EGFL8 CRISPR Activation Plasmid (m): sc-429875-ACT

Egfl8 encodes EGFL8, a secreted extracellular matrix–associated protein containing EGF-like domains that supports cell–cell and cell–matrix communication. In mouse tissues, EGFL8 expression has been linked to regulation of developmental patterning, epithelial and endothelial behavior, and extracellular remodeling processes that influence adhesion, migration, and tissue organization. Through these roles, EGFL8 is relevant to signaling contexts that coordinate morphogenesis and stromal–parenchymal interactions, including pathways that interface with growth factor–responsive programs. Altered EGFL8 activity or expression is therefore studied in models of dysregulated tissue remodeling and proliferative disease biology, where changes in the microenvironment can shape cell fate and invasion-associated phenotypes.

EGFL8 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Egfl8 expression without altering the underlying DNA sequence.

EGFL8 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Egfl8 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 Egfl8 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous EGFL8 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Egfl8 locus and enabling the study of EGFL8-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of EGFL8 pathway restoration in tumor cells with silenced or reduced Egfl8 expression.

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