CRSP77 CRISPR Activation Plasmid (h): sc-402693-ACT

MED17 encodes CRSP77, a core subunit of the human Mediator complex that couples sequence-specific transcription factors to RNA polymerase II to coordinate transcription initiation and promoter-specific gene regulation. Through Mediator-dependent integration of signaling inputs, CRSP77 contributes to control of cell-cycle programs, differentiation, and stress-responsive transcriptional outputs. Dysregulation of Mediator components, including MED17, has been associated with altered global transcriptional homeostasis and has been explored in the context of neurodevelopmental and proliferative disease mechanisms. As a result, MED17/CRSP77 is frequently studied to understand how transcriptional co-regulators shape pathway-specific gene expression networks.

CRSP77 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MED17 expression without altering the underlying DNA sequence.

CRSP77 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MED17 locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the MED17 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CRSP77 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MED17 locus and enabling the study of CRSP77-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CRSP77 pathway restoration in tumor cells with silenced or reduced MED17 expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.