MBD2 CRISPR Activation Plasmid (h): sc-401561-ACT

Human MBD2 (methyl-CpG binding domain protein 2) is a reader of DNA methylation that binds methylated CpG dinucleotides and helps translate epigenetic marks into transcriptional outcomes. Through interactions with chromatin remodeling and histone deacetylase complexes such as NuRD, MBD2 contributes to gene silencing, chromatin compaction, and regulation of lineage-specific programs. It influences processes including cell differentiation, immune signaling, and maintenance of genome-wide epigenetic stability. Dysregulated MBD2 activity and altered methylation-dependent repression have been associated with aberrant transcriptional networks observed in cancer and other disorders involving epigenetic imbalance.

MBD2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MBD2 expression without altering the underlying DNA sequence.

MBD2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MBD2 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 MBD2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MBD2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MBD2 locus and enabling the study of MBD2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MBD2 pathway restoration in tumor cells with silenced or reduced MBD2 expression.

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