Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 112, Issue 10, Pages 3002-3007Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1420024112
Keywords
4D nucleome; telomeres; pericentromeric DNA; chromosomes
Categories
Funding
- US National Science Foundation [MCB 0445841]
Ask authors/readers for more resources
The intranuclear location of genomic loci and the dynamics of these loci are important parameters for understanding the spatial and temporal regulation of gene expression. Recently it has proven possible to visualize endogenous genomic loci in live cells by the use of transcription activator-like effectors (TALEs), as well as modified versions of the bacterial immunity clustered regularly interspersed short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system. Here we report the design of multicolor versions of CRISPR using catalytically inactive Cas9 endonuclease (dCas9) from three bacterial orthologs. Each pair of dCas9-fluorescent proteins and cognate single-guide RNAs (sgRNAs) efficiently labeled several target loci in live human cells. Using pairs of differently colored dCas9-sgRNAs, it was possible to determine the intranuclear distance between loci on different chromosomes. In addition, the fluorescence spatial resolution between two loci on the same chromosome could be determined and related to the linear distance between them on the chromosome's physical map, thereby permitting assessment of the DNA compaction of such regions in a live cell.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available