Insulin CRISPR Activation Plasmid (h): sc-400149-ACT

Human INS encodes insulin, a peptide hormone synthesized as preproinsulin in pancreatic β cells and processed through ER–Golgi secretory trafficking into mature insulin and C-peptide. Insulin receptor engagement activates PI3K–AKT and MAPK signaling, coordinating glucose uptake, glycogen and lipid metabolism, and broader anabolic programs across liver, muscle, and adipose tissue. Dysregulation of INS expression, proinsulin processing, or insulin signaling contributes to impaired glucose homeostasis and is central to diabetes-related pathophysiology and β-cell stress responses. INS is also widely used as a marker of endocrine identity and stimulus-secretion coupling in models of pancreatic development, metabolic inflammation, and cellular differentiation.

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

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

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