Nebulette CRISPR Activation Plasmid (h): sc-406565-ACT

NEBL encodes nebulette, a striated muscle–specific actin-binding protein that localizes to the Z-disc and helps organize thin filament architecture in cardiomyocytes. Through interactions with actin and Z-disc scaffolding components, nebulette supports sarcomere assembly, mechanotransduction, and stabilization of contractile function. Perturbation of NEBL expression or protein integrity is linked to disrupted myofibrillar organization and altered cardiac muscle physiology, making it relevant to studies of cytoskeletal remodeling and cardiomyopathy-associated pathways. As a human gene, NEBL is frequently examined in models of cardiac development, stress responses, and sarcomeric signaling networks.

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

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

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