VGF CRISPR Activation Plasmid (h): sc-402030-ACT

VGF encodes a neuroendocrine secretory granule protein that is proteolytically processed into bioactive peptides regulating synaptic plasticity, energy balance, and neuroendocrine signaling. In neurons and endocrine cells, VGF-derived peptides influence activity-dependent transcriptional programs and intracellular signaling linked to secretion, metabolism, and stress responses. VGF expression is responsive to neurotrophin and neuronal activity cues and is often studied in the context of circuits controlling feeding behavior and mood-related phenotypes. Dysregulated VGF signaling has been associated with neuropsychiatric and metabolic disease-relevant processes, supporting its use as a mechanistic node in pathway interrogation studies.

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

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

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