Nanog CRISPR Activation Plasmid (h): sc-418033-ACT

Human NANOG encodes the homeobox transcription factor Nanog, a central regulator of pluripotency that maintains embryonic stem cell self-renewal and suppresses lineage commitment. Nanog operates within the core pluripotency network with OCT4 and SOX2, coordinating transcriptional programs and chromatin states that influence differentiation, epigenetic remodeling, and cell fate decisions. Dysregulated NANOG expression is linked to stem-like phenotypes, altered developmental signaling, and transcriptional reprogramming observed across diverse disease models, including cancer biology contexts where tumor cell plasticity and resistance-associated states are studied. As a nuclear DNA-binding factor, Nanog provides a tractable node for interrogating transcriptional circuitry underlying development and cell identity.

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

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

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