CDKN3 CRISPR Activation Plasmid (h): sc-403243-ACT

CDKN3 (cyclin-dependent kinase inhibitor 3) encodes a dual-specificity phosphatase that modulates cell-cycle progression by dephosphorylating cyclin-dependent kinases, with prominent roles in controlling CDK1/CDC2 activity at the G2/M transition. Through regulation of phosphorylation-dependent checkpoints, CDKN3 influences mitotic entry, proliferation rate, and genome stability programs linked to DNA damage responses. Dysregulated CDKN3 expression has been associated with altered cell-cycle control in multiple cancer contexts, making it a useful molecular node for studying proliferative signaling and checkpoint adaptation. In human cells, perturbing CDKN3 levels can help dissect how phosphatase-mediated CDK tuning shapes mitotic fidelity and stress-responsive transcriptional states.

CDKN3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CDKN3 expression without altering the underlying DNA sequence.

CDKN3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CDKN3 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 CDKN3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CDKN3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CDKN3 locus and enabling the study of CDKN3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CDKN3 pathway restoration in tumor cells with silenced or reduced CDKN3 expression.

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