FLASH CRISPR Activation Plasmid (h): sc-403170-ACT

CASP8AP2 encodes FLASH, a large nuclear protein that coordinates apoptosis-associated signaling and RNA processing. FLASH is implicated in death receptor–linked caspase-8 pathway modulation and functions in histone mRNA 3′-end processing through interactions with the U7 snRNP and the histone locus body, linking cell-cycle progression to chromatin and transcriptional programs. By influencing S-phase–coupled histone biogenesis, genome stability, and stress responses, FLASH has been studied in contexts of dysregulated proliferation and altered apoptotic thresholds. Changes in CASP8AP2/FLASH activity or expression have been associated with cancer biology and hematologic disease mechanisms, supporting its use as a target for pathway interrogation in human cell models.

FLASH CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CASP8AP2 expression without altering the underlying DNA sequence.

FLASH CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CASP8AP2 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 CASP8AP2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous FLASH expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CASP8AP2 locus and enabling the study of FLASH-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of FLASH pathway restoration in tumor cells with silenced or reduced CASP8AP2 expression.

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