Using CRISPR-dCas9-based methods in living and fixed plant cells: Making genomic sequences visible

Abstract

Enzymatically inactive dCas9 can be used with corresponding gRNAs to stain specific sequences in microscopic preparations in situ. For this purpose, dCas9 can be coupled directly with GFP, the sgRNA can be loaded with a fluorophore or bound to GFP via an aptamer in the gRNA. A major advantage of the method is that observations can be made in vivo and thus the dynamics of the labelled sequences in the cell nucleus can also be observed. With the help of CID (chromogenic in situ detection), the method can also be carried out without an expensive fluorescence microscope, e. g. in schools.