De novo assembly and delivery of synthetic megabase-scale human DNA into mouse early embryos

作者：Yue Liu, Jianting Zhou, Duo Liu, Xiaoyu Hu, Lin Yang, Xueru Song, Xiaodong Jin, Wei Xie, Luhan Yang, Zichuan Liu, Ying‐Jin Yuan · 发表于：Nature Methods · 年份：2025 · DOI：10.1038/s41592-025-02746-8 · 被引用次数：14 · 研究领域：CRISPR and Genetic Engineering、Animal Genetics and Reproduction、Pluripotent Stem Cells Research

Epigenetic modifications on natural chromosomes are inherited and maintained in a default state, making it challenging to remove intrinsic marks to study the fundamental principles of their establishment and further influence on transcriptional regulation. In this study, we developed SynNICE, a method for assembling and delivering intact, naive, synthetic megabase (Mb)-scale human DNA into early mouse embryos, to study de novo epigenetic regulation. By assembling and delivering a 1.14-Mb human AZFa (hAZFa) locus, we observed the spontaneous incorporation of murine histones and the establishment of DNA methylation at the one-cell stage. Notably, DNA methylation from scratch strongly enriches at repeat sequences without H3K9me3 reinforcement. Furthermore, the transcription of hAZFa initiated at the four-cell stage is regulated by newly established DNA methylation. This method provides a unique platform for exploring de novo epigenomic regulation mechanisms in higher animals. This work presents a strategy for synthesizing and assembling megabase-scale DNA, along with a delivery method for introducing the synthetic DNA into mouse embryos, offering a valuable tool for investigating de novo epigenetic regulation.