PHOSPHO1 CRISPR Activation Plasmid (m): sc-433594-ACT

Phospho1 encodes PHOSPHO1, a phosphatase enriched in mineralizing tissues that hydrolyzes phosphoethanolamine and phosphocholine to generate inorganic phosphate, supporting matrix vesicle–mediated initiation of hydroxyapatite deposition. In mouse, PHOSPHO1 activity integrates with phosphate homeostasis and skeletal development programs, acting alongside mineralization regulators such as alkaline phosphatase to shape osteoblast and chondrocyte function. Disruption or dysregulation of PHOSPHO1 is linked to impaired bone mineralization phenotypes and altered extracellular matrix maturation, making it relevant to studies of skeletal fragility and developmental bone disorders. As a node in mineralization biology, it is frequently examined in pathways governing osteogenesis, cartilage-to-bone transition, and local phosphate handling.

PHOSPHO1 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Phospho1 expression without altering the underlying DNA sequence.

PHOSPHO1 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Phospho1 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 Phospho1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PHOSPHO1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Phospho1 locus and enabling the study of PHOSPHO1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PHOSPHO1 pathway restoration in tumor cells with silenced or reduced Phospho1 expression.

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