HPRT CRISPR Activation Plasmid (h): sc-417332-ACT

HPRT1 encodes hypoxanthine-guanine phosphoribosyltransferase (HPRT), a key enzyme in the purine salvage pathway that catalyzes the conversion of hypoxanthine and guanine to IMP and GMP using PRPP. By maintaining nucleotide pool homeostasis, HPRT supports DNA/RNA synthesis and influences cellular responses to metabolic stress and replication demand. HPRT1 activity is central to purine metabolism, intersects with PRPP-dependent biosynthetic flux, and is widely leveraged as a selectable marker locus in mammalian cell genetics. Loss-of-function variants in HPRT1 disrupt purine recycling and are associated with inborn errors of metabolism and neurobehavioral phenotypes, providing a tractable model for studying metabolic regulation and genotype–phenotype relationships.

HPRT CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous HPRT1 expression without altering the underlying DNA sequence.

HPRT CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the HPRT1 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 HPRT1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous HPRT expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native HPRT1 locus and enabling the study of HPRT-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of HPRT pathway restoration in tumor cells with silenced or reduced HPRT1 expression.

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