dCas9-VP64 Transcriptional Activator

Product Introduction

dCas9-VP64 is an artificial chimeric protein formed by fusing nuclease-dead Cas9 (dCas9) with the VP64 transcriptional activation domain, representing the core component of the first-generation CRISPR activation (CRISPRa) system. The dCas9 moiety of this protein carries the D10A and N863A double mutations, completely abolishing DNA cleavage activity while retaining the ability to precisely bind target DNA sequences guided by sgRNA. The VP64 activation domain consists of four tandem copies of the herpes simplex virus VP16 transcriptional activation domain. VP16 itself is a potent eukaryotic transcriptional activator capable of efficiently recruiting endogenous cellular transcriptional complexes.

When a specific sgRNA guides dCas9-VP64 to bind to the promoter region of a target gene, the VP64 domain recruits transcriptional machinery components such as the pre-initiation complex, histone acetyltransferases, and the Mediator complex. This promotes the assembly of RNA polymerase II and transcription initiation, thereby achieving transcriptional upregulation of the target gene. This system typically achieves basal gene activation levels of 2- to 5-fold, with activation efficiency influenced by factors such as target site selection, chromatin accessibility, and cell type. To overcome the limitations of single sgRNA activation efficiency, researchers often employ multi-sgRNA strategies—designing multiple sgRNAs targeting different regions of the same gene promoter to significantly enhance transcriptional activation through synergistic effects. Some optimized systems can even achieve hundreds-fold gene upregulation.