Rab 6 CRISPR Activation Plasmid (h): sc-417258-ACT

RAB6A encodes the small GTPase Rab6, a central regulator of membrane trafficking within the Golgi apparatus and along retrograde transport routes to the endoplasmic reticulum. By cycling between GTP- and GDP-bound states, Rab6 coordinates vesicle tethering, cargo sorting, and microtubule-dependent transport through interactions with effectors involved in Golgi maintenance and secretory pathway dynamics. RAB6A activity influences organelle organization, protein secretion, and endosomal trafficking, processes frequently remodeled in cancer and neurodegeneration. Altered Rab6-dependent transport has also been linked to pathogen exploitation of host trafficking and to cellular stress responses that impact proteostasis.

Rab 6A CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RAB6A expression without altering the underlying DNA sequence.

Rab 6A CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RAB6A 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 RAB6A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Rab 6A expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RAB6A locus and enabling the study of Rab 6A-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Rab 6A pathway restoration in tumor cells with silenced or reduced RAB6A expression.

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