BAF CRISPR Activation Plasmid (h): sc-401654-ACT

BANF1 encodes barrier-to-autointegration factor (BAF), a conserved DNA-binding protein that bridges double-stranded DNA and LEM-domain nuclear envelope proteins to coordinate nuclear assembly and chromatin organization. BAF participates in post-mitotic nuclear envelope reformation, higher-order genome architecture, and regulation of transcriptional programs through interactions with lamins and chromatin-associated factors. Through these roles, BANF1 integrates pathways governing mitotic exit, genome stability, and DNA damage responses. Dysregulation of BANF1/BAF has been linked to laminopathy-related phenotypes and defects in nuclear morphology, making it a useful node for studying nuclear envelope biology and epigenome control in human cells.

BAF CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous BANF1 expression without altering the underlying DNA sequence.

BAF CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the BANF1 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 BANF1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous BAF expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native BANF1 locus and enabling the study of BAF-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of BAF pathway restoration in tumor cells with silenced or reduced BANF1 expression.

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