HURP CRISPR Activation Plasmid (h): sc-402206-ACT

DLGAP5 encodes HURP, a microtubule-associated protein that accumulates on kinetochore fibers and stabilizes spindle microtubules to support chromosome congression and accurate segregation during mitosis. HURP is regulated by cell cycle kinases and the Ran-GTP/importin pathway, integrating cues that coordinate spindle assembly with progression through G2/M. Through its roles in spindle dynamics and mitotic checkpoint integrity, dysregulated HURP expression is linked to proliferative phenotypes and genomic instability observed across multiple cancer contexts. In human cell models, DLGAP5/HURP is frequently studied as a readout of mitotic control, aneuploidy tolerance, and microtubule-targeting stress responses.

HURP CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous DLGAP5 expression without altering the underlying DNA sequence.

HURP CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the DLGAP5 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 DLGAP5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous HURP expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native DLGAP5 locus and enabling the study of HURP-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of HURP pathway restoration in tumor cells with silenced or reduced DLGAP5 expression.

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