Cytokeratin 3 CRISPR Activation Plasmid (h): sc-402161-ACT

KRT3 encodes cytokeratin 3, a type II intermediate filament protein that partners with keratin 12 to form the core cytoskeletal network of differentiated corneal epithelial cells. Cytokeratin 3 contributes to mechanical stability, cell shape, and resilience to shear stress by integrating into intermediate filament assembly and coordinating with desmosome- and hemidesmosome-associated structures. Its expression reflects epithelial maturation programs and is linked to processes such as barrier maintenance, wound repair, and epithelial remodeling. Dysregulated keratin expression patterns, including altered KRT3, are used to study corneal epithelial differentiation states and ocular surface pathology in experimental models.

Cytokeratin 3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous KRT3 expression without altering the underlying DNA sequence.

Cytokeratin 3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the KRT3 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 KRT3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Cytokeratin 3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native KRT3 locus and enabling the study of Cytokeratin 3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Cytokeratin 3 pathway restoration in tumor cells with silenced or reduced KRT3 expression.

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