ZFR CRISPR Activation Plasmid (h): sc-407242-ACT

Human ZFR (zinc finger RNA-binding protein) is a predominantly nuclear RNA-binding factor implicated in post-transcriptional gene regulation, including modulation of pre-mRNA splicing and RNA processing through interactions with spliceosomal and ribonucleoprotein complexes. By shaping transcript isoform usage and RNA fate, ZFR can influence cell-state programs such as proliferation, differentiation, and stress-responsive gene expression. Dysregulated RNA processing is a common feature of cancer and neurodevelopmental and neurodegenerative disorders, making ZFR a relevant target for mechanistic studies of RNA regulatory networks. Investigating ZFR function can help clarify how splicing-linked pathways and RNA binding proteins coordinate gene expression outputs in human cells.

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

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

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