p47phox CRISPR Activation Plasmid (h): sc-417916-ACT

NCF1 encodes p47phox, a cytosolic organizer subunit of the phagocyte NADPH oxidase (NOX2) complex that is essential for stimulus-dependent assembly and activation of superoxide production. Upon phosphorylation, p47phox translocates to membranes and coordinates interactions among p67phox, p40phox, and membrane-bound cytochrome b558 (gp91phox/NOX2 and p22phox), enabling reactive oxygen species generation during innate immune responses. This oxidative burst supports antimicrobial defense, redox signaling, and inflammatory pathway modulation, linking NCF1 activity to neutrophil and macrophage function. Altered NCF1/p47phox function is associated with immunodeficiency phenotypes such as chronic granulomatous disease and has been studied in the context of dysregulated inflammation and autoimmunity.

p47phox CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous NCF1 expression without altering the underlying DNA sequence.

p47phox CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the NCF1 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 NCF1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous p47phox expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native NCF1 locus and enabling the study of p47phox-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of p47phox pathway restoration in tumor cells with silenced or reduced NCF1 expression.

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