Nkx-2.3 CRISPR Activation Plasmid (h): sc-406709-ACT

NKX2-3 encodes the homeobox transcription factor Nkx-2.3, a sequence-specific DNA-binding regulator that helps establish and maintain regional identity programs during development and in adult tissues. It influences transcriptional networks controlling epithelial and mesenchymal differentiation, tissue patterning, and cell fate decisions, with downstream effects on lineage-restricted gene expression and microenvironmental signaling. Altered NKX2-3 activity has been associated with dysregulated mucosal immune homeostasis and intestinal biology, supporting its relevance in studies of inflammation-linked gene regulation and disease-associated transcriptional circuits. As a nuclear factor operating at promoter and enhancer elements, Nkx-2.3 serves as a useful node for dissecting context-dependent transcriptional control mechanisms.

Nkx-2.3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous NKX2-3 expression without altering the underlying DNA sequence.

Nkx-2.3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the NKX2-3 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 NKX2-3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Nkx-2.3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native NKX2-3 locus and enabling the study of Nkx-2.3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Nkx-2.3 pathway restoration in tumor cells with silenced or reduced NKX2-3 expression.

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