cGAS CRISPR Activation Plasmid (h): sc-403354-ACT

Human MB21D1 encodes cyclic GMP-AMP synthase (cGAS), a cytosolic DNA sensor that catalyzes synthesis of 2′3′-cGAMP to trigger STING (TMEM173) signaling and downstream TBK1–IRF3 and NF-κB activation. This pathway coordinates type I interferon and proinflammatory transcriptional programs in response to pathogen-derived or mislocalized self DNA, linking innate immunity with DNA damage responses and mitochondrial stress. Dysregulated cGAS–STING activity has been associated with autoinflammatory phenotypes, aberrant inflammatory signaling in cancer and senescence, and altered responses to DNA viruses, making MB21D1 a key node for studying immune surveillance and sterile inflammation. cGAS also influences chromatin-associated processes and micronuclei sensing, supporting mechanistic studies of genome instability–driven immune activation.

cGAS CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MB21D1 expression without altering the underlying DNA sequence.

cGAS CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MB21D1 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 MB21D1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous cGAS expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MB21D1 locus and enabling the study of cGAS-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of cGAS pathway restoration in tumor cells with silenced or reduced MB21D1 expression.

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