robo1 CRISPR Activation Plasmid (h): sc-402263-ACT

ROBO1 encodes the Roundabout guidance receptor 1 (robo1), a transmembrane receptor for SLIT ligands that regulates axon guidance, neuronal migration, and tissue patterning through SLIT–ROBO signaling. In human cells, ROBO1 influences cytoskeletal dynamics and cell motility via downstream Rho-family GTPase pathways, impacting processes such as directional migration and boundary formation. Dysregulated ROBO1 activity or expression has been associated with aberrant developmental signaling and altered cell invasion programs, making it relevant to studies of neurodevelopmental mechanisms and tumor biology. ROBO1 is also used as a marker and functional node in investigations of guidance cue signaling networks and their cross-talk with growth factor and adhesion pathways.

robo1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ROBO1 expression without altering the underlying DNA sequence.

robo1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ROBO1 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 ROBO1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous robo1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ROBO1 locus and enabling the study of robo1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of robo1 pathway restoration in tumor cells with silenced or reduced ROBO1 expression.

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