MBD3 CRISPR Activation Plasmid (h): sc-401072-ACT

Human MBD3 encodes methyl-CpG-binding domain protein 3, a core component of the NuRD (nucleosome remodeling and deacetylase) complex that integrates ATP-dependent chromatin remodeling with histone deacetylation to regulate transcriptional programs. MBD3 contributes to epigenetic control of cell identity, including differentiation and pluripotency transitions, by coordinating gene repression/poising at promoters and enhancers and influencing chromatin accessibility. Through its role in chromatin organization, MBD3 intersects with pathways governing DNA damage responses, replication timing, and developmental gene networks. Dysregulated NuRD/MBD3 activity has been linked to aberrant transcriptional states in cancer and neurodevelopmental phenotypes, making it relevant for studies of epigenetic mechanisms underlying disease-associated gene expression changes.

MBD3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MBD3 expression without altering the underlying DNA sequence.

MBD3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MBD3 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 MBD3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MBD3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MBD3 locus and enabling the study of MBD3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MBD3 pathway restoration in tumor cells with silenced or reduced MBD3 expression.

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