MFG-E8 CRISPR Activation Plasmid (h): sc-402282-ACT

Human MFGE8 encodes milk fat globule‑EGF factor 8 (MFG‑E8), a secreted glycoprotein that binds phosphatidylserine on apoptotic cells and bridges them to phagocytes via integrins such as αvβ3 and αvβ5, promoting efferocytosis and resolution of inflammation. Through regulation of apoptotic cell clearance, MFG‑E8 influences innate immune signaling, tissue remodeling, and macrophage polarization, with downstream effects on cytokine tone and extracellular matrix homeostasis. MFGE8 activity intersects with angiogenic and wound-repair programs and is often studied in contexts of chronic inflammation, fibrosis, and tumor microenvironment biology where defective clearance or altered stromal signaling can contribute to disease-associated phenotypes.

MFG-E8 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MFGE8 expression without altering the underlying DNA sequence.

MFG-E8 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MFGE8 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 MFGE8 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MFG-E8 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MFGE8 locus and enabling the study of MFG-E8-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MFG-E8 pathway restoration in tumor cells with silenced or reduced MFGE8 expression.

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