Spo11 CRISPR Activation Plasmid (h): sc-404432-ACT

Human SPO11 encodes Spo11, a meiosis-specific topoisomerase-like endonuclease that catalyzes programmed DNA double-strand breaks to initiate homologous recombination. By creating breaks at meiotic recombination hotspots, Spo11 promotes synapsis, crossover formation, and accurate chromosome segregation during gametogenesis, linking it to pathways governing DNA repair, recombination, and genome stability. Disruption or dysregulation of SPO11 activity can perturb meiotic progression and recombination outcomes, making it relevant to studies of infertility, aneuploidy, and mechanisms that safeguard chromosomal integrity. SPO11 is also used as a model factor for dissecting controlled DNA break formation and repair pathway choice in specialized cell states.

Spo11 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SPO11 expression without altering the underlying DNA sequence.

Spo11 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SPO11 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 SPO11 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Spo11 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SPO11 locus and enabling the study of Spo11-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Spo11 pathway restoration in tumor cells with silenced or reduced SPO11 expression.

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