Bcl-2 CRISPR Activation Plasmid (h): sc-400025-ACT

BCL2 encodes Bcl-2, an integral mitochondrial outer membrane protein that suppresses apoptosis by restraining mitochondrial outer membrane permeabilization and limiting cytochrome c release. By sequestering pro-apoptotic BH3-only proteins and antagonizing BAX/BAK activation, Bcl-2 functions as a key node in the intrinsic apoptosis pathway and shapes cellular responses to stress, growth factor withdrawal, and DNA damage. BCL2 activity intersects with survival signaling networks including PI3K–AKT and MAPK pathways and influences mitochondrial dynamics and redox homeostasis. Dysregulated BCL2 expression is frequently associated with impaired apoptotic priming in cancer and contributes to aberrant survival of malignant and immune cell populations, making it broadly relevant to studies of oncogenesis, treatment response, and immune homeostasis.

Bcl-2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous BCL2 expression without altering the underlying DNA sequence.

Bcl-2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the BCL2 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 BCL2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Bcl-2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native BCL2 locus and enabling the study of Bcl-2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Bcl-2 pathway restoration in tumor cells with silenced or reduced BCL2 expression.

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