Recoverin CRISPR Activation Plasmid (h): sc-404106-ACT

Human RCVRN encodes recoverin, a retina-enriched EF-hand Ca2+-binding protein that functions as a Ca2+-myristoyl switch to modulate phototransduction. Upon changes in intracellular Ca2+ during light adaptation, recoverin regulates the activity of rhodopsin kinase (GRK1), shaping rhodopsin phosphorylation dynamics and recovery kinetics in rod and cone photoreceptors. Through its role in GPCR signaling, Ca2+ homeostasis, and cGMP-dependent visual signaling, recoverin supports normal retinal signal transduction and adaptation. Altered recoverin expression or immunoreactivity has been associated with retinal dysfunction contexts, making RCVRN a relevant target for mechanistic studies of photoreceptor biology and visual signaling networks.

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

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

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