Project Summary/Abstract Chromatin organization and epigenomic modifications such as cytosine methylation play instrumental roles in gene regulation during human development and in healthy and disease states. Although the genome is nearly identical in each cell in the human body, chromatin conformation and cytosine DNA methylation are highly dynamic across cell types and during development. Previous studies showed that chromatin looping can be regulated by cytosine methylation. However, our current state of knowledge of the interactions between chromatin organization and DNA methylation is built on analyzing cultured cells and bulk tissues. The interaction between cell-type specific chromatin looping and methylation in heterogeneous tissues remain largely unexplored. This project will develop single-cell multi-omic methods to jointly analyze chromatin conformation and cytosine methylation from the same cell. The methods will lead to high quality cell-type specific chromatin conformation maps of human tissue, and the analysis of the relationship between chromatin conformation and cytosine methylation. If successful, the proposed methods will greatly facilitate the study of gene regulation in complex human tissues and in heterogeneous diseases such as cancer.