Ankyrin G CRISPR Activation Plasmid (h): sc-400421-ACT

Human ANK3 encodes ankyrin G, a multifunctional adaptor that links integral membrane proteins to the spectrin–actin cytoskeleton, organizing specialized membrane domains and stabilizing protein complexes. Ankyrin G is essential for neuronal polarity and excitability through its role in clustering voltage-gated sodium channels and other components at the axon initial segment and nodes of Ranvier, and it also contributes to epithelial membrane organization. Through these scaffolding functions, ankyrin G influences cytoskeletal remodeling, membrane trafficking, and signal transduction pathways that coordinate cell architecture and communication. Genetic and expression studies have associated ANK3 dysregulation with neuropsychiatric and neurodevelopmental phenotypes, supporting its relevance for mechanistic research in neuronal circuitry and membrane domain biology.

Ankyrin G CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ANK3 expression without altering the underlying DNA sequence.

Ankyrin G CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ANK3 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 ANK3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Ankyrin G expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ANK3 locus and enabling the study of Ankyrin G-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Ankyrin G pathway restoration in tumor cells with silenced or reduced ANK3 expression.

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