Study of chromatin dynamics through live visualization of chromosomal domains

Recent studies have suggested that chromosomes are hierarchically organized into topologically associated domains. Direct visualization of the genomic elements in living cells is required to explore the relationship between such structural organization and its functional roles. To study the structural dynamics of genomic elements, we have developed an improved CRISPR-based imaging system for visualization of arbitrary non-repetitive gene loci in living cells. Use of split-fluorescent proteins nearly eliminated background fluorescence and signals from non-specific aggregation, allowing reliable long-term tracking of genomic loci. Stochastic optical reconstruction microscopy of genomic loci in living cells revealed ultrastructural dynamics within the loci, suggesting further extension of the method to live super-resolved imaging of topologically associated domains. These approaches provide novel tools to study the mechanism of reorganizing chromosomes and its relevance in gene regulation