M33 CRISPR Activation Plasmid (h): sc-402604-ACT

CBX2 encodes the Polycomb group protein M33, a chromobox family component of canonical Polycomb repressive complex 1 (PRC1) that recognizes H3K27me3 and helps maintain transcriptional silencing through chromatin compaction and ubiquitination of histone H2A. By enforcing stable epigenetic repression programs, CBX2 contributes to cell fate specification, developmental gene control, and regulation of proliferation-associated transcriptional networks. Dysregulated CBX2 expression or PRC1 activity is linked to altered differentiation states and oncogenic transcriptional programs in multiple tumor contexts, making it a useful node for studying epigenetic mechanisms of disease-relevant gene regulation. CBX2/M33 also intersects with broader chromatin regulatory pathways coordinated by PRC2-mediated H3K27 methylation and other histone modification readers and writers.

M33 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CBX2 expression without altering the underlying DNA sequence.

M33 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CBX2 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 CBX2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous M33 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CBX2 locus and enabling the study of M33-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of M33 pathway restoration in tumor cells with silenced or reduced CBX2 expression.

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