核移植胚胎往往存在明显的体内外发育缺陷。近年来，随着微量组学技术的发展，核移植胚胎中表观修饰的不完全重编程机制已经逐渐被揭示。但是受限于材料及技术瓶颈等问题，核移植胚胎中重要转录因子的调控机制，迄今尚未有研究。本研究将充分利用课题组已优化完善的少量细胞CUT&RUN-seq技术和相关分析方法，系统揭示核移植胚胎发育过程中转录因子CTCF在全基因组水平上的分布特征和动态变化规律。结合多种组学数据，对具有不同发育命运的核移植胚胎进行CTCF结合情况的比较，筛选出其中与胚胎发育命运密切相关的CTCF结合缺陷位点。并运用基因及表观修饰编辑技术，探讨CTCF结合缺陷的靶向修复方案对核移植胚胎发育的影响。本研究将有助于更深入的了解体细胞核移植介导的遗传和表观遗传重塑的分子机制，为进一步优化现有核移植技术提供重要的理论基础。

Somatic cell nuclear transfer (SCNT) enables cloning of differentiated cells by reprogramming their nuclei to a totipotent state. However, successful full-term development of SCNT embryos is a low-efficiency process and cloned embryos frequently exhibit severe abnormalities. Recent technical advances, particularly low-input sequencing techniques, have enabled analysis of transcriptome and epigenetic changes during SCNT reprogramming. Despite high resolution and robustness of the traditional chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq), millions of cells are needed, thereby limiting its application in rare biological samples. Therefore, how key transcriptional factors serve as critical barriers in SCNT-mediated reprogramming is unknown. In the present study, we will make full use of our advanced small-scale cleavage under targets and release using nuclease sequencing (CUT&RUN-seq) technology and bioinformatics method to generate, for the first time ever to our knowledge, a genome-wide CTCF-binding roadmap from mouse pre-implantation SCNT embryos. In addition, we will identify and rescue the widespread CTCF-binding defects by using targeted gene editing and/or epigenetic editing systems. Moreover, we will explore the synergistic effect of CTCF and epigenetic modifications in SCNT-mediated reprogramming process. Our study will significantly advance the understanding of genetic and epigenetic remodeling mechanism during nuclear transfer embryo development, and eventually facilitate in achieving higher cloning efficiency.