SRP54 CRISPR Activation Plasmid (h): sc-403655-ACT

SRP54 encodes the 54 kDa subunit of the signal recognition particle (SRP), a core ribonucleoprotein complex that recognizes N-terminal signal peptides and couples translating ribosomes to the endoplasmic reticulum membrane. SRP54 functions as a GTPase that coordinates co-translational targeting through interaction with the SRP receptor and subsequent engagement of the SEC61 translocon, supporting secretion and membrane protein biogenesis. By governing ER import and proteostasis, SRP54 impacts pathways linked to ER stress responses and secretory pathway homeostasis. Disruption or dysregulation of SRP54 has been associated with disease-relevant defects in protein targeting and cellular stress phenotypes, including hematologic and immunologic dysfunction reported in human genetic studies.

SRP54 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SRP54 expression without altering the underlying DNA sequence.

SRP54 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SRP54 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 SRP54 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SRP54 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SRP54 locus and enabling the study of SRP54-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SRP54 pathway restoration in tumor cells with silenced or reduced SRP54 expression.

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