COPA CRISPR Activation Plasmid (h): sc-404020-ACT

COPA encodes the alpha subunit of the coatomer complex I (COPI), a core mediator of vesicular trafficking between the Golgi apparatus and endoplasmic reticulum and within Golgi stacks. By supporting retrograde transport and cargo sorting, COPA contributes to maintenance of organelle homeostasis, protein processing, and secretory pathway integrity. COPA-dependent trafficking interfaces with cellular stress responses, including ER stress and unfolded protein response signaling, and influences immune and inflammatory signaling through effects on protein localization. Genetic perturbation of COPA has been associated with immune dysregulation and inflammatory disease phenotypes, making it a relevant target for mechanistic studies of intracellular transport and immunobiology.

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

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

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