Development of a high-throughput technology capable of detecting in situ DNA-RNA interactions and transcriptomes in single cells

While methods for detecting single cell transcriptome and genome architecture have been established, there are currently no technologies for detecting single-cell DNA-RNA interactions. Thus, while a regulatory role for chromatin and RNA organization has been hypothesized, we still lack a comprehensive understanding of where and how chromatin and RNA interact, as well as the consequences for gene regulation.In this thesis, I describe a breakthrough technology called single cell in situ Mapping of RNA-Genome Interactome (sciMARGI). Through integrating two promising technologies, combinatorial indexing and microdroplet methods, we can decode single-cell single-complex information by labeling interacting DNA and RNA molecules with unique cell and cluster barcodes. Currently, a DNA library based on human brain tissue has been successfully generated and shown to have a proper single cell cluster distribution and DNA-DNA interaction map. While the RNA library still needs additional adjustments, the libraries as a whole demonstrated promising results. When fully developed, the sciMARGI technology will not only detect three-way in situ interactions between DNA and DNA, DNA and RNA, and RNA and RNA, but also quantify RNA within the single nucleus, establishing a direct link between chromatin-RNA organization and gene expression. Due to its exceptional versatility, sciMARGI can be applied to not only cell lines but also complex tissues, allowing us to resolve some of the most critical obstacles regarding gene expression regulatory processes