CCDC111 CRISPR Activation Plasmid (h): sc-404310-ACT

PRIMPOL (also known as CCDC111) encodes PrimPol, a primase–polymerase that enables replication restart and DNA damage tolerance when replication forks encounter lesions or secondary DNA structures. PrimPol supports repriming downstream of stalled forks and contributes to post-replicative gap filling, linking its activity to genome maintenance pathways that intersect with ATR-dependent replication stress responses. Altered PRIMPOL function has been associated with increased mutagenesis and sensitivity to genotoxic stress, making it relevant to studies of replication fidelity and cellular responses to endogenous and exogenous DNA damage. As a nuclear enzyme acting at challenged replication forks, PRIMPOL is frequently examined in the context of replication stress, fork stability, and maintenance of chromosomal integrity.

CCDC111 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PRIMPOL expression without altering the underlying DNA sequence.

CCDC111 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PRIMPOL 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 PRIMPOL transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CCDC111 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PRIMPOL locus and enabling the study of CCDC111-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CCDC111 pathway restoration in tumor cells with silenced or reduced PRIMPOL expression.

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