3PGDH CRISPR Activation Plasmid (h): sc-401405-ACT

Human PHGDH encodes 3-phosphoglycerate dehydrogenase (3PGDH), the first and rate-limiting enzyme of the de novo serine biosynthesis pathway that diverts the glycolytic intermediate 3-phosphoglycerate toward L-serine and downstream one-carbon metabolism. Through control of serine and glycine pools, PHGDH influences nucleotide synthesis, methylation reactions, and cellular redox balance via NADH/NADPH-coupled metabolism. PHGDH dysregulation has been linked to metabolic reprogramming in proliferative states and to neurometabolic phenotypes where serine availability constrains protein, lipid, and neurotransmitter-related biosynthetic processes. As a pathway node connecting glycolysis to amino acid and folate-cycle flux, 3PGDH is frequently interrogated in studies of nutrient dependence, mitochondrial–cytosolic metabolic crosstalk, and stress adaptation.

3PGDH CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PHGDH expression without altering the underlying DNA sequence.

3PGDH CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PHGDH 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 PHGDH transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous 3PGDH expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PHGDH locus and enabling the study of 3PGDH-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of 3PGDH pathway restoration in tumor cells with silenced or reduced PHGDH expression.

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