GABA_A Rε CRISPR Activation Plasmid (h): sc-405545-ACT

GABRE encodes the ε subunit of the human GABA\_A receptor, a ligand-gated chloride channel that mediates fast inhibitory neurotransmission in the central nervous system. Incorporation of the ε subunit can alter receptor gating, pharmacology, and desensitization kinetics, shaping neuronal excitability and synaptic inhibition. GABA\_A receptor signaling is a core component of neurotransmitter-gated ion channel pathways that regulate membrane potential, network oscillations, and activity-dependent plasticity. Dysregulated GABAergic tone and altered receptor subunit composition have been associated with neuropsychiatric and seizure-related phenotypes, motivating mechanistic studies of subunit-specific function.

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

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

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