Cdc6 CRISPR Activation Plasmid (h): sc-400796-ACT

CDC6 encodes the human Cdc6 ATPase, an essential licensing factor for DNA replication that cooperates with ORC and CDT1 to load the MCM2–7 helicase at replication origins during G1 phase. By coordinating origin licensing, Cdc6 helps enforce once-per-cell-cycle replication and interfaces with S-phase entry control, replication stress signaling, and checkpoint pathways including ATR/CHK1. Dysregulated CDC6 expression can drive aberrant origin firing, genomic instability, and altered cell-cycle dynamics, features commonly observed in tumor biology and other proliferation-linked pathologies. As a core component of replication initiation machinery, CDC6 is frequently studied in the context of DNA damage responses, oncogene-induced replication stress, and mechanisms that couple replication licensing to chromatin state.

Cdc6 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CDC6 expression without altering the underlying DNA sequence.

Cdc6 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CDC6 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 CDC6 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Cdc6 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CDC6 locus and enabling the study of Cdc6-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Cdc6 pathway restoration in tumor cells with silenced or reduced CDC6 expression.

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