EphA2 CRISPR Activation Plasmid (h): sc-400535-ACT

EPHA2 encodes EphA2, a receptor tyrosine kinase of the ephrin family that mediates contact-dependent signaling to coordinate cell–cell communication, cytoskeletal remodeling, and tissue boundary formation. Upon ephrin-A ligand engagement, EphA2 influences downstream cascades including Rho family GTPases, MAPK/ERK, PI3K/AKT, and focal adhesion dynamics, shaping adhesion, migration, and proliferation programs. In epithelial and endothelial contexts, EphA2 contributes to morphogenesis and barrier organization, and its dysregulation is frequently studied in relation to oncogenic signaling, invasion-associated phenotypes, and altered angiogenic responses. As a surface receptor integrating microenvironmental cues, EPHA2 serves as a tractable node for dissecting receptor-driven signaling networks and membrane-proximal pathway cross-talk.

EphA2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous EPHA2 expression without altering the underlying DNA sequence.

EphA2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the EPHA2 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 EPHA2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous EphA2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native EPHA2 locus and enabling the study of EphA2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of EphA2 pathway restoration in tumor cells with silenced or reduced EPHA2 expression.

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