Cdc16 CRISPR Activation Plasmid (h): sc-404163-ACT

CDC16 encodes Cdc16, a core subunit of the anaphase-promoting complex/cyclosome (APC/C) that functions as an E3 ubiquitin ligase controlling ordered proteolysis during mitosis and G1. By supporting APC/C assembly and substrate recognition, Cdc16 contributes to timely degradation of cyclins and other cell-cycle regulators, thereby enforcing spindle checkpoint fidelity and proper chromosome segregation. Perturbation of APC/C regulation is linked to replication stress, aneuploidy, and altered proliferative capacity, making CDC16 relevant to studies of genome stability and cancer-associated cell-cycle dysregulation. CDC16 is also used as a node for dissecting ubiquitin–proteasome pathway control of mitotic exit and checkpoint adaptation.

Cdc16 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CDC16 expression without altering the underlying DNA sequence.

Cdc16 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CDC16 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 CDC16 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Cdc16 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CDC16 locus and enabling the study of Cdc16-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Cdc16 pathway restoration in tumor cells with silenced or reduced CDC16 expression.

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