Orai2 CRISPR Activation Plasmid (h): sc-403535-ACT

Human ORAI2 encodes Orai2, a core subunit of the calcium release–activated calcium (CRAC) channel that mediates store-operated calcium entry following ER Ca2+ depletion. By controlling cytosolic Ca2+ influx, Orai2 tunes downstream signaling programs including calcineurin–NFAT transcription, MAPK activity, and other Ca2+-dependent pathways that shape immune activation, secretion, and broader stimulus–response coupling. Altered ORAI2 expression or CRAC channel balance can shift calcium signaling thresholds, contributing to dysregulated inflammatory signaling, aberrant cell proliferation, and context-dependent stress responses. As a result, ORAI2 is frequently studied in models of immune cell function, epithelial signaling, and mechanisms linking Ca2+ homeostasis to disease-relevant phenotypes.

Orai2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ORAI2 expression without altering the underlying DNA sequence.

Orai2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ORAI2 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 ORAI2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Orai2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ORAI2 locus and enabling the study of Orai2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Orai2 pathway restoration in tumor cells with silenced or reduced ORAI2 expression.

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