RFX1 CRISPR Activation Plasmid (h): sc-403144-ACT

RFX1 (regulatory factor X1) is a DNA-binding transcription factor that recognizes X-box motifs and modulates promoter activity across diverse gene networks. In human cells, RFX1 contributes to regulation of transcriptional programs linked to cell-cycle control, chromatin state, and immune-related gene expression, with downstream effects on differentiation and cellular stress responses. Altered RFX1 activity has been investigated in the context of dysregulated transcriptional control observed in cancer biology and immune-mediated processes, where shifts in expression can rewire pathway output. Because it sits upstream of multiple regulatory cascades, RFX1 is frequently used as a node for studying promoter architecture, transcription factor occupancy, and gene network plasticity.

RFX1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RFX1 expression without altering the underlying DNA sequence.

RFX1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RFX1 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 RFX1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RFX1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RFX1 locus and enabling the study of RFX1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RFX1 pathway restoration in tumor cells with silenced or reduced RFX1 expression.

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