RBM35B CRISPR Activation Plasmid (h): sc-408008-ACT

Human ESRP2 (RBM35B) encodes an epithelial splicing regulatory protein that binds pre-mRNA and directs alternative splicing programs that help maintain epithelial identity and cell–cell junctional architecture. Through regulation of splice isoforms in pathways governing cytoskeletal dynamics, adherens junctions, and epithelial–mesenchymal transition (EMT), ESRP2 contributes to context-dependent control of differentiation, migration, and signaling output. Altered ESRP2 expression or splicing activity has been associated with aberrant EMT-related transcriptomes and dysregulated RNA processing observed across multiple tumor and developmental models. As a node linking RNA-binding specificity to isoform-resolved gene regulation, ESRP2 is widely used to study splicing-driven phenotypic switching and pathway rewiring.

RBM35B CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ESRP2 expression without altering the underlying DNA sequence.

RBM35B CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ESRP2 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 ESRP2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RBM35B expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ESRP2 locus and enabling the study of RBM35B-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RBM35B pathway restoration in tumor cells with silenced or reduced ESRP2 expression.

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