CRISPRa and CRISPRi Gene Expression Modulation

CRISPR gene expression activation and gene expression interference

CRISPR/Cas9 has recently adapted to generate two new technologies that modulates gene expression: CRISPRa (CRISPR activation) for gene activation and CRISPRi (CRISPR interference) for gene expression interference. In both CRISPRa and CRISPRi systems, an enzymatically deficient Cas9 (dCas9) is fused or interacts with transcriptional effector(s). dCas9 contains mutations in two active endonuclease domains, losing the capability to cut DNA. However, dCas9 can still target a specific DNA location when coupled with a gRNA. In the CRISPRa system, dCas9 is fused or interacts with transcriptional activators leading to gene expression upregulation. In the CRISPRi system, dCas9 is fused with transcriptional repressor(s) leading to gene expression repression. 

Diagram of CRISPRa and CRISPRi. dCas9 - activator leads to activation; dCas9 - repressor leads to repression.

CRISPRa – CRISPR/Cas9 SAM Synergistic Activation Mediator

CRISPRa SAM is a robust CRISPR gene activation system to activate gene expression using CRISPR technology. The CRISPRa SAM consists of dCas9-VP64, a modified gRNA containing MS2 RNA aptamers, and MS2-p65-HSF1 activation domains. VP64 has four copies of VP16, a viral protein that is used for transcriptional activation. p65 and HSF1 are transcription activation domains. When p65 and HSF1 are brought in close proximity to dCAs9-VP64 via interaction of MS2 with MS2 RNA aptamers in gRNA, the three transactivators can synergistically upregulate gene expression. CRISPRa SAM system can robustly activate both coding and non-coding RNA (lincRNA).

• Robust CRISPR gene activation system
• Synergistic activation by three activation domains, VP64, p65 and HSF1
• Two vector system, dCas9-VP64-gRNA (MS2) and MS2-p65-HSF1
• Complement to gene overexpression with cDNA clones

Diagram of CRISPRa SAM – CRISPR/Cas9 activation system. MS2 in MS2-p65-HSF1 fusion protein binds to MS2 RNA aptamer in the gRNA, bringing p65 and HSF1 to VP64; VP64, p65 and HSF1 then synergistically activate gene expression.

CRISPRa Constructs and Genome-wide Human and Mouse CRISPRa Activation Kits
In addition to the CRISPRa constructs below, genome-wide human and mouse CRISPRa activation kits containing 3 gene-specific gRNAs, 1 scramble control and 1 enhancer vector are available. Please contact us for more information.

CRISPRi – dCas9-KRAB-MeCP2 Gene Interference

In this CRISPRi system, dCas9 is fused with KRAB and MeCP2 repression domains to carry out robust gene repression. Krüppel-associated box (KRAB) is a well-known transcriptional repressor domain. MeCP2 has been shown to bind to methylated DNA.

• All-in-one CRISPRi vector system
• gRNA driven by U6 promoter (after target sequence cloned)
• dCas9-KRAB-MeCP2 fusion protein expression by CMV promoter

CRISPRi Constructs and Genome-wide Human and Mouse CRISPRi Interference Kits
In addition to the CRISPRi constructs below, genome-wide human and mouse CRISPRi interference kits containing 3 gene-specific gRNAs and 1 scramble control vector are available. Please contact us for more information.