Nkx-6.1 CRISPR Activation Plasmid (h): sc-401997-ACT

NKX6-1 encodes the homeobox transcription factor Nkx-6.1, a key regulator of pancreatic endoderm patterning and β-cell lineage specification. Nkx-6.1 coordinates transcriptional programs that support endocrine differentiation, insulin secretory capacity, and maintenance of β-cell identity through interactions with core developmental networks and enhancer-driven gene regulation. Altered NKX6-1 expression or function is associated with impaired β-cell maturation and dysregulated glucose homeostasis, making it relevant to mechanistic studies of diabetes susceptibility and β-cell failure. As a lineage-defining factor, NKX6-1 is also used as a marker and functional node in stem cell–derived pancreatic differentiation and islet cell state transitions.

Nkx-6.1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous NKX6-1 expression without altering the underlying DNA sequence.

Nkx-6.1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the NKX6-1 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 NKX6-1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Nkx-6.1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native NKX6-1 locus and enabling the study of Nkx-6.1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Nkx-6.1 pathway restoration in tumor cells with silenced or reduced NKX6-1 expression.

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