DPYD CRISPR Activation Plasmid (h): sc-403396-ACT

Human DPYD encodes dihydropyrimidine dehydrogenase, a cytosolic flavoprotein that catalyzes the rate-limiting reduction of uracil and thymine, initiating pyrimidine catabolism. By controlling pyrimidine turnover, DPYD influences nucleotide homeostasis, RNA/DNA precursor balance, and cellular responses to metabolic stress. Altered DPYD activity has been associated with inborn errors of pyrimidine metabolism and variability in fluoropyrimidine handling, linking the pathway to neurodevelopmental and pharmacogenomic research contexts. DPYD is therefore a useful node for studying metabolic regulation, redox biochemistry, and pathway-level effects on proliferation and cellular fitness.

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

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

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