GALE CRISPR Activation Plasmid (h): sc-408127-ACT

Human GALE (UDP-galactose-4-epimerase) catalyzes the reversible interconversion of UDP-galactose and UDP-glucose, as well as UDP-N-acetylgalactosamine and UDP-N-acetylglucosamine, linking the Leloir pathway to broader nucleotide-sugar metabolism. Through these activities, GALE helps maintain carbohydrate homeostasis and supplies activated sugars required for glycosylation processes that shape protein folding, trafficking, and cell–cell signaling. Disruption of GALE-dependent flux can alter glycan composition and impact ER and Golgi-associated biosynthetic programs. GALE dysfunction is associated with epimerase-deficiency galactosemia and is relevant to studies of metabolic stress responses and glycosylation-linked phenotypes.

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

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

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