PGD CRISPR Activation Plasmid (h): sc-404842-ACT

Human PGD encodes 6-phosphogluconate dehydrogenase, a cytosolic enzyme of the oxidative pentose phosphate pathway that converts 6-phosphogluconate to ribulose-5-phosphate while generating NADPH. By supporting NADPH-dependent redox homeostasis and providing ribose-5-phosphate for nucleotide biosynthesis, PGD links central carbon metabolism to antioxidant defense and proliferative capacity. PGD activity influences cellular responses to oxidative stress and can shape metabolic rewiring associated with oncogenic growth and survival. Dysregulated pentose phosphate pathway flux involving PGD has been investigated across cancer metabolism, highlighting its relevance for studying metabolic dependencies and redox-buffering mechanisms.

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

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

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