Raftlin CRISPR Activation Plasmid (h): sc-406009-ACT

RFTN1 encodes Raftlin, a lipid raft–associated scaffold protein enriched in immune cells where it helps organize membrane microdomains that coordinate receptor proximal signaling. Raftlin contributes to B cell receptor and other immunoreceptor pathways by regulating localization and stability of signaling complexes within cholesterol-rich rafts, influencing downstream MAPK and NF-κB–linked transcriptional programs. Through these roles, RFTN1 is studied in the context of immune activation, antigen receptor signaling dynamics, and inflammatory network regulation, with reported associations to immune-mediated disease phenotypes and altered immune cell function. Its membrane-proximal positioning makes it a useful node for dissecting how raft organization shapes signal transduction and cellular responses.

Raftlin CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RFTN1 expression without altering the underlying DNA sequence.

Raftlin CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RFTN1 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 RFTN1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Raftlin expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RFTN1 locus and enabling the study of Raftlin-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Raftlin pathway restoration in tumor cells with silenced or reduced RFTN1 expression.

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