Quantifying the global binding and target-search dynamics of epigenetic regulatory factors using live-cell single-molecule tracking

STAR Protocols

Xiaojun Ren¹  Samantha Kent¹  Kyle Brown¹ 

[1] University of Colorado Denver Chemistry Department, 1150 12th St., Denver, CO 80204, USA;

关键词: Biophysics;    Single-molecule Assays;    Cell Biology;    Cell-based Assays;    Microscopy;    Gene Expression;   

DOI  :  

来源: DOAJ

【 摘 要 】

Summary: This protocol provides instructions to track the global dynamics of single epigenetic regulatory factors in live cells. We describe an approach to generate cell lines that stably express HaloTag-fused proteins. We then use live-cell single-molecule tracking to obtain kinetic populations and residence times. The kinetic parameters obtained can be used to determine important aspects of transcriptional regulation such as target-search time, 3D free diffusion time, and number of non-specific sites sampled before reaching a specific site and compare behaviors across different nuclear environments.For complete details on the use and execution of this protocol, please refer to Kent et al. (2020).

【 授权许可】

Unknown