A gene activation system based on a fusion of inactive Cas9 or dCas9 to the catalytic histone acetyltransferase (HAT) core domain of the human E1A-associated protein p300. The dCas9-p300 CRISPR Gene Activator with the p300 domain represents a distinct mechanism of action relative to dCas9-VP64 or other similar gene activation motifs. The p300 histone acetyltransferase protein opens a transcriptional highway by releasing the DNA from its heterochromatin state and allowing for continued and robust gene expression by the endogenous cellular machinery. The Cas9 expression plasmids use the CMV promoter for strong transient expression of Cas9. Alternate promoters can be substituted by replacement of CMV using MluI and NheI. Also, the Cas9 expression plasmids can be linearized using XbaI for T7-based mRNA production.
The Sigma CRISPR dCas9p300 plasmid co-expresses p300-HAT and GFP, to easily monitor delivery and expression in your target cell type (figure 1)gRNAs can successfully direct nuclease-deficient Cas9 (dCas9) fused to p300 HAT catalytic domain to increase levels of histone acetylation and endogenous gene expressionThe dCas9-p300 histone acetylation approach represents a distinct mechanism of action relative to dCas9-VP64 or other similar gene activation motifs