CRSP70 CRISPR Activation Plasmid (h): sc-402381-ACT

MED26 encodes CRSP70, a core subunit of the Mediator complex that links sequence-specific transcription factors to RNA polymerase II and helps coordinate transcription initiation with elongation. By engaging the CDK8 module and the super elongation complex, MED26 contributes to promoter-proximal pause release and dynamic regulation of gene expression programs involved in cell identity and stress responses. Altered Mediator function, including dysregulation of MED26-associated transcriptional networks, is relevant to studies of developmental disorders and cancer-associated transcriptional rewiring. As a nuclear regulator of Pol II transcription, CRSP70 is frequently examined in pathway-level analyses of enhancer–promoter communication, chromatin-dependent gene control, and context-specific signaling-to-transcription coupling.

CRSP70 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MED26 expression without altering the underlying DNA sequence.

CRSP70 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MED26 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 MED26 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CRSP70 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MED26 locus and enabling the study of CRSP70-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CRSP70 pathway restoration in tumor cells with silenced or reduced MED26 expression.

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