Advancing Locus-specific Chromatin Isolation Methods for Multi-omic Discoveries

Abstract

Chromatin functions, such as gene expression, DNA replication and repair, and 3D genome architecture, are mediated by the concerted action of the unique molecular repertoire of interacting proteins and DNAs at the gene locus of interest. Thus, a comprehensive functional understanding of a locus requires the thorough characterization of its local chromatin composition. As a prerequisite to unbiased identification of locus-bound factors, the purification of specific gene loci has attracted considerable scientific attention as an important technical challenge that if solved, promises crucial biological discoveries. During my dissertation work, I have contributed to the development of locus-specific chromatin isolation methods easily adaptable to large, repetitive loci as well as small, single-copy ones. The work I describe here includes the development of "DNA O-MAP", a labeling method that biotinylates proteins around a gene locus using peroxidase-conjugated oligonucleotides. In collaboration with colleagues, I demonstrate that DNA O-MAP, coupled with sample multiplexed quantitative proteomics and next-generation sequencing, can generate well-annotated DNA-protein networks at specific gene loci.