ZEB CRISPR Activation Plasmid (h): sc-400201-ACT

Human ZEB1 encodes the zinc finger E-box binding homeobox protein ZEB, a sequence-specific transcription factor that binds E-box-like motifs to coordinate gene regulatory programs controlling epithelial–mesenchymal transition (EMT), cell polarity, and differentiation. ZEB1 integrates signals from TGF-β/SMAD, Wnt/β-catenin, Notch, and MAPK pathways and interfaces with microRNA networks such as the miR-200 family to modulate transcriptional repression and activation. Altered ZEB1 activity is frequently linked to changes in cell adhesion, motility, and lineage plasticity, making it relevant to studies of fibrosis, developmental disorders, and tumor biology. As a central regulator of EMT-associated transcriptional states, ZEB1 is widely used as a molecular node to interrogate context-dependent gene expression programs.

ZEB1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ZEB1 expression without altering the underlying DNA sequence.

ZEB1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ZEB1 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 ZEB1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous ZEB1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ZEB1 locus and enabling the study of ZEB1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of ZEB1 pathway restoration in tumor cells with silenced or reduced ZEB1 expression.

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