POL H CRISPR Activation Plasmid (h): sc-401794-ACT

POLH encodes DNA polymerase eta (Pol η), a specialized Y-family polymerase that performs translesion DNA synthesis to bypass UV-induced cyclobutane pyrimidine dimers and other bulky DNA adducts during S phase. By extending stalled replication forks, POLH helps maintain genome stability and coordinates with the DNA damage response, nucleotide excision repair, and post-replication repair pathways. Defects in POLH compromise lesion bypass fidelity and increase mutation burden, linking its dysfunction to UV sensitivity and cancer predisposition phenotypes, including xeroderma pigmentosum variant. POLH is therefore widely studied in replication stress biology, mutagenesis mechanisms, and pathway crosstalk that governs DNA repair choice.

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

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

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