Topo IIβ CRISPR Activation Plasmid (h): sc-400773-ACT

TOP2B encodes human DNA topoisomerase IIβ (Topo IIβ), an ATP-dependent enzyme that resolves DNA supercoiling and catenation by creating transient double-strand breaks and re-ligating DNA during transcription and chromatin remodeling. Topo IIβ supports genome organization and long-range gene regulation, contributing to RNA polymerase II–dependent transcription programs and maintenance of genomic stability. Its activity intersects with DNA damage response signaling, cell cycle–linked topological control, and differentiation-associated transcriptional switches. Dysregulated TOP2B function or expression has been associated with altered transcriptional landscapes, genome instability phenotypes, and disease-relevant stress responses studied in cancer biology, neurobiology, and cardiotoxicity mechanisms.

Topo IIβ CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TOP2B expression without altering the underlying DNA sequence.

Topo IIβ CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TOP2B 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 TOP2B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Topo IIβ expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TOP2B locus and enabling the study of Topo IIβ-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Topo IIβ pathway restoration in tumor cells with silenced or reduced TOP2B expression.

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