Pitx1 CRISPR Activation Plasmid (h): sc-402911-ACT

PITX1 encodes the human paired-like homeodomain transcription factor Pitx1, a nuclear regulator that binds DNA to coordinate gene programs controlling cell fate specification, differentiation, and tissue patterning. Pitx1 participates in transcriptional networks that shape developmental processes and maintain lineage identity through modulation of promoter and enhancer activity. Altered PITX1 expression or regulatory disruption has been linked to dysregulated growth control and changes in differentiation state in multiple disease-relevant contexts, making it a useful node for studying transcriptional rewiring. Because Pitx1 influences broad gene expression outputs, it is frequently examined in pathway studies connecting developmental transcription factors to proliferation, migration, and cellular identity programs.

Pitx1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PITX1 expression without altering the underlying DNA sequence.

Pitx1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PITX1 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 PITX1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Pitx1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PITX1 locus and enabling the study of Pitx1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Pitx1 pathway restoration in tumor cells with silenced or reduced PITX1 expression.

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