Imaging and Tracing Multiple Genetic Elements via Multiplexed CRISPR Imaging

Abstract

Higher order chromatin structures and nuclear genome organization play a key role in regulating gene functions in addition to other epigenetic markers such as DNA and histone modifications, which have been extensively studied. Genetic variations in the human genome are known to lead to several human diseases but little is known yet about how the structural organization of the chromatin fibers affects gene regulation, cell differentiation, and disease pathways. Understanding the dynamics of native chromatin in living cells by imaging and tracing specific gene loci and nuclear transcripts would allow us to uncover the correlation between chromosome organizations and transcriptional activity as well as the functional mechanism of sub-nuclear distribution of chromosomal segments induced by genetic defects. To study the dynamics of multiple genetic elements, we have developed a method to target gene loci and nuclear transcripts with minimally interrupting labels based on CRISPR system. Here we describe the application of multiplexed CRISPR imaging to study the nuclear organization of genes and discuss the approaches to study genome-wide chromatin compaction and physical map of chromosome territories.Keyword: Multiplexed CRISPR imaging, single cell, chromatin dynamics