TFPI CRISPR Activation Plasmid (m): sc-423361-ACT

Mouse Tfpi encodes tissue factor pathway inhibitor (TFPI), a Kunitz-type serine protease inhibitor that restrains initiation of the extrinsic coagulation cascade by inhibiting the TF–FVIIa complex and factor Xa. TFPI helps maintain vascular hemostatic balance and modulates protease signaling at the endothelial surface, linking coagulation to inflammatory and barrier-regulatory processes. Altered TFPI expression or function is associated with prothrombotic phenotypes and vascular dysfunction, and it is frequently studied in the context of sepsis-associated coagulopathy, atherosclerosis, and endothelial injury models. Tfpi regulation also intersects with lipid-associated pathways via endothelial interactions and circulating lipoprotein binding, making it relevant to cardiometabolic research.

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

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

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