Multiplexed superresolution crispr imaging of chromatin in living cells

Abstract

Visualization of the spatiotemporal organization of chromatin is highly desirable in the study of genome function regulations. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPRassociated endonuclease system has shown great promise for application in real-time chromatin imaging due to its DNA targeting ability in living cells. Previous studies typically used fluorescent proteins to generate fluorescent signals which, however, have trade-offs among signal intensity, multiplexibility, and simplicity. Here, we developed a CRISPR-based fluorescence probe for real-time imaging of chromatin in living cells. We demonstrated multicolor imaging of chromatin elements, including telomeres and major satellites with an approximately fivefold enhancement of the signal-to-background ratio. We also realized real-time monitoring of the dynamics of the target loci in living cells. The high brightness of this probe allowed sub-diffraction-limit imaging of telomeres, using a stimulated emission depletion microscopy. We concluded that our new CRISPR imaging system holds great potential for studying intracellular structures and functions with high spatiotemporal resolution.