KIR4.1 CRISPR Activation Plasmid (h): sc-402244-ACT

KCNJ10 encodes the inwardly rectifying potassium channel KIR4.1, a key determinant of membrane potential and potassium buffering in glial and epithelial contexts. In the central nervous system, KIR4.1 supports extracellular K+ clearance and spatial buffering linked to neuronal excitability, astrocyte homeostasis, and ion/water coupling processes. Channel activity interfaces with electrochemical gradient maintenance and influences downstream signaling through changes in resting potential and cellular excitability. Genetic and functional perturbations of KCNJ10 have been associated with neurodevelopmental and seizure-related phenotypes, supporting its relevance in studies of ion channel dysfunction and glial biology.

KIR4.1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous KCNJ10 expression without altering the underlying DNA sequence.

KIR4.1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the KCNJ10 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 KCNJ10 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous KIR4.1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native KCNJ10 locus and enabling the study of KIR4.1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of KIR4.1 pathway restoration in tumor cells with silenced or reduced KCNJ10 expression.

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