PHF5A CRISPR Activation Plasmid (h): sc-407986-ACT

PHF5A (PHD finger protein 5A) is a conserved component of the SF3b subcomplex of U2 snRNP and supports spliceosome assembly and pre-mRNA splicing fidelity. By stabilizing recognition of the branch point and 3′ splice site, PHF5A contributes to transcript isoform selection that impacts cell-cycle progression, DNA damage responses, and differentiation programs. Perturbation of PHF5A-dependent splicing can reshape gene expression networks and has been linked to vulnerabilities in contexts of dysregulated RNA processing, including oncogenic states where spliceosome function is stressed. As a core splicing factor, PHF5A is frequently used to interrogate RNA maturation pathways and splicing-driven mechanisms underlying phenotypic change.

PHF5A CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PHF5A expression without altering the underlying DNA sequence.

PHF5A CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PHF5A 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 PHF5A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PHF5A expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PHF5A locus and enabling the study of PHF5A-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PHF5A pathway restoration in tumor cells with silenced or reduced PHF5A expression.

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