GRB2 CRISPR Activation Plasmid (h): sc-400482-ACT

GRB2 (growth factor receptor-bound protein 2) is a ubiquitously expressed adaptor protein that couples activated receptor tyrosine kinases to downstream signaling through its SH2 and SH3 domains. By linking phosphorylated receptors and docking proteins to SOS and other effectors, GRB2 promotes RAS–RAF–MEK–ERK cascade activation and coordinates crosstalk with PI3K/AKT and additional mitogenic pathways. GRB2-dependent signaling influences proliferation, differentiation, survival, cytoskeletal remodeling, and receptor endocytosis across many human cell types. Dysregulated GRB2-centered networks are frequently investigated in oncogenic growth factor signaling, aberrant MAPK pathway activity, and other signaling-driven disease phenotypes relevant to cell state control.

GRB2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous GRB2 expression without altering the underlying DNA sequence.

GRB2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the GRB2 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 GRB2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GRB2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native GRB2 locus and enabling the study of GRB2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GRB2 pathway restoration in tumor cells with silenced or reduced GRB2 expression.

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