Several research groups have harnessed the specificity and easy re-programmability of the CRISPR/Cas9 system to create targetable CRISPR/Cas9 ribonucleoprotein complexes that can either activate (CRISPRa) or interfere (CRISPRi) with transcription of any desired coding region within a genome.

These systems fuse dCas9 to a well-characterized transcription-regulatory domain, using pre-designed guide RNAs to direct the complex upstream of the transcription start site. By using inactivated dCas9 protein, the complex can be targeted to specific loci without cleaving or altering the genomic DNA. After Cas9 binds the targeted DNA sequence, the fused transcription-regulatory domains are then able to recruit repressive or activating effectors to modify gene expression.

The laboratory of Feng Zhang at the Broad Institute has pioneered use of the CRISPR/Cas9 Synergistic Activation Mediator (SAM) system in gene activation assays. The SAM system enables robust transcriptional activation of endogenous genes targeted by guide RNAs that bind within 200 bp upstream of the transcription start site.

Studies using the SAM system to activate gene expression show an increase in transcription of up to 3,000. The SAM system has multiplexing gene activation ability, and has been shown to activate the transcription of as many as 10 genes concurrently. In addition, SAM has been demonstrated to activate non-coding elements, such as long intergenic non-coding RNAs.

SAM can also be used for discovery research to identify the genes that drive phenotypes of interest in disease models or developmental/differentiation process by using a genome-wide SAM gRNA library for gain-of-function screening. The screening process is similar to the GeCKO loss-of-function library screening techniques, but the library is designed to activate transcription rather than edit the genome. The human genome-wide SAM library contains 3 distinct guide RNAs each targeting one of 23,430 coding gene isoforms with a unique transcription start site in the human reference genome, for a total of 70,290 guides.

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