PYGB CRISPR Activation Plasmid (h): sc-406294-ACT

Human PYGB encodes the brain isoform of glycogen phosphorylase, a rate-limiting enzyme that catalyzes glycogen breakdown to generate glucose-1-phosphate and sustain cellular energy supply. By linking glycogen mobilization to glycolysis and broader carbohydrate metabolism, PYGB supports metabolic flexibility during fluctuating nutrient or oxygen availability. PYGB activity intersects with signaling networks that tune energy homeostasis, including AMP/ATP sensing and stress-responsive pathways that influence cell survival and proliferation. Dysregulated glycogen metabolism and altered PYGB expression have been observed in contexts of metabolic adaptation and cancer-associated rewiring of energy utilization, making it a useful target for mechanistic studies of bioenergetics.

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

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

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