CDKN2A/p16 CRISPR Activation Plasmid (m): sc-419610-ACT

Mouse Cdkn2a encodes the CDKN2A/p16 tumor suppressor, a cyclin-dependent kinase inhibitor that restrains CDK4/6 activity to preserve RB1 in its hypophosphorylated state and enforce the G1/S checkpoint. By limiting E2F-driven transcription, p16 integrates proliferative cues with cell-cycle arrest programs and is frequently engaged during replicative senescence and oncogene-induced stress responses. Dysregulation of the Cdkn2a locus is closely linked to aberrant proliferation and altered senescence states, making it a central node in studies of cell-cycle control, aging-associated phenotypes, and cancer biology in mouse models.

CDKN2A/p16 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Cdkn2a expression without altering the underlying DNA sequence.

CDKN2A/p16 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Cdkn2a 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 Cdkn2a transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CDKN2A/p16 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Cdkn2a locus and enabling the study of CDKN2A/p16-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CDKN2A/p16 pathway restoration in tumor cells with silenced or reduced Cdkn2a expression.

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