PROJECT SUMMARY / ABSTRACT The epigenetic status of genomes including nucleosome occupancy, chromatin accessibility and DNA methylation, is highly relevant to the regulation of DNA-template biological processes from transcription to DNA replication and repair. Most existing techniques for characterizing epigenome utilize Next Generation Sequencing (NGS) following the biochemical reactions that capture the signals (e.g., bisulfite conversion, cross-linking and chromatin immunoprecipitation/ChIP). The development of these techniques has dramatically accelerated the research of different epigenetics events and has led to many important biological findings. As the new technique nanopore sequencing has been optimized to convey robust sequencing data of single DNA molecules with long read length, it brings in new discernible information that is useful for addressing certain challenging but important epigenetics problems. We develop an experimental protocol MeSMLR-seq and a series of bioinformatics methods to define the multiple types of epigenetic events, including nucleosome occupancy, chromatin accessibility and DNA methylation, at the rarely explored genome regions and biological context by leveraging the unique information of nanopore sequencing. Utilizing these experimental and bioinformatics methods, we aim to interrogate the following problems during the epigenetic reprogramming of early embryonic development and primordial germ cell development: in Aim 1, we will study the epigenetic regulation of transposable element expression and transposition. Aim 2 will construct the allele-specific epigenome and identify the genome loci with significant epigenome difference between alleles. Aim 3 will identify significant difference of the epigenomes between DNA strands and investigate their regulatory roles and dynamics during epigenetic reprogramming. These studies are anticipated to provide the first experimental and bioinformatics platform for improve our understanding of epigenome with complex biomedical context in a comprehensive manner.

PROJECT NARRATIVE / PUBLIC HEALTH RELEVANCE STATEMENT Our innovative experimental approach using nanopore sequencing provides an unparalleled opportunity to study complex epigenetics events at stem cell biology and germ cell biology. In addition, our sophisticated bioinformatics platform will provide a set of robust and handy tools to fully analyze nanopore sequencing epigenome data, such that this cutting-edge technology can be feasible in biomedical research laboratories of all sizes. The analysis of nanopore sequencing epigenome data can identify epigenetic events at each single long DNA molecules, providing a solid foundation for precision medicine.