FIGURE 1.

Schematic of the CRISPRi approach. Left, Cartoon representation of the CRISPR-mediated regulation of gene expression. The wild-type Cas9 protein binds to the sgRNA and forms a protein–RNA complex. Once Cas9–sgRNA complex binds to specific DNA target adjacent to PAM, it leads to the cleavage of the target DNA due to the nuclease activity of the Cas9 protein. Right, Cartoon depicting the CRISPRi-mediated interference of gene expression caused by nuclease-deficient dCas9. The nuclease-deficient dCas9 contains two substitutions in the nuclease domains (D10A and H840A, blue dots), and thus lose the endonuclease activity. If the target DNA sequence locates inside an open reading frame, the dCas9–sgRNA–DNA complex will block the movement of RNAP and subsequent transcription elongation, resulting in transcription inhibition of the target gene.