In situ Transcriptome and Genome Imaging in Single Cells
Howard Hughes Medical Institute, Harvard University, Cambridge, MA, USA
In situ transcriptomic analysis of single cells promise to transform our understanding in many areas of biology, such as regulation of gene expression, development of cell fate, and organization of distinct cell types in complex tissues. We developed a single-cell transcriptome imaging method, multiplexed error-robust fluorescent in situ hybridization (MERFISH), which uses combinatorial labeling and sequential imaging to massively multiplex single-molecule FISH measurements and error-robust encoding schemes to minimize measurement error, enabling RNA imaging and profiling at the transcriptomic scale. Specifically, we assign error-robust barcodes to individual RNA species, label RNAs combinatorically with oligonucleotide probes that represent these barcodes, and read out these barcodes through sequential rounds of imaging. Using this approach, we have imaged hundreds to thousands of RNA species in individual cells both in culture and in complex tissues with high throughput. By enabling single-cell transcriptomic analysis in the native context of cells and tissues, MERFISH facilitates the delineation of gene regulatory networks, the mapping of RNA distributions inside cells, and the mapping of distinct cell types in complex tissues. In this presentation, I will talk about our technology development and recently applications of MERFISH.
I will also talk about a multiplexed FISH method that we developed for tracing the 3D conformation of chromosomes in single cells. The spatial organization of genome plays an important role in many essential genome functions from gene regulation to genome replication.However, many gaps remain in our understanding of the 3D organization of chromatin in the nucleus, partly because of the lack of proper imaging tools. Our multiplex FISH method allows numerous genomic loci to be imaged and localized. Using this approach, we traced the 3D conformation of chromatin in cell nucleus with high resolution, and revealed novel spatial organization of chromatin domains and compartments in individual chromosomes.