XPF CRISPR Activation Plasmid (h): sc-401692-ACT

ERCC4 encodes the human structure-specific endonuclease XPF, which forms a heterodimer with ERCC1 to execute 5′ incision during nucleotide excision repair (NER) and to process DNA interstrand crosslinks via the Fanconi anemia/HR network. XPF recognizes branched DNA structures and contributes to resolution of stalled replication forks, maintaining genome stability under endogenous and genotoxic stress. Disruption of ERCC4/XPF function is linked to DNA repair deficiency syndromes, including xeroderma pigmentosum and related progeroid phenotypes, and is relevant to carcinogenesis studies through impaired lesion removal and elevated mutational burden. As a result, ERCC4 is frequently interrogated in models of UV damage responses, crosslink repair, replication stress, and checkpoint signaling.

XPF CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ERCC4 expression without altering the underlying DNA sequence.

XPF CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ERCC4 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 ERCC4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous XPF expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ERCC4 locus and enabling the study of XPF-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of XPF pathway restoration in tumor cells with silenced or reduced ERCC4 expression.

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