RecQL4 CRISPR Activation Plasmid (h): sc-403224-ACT

RECQL4 encodes the human RecQ DNA helicase RecQL4, a genome maintenance factor required for DNA replication initiation, stabilization of stalled replication forks, and multiple DNA repair mechanisms including homologous recombination and base excision repair. RecQL4 functions within the broader DNA damage response network to preserve chromosomal integrity during S phase and after genotoxic stress. Dysregulated RECQL4 activity is linked to replication stress, chromosomal instability, and altered sensitivity to DNA-damaging agents, making it relevant for studying genome stability pathways. Pathogenic variants in RECQL4 are associated with inherited disorders featuring developmental defects and cancer predisposition, supporting its use in mechanistic models of disease-associated DNA repair dysfunction.

RecQL4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RECQL4 expression without altering the underlying DNA sequence.

RecQL4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RECQL4 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 RECQL4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RecQL4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RECQL4 locus and enabling the study of RecQL4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RecQL4 pathway restoration in tumor cells with silenced or reduced RECQL4 expression.

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