SPAG4 CRISPR Activation Plasmid (h): sc-404154-ACT

SPAG4 (sperm-associated antigen 4), also known as SUN4, is a testis-enriched nuclear envelope protein implicated in LINC complex–mediated coupling between the nucleus and cytoskeleton. In germ cells, SPAG4 supports spermatid nuclear shaping and cytoskeletal organization during spermiogenesis, reflecting roles in nuclear positioning and mechanotransduction at the nuclear periphery. Disruption or altered expression of SPAG4 has been linked to male fertility phenotypes and is also reported as aberrantly expressed in multiple tumor contexts, where it is investigated as a cancer-testis antigen. These properties make SPAG4 a useful model for studying nuclear envelope dynamics, germline differentiation programs, and dysregulated antigen expression in disease-relevant cell states.

SPAG4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SPAG4 expression without altering the underlying DNA sequence.

SPAG4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SPAG4 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 SPAG4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SPAG4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SPAG4 locus and enabling the study of SPAG4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SPAG4 pathway restoration in tumor cells with silenced or reduced SPAG4 expression.

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