EPHX2 CRISPR Activation Plasmid (h): sc-402095-ACT

EPHX2 encodes soluble epoxide hydrolase (sEH), a bifunctional enzyme that converts bioactive epoxy-fatty acids such as epoxyeicosatrienoic acids (EETs) into less active diols, shaping lipid mediator tone in vascular, renal, and immune contexts. Through this role in arachidonic acid and broader oxylipin metabolism, EPHX2 influences signaling networks linked to inflammation resolution, endothelial function, oxidative stress responses, and blood pressure regulation. Altered sEH activity and EPHX2 expression patterns have been associated with cardiometabolic traits, hypertension-related pathways, and inflammatory disease mechanisms, motivating mechanistic studies of lipid-epoxide turnover. In cellular models, EPHX2 modulation is used to probe cross-talk between lipid signaling, cytokine programs, and stress-activated transcriptional responses.

sEH CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous EPHX2 expression without altering the underlying DNA sequence.

sEH CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the EPHX2 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 EPHX2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous sEH expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native EPHX2 locus and enabling the study of sEH-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of sEH pathway restoration in tumor cells with silenced or reduced EPHX2 expression.

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