Barttin CRISPR Activation Plasmid (h): sc-404023-ACT

Human BSND encodes barttin, an essential accessory subunit for the CLCNKA and CLCNKB chloride channels that supports their trafficking, stability, and chloride conductance at the plasma membrane. Barttin-dependent Cl⁻ transport is critical for epithelial ion homeostasis in the kidney and inner ear, linking BSND to transepithelial salt handling and endolymph ionic balance. Through regulation of chloride channel function, barttin contributes to electrochemical gradients that influence coupled Na⁺/K⁺ transport processes and fluid balance. Dysregulation of BSND is associated with inherited salt-wasting and sensorineural hearing phenotypes, making it a relevant target for studying epithelial transport mechanisms and genotype–phenotype relationships.

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

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

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