Cytokeratin 6A CRISPR Activation Plasmid (h): sc-401650-ACT

KRT6A encodes cytokeratin 6A, a type II intermediate filament protein that heterodimerizes with type I keratins to build the keratin cytoskeleton in stratified epithelia. It supports epithelial mechanical integrity, cell stress responses, and wound-associated keratinocyte activation, integrating with cytoskeletal remodeling programs that influence adhesion, migration, and differentiation. Cytokeratin 6A expression is dynamically regulated during hyperproliferative states and tissue repair, and altered KRT6A activity has been linked to keratinization phenotypes and epithelial barrier dysfunction. These properties make KRT6A a useful node for studying epidermal homeostasis, keratin network assembly, and epithelial stress-adaptation pathways.

Cytokeratin 6A CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous KRT6A expression without altering the underlying DNA sequence.

Cytokeratin 6A CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the KRT6A 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 KRT6A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Cytokeratin 6A expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native KRT6A locus and enabling the study of Cytokeratin 6A-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Cytokeratin 6A pathway restoration in tumor cells with silenced or reduced KRT6A expression.

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